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cardio pulmonary resuscitation (cpr) is an emergency procedure performed on people under cardiac arrest and on people who stop breathing due to reasons such as drowning. cpr 's main benefit is that it maintains blood flow, which prevents tissue and brain damage. the procedure involves creating artificial blood circulation by applying rhythmic pressure to a person 's chest. references to resuscitation attempts can be found in ancient texts that date back thousands of years, but the first known attempts at resuscitation in modern times occurred in the 18th century. practitioners, then, used various techniques to resuscitate a person who was unconscious or not breathing. these included blowing air into the mouth, massaging the chest, tickling the throat, or applying manual pressure to the abdomen. these methods were most effective when used for drowning victims. over the years, practitioners refined the techniques, and until 1950s, the accepted resuscitation method was applying back pressure and arm lifting. he along with dr. peter safar demonstrated the superiority of their cpr method to earlier methods. their method used chest compressions in combination with periodic mouth - to - mouth breathing. latest guidelines from the american heart association have modified the elam and safar cpr approach ; the aha recommends using continuous chest compression (ccr) because this approach works better than periodically stopping compressions for mouth - to - mouth breathing,. the first organized attempt to make citizens a part of the emergency procedure in cases of cardiac arrest was made in seattle in march of 1970,. fire department personnel were trained in cpr so that they could perform it on the victim before paramedics arrived to attempt defibrillation. the data gathered from this exercise proved that when cpr was started within 23 minutes of the event, survival chances increased. in 1972, community - based cpr training of general public has expanded across the united states. in 1981, effective cpr consists of the following procedure, : lay the person flat on his back.place your hand flat on the person 's upper chest between the nipples. for infants only two fingers are used - the middle finger and the index finger. for adults only, place your second hand above the first hand (for children only one hand is used).start applying pressure to compress the chest.the recommended rate of chest compressions is about 100 per minute. for infants only two fingers are used - the middle finger and the index finger. for adults only, place your second hand above the first hand (for children only one hand is used). the depth of chest compression is about 2 - 2.5 inches for adults, about 1 - 1.5 inches for children, and about 1/3 inch for infants. original aha guidelines emphasize a - b - c as a cpr guideline. in the acronym, a stands for airways, meaning that the person giving cpr needs to make sure that the airways are open ; b stands for breathing, meaning that the person giving cpr does mouth - to - mouth breathing ; and c stands for chest compressions. in traditional methods, periodic mouth - to - mouth breathing is also done to replenish oxygen supply, but newer guidelines suggest that continued compression is more important. consequently, mouth - to - mouth breathing has now become the third, optional, portion of cpr,. the primary reason for the change is that most bystanders or paramedics hesitate to use mouth - to - mouth breathing with unknown people because mouth - to - mouth breathing may cause spread of infectious diseases. apart from concerns over infections, there has also been discussion on how often to give mouth to mouth breathing. normally there is enough oxygen in the blood stream to only do continuous chest compressions. since an oximeter may not be available at that moment, there is no way to determine the oxygen level ; therefore, it is difficult to determine whether mouth - to - mouth breathing is required. making mouth - to - mouth breathing experts are also debating the need to give mouth - to - mouth breathing in cases where the blood oxygen saturation level falls. a person 's blood oxygen saturation level (bos) indicates how efficiently a body 's blood cells retain oxygen. cardiopulmonary resuscitation is performed to force the movement of oxygenated blood through the circulatory system and prevent the damage of vital organs in the body. the level is measured by analyzing the ratio between the amount of oxygenated hemoglobin and the total amount of hemoglobin present. bos level ranges can help to determine a person 's risk of lung disease and tissue death. this paper, however, focuses on the bos level 's ability to determine the cardiopulmonary resuscitation 's efficiency. with the knowledge of a victim 's blood oxygen saturation level, the decision to give mouth - to - mouth breathing may be necessary to keep oxygenation at a healthy level. over the years, these devices improve the quality of cpr by providing feedback on proper placement of hands on chest and the correct frequency and depth of compressions. mechanical devices which give accurate frequency and depth of chest compression provide automatic cpr. studies have shown that these automated cpr devices improve the survivability of patients who need out - of - hospital cpr. during an emergency it is likely that a person trained in cpr may not be available. in such situations 9 - 1 - 1 operators help the caller to administer cpr by giving instructions over a phone. in such instances, a readily available technology would be useful in ensuring that people untrained could deliver cpr properly. if the application is not available, a 9 - 1 - 1 operator can help in downloading that application. however, having to download the application and then watch the video seriously reduces the window of survivability for the injured person. alternatively, there exist devices that give real - time feedback on the quality of cpr. during an emergency situation, it is highly likely that persons trained in cpr are unavailable. even though devices can provide automatic cpr, these devices are highly unlikely to be accessible at the time of need. in such cases 9 - 1 - 1 operators provide cpr instructions over a phone. however, the success of such an approach depends upon the emotional and physical capabilities of the person actually administering the cpr. with newer technology, the operator may even attempt to send video instructions, which may assist in improving the cpr given. however, again, the 9 - 1 - 1 operator may not have all the information necessary to determine whether the cpr is being done efficiently and helping the injured person. currently, 9 - 1 - 1 operators can not evaluate the results of cpr remotely and, in fact, there is no proven method to evaluate cpr effectiveness even when a trained person is giving the cpr. a recent report from american heart association strongly recommends that methods and technology should be developed to evaluate the quality metrics of cpr. some of the highlights of their recommendations include the use of at least 1 modality of monitoring cpr performance, i.e. the compression depth and the frequency ; use at least one modality to monitor patient 's physiological response to resuscitative efforts ; and continually adjust resuscitative efforts based on the patient 's physiological response. in this paper, we present a method to evaluate cpr performance in real - time without the need of special devices. we also show that many of the recommendations can be met by using the methods we propose. using the sensors in a smartphone, such as an accelerometer, this paper describes an application that can evaluate and guide a person in giving effective cpr while providing timely feedback to the 9 - 1 - 1 operator. the smartphone 's sensors provide feedback about chest compression frequency and depth, and about blood oxygen saturation levels. the smartphone 's sensors provide feedback about chest compression frequency and depth, and about blood oxygen saturation levels. a smartphone controller (box 1) holds an algorithm to evaluate data from sensors (box 3). the system (box 2) consists of the affected person and the person giving the cpr. as mentioned in prior sections, continuous chest compressions have been emphasized over cardiopulmonary resuscitation as the most critical cpr procedure to perform in an emergency. that said, mouth - to - mouth breathing remains a viable option in certain cases, especially when trained personnel are present. we present a smartphone application which measurers the blood 's oxygen saturation without specialized equipment. in conjunction, one smartphone can be used by the person giving the cpr where it measures the frequency and depth of compressions. the data from these smartphones is continuously reported to the 9 - 1 - 1 operator who can use the information to guide the cpr giver. a patent has been filed by one of the authors for devices that can measure the vital signs using sensors of the smartphones. figure 2.cpr and its evaluation. the compression frequency, depth and the oxygen saturation levels are reported to the 9 - 1 - 1 operator. the person giving cpr has a phone tied to her hands, as shown in the picture below the arrow. a smartphone is also placed near the hand of the person receiving the cpr with finger on the camera lens as shown in the second picture below the arrow. the compression frequency, depth and the oxygen saturation levels are reported to the 9 - 1 - 1 operator. the person giving cpr has a phone tied to her hands, as shown in the picture below the arrow. a smartphone is also placed near the hand of the person receiving the cpr with finger on the camera lens as shown in the second picture below the arrow. it involves integrating acceleration readings to compute velocity and further integration of the calculated velocity to find the displacement or distance of movement. several sources of error may arise in this process : errors inherent in the accelerometer or caused by noise from the electronic signal.external errors errors caused by force applied to the accelerometer such as lateral movements of the hands during cpr or movement in a vehicle when a patient is being transported while receiving cpr).error due to drift : these errors are introduced during the compression of the chest. for example, the chest may not fully recover to its normal position before the next compression is started. errors inherent in the accelerometer or caused by noise from the electronic signal. external errors errors caused by force applied to the accelerometer such as lateral movements of the hands during cpr or movement in a vehicle error due to drift : these errors are introduced during the compression of the chest. for example, the chest may not fully recover to its normal position before the next compression is started. unfortunately the process of double - integration on these readings compounds these accelerometer reading errors even a small error can produce large variation in calculated displacement. measuring blood oxygenation levels using a smartphone is even trickier because the method has to be non - invasive, should not require additional devices, and should be simple and quick for anyone to use, even if not a health professional. this section discusses the methods to measure the frequency and depth of chest compressions and the oxygen saturation levels using smartphones. these three figures can guide persons administering cpr, even when they lack cpr training. frequency of compressions : an accelerometer measures acceleration of movement in the x, y, and z axes. when people lie on their backs, chest compressions are in the direction of the z axis. so, each upward movement is considered to be negative acceleration ; each downward movement is considered to be positive acceleration. a complete up and down movement counts as one compression. we can, therefore, calculate the frequency of compressions as up - down movements. their straight - forward approach is to : 1)calculate velocity from acceleration as follows : given acceleration (\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(a)\) \end{document } and a period of time (\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(t)\) \end{document }, it is possible to calculate the change in velocity during the relevant time period. if the original velocity is available, the change in velocity at the end of the time period can be calculated using the equation:\documentclass[12pt]{minimal } 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is:\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \begin{equation } d = \delta d + d_{0}.\end{equation } \end{document } calculate velocity from acceleration as follows : given acceleration (\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(a)\) \end{document } and a period of time (\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(t)\) \end{document }, it is possible to calculate the change in velocity during the relevant time period. if the original velocity is available, the change in velocity at the end of the time period can be calculated using the equation:\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \begin{equation } \delta \textit{v } = at;\end{equation } \end{document } where \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(\delta \) \end{document}v is the change in velocity during time \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(t\) \end{document}. if the velocity at the start of the time 0 is \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(v_{0}\) \end{document }, then velocity at time t is:\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \begin{equation } \textit{v}= \textit{v}_{0}+ \delta \textit{v}\end{equation } \end{document } calculate the distance as follows:\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \begin{equation } \delta d = \frac { (\textit{v}_{0}+\textit{v})}{2 } \ast t.\end{equation } \end{document } where \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(\delta d\) \end{document } is the change in distance, \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(v_{0}\) \end{document } is the velocity at time \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(0,\) \end{document } and \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(v\) \end{document } is the velocity at the time \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(t\) \end{document}. if \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(d_{0}\) \end{document } is the distance at time 0, the distance at the time \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(t\) \end{document } is:\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \begin{equation } d = \delta d + d_{0}.\end{equation } \end{document } unfortunately, these calculations assume a straight - line motion, which is not the case for cpr. this means we need to find velocity and displacement while the accelerometer readings are still being logged, and so we need to use numerical methods that allow for integration on data readings as they are logged. one method, trapezoidal rule, offers an approximation technique for calculating the integration. the existing literature on calculating displacement from accelerometer readings is based on devices that are dedicated to cpr compressions. may not be easily accessible, but a smartphone with an accelerometer is more likely to be available. our project assumed that an untrained person administers the cpr and does not have access to such dedicated devices. for our experiments, we collected data by placing the smartphone in the middle of the chest. blood oxygen saturation level : we determined blood oxygen saturation levels using the camera lens of the smartphone. pulse oximeters measure the visible and infra - red spectrum of the oxy - hemoglobin and de - oxy hemoglobin, respectively. a pulse oximeter works by exploiting particular properties of light. when light passes through a substance (such as blood), the substance absorbs a certain amount of light. the amount of absorbed light depends on the sample 's concentration, the sample 's absorbance capacity, and the light 's path length. we calculated the bos level using beer - lambert 's law:\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \begin{equation } a = abc\end{equation } \end{document } where, \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(a\) \end{document } is absorption ; \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(a\) \end{document } is molar absorptivity of the sample, \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(c\) \end{document } is concentration of the sample, and \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(b\) \end{document } is path length. oxy - hemoglobin (hbo2) and deoxy - hemoglobin (hb) absorb light at different wavelengths. oxy - hemoglobin (hbo2) absorbs more infra - red light than deoxy - hemoglobin (hb) ; hb absorbs more red light. oxygen saturation is calculated using the following relation:\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \begin{equation } \mathrm { spo2 = f(\varphi) ; \;\;\ ; { \rm where}\;\;\ ; \varnothing = } \left (\frac { ac_{red}/dc_{red}}{ac_{ir}/dc_{ir}}\right) \!.\end{equation } \end{document } in this section we discuss issues we encountered during data collection and the steps we took to resolve them. one of the first problems resolved concerned placement of the smartphone on the person receiving cpr. such a placement can be extremely uncomfortable and can cause injury if the smartphone is pressed against the chest during cpr. placing the smartphone between the palms of the person giving cpr the screen may crack under pressure. additionally, this placement is uncomfortable for the person giving the cpr. the ideal solution was to place the phone above the hands of the person giving the cpr (see figure 2). in these experiments, we tied the smartphone to the hands so that it did not fall off. this can be done by any piece of cloth such as a shirt or undershirt. there are two kinds of manikins used for training - a soft - chest (sponge) manikin and a hard - chest manikin. figure 3 shows the accelerometer plot for a student performing cpr on a soft - chest manikin ; figure 4 shows a similar plot for an instructor performing cpr on a soft - chest manikin. it may be observed that the best results were from the instructor doing the cpr on a hard - surface manikin. figure 4.accelerometer readings for an instructor doing cpr using a sponge manikin. the compression frequency and depth are not uniform. when the accelerometer moves towards gravity, the acceleration is considered positive and when it moves against the gravity, it is considered as the negative direction. we can count the number of times the accelerometer readings show a change in the sign of magnitude from positive to negative number. however, calculating depth of compression is subject to errors from several sources, and so our algorithm had to be fine - tuned to reduce the influence of such errors on our results. in this section we document efforts to improve the accuracy of calculation of depth of cpr compressions. when we used a simple method of double integration to calculate the compression depth (see figure 6), the depth of compression varied up to 40 cm. the integration function uses the magnitude of acceleration and velocity over the time period between two successive readings. so, if the time between two successive readings was large, the algorithm calculated too large a displacement value. the first change used the fact that cpr involves restricted and repeated movements in a vertical direction. we found even a minor error in the acceleration magnitude can result in a large error in displacement measurements. we modified our algorithm to calculate a rolling average after a number of accelerometer readings were logged. figure 7 shows a plot of the acceleration magnitude and the corresponding velocity and the calculated distance. this solution localized errors of magnitude to within one compression, which provided further correction to errors that build over time. dots indicate points where velocity is reset to zero. the y - axis is m / s\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(^{2}\) \end{document } for acceleration, dm / s for velocity and cm for distance. the y - axis is m / s\documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(^{2}\) \end{document } for acceleration, dm / s for velocity and cm for distance. in this section the experimental setup consists of the following steps : write an android application to calculate the cpr depth from the smartphone 's accelerometer.use a commercially - available manikin designed for cpr training to collect data. the force required to press the manikin 's chest must match the actual force required to press a human 's chest during a real cpr.use a mobotix camera to record cpr compressions. the professional security mobotix camera allows us to study video frame by frame and determine actual depth of compression.compare the calculated cpr depth for each compression with the actual movement observed on video. write an android application to calculate the cpr depth from the smartphone 's accelerometer. use a commercially - available manikin designed for cpr training to collect data. the force required to press the manikin 's chest must match the actual force required to press a human 's chest during a real cpr. the professional security mobotix camera allows us to study video frame by frame and determine actual depth of compression. compare the calculated cpr depth for each compression with the actual movement observed on video. we overlaid a scale on the image to allow us to calibrate the actual movement during cpr as compared to the observed movement on the video frame. after recording 40 seconds of cpr, the video file was played frame by frame and the actual depth of cpr was measured. our algorithm then compared the two to determine the difference between the actual depth of compression (as observed on the video frame) and the depth calculated by our application. lines of scale are superimposed to calibrate movement as the manikin chest is compressed. in this section the application prompts the cpr giver to increase or decrease the depth of chest compression to meet the 2 - 2.5 depth requirement. similarly, our application gives a prompt when the frequency of compressions is not within a range of 90100. the number of compressions volunteers performed during these 30 seconds ranged from 30 to 50. it shows the accuracy of the depth calculation done by our application as compared to the actual depth as observed in the mobotix video. our application 's accuracy ranged from a low of 57% to a high of 98%. the range is from a low of 57% to a high of 98%. in the previous section we discussed the accuracy of our application to determine the depth of compression. the focus was on how accurately the application can calculate the depth as compared to the actual depth. in this section as has been noted earlier, the person administering the cpr must accurately judge whether the chest has been compressed to the recommended depth before releasing the chest to return to normal. our application gives an alert when the depth of the compression falls outside an acceptable standard range. one of the factors we considered was the accuracy of our application 's calculation. if the accuracy of a particular calculated depth of compression is low, then the decision to provide an alert by the application may be inaccurate. figure 10 shows a bar plot comparing the actual compression depth and the calculated compression depth for the same volunteer as in figure 9. it may be observed that the calculated depth value reaches near 2 inch value for many compressions, but the actual depth never reaches the 2 inch value. so, for these compressions, the algorithm will not give an alert, even though it should have given one. some of the calculated errors have a positive magnitude and some have a negative magnitude. table-1 shows the accuracy of average over different time durations ranging from 1 second to 10 seconds. figure 10.comparison of calculated compression depth and the actual compressions depth for an individual. the rows show accuracy for alerts given for each compression, and for each second between 1 and 10 seconds. total alerts shows the total number of alerts that occur during the specified time period. accuracy range for alerts% shows, in 5% increments, the number alerts with compression accuracy. for example, when an alert is given every second, 6 alerts of the 20 alerts have a compression accuracy of less than 85%. however, when an alert is given every 5 seconds, all the alerts have compression accuracy greater than 85%. the rows show accuracy for alerts given for each compression, and for each second between 1 and 10 seconds. total alerts shows the total number of alerts that occur during the specified time period. accuracy range for alerts% shows, in 5% increments, the number alerts with compression accuracy. for example, when an alert is given every second, 6 alerts of the 20 alerts have a compression accuracy of less than 85%. however, when an alert is given every 5 seconds, all the alerts have compression accuracy greater than 85%. a person requires a few seconds to understand and respond to an alert. by the time the person reacts this leads to confusion and the person may not be able to adjust their compressions accurately and in a timely manner. we did an analysis to decide how frequently the alert should be given (table 1). when our application gave an alert for each compression, out of a total of 38 alerts (for 38 compressions in that session), 21 alerts were less than 90% accurate. this means that for 21 alerts the accuracy of calculated depth as compared to the actual depth was less than 90%. the total of 38 alerts assumes that we give an alert for each compression (assuming no compression is going to be 100% accurate). when our application gave an alert every second, we averaged the calculated depth of all compressions within each second. as table-1 indicates, the total number of alerts for one second will be 20. when our application gave an alert every 6 seconds, then 4 alerts were more than 90% accurate. we continued this analysis through 10 seconds. at 10 seconds, our application gave two alerts, each 100% accurate. we conclude that the accuracy of alerts increases when an alert is given every few seconds rather than for every compression. the accuracy improves because the errors with negative magnitude adjust the errors with positive magnitude with in the time period. so the overall accuracy improves. within the \documentclass[12pt]{minimal } \usepackage{amsmath } \usepackage{wasysym } \usepackage{amsfonts } \usepackage{amssymb } \usepackage{amsbsy } \usepackage{upgreek } \usepackage{mathrsfs } \setlength{\oddsidemargin}{-69pt } \begin{document } } { } \(6 -7\) \end{document } second range, our application 's accuracy is reasonable at more than 90%. but, our experiments suggest an alert every 67 seconds does not provide persons giving cpr enough time to adjust their cpr compression depth. our experiments also suggest that an optimum time for giving alerts is every 10 seconds. as explained, alerts provide feedback to a person giving cpr so that person can adjust the compression depth or frequency to fall within a prescribed range. the application issued alerts when the compression depth fell below 1.5 (low alert) or rose above 2.5 (high alert). the application provided a low alert at 10 seconds and then, again, at 20 seconds. at 30 seconds the compression depth then increased to more than 2 for a 30-second period, so no alerts were issued. at 80 seconds, we concluded that our application achieved its purpose of providing alerts to the cpr giver, which enabled the cpr giver to adjust to more effectively administer cpr. the results show that the accuracy ranges from a low of 77% to a high of 99%. one may have to give cpr as the patient is being transported in a moving vehicle to the hospital. several factors come into play in a moving vehicle that increases the difficulty of calculating the compression depth accurately when using a smartphone accelerometer. if shock absorbers are inadequate, an accelerometer records vehicle movements in the z axis, skewing the z axis motion of the chest compressions. road condition presents another major factor that contributes to increased errors. bumps in the road, lane changes, and traffic turns also affect readings. traffic patterns also add to the randomness of readings. the vehicle may have to be slow at times and then accelerate as the traffic moves. it may have to stop at traffic lights and then accelerate. even if we keep factors constant, such as using the same vehicle, driving on the same road and even driving at the same speed, randomness of a traffic pattern still produces different results each time the cpr is attempted. the road had almost no traffic, required no turns for a few miles, and had no traffic lights. we, then, drove at a constant speed of 30 mph to minimize movements due to vehicle motion. table 3results of cpr in a moving vehicle, showing the depth of compressions in inches. results of cpr in a moving vehicle, showing the depth of compressions in inches. should the injured person 's blood oxygen saturation level drop precipitously, the person may suffer physical deterioration and even death. to reduce this risk, our algorithm must monitor the bos levels while it is measuring compression frequency and depth. in this section we describe a procedure that uses smartphones to measure the bos level using principles of optics. while a person gives cpr, this information can assist in deciding whether to give breathing. however, many prefer to avoid using the technique unless absolutely necessary because of possible exposure to infectious diseases. our algorithm needed to provide a method for the 9 - 1 - 1 dispatcher or the person giving the cpr to determine whether mouth - to - mouth breathing was really necessary. to resolve this issue the surface of the injured person 's fingertip (the area of analysis) is placed on the smartphone 's camera lens. by taking a video, a beam of near - infrared light is passed through the finger. as the light passes through, the smartphone creates video of the area of analysis. the video is then analyzed to determine the rgb values (red green blue). rgb values of the refracted light in the blood are then analyzed for the scattering effect of near - infrared light. the two phones can simultaneously send information to the 9 - 1 - 1 operator. after measuring the oxygen saturation level under normal circumstance, we wanted to confirm that our system can detect reduced levels of oxygen in the blood. we used occlusion spectroscopy, the method of using over - systolic pressure to temporarily stop the flow of blood to the finger to collect data, (figure 12). in our study this allowed us to measure blood flow in the upper layers of the finger.. later when occlusion is removed, the oxygen saturation level reaches the normal value and our system can measure this return to the normal value. the occlusion experiment confirmed that we can indeed measure the depletion of oxygen saturation in the blood using the optical features of a smartphone. in fact, occlusion allowed us to obtain a set of data points instead of one data point. the blood flow to the finger is decreased by applying pressure to it. to establish standardization, we also required a method of dividing the intensity of the blood pulse (ac) by the intensity of the blood color (ad). we applied this to data to analyze both the intensity of green and of red values. the green ac / dc value was then divided by the red ac / dc value. this value corresponds to the percentage of blood oxygen saturation. figure 13 shows the oxygen saturation level as it drops to 50% after occlusion. the graph also shows that the oxygen saturation level recovers back to 100% after occlusion is removed. about 50% occurs. then the saturation level returns to 100% after occlusion is removed. a drop in oxygen saturation level to about 50% occurs. then the saturation level returns to 100% after occlusion is removed., we found that the rate of the decay of oxygen varies with each subject. while some subjects experienced a quick decrease of blood oxygen, other subjects experienced a slow decay. the amount of oxygen within the blood in the finger dropped significantly for the first few seconds and then finally stabilized for each subject. the amount of oxygen within the blood in the finger dropped significantly for the first few seconds and then finally stabilized for each subject.. the application can then provide feedback to the person administering cpr to improve its effectiveness. existing applications available on smartphones simply furnish a short video tutorial on how to perform cpr. our smartphone application prompts the cpr giver in real time on when and how to adjust their frequency and depth of chest compressions to meet cpr guidelines. our experiments ' results show that our smartphone application can be used to effectively administer cpr, even by people who have not been trained to give cpr. in emergency situations, where a trained person may not be easily available and timing is crucial, these devices can mean the difference between life and death. additionally, the devices ' sensors can also help by continuing to provide vital information to paramedics as they rush a patient to a hospital. by measuring oxygen decay using the smartphone camera as occlusion is induced upon the finger, our application allows accurate determination of the blood oxygen saturation level. by using the ubiquitous smartphone, people performing cardiopulmonary resuscitation can also determine when the frequency and depth of their compressions enhance blood flow. for example, the oxygen saturation level may offer a better indicator of cpr effectiveness than the depth or frequency of compressions. the method we have proposed and developed helps in meeting the latest recommendations of the america heart association. our method helps in monitoring the cpr, it provides real time feedback to the person giving cpr and insures that the quality of cpr meets the guidelines. for future work, additional experiments may improve the cpr depth calculations in a moving vehicle, in actual traffic and in uncontrolled conditions. also future studies may help determine a more accurate time between alerts rather than the 10 seconds we used. | timely cardio pulmonary resuscitation (cpr) can mean the difference between life and death. a trained person may not be available at emergency sites to give cpr. normally, a 9 - 1 - 1 operator gives verbal instructions over the phone to a person giving cpr. in this paper, we discuss the use of smartphones to assist in administering cpr more efficiently and accurately. the two important cpr parameters are the frequency and depth of compressions. in this paper, we used smartphones to calculate these factors and to give real - time guidance to improve cpr. in addition, we used an application to measure oxygen saturation in blood. if blood oxygen saturation falls below an acceptable threshold, the person giving cpr can be asked to do mouth - to - mouth breathing. the 9 - 1 - 1 operator receives this information real time and can further guide the person giving cpr. our experiments show accuracy > 90% for compression frequency, depth, and oxygen saturation. |
there are 20 class i genes and in transplantation hla - a, b, and c are the classic genes referenced to when typing the recipient. class i major histocompatibility complex (mhc) molecule processing and loading of peptides occurs in all nucleated cells, where class ii mhc molecule involves primarily b cells, macrophages, and dendritic cells by method of endocytosis and phagocytosis. class ii mhc molecule nomenclature is designated by class (d), family (m, o, p, q, r), and chain (a or b). late antibody - mediated rejection (amr) is a major cause of late kidney transplant failure and several developments in understanding of pathogenesis allow for improvement of diagnosis, treatment, and prevention. this review will concentrate on reviewing the pathogenesis, diagnosis, and treatment of amr. not only can antibodies form against hla molecules, but to endothelial - cell antigens and across abo blood group. blood transfusions can induce humoral immunity by formation of hla alloantibodies and are more likely to occur in individuals who have been previously pregnant. our understanding of how blood transfusions cause sensitization is incomplete, but it is not inevitable and can be attenuated by immunosuppression. in a study performed by eikmans. it was found two weeks after blood transfusion in both nonsensitized and sensitized recipients, increased numbers of ifn--producing cells were produced stimulating cd4 + and nk cells. with de novo hla production, endothelium of donor graft peritubular and glomerular capillaries display mhc molecules which are the target for antibody production. once the endothelium is damaged by antibody, von willebrand factor and p - selectin are released as an inflammatory response. leukocytes adhere to glomeruli or to dilated peritubular capillaries via cytokines (il-1, il-8, and chemokine ligand 2) allowing complement activation. in antibody rejection, c4d is a useful marker of complement activation and is seen commonly in peritubular capillaries with proper immunohistochemical staining technique. chemoattractants c3a and c5a are a part of the compliment cascade which activate c5b allowing for assembly of membrane attack complex (mac) which is composed of c5b - c9. the mac complex once activated will cause local necrosis allowing detachment of endothelial cells from the basement membrane, which is also a characteristic finding upon biopsy indicative of amr. if not treated properly in a timely manner, sequelae can include thrombotic microangiopathy (tma) causing hemorrhage with arterial wall necrosis, and ultimately graft loss. also potentially playing a role in amr pathogenesis are polymorphic mhc - class - i - related chain a (mica) antigens. it has been found that not all patients with amr have anti - hla antibodies, which suggests other possible mechanisms involved in acute or chronic graft damage. in a study conducted by worthington., only five out of 14 patients with renal graft dysfunction and pathology showing c4d deposition in peritubular capillaries were found to have anti - hla antibodies. mica antigens are structurally similar to mhc class i proteins, closely linked to hla - b and c loci. mica antigens can be found on fibroblasts, endothelial cells, dendritic cells, and many tumors which unfortunately makes detection difficult by standard donor and recipient crossmatching. mica eplet repertoire has become the basis for developing hlamatchmaker program which analyzes antibody patterns with single mica alleles on luminex platform. it has been determined so far (duquesnoy.) that there are 38 potentially immunogenic mica eplets associated with mica - reactive serum. mica and hla class i chains are similar in configuration however none of the polymorphic residues are shared. hlamatchmaker program is therefore used to determine the specificity of anti - mica antibodies in sensitized patients since mica is considered as a separate alloantigenic system. in a study by zhang., 52 patients transplanted who were crossmatch negative were placed on triple therapy (cyclosporine / tacrolimus, mycophenolate, and prednisone). once graft dysfunction was diagnosed with positive c4d, both anti - hla and anti - mica antibodies were collected from serum and reviewed via luminex. of the 15 patients found to have anti - mica antibodies, 9 patients were positive for class i and/or class ii anti - hla antibodies. nine out of 15 patients were found to have c4d deposition, and after reviewing patients for the follow - up period (1 year), it was found that egfr decreased in anti - mica and anti - hla antibody positive group. this study suggested that patients with mica antibodies have a more rapid deterioration of graft function versus patients with just anti - hla antibodies. there are several proposed mechanisms for anti - mica antibody activation involving complement or natural killer cells and studies are continuing to determine the pathogenesis. a more thorough understanding of hla typing as well as continuing studies to determine anti - mica and antibody rejection pathogenesis may allow for novel treatment strategies. amr occurs in up to 33% of cases undergoing abo incompatible kidney transplantation. among cases in which amr is triggered in the early post - op period, this is opposed to the case in which gloor found patients who received desensitization to hla antibodies showed more glomerular changes versus abo incompatible transplant grafts not having clinical amr, suggesting subclinical amr may exist. the resistance of the graft to amr in spite of the presence of antibodies against donor endothelium is called accommodation. this is explained either by difference in antigen expression, change in repertoire / level of antibodies in recipient, or decreased susceptibility of injury to endothelium secondary to continual stimulation of antibody and complement components. it is proposed that continued stimulation of endothelial cells by endotoxin or il-1 over time diminishes cellular responses to restimulation. another proposed mechanism explaining accommodation could be a decrease in the recipient 's type blood - type antigen on the endothelium of the graft. koestner. reported histo - blood type change of a blood type b graft after abo - mismatched heart transplantation, where the recipient 's blood - type o antigen was replaced by the donor 's type b antigen after amr graft loss. the mechanism of replacement of the recipient 's antigens remains unclear, and several studies have been attempted to explain occurrence of blood - type chimerism. one explanation is replacement of the recipient type antigens by stem cells derived from injured vessel walls after graft rejection. it is speculated that during amr some blood - type antigens could be shaved off from endothelial surface from immunological reaction, resulting in loss of blood - type antigens on the transplanted mismatched organ. in addition the presence of diffuse c4d without capillary damage or inflammation infers that accommodation may include resistance to the terminal complement cascade. there are some noted differences between hla - sensitized and abo - incompatible renal transplantation in terms of pattern of cell injury after c4d deposition. it has been found that there is more graft loss with positive c4d amr in hla - sensitized patients which may be due to differences in biological responses between non - self - carbohydrate antigens (blood type antigens) and non - self - peptide antigens (hla antigens). crossmatching not only improves long - term graft survival by identifying the presence of antidonor antibody in the recipient, but is used as a predictor of amr. the donor cells are incubated with the recipient serum, and antihuman globulin which is fluorescent dye tagged is placed in the flow cytometer. hla - specific antibody detects antibodies binding to individual hla antigens and allowing determination of antigens to avoid in a donor, coined unacceptable antigens thus avoiding rejection. of patients who are highly sensitized hla - specific antibody detection prior to transplantation and surveillance after transplantation during a rejection episode allows for avoidance of rejection and determination of course of treatment for amr, respectively. the 1997 banff classification was used to classify rejection prior to the meeting in 2001, which further defined pathological classification of amr. gorer determined the role of antibody after much debate in the early days of transplantation, and unfortunately after his death in 1961 the concept of amr was lost. in 1991 - 1992, feucht and halloran described c4d as a marker for amr however this was not uniformly utilized until the report from solez. in 19982000. after international transplant meetings in 1997, amr was an entity recognized by most of the participants and c4d staining raised hope that morphological classification of amr can be further defined. incidence of amr has been reported as 08% in renal transplant recipients in larger centers largely due to increased recognition, detection of dsa, retransplanted patients, as well as increase in positive crossmatch and abo incompatible transplantation for highly sensitized patients. colvin had reviewed studies from massachusetts general hospital which for the first time indicated a clear correlation of peritubular capillary c4d staining pathological features with dsa in amr. a few studies have indicated c4d staining is around 9396% specific, but 3195% sensitive. this raises a concern that additional evidence is needed to diagnose amr such as quantification of dsa or other morphologic feature consistent with amr on pathology. however it should be noted that regele and colleagues presented data from 1 year after amr that kidney transplant patients with positive c4d had higher incidence of graft loss and elevation of serum creatinine. after the banff meeting in 2001 acute tubular injury ; neutrophils and/or mononuclear cells in peritubular capillaries and/or glomeruli, and/or capillary thrombosis ; or arteritis / fibrinoid necrosis in the intima along with intramural / transmural arterial inflammation. c4d evidence for antibody action and/or immunoglobulin in peritubular capillaries, immunoglobulin and complement in arterial fibrinoid necrosis. anti - hla antibody (dsa) circulation in serum or other antidonor endothelial antigens. after the banff meeting in 2009, positive c4d deposition without evidence of rejection was added to criteria suggesting antibody - mediated changes. several studies had reported this finding including mark haas (los angeles) who had seen c4d deposition in biopsies without evidence of rejection. in protocol biopsies from abo incompatible grafts, 21/37 had c4d deposition without evidence of amr lesions or t - cell - mediated rejection which can suggest accommodation. another addition to banff criteria indicating antibody - mediated changes was determined to be positive c4d with presence of circulating antidonor antibodies (no signs of acute or chronic t - cell - mediated rejection or amr / no atn - like minimal inflammation). in normal kidneys, specifically in the glomerular mesangium as well as arterioles at the vascular pole, turnover of immune complexes occurs (c4d). transplanted kidneys in addition show deposition of c4d in peritubular capillaries and can follow a dynamic course. c4d can be in a transitional state and show staining of only a few capillaries indicating either an ongoing humoral attack or resolving previous attack. there is sometimes diffuse staining in every capillary or focal staining in only a few capillaries, but c4d is always attached to endothelial cells and basement membranes. there has also been evidence for c4d negative amr in a couple studies, which in the past few years has brought to the forefront the limitations of c4d as a marker for amr. in one such study by sis., overexpression of 12-individual endothelial - associated transcripts genes (endats) upon transplant biopsies with graft dysfunction was correlated with an increased risk of graft loss. high endat expression and endothelial activation with dsa was found to be strongly associated with the presence of transplant glomerulopathy. interestingly, fewer than 50% of the biopsies performed (n = 173) with dsa and endats had diffuse c4d staining in peritubular capillaries. investigators also have found that c4d negative form of amr is less severe than c4d positive amr but is associated with chronic changes within the graft, such as transplant glomerulopathy. it has been postulated that dsa binding to endothelial cells triggers natural - killer cells to release interferon- resulting in granule - associated toxicity. dsa has been found to play a role in diagnosis of amr and can be an independent predictor of allograft loss. it was found when patients had dsa at time of clinical rejection diagnosed by luminex, 50% reduction in dsa within 14 days of diagnosis had higher allograft survival. in a study performed by everly., acute rejection was defined as an increase in serum creatinine by 20% above the baseline upon rejection ; dsa was identified by antigen bead panels by luminex assay. 650 patients were analyzed for acute rejection, 94 of which were identified with biopsy - proven acute rejection by banff criteria. analysis of predictors of allograft loss revealed dsa to be an independent predictor of allograft loss, with a sixfold increase in allograft loss rates. however, it is noted in this study that dsa was present in the absence of c4d staining on biopsy. further studies need to be conducted to determine if complete elimination of dsa is necessary. however, from this study it should be deduced that dsa reduction should receive consideration as a potential therapeutic goal for rejection therapy. suppression of dsa with antihumoral therapies may provide a means for improving long - term renal allograft survival. bortezomib, a new anti - b - cell agent that targets antibody producing cells (plasma cells) has shown promise as an effective means for reducing dsa as well. it has also been postulated that surveillance of dsa can be a means of preventing rejection, especially for highly sensitized patients. it has been discussed that high - risk patients (presensitized and retransplants) may benefit from more dsa surveillance in the first 3 months after transplant to detect an early immune response ; however this is not rigorously proven to improve outcomes. plasmapheresis (pp) removes alloantibodies from the circulation and is the fastest and most effective method for elimination of dsa. usually 1.01.5 full - volume exchanges are performed using albumin solution daily or alternate days, continued until serum creatinine decreases 30% from the baseline value. although pp is effective in removing alloantibodies (dsa) from the circulation, rebound synthesis of de novo alloantibodies can occur. ivig is used in combination with pp to neutralize antibodies and potentially block complement activity as well as agents used for suppression of antibody (cacineurin inhibitors, mycophenolate, or rituximab). immunoadsorption (ia) is a substance derived from the cowan strain of staphylococcus aureus. the protein a coating the column beads has a strong affinity for the fc portion of immunoglobulin and immune complexes. for the past decade, protein a ia was found to remove harmful antibodies and immune complexes efficiently and has been used as treatment of amr of highly sensitized patients. it was found in a study by qing. that classes i and ii panel - reactive antibody (pra) as well as immunoglobulin and complement (c3 and c4) were reduced significantly. however, it was found that within 6 to 8 hours rebound of antibodies was seen. it has been postulated that ia may cause a conformation change of the structure of the antibodies and immune complexes making them more immunogenic, and immunostimulating to regulatory anti - idiotype antibodies. these antibodies play a role in the control of autoimmune mechanisms and reverse the continuing cycle of abnormal immune balance. the long lasting response of ia therapy may be explained by phagocytosis of the reticuloendothelial system and activation of complement. ivig is prepared by human plasma from approximately 50,000100,000 of healthy donors, composed of 90% intact igg, a few dimers, fabs (fragment antigen - binding) and traces of igm and iga. the fc region in igg allows it to interact and signal through fc - gamma receptors on phagocytes, b cells, and other cells as well as with fc - binding plasma proteins, such as components of the complement system. immune globulin also prevents damage mediated by immune complexes containing c3b into an inactive form ic3b as well as neutralization of some cytokines. ivig can neutralize autoantibodies and down regulate synthesis of antibodies by b cells expressing the relevant idiotype. there are some studies with usage of ivig alone or in combination with pp, and when ivig is used alone a high dose is given (1 - 2 g / kg). it was found ivig was effective in reversing rejection within 25 days of infusion with no recurrence in kidney transplant recipients. the usual recommended dose is 100 mg / kg of ivig after each pp session and 300400 mg / kg for 1 - 2 days after the last pp with a cumulative dose of 1000 mg / kg. a benefit of ivig is its ability to replenish gammaglobulin lost during pp, decreasing infection risk. adverse effects of ivig which are common are headache, fever, chills, myalgias, and hypotension / hypertension which can be reduced by slowing the infusion rate. serious adverse effects are rare and can include aseptic meningitis, acute renal failure, thrombotic events, and anaphylaxis. jin. had used thymoglobulin (atg) 0.75 mg / kg / day for 510 days in combination with plasma exchange in 7 patients with amr and showed 85% reversal in amr. atg eliminates cd4 + t - cell and b - cell interaction causing b - cell toxicity / apoptosis and modulation of alloantibody production. if both cellular and humoral features are seen on biopsy, atg is often used along with steroids for treatment of amr. atg can be given in 3 - 4 doses to a cumulative dose of 6 mg / kg, and it is noted that leukopenia is a side effect which should be monitored to guide specific treatment for each patient. the spleen is the largest lymphoid organ in the body and has been found to play a role in alloantibody generation. splenectomy is used in desensitization protocols for abo incompatible transplants as well as positive crossmatches, however it has been found to surprisingly treat amr refractory to treatment. there are a few case studies, one in particular where abundant clusters of cd138 + plasma cells were found upon review of pathology of the spleen. usually in traumatic splenectomy controls, cd138 + cells she had been treated with pp / ivig and then underwent emergent splenectomy. immediately afterward the mechanism of action is not entirely clear as to why splenectomy treats amr, but several case reports have indicated that it could be a potential treatment for refractory amr to pp / ivig. it is not certain whether the cd138 + cells found in the spleen had produced anti - dr51 antibodies in this particular case, but provides a novel insight for further studies in the role of the spleen with amr. of course, splenectomy has risks associated with it which include increased risk of infections and the risks associated with surgery. therefore these risks must be discussed with the patient before proceeding, as this is not a conventional treatment for amr. there is lifelong risk with encapsulated microorganisms ; therefore patients must receive haemophilus influenza vaccine, pneumococcal, and meningococcal vaccines. most episodes of amr are associated by evidence of early complement activation by the presence of c4d+ staining of the peritubular capillaries. eculizumab is a humanized monoclonal antibody with high affinity for c5 and blocks the activation of terminal complement and is fda approved for the treatment of paroxysmal nocturnal hemoglobinuria. serious adverse effects of this medication are an increased risk of infections, particularly to encapsulated bacteria. it is recommended to immunize patients at least two weeks before the administration of eculizumab to meningococcus and pneumococcus. the transplant community is looking towards the use of this medication because of its highly selective mechanism of action. there are several case reports using eculizumab for desensitization before transplant and to treat amr after transplant. stegall and colleges reported using eculizumab in sensitized renal transplant recipients with high levels of dsa to determine if the incidence of amr in the first 3 months after transplant was decreased compared to historical controls. these patients were immunized one month prior to transplant, received plasmapheresis treatments before and after transplant based on the crossmatch channel shifts, and received the same induction and maintenance immunosuppression. the eculizumab dosing regimen included 600 mg on postoperative day 1, and 600 mg weekly thereafter for 4 weeks. at week 4, assessment of dsa levels was performed to determine if patients would continue treatment based on dsa levels. the study found the incidence of amr was statistically significanty lower in the eculizumab group compared to the control group and reduced the need for splenectomy, reducing transplant glomerulopathy. considering the cost of this medication, this dosing regimen and prophylactic use might be unrealistic for smaller transplant centers. a few case reports used eculizumab in treating renal thrombotic microangiopathy (tma) and reversal of complement activation after abo - incompatible transplantation [2628 ]. the use for tma involved a patient who underwent a living related kidney transplant with a history of lupus developed severe tma, glomerular scarring, and diffuse tubulointerstitial fibrosis with positive apl antibodies. the patient received dialysis and plasmapheresis sessions with no improvement in kidney function. the patient received weekly infusions of eculizumab without complication and renal function improved with subsequent biopsy revealed complete resolution of tma. the transplant community has recognized the potential use of an agent that can directly target plasma cells. traditional treatments have been successful in removing antibodies, inhibiting antibody activity or lowering production but no treatments have been effective in targeting mature antibody production in plasma cells. amr involves the production of high levels of dsa by plasma cells either newly made from memory or nave b cells or from those that existed prior to transplant. bortezomib is a proteasome inhibitor which is fda approved for the treatment of multiple myeloma. this medication has been shown to cause apoptosis of normal plasma cells which in turn decreases alloantibody production in sensitized patients. the university of cincinnati reported the first study on the use of bortezomib in amr. everly and colleagues treated six kidney transplant recipients with refractory amr and concomitant acr with bortezomib and found fast rejection reversal, reductions in dsa levels, improved renal allograft function, and suppression of recurrent rejection. the same group reported two patients with amr who received bortezomib and these patients experienced prompt amr reversal and dsa elimination in 14 days. the use of bortezomib has also been published in five patients with a combination of acr and amr. these patients were given four doses of bortezomib and reversal of both acr and amr occurred with reduction in dsa. other studies have shown positive and mixed results regarding the use of this medication in amr and dsa levels [31, 32 ]. rituximab is a chimeric anti - cd 20 (anti - b cell) monoclonal antibody that is currently fda approved for the treatment of lymphoma. the cd 20 antigen is expressed early in b - cell cycle but is absent on mature plasma cells. the variable region of rituximab binds to cd 20 through three different mechanisms including antibody - dependent cell cytotoxicity, complement - dependent cell killing, and induction of apoptotic cell death. these mechanisms mark cells for destruction and leads to sustained depletion of circulating b cells [32, 33 ]. genberg and colleagues reviewed the pharmacodynamics after a single dose of rituximab given to renal transplant patients and showed b - cell elimination was rapid and occurred in the peripheral blood over 1 - 2 days. the effect on the b - cell population was prolonged and did not reemerge for 1 year and remained suppressed for 2 years. the first report of using rituximab in amr evaluated 27 patients with refractory rejection and received a single dose of rituximab. three grafts were lost and the remaining grafts had good function at the time of discharge. kaposztas and colleagues published a retrospective study involving 54 patients with amr and were split into two groups. group a (n = 26) received plasmapheresis and rituximab and group b (n = 28) received plasmapheresis alone. the two year graft survival was significantly better in the rituximab group. since these two large case reports, there have been many case series, and so forth published on using rituximab with any of the studies for desensitization and for amr [3740 ]. with many of studies the patient population is small, with incomplete criteria for amr diagnosis, and other treatments used in conjunction with rituximab including ivig, plasmapheresis, and steroids. however it should be noted in other studies particularly by pescovitz, a small number of patients failed to show a decrease in pra with rituximab alone. | antibody - mediated rejection (amr) is a major cause of late kidney transplant failure. it is important to have an understanding of human - leukocyte antigen (hla) typing including well - designed studies to determine anti - mhc - class - i - related chain a (mica) and antibody rejection pathogenesis. this can allow for more specific diagnosis and treatment which may improve long - term graft function. hla - specific antibody detection prior to transplantation allows one to help determine the risk for amr while detection of dsa along with a biopsy confirms it. it is now appreciated that biopsy for amr does not have to include diffuse c4d, but does require a closer look at peritubular capillary microvasculature. although plasmapheresis (pp) is effective in removing alloantibodies (dsas) from the circulation, rebound synthesis of alloantibodies can occur. splenectomy is used in desensitization protocols for abo incompatible transplants as well as being found to treat amr refractory to conventional treatment. also used are agents targeted for plasma cells, b cells, and the complement cascade which are bortezomib rituximab and eculizumab, respectively. |
science aims to provide simple and general explanations for natural phenomena, and all sciences must deal with the problem of heterogeneity. variation and heterogeneity are the hallmarks of biological diversity and capture the attention of anyone interested in trying to unravel the mysteries of the biological world. explaining biological diversity was indeed the problem for which darwin provided a simple but revolutionary solution. variation and adaptation are the first two words that come to mind in relation to living organisms, and it was darwin 's genius that in using these two observations he was able to formulate the theory of natural selection to explain the diversity that we see reflected in the millions of different kinds of organisms or species on this planet. given its spectacular success in providing a causal explanation for organismic change (evolution within lineages), it is equally remarkable that darwin was unable to provide a causal mechanism of speciation (evolution between lineages). such a causal theory had to wait nearly a century after the publication of darwin 's origin of species and it materialized only after the evolutionary synthesis of the 1940s after population genetics had developed a theoretical framework [2, 3 ]. diversity is a problem in biology in two ways : the most obvious of which is that diversity needs to be explained ; the other is that it can thwart our efforts in elucidating precise and simple explanations for the complexity of biological phenomena. as a consequence, we trade precision for generality and a rich theoretical base has been developed, which at first appears to ignore diversity. population genetics is a good example as it developed a significantly elaborate theory of how genes change in evolution without prior knowledge of the material / chemical basis of the gene or of genetic variation. the allopatric theory of speciation relied on geographic isolation and differentiation of populations and cumulative effects of gene differences and gene incompatibility [2, 5 ] without specifying anything about the nature of the genes involved. it was therefore surprising that when gel electrophoresis made its debut in scientific methods, genetic differences between closely related species turned out to be minimal. much of this had to do with the fact that the genes being investigated by molecular biologists were of the general cell metabolism category (i.e., allozymes) and had no direct relation to reproductive biology the crime scene of reproductive isolation. on the other hand, the mendelian approach to studying speciation focused on the right phenotype the mendelian approach therefore eventually became more successful in speciation research by uncovering regions of chromosomes and discrete genes with large effects that played a role in causing hybrid sterility / inviability. provided a glimpse into the nature and the variety of genes that effect postzygotic reproductive isolation, but by the virtue of deliberately being chosen as large effect genes (for the ease of mapping), they may or may not represent the pool of genetic variation that is the basis of speciation. it is for this reason that another, parallel approach to investigating the nature of genetic variation affecting reproductive isolation was needed. to understand the genetic basis of reproductive isolation, a more systematic methodology was needed to screen genes and genetic variation associated with the reproductive system. a systematic genomic / proteomic approach was essential because not all genes in the reproductive system would affect reproductive isolation ; many are indeed essential for development and reproductive biology. we needed to find and target the genes and proteins in the reproductive system that matter genes with minor or large effects that are most likely involved in the early stages of species isolation. it was indeed this realization that led us to the idea of investigating genetic variation in the reproductive tracts of drosophila. the idea of using 2d electrophoresis to examine reproductive proteins seemed exciting but there was some hesitation due to the technical difficulties associated with the technique. mike coulthart, a graduate student at that time, was up to the task as he had what was needed : technical precision, patience, and perseverance. he investigated the levels of genetic variation and genetic differentiation, respectively, within and between sibling species of the drosophila melanogaster group. the 2d results were surprising and somewhat un - interpretable at the time. by separating over 250 protein spots from the reproductive tract and comparing them between species, mike found little genetic variation within species but high genetic divergence between species [810 ]. under the neutral theory we expected parity between the levels of within - species and between - species variation, which was indeed the case in the massive amount of data that had accumulated using one - dimensional gel electrophoresis [6, 11 ]. the 2d results were therefore interesting and revolutionary given the dominant framework of neutrality and the expected constant and slow rate of evolution. this was mainly because reproductive tract proteins were considered essential to the organism and therefore not expected to evolve rapidly. the unusual nature of these results called for more investigations and research on sex and reproduction - related (srr) molecules began. the ensuing series of experiments involving 2d electrophoresis showed that (1) nonenzymatic, abundant proteins were generally less polymorphic than enzymes ; (2) reproductive tissue proteins were more diverged between species than nonreproductive tissue proteins, such as those of the brain ; (3) testes and ovary proteins showed higher levels of species divergence than nonreproductive proteins ; (4) reproductive proteins (and reproductive morphological traits) showed more gene expression breakdown in species hybrids than nonreproductive proteins [1316 ]. these data and particularly civetta and singh 's [13, 14 ] research, which showed that sex and reproduction - related (srr) genes evolve faster than nonreproductive genes, unveiled the importance of studying the evolution of srr molecules in speciation research [14, 1720 ]. while the average rates of evolutionary change per gene may be small, genes can evolve rapidly depending upon the environmental conditions and the selection pressure. the dynamics of selection acting on each locus will determine its rate and pattern of evolution. some groups of genes may evolve rapidly by virtue of their functions as is the case with the immune response genes in mammals. immune response genes are an example of a coevolutionary system where evolution of immunity or resistance in hosts is countered by the evolution of virulence in pathogens and/or parasites. immune system genes and virulence genes are locked in an antagonistic coevolutionary arms race and are expected to evolve rapidly. sexual system genes provide another example of a coupled coevolutionary system, in this case involving the interactions between males and females of the same species. advances in molecular technology particularly dna amplification and sequencing propelled srr gene research and a remarkable trend of pervasive rapid srr gene evolution emerged at several levels. some sex determining genes, assumed to be conserved due to their important functions during early development, were shown to evolve rapidly. when genes expressed in testis, ovary, and nonreproductive tissues were screened for rates of evolution it became clear that a substantial proportion of these genes evolved more rapidly than genes expressed in nonreproductive tissues [24, 25 ]. a divergence trend of testis > ovary > somatic genes emerged suggesting male and female srr genes evolve under different selective pressures. microarray and computational methods using entire tissue - specific transcriptomes showed that testis that expressed srr genes were more likely to break down in species hybrids [27, 28 ]. sperm proteins were shown to evolve rapidly and divergently in invertebrates and mammals [2932 ]. proteins in the seminal fluid of drosophila were also found to be evolving rapidly and were shown to confer specific physiological and behavioural modifications in the female [3337 ]. these studies not only indicate that sexual reproduction provides an opportunity for exerting constant selection pressure generation after generation but also that differences in the evolutionary dynamics of male and female reproductive systems are presumably due to intersexual selection pressure arising from male - female interaction in each generation. the relationship between rapid evolution of srr genes and reproductive isolation is attested by the fact that the known candidate speciation genes are either srr genes or autosomal genes that, via incompatible interactions with genes on the x chromosome, affect hybrid dysfunction (reviewed in [3841 ]). in addition, genome - wide evidence of rapid evolution of srr genes provided a mechanistic framework to discuss the nature of genetic changes that may occur during speciation [14, 17, 42 ]. the discovery of jingwei and sdic opened up investigations into the origin of new srr genes marking yet another important step into understanding the evolutionary dynamics of genetic systems [4548 ]. novel genes arise through a variety of molecular mechanisms, including being derived from previously noncoding dna and may be important in functional diversification. what is extremely interesting is that the majority of novel genes or gene copies that have evolved novel functions have also evolved testis - specific expression. interestingly, odysseus (odsh), a hybrid sterility gene, evolved as a duplicate of the neuron expressed unc-4 gene and has taken up a testicular expression and role [50, 51 ]. another example is ms(3)k81, a gene that evolved by duplication and retroposition from a previously ubiquitously expressed copy to acquire a male - germline - specific expression and function. ms(3)k81 is only found in the 9 species of the melanogaster subgroup and in its new functional role is crucial for zygote viability. accessory gland proteins (acps) are a prime example of male - specific genes in drosophila that have taken up a variety of reproduction - related functions and have important physiological effects in the female reproductive system [3437, 5356 ]. retrotransposition is another important means of gene copying and shuffling that can be important in the evolution of new functions [45, 46 ]. interestingly more genes moving from the x chromosome to autosomes have been retained (active) than genes moving in the opposite direction [7, 47, 5761 ]. again, testis - specific expression and rapid evolution appear to be common amongst retroposed genes. thus for some yet unexplained reason, it appears that the testes act as cauldrons of retaining, if not manufacturing, new / refugee genes. in fact, it turns out that not only the evolution of new genes but also gene loss (loss of orthology) is also elevated in male - biased genes as compared to female - biased genes in drosophila species. the genetic machinery of the sexual system shows faster rates of turnover and it points to the role of sexual selection acting preferentially through the males (male - driven sexual selection) [62, 63 ]. the evolution of new genes and novel functions can be a potent driving force of reproductive isolation as exemplified by odysseus (odsh). sexual selection, strictly speaking, is only a small part of the total selection pressure that the organism is exposed to in relation to sex. classical theories of sexual selection apply only to those traits and genes that are influenced, directly or indirectly, by female choice. on the contrary, selection in relation to sex, or what has been called sexual selection in the broad sense, applies to all aspects of reproductive biology from soliciting mates, courting, and mating, to production of offspring. a large proportion of the genome is involved in the development and maintenance of reproductive systems and a significant proportion of genes (~30%) in the drosophila genome shows sex - biased gene expression, most of which is reproductive tissue specific [65, 66 ]. this raises the possibility of an unexpected level of conflict between natural and sexual selection. current studies of sexual selection have expanded to different aspects of reproductive biology and constitute a major area of research in evolutionary biology. genetically identical (with the exception of the y - chromosome), males and females are expected to differ in genes associated with primary sexual characteristics such as ovary, testes, and copulatory organs. traditionally thought to be associated to few genes on the sex chromosome, it turns out that the breadth and complexity of sexual interaction between the two sexes has become so elaborate that a large number of genes controlling a variety of traits have become associated in a sex - specific manner (sex - biased and sex - enriched genes) expanding the level of sexual dimorphism [66, 67 ]. in the drosophila genome a substantial proportion of genes show sex - biased expression [65, 66 ]. male - biased genes are underrepresented on the x chromosome and female - biased genes are enriched on the x chromosome. these genes are expressed in a tissue - specific manner (e.g., somatic tissues, ovary, and testis) and they even show sex - specific elevated levels of movement between sex chromosome and autosomes [59, 60, 6772 ]. in the last decade, several theories including sexual antagonism, dosage compensation, meiotic sex chromosome inactivation, and meiotic drive have been proposed to explain the paucity of male - biased gene on the x chromosome and the driving force responsible for the evolution of sex chromosomes, their gene content, and expression patterns [58, 67, 7274 ]. meiotic sex chromosome inactivation (msci) pioneered first by lifschytz and lindsley and then shown at a genomic scale [58, 72 ] appeared to be convincing in drosophila ; however, recent evidence shows otherwise and is currently under debate [74, 7679 ]. while explaining the relocation and expression pattern of sex - biased genes will remain a prominent research area, it is noteworthy that, with respect to rates of evolution, genes with sex - biased expression and particularly male - biased sex genes show unusually high rates of evolution [67, 80, 81 ]. it is not surprising then that the sexes differ in their rates of evolution of sterility and inviability during speciation as pointed out by haldane. sea urchins have traditionally been a model organism for development biology and reproductive biology but have recently received considerable attention from an evolutionary standpoint particularly in the evolution of reproductive isolation to explain speciation in the sea [83, 84 ]. in most internally fertilizing animals, specific courtship behaviours mediate male and female interactions ensuring species - specific copulation and fertilization. in contrast, in externally fertilizing organisms with little or no such mating behaviours (e.g., sea urchins) gametes are shed into the sea where species - specific fertilization occurs. several molecules on the surface of gametes that mediate various stages of sperm - egg interaction have been characterized [8587 ]. studies on two important proteins exemplify the evolutionary dynamics of gametic molecules in externally fertilizing marine organisms (see for a recent review). studies on the abalone sperm molecule lysin and its egg receptor verl demonstrate the fact that male and female gametic proteins coevolve species - specific structures and affinities to maintain species - specific interactions and avoid cross - fertilization [83, 84, 8890 ]. the sperm molecule bindin and its egg receptor are another classic example from sea urchins [83, 84, 88, 91 ]. while the evolutionary dynamics of the egg receptor for bindin remains obscure, the sperm protein bindin evolves rapidly and divergently in some genera but not in others [83, 84 ]. however, bindin does appear to have some involvement in reproductive isolation since its divergence correlates with the degree of gametic incompatibility between but not with time since species divergence. in all likelihood, other molecules must be involved and there is a need for further characterization of such gametic and other sex and reproduction - related molecules in broadcast spawners. once such molecules are identified, it will be interesting to correlate the evolution of egg - sperm interacting molecules to the patterns of hybrid incompatibility in these organisms. external fertilizing systems may provide a unique opportunity to assess the relative roles and genetic consequences of sexual selection and conflict in driving divergence of reproductive molecules and speciation. while research into reproductive molecules is at its inception in externally fertilizing systems, the sperm proteome of drosophila has opened up exciting venues of research in reproductive biology [9395 ]. as with other male - biased genes, sperm genes are underrepresented on the x chromosome and are nonrandomly clustered in the genome. while certain groups of sperm proteins, such as binding factors, do evolve rapidly, overall, the sperm proteome does not appear to be evolving fast, there is little evidence of positive selection, and there is widespread functional and structural constraint. this is in stark contrast to seminal fluid proteins that evolve fast and are under selection. the contrasting evolutionary patterns of the two groups of male ejaculate proteins are interesting and are indicative of the complexity of reproductive processes, where crucial sperm - egg interacting proteins are sheltered but seminal fluids that accompany them interact with the general environment of the female reproductive tract and proteins therein are under strong selection. future research on sperm - egg interacting proteins promises to increase our knowledge about the functional evolution of the male and female fertilization machinery and, more broadly, the evolutionary origins of sexual reproduction. haldane 's rule (of speciation) points to the preferential appearance of hybrid sterility and and/or inviability in the heterogametic sex. in flies and mammals, it is the males who are affected while in moths and birds it is the females [39, 96, 97 ]. the genetics of hybrid sterility and inviability have been of intense focus in speciation studies and a great deal of effort has been made in mapping and characterizing genes involved in hybrid breakdown [38, 39, 50, 98106 ]. the evolution of hybrid sterility / inviability is explained by the bateson - dobzhansky - muller incompatibility model which states that incompatibility is the result of independent evolution of genes in isolated populations [7, 106 ]. haerty and singh showed that the genes showing breakdown in the hybrids are preferentially sex - related genes and that these genes evolve faster both in sequence and gene expression [27, 28, 107 ]. in the light of this it is interesting to note that all but a handful of the so - called speciation genes and hybrid - sterility genes are characterized by high sequence divergence ; they are often sex - related genes or somatic genes that affect the sexual system [7, 38, 40, 41, 108 ]. the effects of genes are prone to change in response to incorporation of new mutants and during the course of speciation earlier mutations would have fewer interactions than older mutations (cascade effect). speciation genes may have started as small - effect minor genes and have become elaborate in their genic interactions later through species - specific adaptation and evolution. in this scenario thus the effect of cascade evolution is not only that speciation would occur rapidly and precipitously but also that speciation genes would evolve in their average effect from being minor to major genes. so while in reality speciation may occur in an incremental manner through a combination of many minor genes, in postspeciation genetic investigations genes would often appear as major genes. this is an interesting scenario and we must find a way to approach this problem experimentally. srr genes provide new opportunities to mount comparative studies of the role of selection versus developmental constraints in evolution. for example, srr genes have provided an excellent example of testing alternative explanations for von baer 's third law. this was later interpreted to be the result of selection against changes in earlier stages of development, which could have cascading, deleterious developmental repercussions. darwin on the other hand explained the conservation of morphology in earlier stages as being due to lack of opportunity for natural selection to act. since natural selection results from changes in the environment, it follows that earlier, sessile stages that have not fully developed will have little opportunity to experience variation in the environment. darwin further pointed out that secondary / sex - specific sexual traits appear when they are needed and this can be seen in the secondary sexual traits in animals. two recent studies [28, 107 ] explored the relationship between expression level over ontogeny and rates of divergence and found support for both selection against deleterious cascading effects and darwin 's hypothesis : genes expressed during early stages show reduced divergence. however, the more rapid divergence of later, adult stages, is dominated by genes expressed in adult males, which are, as noted above, presumably diverging under the effect of directional (sexual) selection. as a result of these observations, artieri. proposed a model of divergence involving two factors playing dominant roles during different periods of development : conservation early and opportunity late. more of such studies should shed light on the relationship between evolution and development [110113 ] as well as on the broader aspects of speciation and macroevolution beyond reproductive isolation. while srr research has provided a useful, complementary approach to study speciation, there is a need to explain the evolutionary forces driving pervasive rapid evolution of male and female reproductive genes. initially rapid srr gene evolution invoked the role of sexual selection by female choice but recent developments on parker 's original theses revived the role of sexual conflict in explaining evolutionary changes in sexual systems. sexual dimorphism in higher organisms leads to sex - specific life styles, reproductive behaviours, and investments in sexual interactions. it is expected that these differences will lead to conflicts of interest between males and females and this conflict may work as a stimulus for retaliatory evolutionary changes known as sexual arms races [115117 ]. a sexual arms race would require action and reaction on the part of both partners and thus it provides a test of the role of female choice, male - driven sexual selection, and of sexual arms race theories. increasing empirical evidence suggests that sexual conflict may be pervasive [90, 115, 118123 ] but much remains to be done particularly at the level of genes to substantiate how sexual conflict and sexual selection affect male and female genetic systems differently. genomics provides the means to explore the molecular consequences of sexual arms races and associated sexual selection theories. intersexual interactions, be they mutualistic or antagonistic, have the potential to drive population divergence in a self - accelerating manner and this may be one of the reasons why origins of diversity and speciation are much higher in higher organisms. evo - devo studies will also help to answer the perennial question : during evolution and speciation what comes first reproductive isolation or adaptive radiation ? since its inception 25 years ago, srr gene research has rapidly evolved into a large coherent field in evolutionary biology, particularly influencing reproductive biology and speciation. the focus on srr system studies has provided valuable mechanistic frameworks that directly relate to theories of how speciation occurs. it has emphasized the role of sexual selection in evolution, propelling research on how sexual selection and sexual conflict work at the population level. the genomics era has revolutionized srr gene research and resulted in the characterization of rates of evolution and patterns of gene expression in reproductive transcriptomes. rapid evolution is now commonly associated with reproductive genes but, in the future, work will be needed to understand the functions of rapidly evolving srr genes and details of why they evolve rapidly and how their rapid evolution affects the rest of organismal biology. it will call for an integrated approach, unifying disparate fields of science, particularly biochemistry, genetics, ecology, and molecular biology. a key issue will be to explore the relationship between changes in gene sequence, gene expression, protein syntheses, and protein function in reproductive systems. fundamental behavioural and ecological studies will be essential in explaining the nature of molecular changes associated with the reproductive systems. selection in relation to sex, encompassing sexual selection in the strict sense, and in the broad sense is a large and growing area of research in evolutionary biology. investigating the molecular consequences of sexual interaction and their role in speciation stands to open one of the most important areas of research bearing on the biology of sexual reproduction. | the protein electrophoresis revolution, nearly fifty years ago, provided the first glimpse into the nature of molecular genetic variation within and between species and showed that the amount of genetic differences between newly arisen species was minimal. twenty years later, 2d electrophoresis showed that, in contrast to general gene - enzyme variation, reproductive tract proteins were less polymorphic within species but highly diverged between species. the 2d results were interesting and revolutionary, but somewhat uninterpretable because, at the time, rapid evolution and selective sweeps were not yet part of the common vocabulary of evolutionary biologists. since then, genomic studies of sex and reproduction - related (srr) genes have grown rapidly into a large area of research in evolutionary biology and are shedding light on a number of phenomena. here we review some of the major and current fields of research that have greatly contributed to our understanding of the evolutionary dynamics and importance of srr genes and genetic systems in understanding reproductive biology and speciation. |
assemble the chamber (design illustrated in figure 1). place two beads of vacuum grease, spaced ~8 - 10 mm apart, across the recording chamber to form a channel for superfusate and in which to embed the retinal slices. add a second bead of grease a few millimeters farther out beyond each of these two beads of grease to act as a levee and limit spillover. place a small triangular piece of kimwipe at the end of the chamber to ensure fluid contact with the reference electrode. press a piece of nitrocellulose membrane (~ 5 x 10 mm ; 0.8 m pores) flat against the glass microscope slide into two small beads of vacuum grease. avoid putting grease directly beneath the center of the nitrocellulose membrane, as this can prevent the retina from adhering. to prepare the tissue slicer, break a double edge razor blade into 4 pieces and attach one to the slicing arm. cut a thin slice of nitrocellulose membrane to ensure that the cutting edge of the razor blade lays flat against the recording chamber and therefore cuts cleanly through the nitrocellulose membrane. keep a small beaker of amphibian saline solution (table 1) on ice at the dissection station. place half of the head on a piece of cotton moistened with amphibian saline atop a linoleum block. the other half of the head can be wrapped with a moist paper towel and stored at 4 c for use later in the day. cut the skin connecting the eye to the surrounding orbit. after freeing the front of the eye from the surrounding orbital tissue, pull the eye forward and slide the scissors under the eye to cut through the eye muscles and optic nerve, freeing the eye from the orbit. make a small incision in the center of cornea with a sharp surgical blade. remove the cornea by sliding fine vannas scissors into the incision and extending the cut radially out toward the ora serrata. cut circumferentially around the ora serrata by rotating the linoleum block or the cotton between cuts. after cutting all the way around the eye, remove the cornea and lens by pulling them out of the side of the eyecup. move the resulting eyecup onto a hard surface of the linoleum block moistened with amphibian saline solution. cut it into thirds with a sharp razor blade, using a fine sawing motion to ensure that you have cut all the way through the sclera. place one or two pieces of eyecup on the nitrocellulose membrane with the retinal surface facing down. submerge the remaining pieces with additional saline and place them in the refrigerator at ~4 c. gently press the piece of eyecup against the nitrocellulose membrane with fine forceps. submerge the nitrocellulose membrane and eyecup piece with several drops of cold amphibian saline and blot at the edges with kimwipe to help the retina to adhere. once again, submerge the eyecup and nitrocellulose membrane with several drops of cold amphibian saline solution and peel away the sclera / choroid / retinal pigment epithelium to isolate the retina (which can appear pink due to the presence of unbleached rhodopsin). if the retina has not adhered tightly, drain away the saline with a kimwipe to pull the retina more firmly down onto the nitrocellulose membrane. replace the saline. fill the chamber with cold amphibian saline and transfer it to the stage of the tissue slicer. slice the retina and nitrocellulose membrane into thin strips, working from one end to the other by turning the vernier micrometer in 125 m increments. transfer the retinal slices by moving strips of nitrocellulose membrane to the main channel of the recording chamber. lift a strip of membrane free and then hold it in place while moving the chamber beneath it, being sure to keep the slices submerged. embed the edges of the nitrocellulose membrane in the strips of vacuum grease, rotating them 90 degrees to view the retinal layers. press the nitrocellulose membrane flat against the glass surface. even if there is not retina on every piece, place strips of nitrocellulose membrane at regular intervals (~1 mm apart) along the entire length of the perfusion channel to help break up the surface tension and improve fluid flow. after all of the slices have been transferred, move the recording chamber to the stage of an upright, fixed stage microscope and attach the reference electrode lead. focus on the slices using a long - working distance, water immersion, 40 - 60x objective. the microscope should be placed on an air table to dampen vibrations and enclosed in a faraday cage to reduce electrical interference. superfuse the slices continuously at a rate of 1 ml / min with amphibian saline solution bubbled with 100% o2. outflow can be regulated by rotating the beveled end of the suction needle or by moving the kimwipe at the end of the chamber closer or farther away from the outflow needle. the preparation can be kept at room temperature or cooled with a peltier device or by simply setting an ice pack on the microscope stage. examine slices under dim or infra - red light and identify a pair of cells - a photoreceptor (rod or cone) and nearby horizontal or bipolar cell - to target for whole - cell recording. rods can be identified by their large cell bodies and prominent rod - like outer segments (figure 2a). bipolar cell and horizontal cell somas are in the outermost row of cell bodies in the inner nuclear layer (inl ; figures 2b and 2c). before preparing slices, use a pipette puller to manufacture micropipettes from borosilicate glass (1.2 mm outer diameter, 0.95 mm inner diameter with a glass filament). using a non - metallic filling needle (e.g. one manufactured from a 1 cc syringe or a microfil), fill pipettes with the intracellular solution (table 1) and attach to the electrode holder. position the photoreceptor pipette beneath the objective and then lower it so that the tip is positioned just above the slices. adjust any offset in the baseline current level on the amplifier. check the pipette resistance with a 5 - 10 mv depolarizing pulse. we typically use pipettes that range from 10 - 15 m, the result of the long taper of the shaft and low osmolarity of the amphibian pipette solutions. with higher osmolarity mammalian solutions, these same pipettes exhibit resistance values of ~8 - 12 m. while we have used larger tip diameters with resistance values of 3 - 4 m in amphibian solutions, the advantages provided by a lower access resistance are offset by a greater difficulty in sealing onto cell membranes and a more rapid rundown of calcium currents and other second messenger - sensitive responses. while applying slight positive pressure, position the post - synaptic pipette so that it contacts the horizontal or bipolar cell body. then position the presynaptic pipette so that it contacts the cell body of a rod or cone photoreceptor. recordings appear to be more stable when pipette tips contact the inner segment rather than the soma, especially in cones. while monitoring the resistance, release the positive pressure on the post - synaptic pipette. sometimes, the release of positive pressure is sufficient to form a gigaohm seal. if not, apply gentle suction with a 1 ml syringe or by mouth. after the tip resistance has grown to > 100 m, apply a holding potential of -60 mv. after obtaining a gigaohm seal, null out any pipette capacitance transients and repeat the sealing procedure for the photoreceptor pipette, applying a holding potential of -70 mv. rupture the patch by using your mouth or a syringe to apply suction to each cell in turn. rods, cones, and bipolar cells will typically rupture with gentle suction. obtaining whole - cell configuration with a horizontal cell may require greater suction (i.e. with a 3 cc syringe) in combination with strong quick voltage pulses delivered with the " zap " feature of the patch clamp amplifier. rupture of the membrane and establishment of whole - cell configuration will be evident by the appearance of whole - cell capacitance transients. confirm identity of the post - synaptic cell physiologically by applying a light flash and delivering a series of voltage steps from -120 to + 40 mv in 20 mv increments (figures 3 and 4). to assess if the pair of cells are synaptically connected, deliver a brief (25 - 100 msec), 60 mv step depolarization to the photoreceptor (to -10 mv, near the peak of the l - type voltage - gated calcium current) and look for post - synaptic currents in the second order neuron (figure 5). a strong depolarizing step should evoke a fast, transient inward post - synaptic current in the post - synaptic horizontal or off bipolar cell caused by a burst of vesicle release from the cone (figure 5). assemble the chamber (design illustrated in figure 1). place two beads of vacuum grease, spaced ~8 - 10 mm apart, across the recording chamber to form a channel for superfusate and in which to embed the retinal slices. add a second bead of grease a few millimeters farther out beyond each of these two beads of grease to act as a levee and limit spillover. place a small triangular piece of kimwipe at the end of the chamber to ensure fluid contact with the reference electrode. press a piece of nitrocellulose membrane (~ 5 x 10 mm ; 0.8 m pores) flat against the glass microscope slide into two small beads of vacuum grease. avoid putting grease directly beneath the center of the nitrocellulose membrane, as this can prevent the retina from adhering. to prepare the tissue slicer, break a double edge razor blade into 4 pieces and attach one to the slicing arm. cut a thin slice of nitrocellulose membrane to ensure that the cutting edge of the razor blade lays flat against the recording chamber and therefore cuts cleanly through the nitrocellulose membrane. keep a small beaker of amphibian saline solution (table 1) on ice at the dissection station. place half of the head on a piece of cotton moistened with amphibian saline atop a linoleum block. the other half of the head can be wrapped with a moist paper towel and stored at 4 c for use later in the day. cut the skin connecting the eye to the surrounding orbit. after freeing the front of the eye from the surrounding orbital tissue, pull the eye forward and slide the scissors under the eye to cut through the eye muscles and optic nerve, freeing the eye from the orbit. make a small incision in the center of cornea with a sharp surgical blade. remove the cornea by sliding fine vannas scissors into the incision and extending the cut radially out toward the ora serrata. cut circumferentially around the ora serrata by rotating the linoleum block or the cotton between cuts. after cutting all the way around the eye, remove the cornea and lens by pulling them out of the side of the eyecup. move the resulting eyecup onto a hard surface of the linoleum block moistened with amphibian saline solution. cut it into thirds with a sharp razor blade, using a fine sawing motion to ensure that you have cut all the way through the sclera. place one or two pieces of eyecup on the nitrocellulose membrane with the retinal surface facing down. submerge the remaining pieces with additional saline and place them in the refrigerator at ~4 c. gently press the piece of eyecup against the nitrocellulose membrane with fine forceps. submerge the nitrocellulose membrane and eyecup piece with several drops of cold amphibian saline and blot at the edges with kimwipe to help the retina to adhere. once again, submerge the eyecup and nitrocellulose membrane with several drops of cold amphibian saline solution and peel away the sclera / choroid / retinal pigment epithelium to isolate the retina (which can appear pink due to the presence of unbleached rhodopsin). if the retina has not adhered tightly, drain away the saline with a kimwipe to pull the retina more firmly down onto the nitrocellulose membrane. replace the saline. fill the chamber with cold amphibian saline and transfer it to the stage of the tissue slicer. slice the retina and nitrocellulose membrane into thin strips, working from one end to the other by turning the vernier micrometer in 125 m increments. transfer the retinal slices by moving strips of nitrocellulose membrane to the main channel of the recording chamber. lift a strip of membrane free and then hold it in place while moving the chamber beneath it, being sure to keep the slices submerged. embed the edges of the nitrocellulose membrane in the strips of vacuum grease, rotating them 90 degrees to view the retinal layers. press the nitrocellulose membrane flat against the glass surface. even if there is not retina on every piece, place strips of nitrocellulose membrane at regular intervals (~1 mm apart) along the entire length of the perfusion channel to help break up the surface tension and improve fluid flow. after all of the slices have been transferred, move the recording chamber to the stage of an upright, fixed stage microscope and attach the reference electrode lead. focus on the slices using a long - working distance, water immersion, 40 - 60x objective. the microscope should be placed on an air table to dampen vibrations and enclosed in a faraday cage to reduce electrical interference. superfuse the slices continuously at a rate of 1 ml / min with amphibian saline solution bubbled with 100% o2. outflow can be regulated by rotating the beveled end of the suction needle or by moving the kimwipe at the end of the chamber closer or farther away from the outflow needle. the preparation can be kept at room temperature or cooled with a peltier device or by simply setting an ice pack on the microscope stage. examine slices under dim or infra - red light and identify a pair of cells - a photoreceptor (rod or cone) and nearby horizontal or bipolar cell - to target for whole - cell recording. rods can be identified by their large cell bodies and prominent rod - like outer segments (figure 2a). bipolar cell and horizontal cell somas are in the outermost row of cell bodies in the inner nuclear layer (inl ; figures 2b and 2c). before preparing slices, use a pipette puller to manufacture micropipettes from borosilicate glass (1.2 mm outer diameter, 0.95 mm inner diameter with a glass filament). the tip of each micropipette should be ~1 - 2 microns in diameter. using a non - metallic filling needle (e.g. one manufactured from a 1 cc syringe or a microfil), fill pipettes with the intracellular solution (table 1) and attach to the electrode holder. position the photoreceptor pipette beneath the objective and then lower it so that the tip is positioned just above the slices. we typically use pipettes that range from 10 - 15 m, the result of the long taper of the shaft and low osmolarity of the amphibian pipette solutions. with higher osmolarity mammalian solutions, these same pipettes exhibit resistance values of ~8 - 12 m. while we have used larger tip diameters with resistance values of 3 - 4 m in amphibian solutions, the advantages provided by a lower access resistance are offset by a greater difficulty in sealing onto cell membranes and a more rapid rundown of calcium currents and other second messenger - sensitive responses. while applying slight positive pressure, position the post - synaptic pipette so that it contacts the horizontal or bipolar cell body. then position the presynaptic pipette so that it contacts the cell body of a rod or cone photoreceptor. recordings appear to be more stable when pipette tips contact the inner segment rather than the soma, especially in cones. while monitoring the resistance, release the positive pressure on the post - synaptic pipette. sometimes, the release of positive pressure is sufficient to form a gigaohm seal. if not, apply gentle suction with a 1 ml syringe or by mouth. after the tip resistance has grown to > 100 m, apply a holding potential of -60 mv. after obtaining a gigaohm seal, null out any pipette capacitance transients and repeat the sealing procedure for the photoreceptor pipette, applying a holding potential of -70 mv. rupture the patch by using your mouth or a syringe to apply suction to each cell in turn. rods, cones, and bipolar cells will typically rupture with gentle suction. obtaining whole - cell configuration with a horizontal cell may require greater suction (i.e. with a 3 cc syringe) in combination with strong quick voltage pulses delivered with the " zap " feature of the patch clamp amplifier. rupture of the membrane and establishment of whole - cell configuration will be evident by the appearance of whole - cell capacitance transients. confirm identity of the post - synaptic cell physiologically by applying a light flash and delivering a series of voltage steps from -120 to + 40 mv in 20 mv increments (figures 3 and 4). to assess if the pair of cells are synaptically connected, deliver a brief (25 - 100 msec), 60 mv step depolarization to the photoreceptor (to -10 mv, near the peak of the l - type voltage - gated calcium current) and look for post - synaptic currents in the second order neuron (figure 5). a strong depolarizing step should evoke a fast, transient inward post - synaptic current in the post - synaptic horizontal or off bipolar cell caused by a burst of vesicle release from the cone (figure 5). representative traces of light responses from neurons in vertical slices of salamander retina are shown in figure 3. the cone, horizontal cell, and off bipolar cell all display an outward current in response to light onset. the prominent inward current following the light flash in the horizontal and bipolar cell recordings is caused by the increased release of glutamate from photoreceptors as they depolarize at light offset. the on bipolar cell responds with an inward current at light onset resulting from a sign - inverting metabotropic glutamate receptor signaling cascade and activation of trpm1 channels. horizontal cells and bipolar cells can be distinguished from one another by their i - v relationships (figure 4). horizontal cells typically have a linear or inwardly rectifying i - v and low input resistance (< 500 m ; figure 4a) while bipolar cells have a high input resistance (0.5 - 2 g) and outwardly - rectifying i -- v (figure 4b). figure 5 shows representative results from recordings of a cone - horizontal cell pair (figure 5a) and cone - off bipolar cell pair (figure 5b). in each, depolarizing the cone to -10 mv from a holding potential of -70 mv evoked a voltage - gated calcium current in the cone and fast inward epsc in the horizontal or bipolar cell. this strong stimulus is sufficient to empty the readily - releasable pool of ~20 vesicles from each synaptic ribbon, resulting in an epsc of ~47 pa / ribbon. the horizontal cell epsc in figure 5a was 232 pa, suggesting that it received 5 ribbon contacts from the presynaptic cone. a similar estimate from the 178 pa epsc in the off bipolar cell (figure 5b) suggests that it received 4 ribbon contacts from the presynaptic cone. cs - glutamate is made by neutralizing 40 mm l - glutamic acid with 40 mm csoh in the pipette solution. a 1 m stock of cs - gluconate is made by neutralizing a solution of csoh with 45 - 50% d - gluconic acid. the ph should be adjusted with naoh for the extracellular solution and csoh for the pipette solutions. keeping the osmolarity of the pipette solutions just below that of the extracellular solution prevents cell swelling and enhances longevity of the recording. components and parameters for the standard intracellular and extracellular solutions used in this protocol. recording chamber design.(a - c) top, side, and cutaway views of the recording chamber showing dimensions. superfusate enters the chamber through a 10 cm length of teflon tubing (inflow tube ; type 24lw). superfusate is removed by applying mild suction to a beveled 20 gauge metal tube at the other side of the chamber. the lead from this reference electrode is connected to the reference input of the headstage. a small triangle of kimwipe is placed over the reference electrode to keep it in contact with the solution and regulate solution flow into the outflow tube. the base of the chamber is formed by placing a glass microscope slide into the recessed edges of the chamber.. strips of nitrocellulose membrane with retinal slices are embedded in beads of vacuum grease that form a channel for solution to flow. prior to making slices, a 5 x 10 mm piece of nitrocellulose membrane is affixed to the chamber with two small beads of vacuum grease. a piece of the eyecup is placed vitreal side down on this nitrocellulose membrane and lifted away once the retina adheres. dye - filled cell pairs in vertical slice preparation.a) images of a rod and synaptically - coupled horizontal cell that were filled with contrasting fluorescent dyes introduced via the patch pipette during simultaneous whole - cell recording. lucifer yellow (2 mg / ml) was included in the rod pipette solution (yellow) and sulfarhodamine b (1 mg / ml) was included in the horizontal cell pipette solution (purple). fluorescent images were captured using a spinning disc confocal microscope (perkin elmer ultraview lci) equipped with a cooled ccd camera (orca er) and mounted to the fixed stage microscope (nikon e600 fn with 60x, 1.0 na water immersion objective). these images were overlaid on bright field images of the corresponding retinal slices using adobe photoshop. the bipolar cell axon terminal ramifies in the outermost (s1) sublamina of the inner plexiform layer near the border with the inner nuclear layer. note that the cone terminals are considerably larger than the axon terminal of the rod. although horizontal cells can be identified by the oblong shape of their cell bodies, cell bodies of on- and off - type bipolar cells in the inl can not be easily distinguished prior to recording. however, one can target displaced cone - driven off bipolar cells which have cell bodies in the onl and can be distinguished from cones by the absence of inner and outer segments. light - evoked currents recorded under voltage clamp from four different retinal neurons in response to a bright 500 msec flash of white light. the cone (a), horizontal cell (b), and off bipolar cell (c) all responded to light with an outward current. (d) the on bipolar cell responded to the same light stimulus with an inward current. the baseline noise exhibited by the off bipolar cell (c) reflects a continual release of synaptic vesicles in darkness that diminishes when photoreceptors hyperpolarize in light. responses from these four cells were obtained in separate recordings using slices prepared under white light. the intensity of the white light stimulus used in these examples produced responses in horizontal and bipolar cells equivalent to 580 nm photon flux of 1 x 10 photons / sec/m. figure 4. current - voltage (i - v) relationships of horizontal and off bipolar cells.(a)top panel, membrane currents evoked by a series of 150 msec voltage steps from -120 to + 40 mv applied in 20 mv increments (bottom panel) to a horizontal cell. horizontal cells typically have a low input resistance and linear or inwardly rectifying i - v relationships. (b) i - v relationship of an off bipolar cell in response to a series of voltage steps. bipolar cells have a higher input resistance and an outwardly rectifying i - v, due to the activation of voltage - gated potassium currents. examples of paired recording data.(a) recording of the cone voltage - gated calcium current (cone ica) in response to a 100 msec step to -10 mv from a holding potential of -70 mv (cone vm). a fast excitatory post - synaptic current (epsc ; hc psc) was recorded simultaneously from a horizontal cell, showing that these two cells are synaptically coupled. (b) an epsc was recorded in an off bipolar cell in response to the same stimulus in another cone. the retinal slice preparation has proven very useful for analyzing the circuitry and mechanisms employed by the retina to process visual information. the ability to obtain whole cell recordings simultaneously from pre- and post - synaptic neurons has been particularly helpful in this endeavor. paired whole cell recordings are much easier to achieve with slices than with flat - mount retina preparations because the different retina layers are exposed. moreover, because of their large retinal neurons, salamanders have a long history as a retinal preparation and therefore provide a particularly well - characterized model system. with practice, healthy slices of salamander retina can be prepared regularly. 1) make sure the razor blade is mounted on the tissue slicer so that it lays flat against the glass surface and slices cleanly though both the tissue and underlying nitrocellulose membrane. if you have made a clean cut through the nitrocellulose membrane, you should hear a faint click as the razor blade strikes the surface of the glass slide. 2) make sure the retina has adhered to the nitrocellulose membrane. otherwise, the retina can float away from the membrane during any step of the procedures. 3) do not expose cut slices to air, as this will damage many of the superficial cells. 4) make sure the slices and nitrocellulose membrane lie flat against the glass slide so that the retinal layers are apparent under the dissecting microscope. 5) balance the rates of superfusate inflow and outflow in order to avoid overflowing the recording chamber. 6) select a healthy pair of cells close to one another. cells with smooth cytoplasm are healthier than cells with grainy cytoplasm. 7) make sure the pipette tip has not broken or brushed against other tissue or debris on the way down to the cells. 8) check the pipette resistance to ensure that it is not clogged with debris or a bubble, both of which can make it difficult to obtain a quality whole - cell recording. rather than attaching the retina to nitrocellulose filter paper, some investigators embed retinas in a block of agar and use a vibratome to cut retinal slices. discuss advantages of both approaches. in their experience, the agar - based approach provides a more consistent yield of flat slices with well - delineated retinal layers but the filter paper - based approach yields healthier photoreceptors. rods and cones are responsible for transducing light into changes in membrane potential. with paired recordings, the membrane potential of rods or cones can be manipulated directly and so the ability to generate light responses, while helpful for identifying cell types, may not be essential. we therefore often prepare slices in white light. however, even when prepared under bright illumination, salamander retinal neurons can generate large light responses as illustrated by the responses in fig. this is partly due to a relatively large reservoir of chromophore in the large outer segment volume but may also reflect the ability of mller cells to regenerate 11-cis - retinal for cones. to obtain fully dark - adapted light responses, one can prepare the slices under infrared illumination. for dissections under infrared light, we attach geniii image intensifiers (nitemate nav3, litton industries, tempe, az) to the oculars of the dissecting microscope and illuminate the tissue with an infrared led flashlight. for slicing and other procedures that are not conducted under the dissecting microscope, we employ a head - mounted image intensifier. for placement of the patch pipettes, we visualize slices using an infrared - sensitive ccd camera (e.g. watec 502h, watec inc., middletown, ny) mounted to the upright, fixed - stage microscope. with these precautions one limitation of working in retinal slices is that long cellular processes of large field retinal neurons may lose many of their dendrites during the slicing procedure. retinal slice preparations are therefore more useful for studying the physiology of cells in which the synaptic contacts involve processes close to the cell body. amphibian and mammalian retinas share many of the same cell types and utilize similar physiological mechanisms. while salamander retina is a good model for many aspects of mammalian retina, one important difference appears to be the presence of a dedicated rod pathway in mammals that involves contacts of specialized rod bipolar cells onto aii amacrine cells. an additional limitation of the salamander retina is the small number of genetic tools developed specifically for this species. however, antibodies and shrna reagents that target well - conserved regions in other species can be used successfully in salamander, as can many small molecule inhibitors and peptide reagents. additionally, with a few modifications in technique, retinal slices can be prepared from other species in which some of these tools are more readily available. beyond its utility for paired whole - cell recording, the salamander retinal slice preparation is also amenable to a variety of other approaches. as discussed above, retinal slices can be used to study light responses in combination with various voltage clamp protocols. retinal neurons can also be loaded with fluorescent dyes sensitive to ca, cl, or na introduced through the patch pipette or by bath - application. a fluorescent peptide that binds to the synaptic ribbon can be introduced through the patch pipette and used for imaging the ribbon or, when conjugated to fluorescein, for acutely and selectively damaging the ribbon. we have also used retinal slices in combination with quantum dots to monitor the movements of individual calcium channels at rod and cone synaptic terminals. thus, the vertical retinal slice is a versatile experimental preparation for studying basic synaptic mechanisms and the unique processing functions performed at the first synapse in the visual signaling pathway. | one of the central tasks in retinal neuroscience is to understand the circuitry of retinal neurons and how those connections are responsible for shaping the signals transmitted to the brain. photons are detected in the retina by rod and cone photoreceptors, which convert that energy into an electrical signal, transmitting it to other retinal neurons, where it is processed and communicated to central targets in the brain via the optic nerve. important early insights into retinal circuitry and visual processing came from the histological studies of cajal1,2 and, later, from electrophysiological recordings of the spiking activity of retinal ganglion cells - the output cells of the retina3,4.a detailed understanding of visual processing in the retina requires an understanding of the signaling at each step in the pathway from photoreceptor to retinal ganglion cell. however, many retinal cell types are buried deep in the tissue and therefore relatively inaccessible for electrophysiological recording. this limitation can be overcome by working with vertical slices, in which cells residing within each of the retinal layers are clearly visible and accessible for electrophysiological recording.here, we describe a method for making vertical sections of retinas from larval tiger salamanders (ambystoma tigrinum). while this preparation was originally developed for recordings with sharp microelectrodes5,6, we describe a method for dual whole - cell voltage clamp recordings from photoreceptors and second - order horizontal and bipolar cells in which we manipulate the photoreceptor 's membrane potential while simultaneously recording post - synaptic responses in horizontal or bipolar cells. the photoreceptors of the tiger salamander are considerably larger than those of mammalian species, making this an ideal preparation in which to undertake this technically challenging experimental approach. these experiments are described with an eye toward probing the signaling properties of the synaptic ribbon - a specialized synaptic structure found in a only a handful of neurons, including rod and cone photoreceptors, that is well suited for maintaining a high rate of tonic neurotransmitter release7,8 - and how it contributes to the unique signaling properties of this first retinal synapse. |
. high rate of spontaneous remissions is known in patients with early disease that present constellation of specific features called lfgren 's syndrome. on the other hand, an insidious onset of the disease without lfgren 's syndrome sarcoidosis progression has not been associated with any particularly specific immunological parameters although sarcoidosis inflammation is generally characterized by elevated production of proteolytic enzymes, cytokines and chemokine ligands / receptors, and other molecules with immune - regulatory functions [1, 2 ]. expression of these proinflammatory factors is modulated by mirnome that composes of numerous small single - stranded (2024 nucleotides long) noncoding rnas termed mirnas (alias micrornas). they bind to complementary mrna sequences within target genes whose expression is subsequently regulated through posttranscriptional repression. the regulatory property of mirnas is generally associated with their cytoplasmic accumulation in a cell. in pulmonary sarcoidosis, the intracellular expression of mirnas has been investigated in bronchoalveolar cells, peripheral blood mononuclear cells, and the lung tissue [46 ]. in addition, some intracellular mirnas have been reported to have altered expression during sarcoidosis progression [4, 5 ]. besides their intracellular accumulation, however, mirnas are known to be present in extracellular fluids [79 ]. in addition, the sparse data should be confirmed with regard to current recommendation for normalisation strategy of extracellular mirnas [7, 10 ]. we have therefore investigated expression stability of serum mirnas in sarcoidosis and then compared serum expression of mirnas in sarcoidosis patients classified according to the presence of lfgren 's syndrome as a hallmark of good prognosis in comparison to group of patients with more advanced disease course. serum samples were obtained from 13 healthy controls and 24 patients with pulmonary sarcoidosis according to a standard protocol. diagnosis of pulmonary sarcoidosis was made according to the criteria of ats / ers / wasog international consensus statement. lfgren 's syndrome was characterized by erythema nodosum, bilateral hilar lymphadenopathy, fever, and polyarthritis. all patients with lfgren 's syndrome had chest x - ray (cxr) stage 1 (n = 12) and all patients without lfgren 's syndrome had cxr stage 3 (n = 12). clinical characteristics and bronchoalveolar cellular profile are provided in tables 1 and 2, respectively. all patients were recruited at the department of respiratory medicine and tbc, university hospital in olomouc, the czech republic. the study was performed with the approval of ethical committees of medical faculty pu & university hospital, olomouc. informed consent for the anonymous usage of all serum samples for the purposes of the study was obtained from all enrolled subjects. to ensure sample quality without erythrocyte mirnas contamination, the level of haemolysis in all serum samples was assessed by spectrophotometry (nanodrop 1000, usa). extracellular rna was isolated from 300 l of serum by using mircury rna isolation kit for biofluids (exiqon, denmark). pcr conditions and all reaction mixes were adopted from the user bulletin by applied biosystems (protocol for creating custom rt and preamplification pools using taqman microrna assays). briefly, multiplex reverse transcription (rt) was performed with taqman microrna reverse transcription kit (applied biosystems, ca, usa) and rt primer pool prepared by mixing 5x rt primers provided in taqman microrna assays (applied biosystems, ca, usa). to preamplify all tested mirnas together, taqman preamp master mix was used with preamp primer pool prepared from 20x taqman microrna assay (applied biosystems, ca, usa). qpcrbio probe mix no - rox (pcr biosystems, united kingdom) and 20x taqman microrna assays (applied biosystems, ca, usa) were used to perform rt - pcr of individual mirnas (rotorgene3000 system corbett research, sydney, australia). all taqman microrna assays are listed in supplementary material (online resource 1 in supplementary material available online at http://dx.doi.org/10.1155/2016/1246129). second derivative method, described previously in our laboratory, was used to assess cq and efficiency of rt - pcr. the particular mirnas were selected based on early published screening data in sarcoidosis [7, 15 ] and our pilot qpcr data that were partly published at european respiratory society congress in 2015. only mirnas with efficiency within 1.72.0 (27 mirnas) were utilised for the subsequent analyses (supplementary material / online resource 1). genorm and normfinder algorithms were used to reveal a normalisation factor with the highest stability [18, 19 ]. whitney u test was used to detect possible differences in relative expressions of mirnas between the study groups. to account for a high false - positive rate possibly caused by a multiple comparison among 3 study groups, correction of p value was performed by using false discovery rate (fdr) method according to benjamini and hochberg where desired fdr equaled 5% and a p value 0.03 was considered to be significant. multivariate analysis was performed with simca p version 13.5.0 (umetrics, ab, ume, sweden) by using principle component analysis (pca) and orthogonal projections to latent structures (opls) analysis. analyses were performed on log2-transformed, quantile - normalised, mean - centered data scaled to unit variance. model performance is reported as cumulative correlation coefficient for the model (r), predictive performance based on 7-fold cross - validation (q), and cross - validated anova (cv - anova) p values for opls - based group separation. 20150312) was used to assess the possible cumulative effect of the dysregulated mirnas on gene expression in sarcoidosis. the fold changes of means (patients / controls) of the altered mirnas were used as an input, including kyoto encyclopedia of genes and genomes (kegg) pathways and an observed - to - expected ratio of greater than 1.0. to balance the reliability of the predictions with a manageable number of records, a number of algorithms predicting the same mirna - gene interaction pair were set at three algorithm hits, including validated mirna - gene pairs. only biological functions / pathways with at least 25 genes and at most 500 genes were analysed. to increase reliability of qpcr data on extracellular mirnas, several normalisation strategies were tested before own statistical analysis comparing possible differences among study groups. briefly, both normfinder and genorm algorithms consistently showed a geometric mean of 23 extracellular mirnas that were expressed in all subjects, to have the lowest expression variability within all samples. in comparison to healthy controls, we consistently observed the increased expressions of mir-146a-5p and mir-16 - 5p and decreased expressions of mir-425 - 5p and mir-93 - 5p in both groups of our sarcoidosis patients with / without lfgren 's syndrome (figures 1(a)1(d)). serum expressions of three mirnas (mir-150 - 5p, mir-1, and mir-212 - 3p) were decreased in our patients without lfgren 's syndrome compared to healthy controls (figures 2(a)2(c)). serum mir-21 - 5p was increased in our patients with lfgren 's syndrome compared to healthy controls. by contrast, mir-340 - 5p was decreased in the same patients with lfgren 's syndrome compared to healthy controls (figures 3(a) and 3(b)). the patients without lfgren 's syndrome had decreased expressions of mir-212 - 3p and mir-21 - 5p and increased expression of mir-340 - 5p in comparison with those with lfgren 's syndrome (figures 2(c), 3(a), and 3(b)). multivariate analysis was performed to investigate an effect of coexpression of several dysregulated serum mirnas in sarcoidosis (supplementary material / online resource 2). both multivariate analyses of 6 and 7 mirnas that were dysregulated either in the patients with lfgren 's syndrome or in the patients without lfgren 's syndrome showed consistently that opls modelling provides a significant separation between our patients with pulmonary sarcoidosis irrespective of disease course and the healthy controls resulting in the predictive power of 72% and 65% (r = 0.737 and r = 0.691702 and q = 0.717 and q = 0.652 ; p = 9.2 10 and p = 9.16 10) based on 7-fold cross - validation (supplementary material / online resource 2). the multivariate modelling with three mirnas that were dysregulated in the univariate analysis between our patients with lfgren 's syndrome and those without lfgren 's syndrome showed a significant separation with a poor reproducibility in our training data set resulting in the poor predictive power of 44% (r = 0.536 and q = 0.443 ; p = 2.2 10 ; supplementary material / online resource 2). to reveal cumulative effect of the dysregulated mirnas on gene expression, pathway analysis with mirsystem database the pathways in cancer was consistently predicted to be targeted with the highest mirsystem score by both expression profiles that were dysregulated in the patients with / without lfgren 's syndrome compared to healthy controls (p = 5.0 10 and p = 9.0 10 ; supplementary material / online resource 3). in this pathway, all 6 mirnas (mir-146a-5p, mir-16 - 5p, mir-425 - 5p, mir-425 - 5p, mir-21 - 5p, and mir-340 - 5p) that were dysregulated in the patients with lfgren 's syndrome were predicted to modulate 103 target genes based on experimental validation according mirsystem (table 3). further, all 7 mirnas (mir-146a-5p, mir-16 - 5p, mir-425 - 5p, mir-425 - 5p, mir-150 - 5p, mir-1, and mir-212 - 3p) that were dysregulated in the patients without lfgren 's syndrome were predicted to modulate 112 target genes based on experimental validation according mirsystem (table 3). the transforming growth factor (tgf)- signalling pathway with its highest mirsystem score among kegg pathways was predicted to be significantly affected (p = 1.9 10 ; supplementary material / online resource 3) by the cumulative effect of three mirnas whose serum expressions were dysregulated between our patients with lfgren 's syndrome and those without lfgren 's syndrome. in this pathway, three mirnas (mir-340 - 5p, mir-212 - 3p, and mir-21 - 5p) targeted 25 experimentally validated genes (table 3) this work attempts to provide an insight into differential expression of extracellular mirnas in sarcoidosis patients classified according to the presence of lfgren 's syndrome as a hallmark of good prognosis in comparison to advanced disease course. our geometric mean - normalised expression showed that serum mir-146a-5p, mir-16 - 5p, mir-425 - 5p, and mir-93 - 5p are consistently dysregulated, regardless of sarcoidosis prognosis, in our patients with pulmonary sarcoidosis compared to healthy controls. specifically, patients without lfgren 's syndrome had dysregulated expressions of mir-150 - 5p, mir-1, and mir-212 - 3p and those with lfgren 's syndrome had dysregulated mir-21 - 5p and mir-340 - 5p in comparison to healthy controls. mirsystem predicted the pathways in cancer to be consistently affected by both of the dysregulated expression profiles in sarcoidosis with / without lfgren 's syndrome. three serum mirnas (mir-21 - 5p, mir-340 - 5p, and mir-212 - 3p) differed between the sarcoidosis patients with lfgren 's syndrome and those without lfgren 's syndrome. their cumulative effect may modulate the transforming growth factor (tgf)- signalling pathway during sarcoidosis progression. because of a lack of current knowledge on stably expressed extracellular mirnas in serum from the patients with sarcoidosis, the best approach of qpcr data normalisation was investigated at the earliest. a geometric mean of all expressed extracellular mirnas showed the best stability and was therefore used to normalise the raw qpcr data in subsequent analysis comparing the study groups. the geometric mean - normalisation is different from that used by jazwa. who measured like our paper serum expressions of mir-16 - 5p, mir-146a-5p, and mir-150 - 5p in sarcoidosis patients. they did not find any of these serum mirnas to be dysregulated in pulmonary sarcoidosis. it is in contrast with our observations and could be explain by the different normalisation approach in combination with different representation of cxr stages among the sarcoidosis patients in the study by jazwa.. in comparison with our healthy controls, dysregulation of 4 mirnas however, we also observed several mirnas to be associated with either the presence of lfgren 's syndrome or its absence. through these differences, the pathways in cancer was consistently predicted to be modulated by cumulative effect of both of the serum expressions profiles in pulmonary sarcoidosis with / without lfgren 's syndrome. remarkably, an increased risk of cancer is discussed in sarcoidosis patients and the presence of sarcoidosis granuloma has been reported in case studies on oncology patients [2426 ]. the dysregulation of serum mirnas in our sarcoidosis patients may therefore result from the inflammation, apoptosis, and angiogenesis frequently accompanying various malignant processes [2830 ] and it likely may not be directly related to presence / absence of lfgren 's syndrome in sarcoidosis. in addition, several particular target genes of the pathways in cancer have been indeed indicated in wet laboratory to be dysregulated at their protein and/or mrna level in sarcoidosis [3136 ]. among them, wnt (wingless and integrase-1)7a, catenin - beta, and transforming growth factor- (tgf-) are concurrently involved into other signalling pathways, including the wnt and tgf- signalling pathways. both of these signalling pathways were already once predicted to be modulated by cumulative effect of several intracellular mirnas that are dysregulated in the peripheral blood lymphocytes and the lung tissue obtained from sarcoidosis patients. in line with the intracellular mirnas - based prediction, the tgf- signalling pathway was predicted here to be affected by the extracellular serum mirnas (mir-21 - 5p, mir-340 - 5p, and mir-212 - 3p) that differed between our patients with lfgren 's syndrome and those without lfgren 's syndrome. taking into consideration a profibrotic character of tgf- action, it is notable that the tgf- signalling pathway was predicted here although only two patients with fibrotic changes (cxr stage iv) were enrolled in this study. besides them, we may therefore speculate on an incipient fibrotic process that involves posttranscriptional regulation beginning before cxr stage iv. on the other hand, our patients with cxr stage 3 did not have dysregulated expression of mir-21 whose elevation has been associated with ipf. thus, comparison among all 5 cxr stages needs to be elucidated to gain a whole insight into the disease course ranging from invisible abnormality of the intrathoracic lymph nodes toward the lung parenchymal involvement and lung fibrosis in the most advanced sarcoidosis. this could even reveal the key regulatory player leading to the disease remission that is known to be common (5590%) in early disease (namely, in lfgren 's syndrome) whereas rare (1020%) and absent in cxr stage iii and iv, respectively. however, three mirnas (mir-21 - 5p, mir-340 - 5p, and mir-212 - 3p) did not provide good model to separate our sarcoidosis patients according to the presence / absence of lfgren 's syndrome. the lack of profound differences between two groups of our patients with sarcoidosis is in line with current poor knowledge on any sufficiently sensitive and specific serum profile for the disease course. this seems not to stand for genetic background as several genetic variants have been reported to be associated with lfgren 's syndrome [4042 ]. it should be noted that some mirnas with plausible relevance for sarcoidosis pathogenesis were not investigated and this could represent a limitation of our study. our multiplex qpcr method did not also allow us to perform any high - throughput screening, although we assessed higher number of serum mirna than a previous study on serum mirnas in sarcoidosis. our selection was based on the current knowledge on intracellular mirnas in pulmonary sarcoidosis [6, 15 ]. taking into consideration different biological properties, for example, the broadly discussed processing during cell - cell communication, intracellular mirnas are unlikely to be dysregulated in parallel with their intracellular expression. thus, a high - throughput screening for extracellular mirnas may reveal new serum biomarkers of sarcoidosis and the disease prognosis. in conclusion, we report several serum mirnas to be associated with pulmonary sarcoidosis and also further differences between our sarcoidosis patients stratified according to the presence / absence of lfgren 's syndrome as a hallmark of good prognosis. in an attempt to link the serum mirnas to certain biological processes using bioinformatics tools, the pathways in cancer was predicted to be related to pulmonary sarcoidosis as a whole whereas the tgf - beta signalling pathway was predicted to be related to the disease course. the complex interplay between the serum mirnas and the predicted target genes of these signalling pathways remains the matter of future experimental investigation to gain detailed insight into the pathological mechanisms underlying the disease and its advancement. the work was supported by grant projects lo1304, cz.1.07/2.3.00/30.0004, and iga pu lf 2015_030, 2016_009. | purpose. pulmonary sarcoidosis is associated with dysregulated expression of intracellular mirnas. there is however only little information on extracellular mirnas and their association with the disease course in sarcoidosis. we therefore assessed serum mirnas in sarcoidosis classified according to the presence of lfgren 's syndrome (ls) as a hallmark of good prognosis in contrast to more advanced disease course. methods. rt - pcr was used to assess 35 mirnas in 13 healthy controls and 24 sarcoidosis patients (12 with x - ray (cxr) stage 1 and ls and 12 with insidious onset and cxr stage 3). results. compared to controls, we consistently observed dysregulated expressions of mir-146, mir-16, mir-425 - 5p, and mir-93 - 5p in both sarcoidosis groups irrespective of disease course. specifically, patients without ls had dysregulated expressions of mir-150 - 5p, mir-1, and mir-212 compared to controls. patients with ls had dysregulated expressions of mir-21 - 5p and mir-340 - 5p compared to controls. bioinformatics predicted consistently pathways in cancer to be modulated by both altered profiles in patients with / without ls. three mirnas (mir-21 - 5p, mir-340 - 5p, and mir-212 - 3p) differed between our patients with ls and those without ls ; their cumulative effect may modulate tgf- signalling pathway. conclusions. further study should focus on possible applications of serum mirnas for diagnostics follow - up and for prognosis. |
thyroid cancer accounts for 12% of all malignancies, representing the most common endocrine malignancy. papillary carcinoma (ptc) is the most frequent among thyroid malignancies, whereas medullary thyroid carcinoma (mtc) accounts for 510% of cases. mtc, on the other hand, originates from parafollicular cells of the ultimobranchial body, derived from the fourth pharyngeal pouch. hereditary mtc can manifest either alone as familial mtc or as part of the multiple endocrine neoplasia type 2 syndromes (men 2). men 2 involves medullary carcinoma and pheochromocytoma with either primary hyperparathyroidism (men 2a) or marfanoid habitus and mucosal neurofibromatosis (men 2b). concurrence of medullary and papillary thyroid carcinomas is an unusual finding. in this report, a case of multifocal thyroid cancer involving two foci of medullary and two foci of papillary microcarcinomas is presented. a 39 year - old female presented for otorhinolaryngologic evaluation at the outpatient clinic of a tertiary referral center (department of otorhinolaryngology, university hospital of heraklion, greece). the examination had been performed as part of a routine assessment for reported difficulty in swallowing and neck discomfort complaints by the patient. it had revealed slightly enlarged thyroid gland lobes, with presence of two nodules in the right lobe of the gland (maximum diameter 0.3 cm and 0.8 cm, respectively) and two nodules in the left lobe (maximum diameter 0.4 cm and 0.6 cm, respectively). ultrasound elastography had been used to complete the imaging study and revealed suspicious findings, with an elastography score of 4 in one, and 5 in all other nodules. patient denied any past exposure to radiation as well as any family history of endocrine disorders. clinical examination of the head and neck was unremarkable, and no masses were noted on palpation of the neck. serum levels of calcium, thyroid stimulating hormone (tsh), free thyroxine and thyroglobulin as well as parathyroid hormone were within normal range. calcitonin levels, on the other hand, were increased (86 pg / ml ; normal values : < 12 pg / ml in women). abdominal ultrasound did not show any adrenal pathology and urinary catecholamines and metanephrine levels were within normal limits. ultrasound guided fine - needle aspiration (fna) cytology of the thyroid was performed twice, but was non - diagnostic. this decision was based on increased levels of calcitonin in addition to findings on elastography that were suggestive of thyroid malignancy. a transverse cervical incision was performed, subplatysmal flaps were raised and strap muscles were separated in the midline. both lobes were completely removed after recognition and meticulous dissection of the recurrent laryngeal nerves. postoperative course was uneventful and the patient was discharged from the hospital two days after surgery. permanent histology of the specimen described the presence of two microscopic foci of medullary carcinoma as well as two papillary microcarcinomas. more specifically, a papillary microcarcinoma (diameter : 7 mm) (fig. 1a) and a microscopic medullary carcinoma (diameter : 1.3 mm) (fig. 1b) were found in the right lobe, while hyperplastic c cells were detected next to the tumors. in the left lobe, a 4 mm papillary carcinoma 1c), as well as a microscopic medullary carcinoma with a diameter of 3.5 mm (fig. eleven lymph nodes had been removed, and all were found free of metastasis on histopathological examination. due to multifocality of the papillary carcinoma, the patient underwent treatment with 100 mci of radioactive iodine (i) six weeks after surgery. ten months later, the patient is under tsh suppression treatment with levothyroxine and retains low levels of thyroglobulin and calcitonin. thyroid cancer is the most common endocrine malignancy, accounting for more than 90% of malignancies of the endocrine glands. well - differentiated thyroid cancer, including papillary and follicular carcinomas, remains the most common thyroid malignancy, whereas medullary and anaplastic carcinomas represent 510% and 1.6% of cases, respectively. the most common mutations described in well - differentiated thyroid carcinomas involve the braf gene, and are observed in 3669% of cases. multifocality of both medullary and papillary components is a rare entity, and only two cases have been previously described in the literature. the concurrence of ptc and mtc is an interesting phenomenon as these tumors have different embryological origin. mtc originates from c cells of the ultimobranchial body, derived from the fourth pharyngeal pouch, whereas ptc originates from follicular epithelial cells, derived from median endodermal analogs. several theories for this coexistence have been proposed. according to the stem cell theory, uncommitted stem cells differentiate into both follicular and c - cell lineages. the divergent differentiation theory states that c cells and thyroid follicles are derived from remnants of the ultimobranchial body and solid cell nests. the field effect theory suggests that simultaneous transformation of both follicular and c cells is a result of common neoplastic stimuli. finally, suggests that two independent tumors are located in the same lesion by simple coincidence. cytological examination of material obtained by fna represents the best single diagnostic test for differentiating malignant from benign thyroid lesions. elastography is a newly developed imaging technique that uses ultrasound to estimate tissue stiffness by measuring the degree of distortion under the application of an external force. it is based upon the principle that softer components of tissues deform easier than harder components under compression, thus allowing an objective determination of tissue consistency. elasticity of thyroid nodules is evaluated according to the rago classification system using a scale of 15. score 1 indicates high elasticity in the whole nodule ; score 2 indicates elasticity in a large part of the nodule ; in score 3 elasticity is detected only at the peripheral part of the nodule ; no elasticity in the nodule is detected in score 4 ; score 5 indicates no elasticity in the nodule as well as the posterior shadowing. scores 13 are generally associated with benign lesions, while scores 45 are suggestive of malignancy. ultrasound elastography demonstrates high sensitivity (97%) and specificity (100%) in the diagnosis of thyroid malignancy, with the predictive value of the examination being independent from the nodule size. thus, elastography can be a reliable, noninvasive tool in the differential diagnosis of thyroid nodules. in the case presented here, the decision to perform an fna and furthermore a thyroidectomy was largely based on the suspicious findings of elastography. total or near total thyroidectomy is the preferred procedure for both ptc and mtc, since thyroid lobectomy is associated with a higher risk of recurrence. therapeutic central neck dissection should be performed in cases of pathologic lymph node involvement noted on preoperative clinical and imaging assessment. on the other hand, thus, a prophylactic central neck dissection should be considered in patients with multifocal ptc, as in the case of our patient. development of multiple, small foci of different histological types of thyroid carcinoma is a rare event. elastography can be a useful and noninvasive tool in the differential diagnosis of small thyroid nodules. written informed consent was obtained from the patient for publication of this case report and accompanying images. a copy of the written consent is available for review by the editor - in - chief of this journal on request. | introductiona few cases of concomitant medullary and papillary carcinoma in the same thyroid nodule have been described in the literature. however, the presence of multiple foci of both types of malignancy in the same gland is very rare.presentation of casea 39 year - old female with multiple thyroid nodules, elevated serum calcitonin levels and elastographic findings suggestive of thyroid malignancy, underwent total thyroidectomy and central neck dissection. histology revealed the presence of one focus of medullary and one focus of papillary carcinoma on each thyroid lobe. subsequently, the patient underwent treatment with radioactive iodine.discussionthis is the third case of synchronous multifocal medullary and papillary thyroid carcinoma reported in the literature. several theories for the simultaneous development of these malignant entities have been proposed.conclusionultrasound elastography can be a useful, noninvasive tool in the assessment of thyroid nodules. |
there is no definition of treatment - resistant bipolar depression in the literature. the international society for bipolar disorders (isbd) task force published their consensus definition of treatment response in bipolar depression1 and proposed a nomenclature for relapse, remission, and recurrence, but did not define treatment resistance. treatment response is defined as > 50% improvement in the core dsm (diagnostic and statistical manual of mental disorders) criteria for depression. treatment - resistant bipolar patients remain symptomatic despite the best available care. here, we propose that treatment resistance in bipolar depression be defined as failure of pharmacotherapy associated with failure of neuromodulation, such as electroconvulsive therapy and repetitive transcranial magnetic stimulation (rtms). the antidepressant effects of psychostimulant drugs have been underappreciated because of conflicting data concerning efficacy, tolerance, and dependence. orr and taylor2 reported that methylphenidate reduced tiredness and enhanced attention and arousal, and recommended its use for treating depressive episodes. candy reported that the main advantage of methylphenidate over antidepressants might be its rapid onset of action within one day, and highlighted that, unlike conventional antidepressants, methylphenidate reduces fatigue. stoll and metz and shader5 showed that the combination of methylphenidate and a selective serotonin reuptake inhibitor antidepressant was effective. in a cochrane meta - analysis,3 candy reported that psychostimulants (excluding modafinil) significantly reduced symptoms of depression in comparison with placebo. emphasizing the role of psychostimulants in increasing dopamine production, stahl6 and nierenberg support the view that psychostimulants are preferable for the more biological depressive disorders (melancholic and bipolar depression) but not for the wider spectrum of depressive conditions (especially nonmelancholic disorders). reconsideration of the overall psychostimulant class is likely to be encouraged by the recent introduction of modafinil, a novel psychostimulant that has been studied in both unipolar and bipolar disorders.8 psychiatrists should always remember the potential usefulness of psychostimulants in the treatment of depression based on available evidence. this paper reports the antidepressant effects of methylphenidate, suggesting that this agent might be highly effective and safe, even after neuromodulation had failed, with no withdrawal syndrome when stopped. thus, methylphenidate might be of some considerable utility in managing patients with treatment - resistant depression. we present a case of bipolar depression in which the patient responded significantly to methylphenidate augmentation, without any side effects, after failure of adjunctive rtms and electroconvulsive therapy. mr u, a 56-year - old man with bipolar i disorder according to dsm - fifth edition criteria,9 experienced his first melancholic depression in 1976, which was followed by five melancholic episodes and four manic episodes. he did not show any comorbid psychiatric condition and had no history of drug or alcohol abuse. his sixth melancholic depressive episode began in march 2008, just after a prison guard workmate had committed suicide. he showed a depressed and anxious mood, negative and obsessive thinking, loss of interest in activities, impairment in activities of daily living, psychomotor retardation, self - blaming behavior, guilty and suicidal thoughts, insomnia, reduced appetite, and a 5 kg loss of body weight. his laboratory examinations included a full blood count, blood biochemistry and urinalysis, electrocardiography, electroencephalography, and structural cerebral magnetic resonance imaging. his initial 21-item hamilton depression rating scale10 score was 37 and his clinical global impression - severity11 score was 6. from april 1, 2008 until june 10, 2010, he received various antidepressant (venlafaxine extended- release, clomipramine or duloxetine, combined with either paroxetine or mirtazapine) and mood stabilizer (olanzapine, aripiprazole, or sodium valproate) combinations with no significant clinical improvement. each medication was prescribed at an optimum and stable dosage for the minimum recommended duration, with no significant clinical improvement. on february 6, 2009, we assessed the patient s whole - brain voxel - based regional cerebral blood flow with tc - ethyl cysteinate dimer single photon emission computed tomography, and hypoperfusion was shown in the bilateral anterior cingulate cortices, the right inferior parietal cortex, and the left dorsolateral and bilateral orbital prefrontal cortices. twenty treatment sessions were administered over a 4-week period (five sessions per week). in mid march, after these sessions were completed, mr u showed no significant clinical improvement. rtms was initiated again, with 20 treatment sessions being administered over a 4-week period until april 15, 2009. his physicians initiated electroconvulsive therapy three times a week on april 18, 2009 for 2 months until june 19, 2009. mr u showed no significant clinical improvement after 24 sessions of electroconvulsive therapy. at that time, he was treated with a combination of fluoxetine 80 mg / day, duloxetine 360 mg / day, mirtazapine 60 mg / day, and sodium valproate 1,000 mg / day. on june 29, 2009, we added methylphenidate at a dose of 10 mg / day for one week. two days after introduction of methylphenidate, mr u reported some mood improvement after a single early morning dose. the second week after its initiation, the dose of methylphenidate was increased to 20 mg / day for one week using an extended - release tablet formulation, with significant improvement. two weeks later, mr u achieved significant and lasting clinical improvement, with hamilton depression rating scale and clinical global impression - severity scores of 8 and 3, respectively. he remains well on sodium valproate monotherapy and is regularly followed up at our bipolar department. this case report suggests that methylphenidate may be an effective and safe medication for treating treatment - resistant bipolar depression. methylphenidate was well tolerated with no drug - related switching to hypomania or mania. during the treatment period, improvements in the clinical global impression - severity and hamilton depression rating scale scores indicated clinically significant improvement. our patient s melancholic depression was not improved satisfactorily by the combination of duloxetine, mirtazapine, clomipramine / fluoxetine, and olanzapine / valproate after adjunctive rtms or electroconvulsive therapy. it was not until the patient received methylphenidate 20 mg / day that his severe depression improved substantially and durably, suggesting that adjunctive methylphenidate may have had an additional therapeutic effect in this difficult - to - treat patient. unfortunately, whole - brain regional cerebral blood flow with single photon emission computed tomography was not performed at that time. theoretical use of methylphenidate has been discussed by stahl,12 who suggests that the dopamine - releasing stimulant properties of methylphenidate allow this agent to be used in combination with other antidepressants. stahl suggests that methylphenidate may be particularly useful in patients with retarded or melancholic depression or those who require an antidepressant concomitantly with a mood stabilizer for bipolar depression. while a number of authors24,6 have reported that methylphenidate is effective for treating treatment - resistant bipolar depression, patkar did not demonstrate any statistically significant benefit from augmentation with methylphenidate in treatment - resistant depression in a randomized, double - blind, placebo - controlled trial. theoretical and clinical use of methylphenydate is broadly consistent with our observations in our bipolar department over recent years, ie, that methylphenidate is useful (as monotherapy or augmentation) in patients with melancholic or bipolar depression, including many patients who do not respond to conventional antidepressant drugs or neuromodulation techniques. our case suggests that methylphenidate can be used as an adjunctive agent for some patients with treatment - resistant bipolar depression or melancholia who do not adequately respond to combination antidepressant therapy, a mood stabilizer, and rtms or electroconvulsive therapy. clearly, placebo - controlled studies are warranted within these diagnostic subgroups to test our clinical impressions. such augmentation studies would be advanced by measuring serum levels of the relevant antidepressants to determine whether psychostimulants act by increasing these levels and/or if they have independent antidepressant activity. | adjunctive use of methylphenidate, a central stimulant, has been considered as a potential therapeutic choice for patients with refractory unipolar, geriatric, or bipolar depression, and depression secondary to medical illness. we present a case of bipolar depression in which the patient responded significantly to augmentation with methylphenidate, without any side effects, after failure of adjunctive repetitive transcranial magnetic stimulation and electroconvulsive therapy. mr u, a 56-year - old man with bipolar i disorder, had melancholic symptoms during his sixth episode of bipolar depression. after failure of repetitive transcranial magnetic stimulation and electroconvulsive therapy, he was treated with fluoxetine 80 mg / day, duloxetine 360 mg / day, mirtazapine 60 mg / day, and sodium valproate 1,000 mg / day, with no improvement. we added methylphenidate at a dose of 10 mg / day for one week, which resulted in mild clinical improvement, and then methylphenidate extended - release 20 mg / day for one week, with significant clinical improvement. he tolerated his medications well. his clinical recovery was stable over one year. the patient s antidepressants and methylphenidate were gradually tapered and finally discontinued after one year with no withdrawal syndrome. to date, he remains well on sodium valproate as monotherapy and is being followed up at our bipolar department. this case suggests that methylphenidate augmentation might be a therapeutic option when treating highly treatment - resistant patients with bipolar depression, even if they had not responded to adjunctive neuromodulation. in these clinical situations, physicians might be interested in prescribing methylphenidate because of its efficacy and safety. |
since childers initially reported the laparoscopic surgical management of 2 cases of stage i endometrial cancer, many studies have advocated the advantages of laparoscopically assisted staging surgery (lass) compared with laparotomic staging surgery in women with endometrial cancer. laparoscopic surgical management has comparable clinical outcomes to those of conventional laparotomy ; it results in less pain and bleeding, shorter hospital stay, and faster recovery to normal activities thus improving quality of life. of gynecologic malignancies, endometrial cancer is the third most prevalent following cervical cancer and ovarian cancer in korean women. in recent years, the incidence of endometrial cancer has gradually increased. in the present study, to examine feasibility, effectiveness, and any complications of lass in korean women with endometrial cancer, we reviewed women who underwent lass and compared our operative data results with those of other studies. of 39 women who were diagnosed with endometrial cancer at kangbuk samsung hospital between august 2003 and november 2007, we excluded 4 women who refused to undergo laparoscopic surgery or were ineligible for laparoscopic surgery because of concurrent cardiopulmonary diseases ; a final number of 35 women were enrolled in the present study. the preoperative workup included medical history, physical examination, pelvic examination, gynecologic ultrasonography, pap smear, endometrial biopsy, and mri. the previous abdominal surgical history and body mass index (bmi) did not affect our decision to perform staging surgery in all women. after being informed of the characteristics depending on the extent of surgery, such as complications and their incidence, and the necessity of staging surgery, each woman gave informed consent. the operating time was defined as the period from trocar insertion to closure of the port site. before august 2005, when american college of obstetricians and gynecologists (acog) clinical guidelines for management of endometrial cancer was published, 10 women with endometrial cancer were managed by laparoscopically assisted vaginal hysterectomy (lavh) and bilateral salpingo - oophorectomy (bso) with laparoscopic pelvic lymphadenectomy (lpl). laparoscopic paraaortic lymphadenectomy (lpal) was then added to lavh and bso with lpl after august 2005. intravenous preoperative prophylactic antibiotics were administered : cefminox 2 g or flomocef 1 g if a woman was allergic to cephalosporins. women were placed in a dorsal lithotomy position and given general anesthesia with endotracheal intubation, and the surgical preparation was performed aseptically using povidone - iodine. the port placement system was established through choi 's 4-trocar method, and the intraabdominal pressure was maintained at 15 mm hg. after the entire abdominal and pelvic cavities were thoroughly examined, peritoneal washing cytology was performed ; multiple biopsies were performed for any suspected area in the abdominal cavity. to prevent the retrograde spread of cancer cells through both tubes, the tube containing both round and ovarian ligaments was ligated by the extracorporeal endosuture technique (suture laploop, sejong medical, seoul, korea.). subsequently, a uterine elevator (uterine grasping forceps by sairges, wolf, germany) was inserted. prior to lavh, lpl was systemically performed from the level corresponding to the deep circumflex iliac vein to the level of the ureter crossing the common iliac artery. while lpal was being performed, neither additional trocar insertion nor positional change of the operator or a monitor, or both, was done. lpal was performed to the level of the left renal vein using a dissector with monopolar coagulator and harmonic shears (ultracision harmonic scalpel, ethicon endo - surgery, cincinnati, oh) (figure 1). the dissected lymphatic tissues were safely removed through the opened vaginal vault after lavh with bso. an actual laparoscopic image after laparoscopic transperitoneal lymphadenectomy. a : (1) left external iliac artery, (2) left external iliac vein, (3) left obturator nerve, (4) left ureter. b : (1) right external iliac artery, (2) right external iliac vein, (3) right obturator nerve, (4) right ureter. c : (1) aorta, (2) inferior vena cava, (3) inferior mesenteric artery, (4) left renal vein, (5) right ovarian vein, and (6) left ovarian vein. following the completion of all surgical procedures, drainage (evacuator barovac, sewoon medical, seoul, korea) was inserted after confirming ureteric peristalsis and that no bleeding was occurring at the trocar site or in the abdominal cavity. to minimize the chimney effect, the trocar sites were dressed with povi - done - iodine, and the sites of skin incision were sutured. the student t test was used to compare the number of harvested lymph nodes between the right and left pelvic lymph node. all statistical analyses were performed using sas version 9.1 (sas institute inc., cary, nc, usa). the student t test was used to compare the number of harvested lymph nodes between the right and left pelvic lymph node. all statistical analyses were performed using sas version 9.1 (sas institute inc., cary, nc, usa). the median age and bmi were 57 years (range, 28 to 81) and 25.8 kg / m (range, 20.9 to 37.2). eighteen women (51.4%) had some concomitant medical disease : hypertension in 12 women (34.3%), diabetes in 8 (22.9%), ischemic heart disease in 1 (2.9%), and other medical diseases in 3 (8.6%). fourteen women (40.0%) had a history of previous abdominal surgery : laparoscopic tubal ligation in 8 women (22.9%), cesarean delivery in 5 (14.3%), appendectomy in 2 (5.7%), tubal reversal in 1 (2.9%), and salpingectomy due to tubal pregnancy in 1 (2.9%). operative techniques included lavh with bso in 1 patient (2.9%) who was a 76-year - old woman with parkinson 's disease, cerebrovascular accident, diabetes, and hypertension ; lavh with bso combined with lpl in 9 (25.7%) ; lavh with bso combined with lpl and lpal in 24 (68.6%) ; hysteroscopic tumorectomy with lpl in 1 (2.9%) for fertility conservation ; laparoscopic modified radical vaginal hysterectomy with bso, lpl, and lpal in one (2.9%) with an mri finding that tumor had invaded the uterine cervix. median operating time and estimated blood loss were 150 minutes (range, 95 to 410) and 250ml (range, 50 to 1,000), respectively. intraoperative complications included 1 case (2.9%) of inferior vena cava laceration, in which laparoscopic primary repair was done using a prolene 50 suture. postoperative complications were noted in 3 women (8.6%) : lymphocyst in 1 woman, lymphedema of the right leg in 1, and a high fever (> 38 c) within 48 postoperative hours in one. in these 3 cases, the symptoms improved with conservative therapy. median length of hospital stay was 8 days (range, 3 to 20). the median number of harvested lymph nodes was 22 (range, 10 to 41) in pelvic lymph nodes ; 12 (range, 5 to 21) in right pelvic lymph nodes ; 11 (range, 4 to 21) in left pelvic lymph nodes ; and 7 (range, 2 to 21) in paraaortic lymph nodes. no statistical difference was found in the number of harvested lymph nodes between the right and left pelvic lymph node (p=0.9523). the detailed histopathological results and figo stage are shown in table 1. in postoperative adjuvant treatment, 5 women in stage iiic were treated through chemo - radiation therapy with paclitaxel and carboplatin. ten women without lymph node metastasis beyond stage ib were treated with external radiation therapy or vault radiation. for one woman who underwent hysteroscopic tumorectomy with lpl for fertility conservation, medroxyprogesterone acetate (500 mg / day) histopathological results median follow - up was 22 months (range, 1 to 47). no vault recurrence or port - site metastases distant metastasis was found in the lung of a woman 34 months postoperatively ; palliative chemotherapy is currently in process. since 1988, laparotomic staging surgery via a midline incision has been done in women with endometrial cancer. the staging surgery included total abdominal hysterectomy with bilateral salpingo - oophorectomy, peritoneal washing cytology, the sampling of paraaortic and pelvic lymph nodes, and the dissection. since then, total vaginal hysterectomy has been considered an alternative surgical modality to laparotomy in a high - risk group of women ; however, it has several disadvantages as a surgical approach. laparoscopic staging surgery has overcome these disadvantages by causing less pain, shorter hospital stay, and in addition, providing the ability to inspect the abdominal cavity. a meta - analysis of many studies concerning laparoscopic surgery for endometrial cancer reports the problems due to the differences in the patient groups, surgical methods, surgeons ' high technical expertise, and the extent and frequency of lymph node dissection. table 2 summarizes results from previous studies regarding figo stage, the extent and frequency of lymph node dissection, the number of harvested lymph nodes, complications, operating time, the amount of bleeding, and the length of hospital stay. despite the increased number of patients who underwent lpl and lpal compared with those in the previous studies, no significant differences were found in the incidence of complications, the amount of bleeding, and the median number of harvested lymph nodes. several factors might be involved in determining the operating time, but the following matters contributed to shortening the operating time in our series. korean women with endometrial cancer have lower bmi than do western women. through the 4-trocar method, laparoscopic surgeons can obtain better surgical vision and manipulate instruments easily ; additionally, despite the extended scope of lymph node dissection, no additional insertion of trocars was necessary, and the operator and monitor were not required to move or be moved for lpal. in our series, median bmi was 25.8kg / m (range, 20.9 to 37.2). one woman (2.9%) had a high degree of obesity (bmi>35kg/ m), and 2 women (5.7%) were obese (bmi>30kg / m). kim reported that mean bmi was 23.1kg / m in korean women 20 years of age. according to these authors, women with a high degree of obesity were rare compared with those in a western population. our findings support these reports, and such findings were assumed to shorten the operating time. presumably, this might be due to the structural problems of the health insurance system as well as the tendency of korean women wanting to be discharged after complete recovery of their normal activities. comparisons of previous studies on laparoscopic surgery of endometrial cancer data are presented as median (range) or mean sd. ln = lymph node ; ioc = intraoperative complication ; poc = postoperative complication ; ebl = estimated blood loss. in addition, some controversial issues have been raised regarding the management of endometrial cancer using lass ; these include the intraabdominal dissemination of cancer cells (eg, the increase in positive peritoneal washing cytology), port - site metastasis, and the recurrence of vaginal vault, which are all caused by intraabdominal leakage due to the use of a uterine elevator. to prevent the intraabdominal leakage of cancer cells and port - site metastasis, we ligated both oviducts by using the extracorporeal endosuture technique before insertion of the uterine elevator. after trocar insertion, the trocars were tightly fixed with precaution to assure affixation, and all the extracts were removed via the vaginal route. to date, no cases of port - site metastasis or vaginal vault recurrence have been reported. although continuing a large, prospective, and randomized study is necessary, we believe that lass can be performed without additional morbidity and complications, and is feasible and effective in korean women with endometrial cancer. | background and objective : in recent years, the incidence of endometrial cancer has gradually increased in korea, and the use of laparoscopically assisted staging surgery (lass) is increasing in this field. we conducted this study to evaluate the feasibility of lass in korean women with endometrial cancer.methods:we conducted a retrospective review of 35 korean women with endometrial cancer who were managed laparoscopically.results:the median age and bmi were 57 years (range, 28 to 81) and 25.8 kg / m2 (range, 20.9 to 37.2), respectively. the median operating time, estimated blood loss, and length of hospital stay were, respectively, 150 minutes (range, 95 to 410), 250 ml (range, 50 to 1000), and 8 days (range, 3 to 20). no conversion to laparotomy was noted. the median number of harvested lymph nodes was 22 (range, 10 to 41) in pelvic lymph nodes and 7 (range, 2 to 21) in paraaortic lymph nodes. no vault recurrence or port - site metastasis was noted until the last follow-up.conclusions:lass can be performed without additional morbidity and complications, and might be feasible in korean women with endometrial cancer. |
a literature search was performed up to march 2011 in 13 international databases : african index medicus, anthropology plus, british nursing index and archive, the cochrane library, embase, epoc, medline, pilots, popline, psycinfo, social services abstracts, sociological abstracts, and wholis. it was supplemented with searches of the databases of six international organizations that are engaged in projects regarding fgm / c, the reference lists of relevant reviews and included studies, and communication with experts involved in fgm / c - related work. study designs eligible for inclusion were cross - sectional quantitative studies, qualitative studies, and mixed - methods studies. the population considered in scope was members of communities practicing fgm / c residing in a western country, defined as a country with a culture of european origin (huntington, 1996). the outcome of interest was the practice of fgm / c ; specifically, the studies had to describe participants perspectives and understandings of the factors perpetuating or hindering the continuation of fgm / c. all publication years and languages were acceptable and when considered likely to meet the inclusion criteria, studies were translated to english. unpublished reports and brief and preliminary reports were considered for inclusion on the same basis as published articles. the processes of literature screening, assessment of methodological quality, and data extraction were first done independently by two reviewers. a final decision was agreed upon after discussing whether there was a discrepancy between the two reviewers. for all processes, differences in opinion were few and were resolved through rereading the publications and consensus. in selecting literature, the reviewers read all titles, abstracts, or both resulting from the search process and obtained full text copies of studies considered relevant. next, they read the full texts and determined whether they inclusion criteria. predesigned inclusion forms were used for each screening level. to assess the quality of included studies, the checklist for cross - sectional quantitative studies (nokc) and the critical appraisal skills programme (casp) appraisal tool for qualitative research (www.sph.nhs.uk/what-we-do/public-health-workforce/resources/critical-appraisals-skills-programme) were used. for mixed - methods studies, both the qualitative component and the quantitative component of the study data from the full texts were extracted using a predesigned data recording form. extracted data pertained to study and participant characteristics and descriptive data of factors perpetuating and hindering fgm / c. for the qualitative research papers, study findings were defined to be all of the text considered results or findings in the publications, whether interpretations made by the authors or statements by the participants (sandelowski & barrows, 2003 ; thomas & harden, 2008). all findings in the form of sentences, phrases, or text units dealing with factors perpetuating and hindering fgm / c were copied verbatim onto the data extraction form. in recognition that the analysis method needs to be appropriate to the aim of the evidence synthesis, the systematic review utilized an integrative evidence approach (figure 1). the approach was largely based on published examples and guidelines from the evidence for policy and practice information and co - ordinating centre (eppi centre ; see, e.g., harden., 2004 ; shepherd., briefly, data from cross - sectional survey studies were combined with data from studies that examined participants perspectives of factors perpetuating and hindering fgm / c. the synthesis was aggregative (dixon - woods., 2006) and focused on summarizing data by pooling conceptually similar data from the quantitative studies and the qualitative views studies. first, a synthesis within study types was performed and then a synthesis between study types. throughout the analysis, the quantitative results were used as the analytic point of departure (shown through capitalization in figure 1), such that the qualitative results were subsumed under the quantitative results and were used to extend the results from the quantitative analysis. with respect to the quantitative analysis, the results from each study were categorized according to whether the factors were perpetuating (continuance) or hindering (discontinuance) factors of fgm / c. the reviewers then calculated the frequencies of these factors in order to create a ranked list of factors. in the next step, similar factors perpetuating and hindering fgm / c were grouped, to facilitate the integration of quantitative factors and thematic categories from the qualitative evidence. the analysis of qualitative evidence was thematic ; that is, the reviewers identified prominent or recurring themes in the literature and summarized the findings of the different studies under thematic headings (dixon - woods, agarwal, jones, young, & sutton, 2005). they organized and assigned descriptive codes to the raw data from each study (level 1 findings). next, findings were grouped into thematic categories, based on commonality of meaning as well as frequency and strength of participants cognitions about fgm / c, thereby developing broader concepts that captured similar themes from different papers (level 2 findings). given that the quantitative evidence served as the analytic point of departure, the reviewers worked by using both a priori codes developed from the included quantitative studies to seek out evidence from the qualitative findings, as well as allowing themes to emerge from the qualitative data. in the last qualitative analysis step, categories were combined to create synthesized themes (level 3 findings). this involved reflecting on the thematic categories as a whole and looking for similarities and differences among the categories. the analyses were first conducted individually, and then the reviewers, through discussion and reflection, agreed on a set of categories and analytic themes. in the last analysis step, once both the quantitative and qualitative sets of data were analyzed, they were integrated. the integration involved creating a matrix in which the list of quantitative factors and thematic categories were juxtaposed. the juxtaposition of findings allowed examination of factors and themes that had been investigated, and factors and thematic categories for which there were more credible evidence due to convergence and corroboration. the accumulation of the analyses and the conclusions were summed in a conceptual model that linked the factors and concepts together and delineated the underlying forces perpetuating and halting the practice. further details about the methods and findings are described in berg and colleagues (2010). two records could not be obtained in full text (black women 's health and family support group, 1994 ; sy, 1993) and one study is forthcoming (kaplan - marcusan., the reviewers read 117 full texts and included 21 studies, 15 of which were qualitative investigations, five were quantitative cross - sectional studies, and one was a mixed - methods study (table 1). there were two dissertations (gali, 1997 ; khaja, 2004), three studies were reports submitted to funding agencies (mwangi - powell, 1999, 2001 ; norman, hemmings, hussein, & otoo - oyortey, 2009), and the remaining studies were published in peer - reviewed journals. application of the checklists showed that nine of the studies had low methodological quality, six moderate, and five high methodological quality. the qualitative and quantitative components of the mixed - methods study were assessed separately, and these were judged as high and moderate, respectively. all quantitative studies lacked documentation about whether the measures were reliable and valid, and most of them failed to explain whether the sample was representative of the population. concerning the qualitative studies, several of them failed adequately to describe consideration of the relationship between the researcher and participants, ethical issues, and rigor of data analysis.. description of included studies (n = 21) legend : med = median ; method. adjunct publication for the study is johansen (2006) ; adjunct publications for the study are johnsdotter (2002) and johnsdotter (2007) ; adjunct publications for the study are khaja. (2009 ; 2010). in total, the studies included 1,741 participants (morris did not report the number of participants in the study), of which 78% were women (table 1). the participants were mostly from northern africa and the horn of africa, especially eritrea, ethiopia, somalia, and sudan. currently, the majority of the participants resided in either scandinavia (n = 634) or canada (n = 603), while the remaining reported residency in england, france, new zealand, or the united states. duration of residency in the west varied, but, across the 10 studies reporting duration of residency in western countries, it was about 10 years. most participants appeared to have been in their thirties or forties at the time of the study, and most considered themselves muslim. all but a few of the women had been subjected to fgm / c, typically infibulation. grouped factors perpetuating fgm / c follow : religion, tradition, marriageability, sexual morals, health benefits, male preference, aesthetics, and social pressure. factors hindering fgm / c included the following : negative health issues, negative personal experiences, illegal, there 's no need to do it, not religious requirement, it 's not natural, and husband is against it. there were no studies that investigated only men 's perspectives, but two studies specified views of men separately from women 's : men expressed a preference for a circumcised wife, but some said they did not view fgm / c as a religious requirement and some that they did not think that uncircumcised women were promiscuous. there were 15 qualitative studies and one mixed - methods study with a qualitative component. no qualitative studies examined only men 's perspectives ; thus men 's views were incorporated with women 's views. from the thematic categories, a set of eight analytic themes was produced that most parsimoniously and accurately captured the content and meaning of all the findings. first, in almost all studies, fgm / c was mentioned as a highly meaningful and valued cultural tradition. for example, this is our tradition, it 's something we should do a participant in johnsdotter (2009, p. 129) said. in almost all studies, the participants described enforcement of the norm through community mechanisms, explaining, there are several social pressures and everyone has a say with regards to circumcising, especially from family and friends and the society as a whole nonetheless, due to exposure to western thought models, migration allowed the participants to question doxic cultural models, including those of fgm / c, a reassessment that helped slow the continuation of the practice : participants in berggren and colleagues study (2006) explained, because of migration, they got rid of most of the female peer pressure to continue all forms of fgc the two closely linked analytic themes of sexual morality and marriage were crucial as facilitators of fgm / c. in almost all studies, participants reasoned that fgm / c decreased women 's sexual desires (an uncut woman will run after men and have sex with anyone, said a participant in johansen [2007, p. 248 ]), thus protecting virginity, which was in many communities seen as prerequisite for marriage : people perform fgc to reduce a girl 's sexual desire to preserve her virginity before marriage all but two of the life history interviewees cited religion as a main reason for fgm / c, viewing it as a practice honoring their muslim faith. one participant said, a girl who is not excised is considered as a bad muslim (allag, abboud, mansour, zanardi, & qureux, 2001, p. 2). conversely, religion appeared also as a factor slowing the continuation of fgm / c in that many saw it merely as a religious option, or even in violation of islam : the most important reason for the women involved in our study for being opposed to pharaonic circumcision is that they are convinced that pharaonic circumcision is contrary to basic islamic principles, johnsdotter (2003, p. 99) concluded. fgm / c was seen as ensuring the hygiene of the genitals, which in their natural form were classified as unclean. asked what they thought to be the reasons behind the practice of fgm / c, one woman replied : some say that the girl who is not circumcised has a bad odour because she is not clean down there a last analytic theme looking specifically at the continuance factors was that some perceived womanhood to be accomplished or activated through fgm / c : as long as she hasn't been through it [excision ] she hasn't become a woman (vissandje, kantibo, levine, & n'dejuru, 2003, p. 118). factors hindering the practice constituted two additional analytic themes, most prominently negative consequences of fgm / c, particularly their loss of sexual pleasure. the health consequences were wideranging, as explained by one woman in norman and colleagues (2009) : harmful effects and complications arise from circumcision, especially the pharaonic type, which has a lot of complications emotional, physical, and health problems. sexual intercourse was hard, painful, especially the first few months (khaja, 2004, p. 113). i feel i miss something explained a third (johansen, 2007, p. 268). some women who themselves had experienced complications following fgm / c did not want to expose their daughters to such risks : i do n't want my daughter to pass through all the pain and suffering that i had stated one woman in lundberg and gerezgiher (2008, p. 221). it was clear in many of the studies that most exiled communities respected western countries laws against the practice. almost all women explained how they perceived the swedish law as supporting them in their decision to protect their daughters from fgc, a few additional factors signaled beliefs perpetuating and countering fgm / c, including the following beliefs : fgm / c enhances sexual pleasure (generally for men), being cut is a sign of honor, and the clitoris is dangerous (kleitorid dangereux). quantitative and qualitative data integration identified six key factors perpetuating and four key factors hindering the practice of fgm / c. a conceptual model summarizes the findings (figure 3), showing the most dominant factors, those with more and credible data through convergence and corroboration (marked through capitalization), perpetuating and hindering the continuation of fgm / c. the center represents any member of a practicing community presently residing in a western country, exposed to myriad influences regarding fgm / c. key drivers perpetuating fgm / c are presented in the upper half of the model, with the most influential factor, cultural tradition, most proximal to the center because there were more data for this factor than the other five key factors situated more distally. when asked why fgm / c is performed, in almost all studies, the participants considered it a meaningful cultural tradition, which functioned both as a form of social control and identity for women, as well as a feature of the ideal girl. it was deeply rooted in their social systems, and the compulsory nature of the practice reflected in community mechanisms enforcing it. extensive collective enforcement of the tradition was strongly linked with honor and avoidance of shame, not just for the girl but also the mother and sometimes the extended family. the second key factor, sexual morals, reflected the common view that fgm / c is a cornerstone of moral virtue. fgm / c, especially infibulation, was believed to reduce sexual lust, which was seen as easily aroused and difficult to control thus likely to lead the uncut woman to sexual promiscuity. together with fgm / c, premarital chastity and marital fidelity were seen to function as proof of morality, granting the woman social respect. related, the factor marriageability was frequently found in both the quantitative and qualitative data sets and converged to a significant extent with sexual morals in that premarital virginity served as a guarantee of moral standards. fgm / c in childhood or young adulthood was considered a prerequisite for good marriage later in life. quantitative results showed that men strongly favored a future wife to have fgm / c, and findings from qualitative data indicated that the partiality concerned preference for a moral, faithful wife. as a fourth important factor influencing the continuation of fgm / c, the practice was commonly expressed as a duty according to the religion of islam. respondents from countries where fgm / c is traditionally performed believed that fgm / c is sanctioned by islamic religion. individuals who did not conform to the practice were considered to be acting against their religion and the qur'an. two additional, less influential factors reported in the included studies were health benefits and male sexual enjoyment. the perception that men preferred cut women for their sexual enjoyment was mentioned both in some quantitative and qualitative studies, perpetuating the view among women that men favored women who had been subjected to fgm / c, specifically infibulation, because they gained greater sexual pleasure from a tight vagina. the lower half of the model shows the four key factors identified as hindering the practice of fgm / c. this model area offers a negated reflection of the top area in that three factors perpetuating fgm / c are similarly found to hinder fgm / c, thus demonstrating that fgm / c among exiled communities is a tradition in transition. one factor hindering fgm / c, health issues, was mentioned most prominently in the data sets. the participants were conscious of the harmful consequences following fgm / c, mentioning pain and women 's reduced sexual responsiveness in particular. there was also strong convergence between quantitative and qualitative findings concerning the second and third factors hindering fgm / c. first, most participants knew the illegal status of fgm / c in their western host countries. the law was not just a deterrent but for many was also a support in their decision to abandon fgm / c. second, many participants stated that fgm / c was not an islamic duty and put this forth as an important reason why they would not follow the tradition. the last key factor tempering fgm / c also influenced the first three factors : migration presented individuals in exile exposure to other cultural models, models that opposed fgm / c, thereby allowing sharper scrutiny of the practice. grouped factors perpetuating fgm / c follow : religion, tradition, marriageability, sexual morals, health benefits, male preference, aesthetics, and social pressure. factors hindering fgm / c included the following : negative health issues, negative personal experiences, illegal, there 's no need to do it, not religious requirement, it 's not natural, and husband is against it. there were no studies that investigated only men 's perspectives, but two studies specified views of men separately from women 's : men expressed a preference for a circumcised wife, but some said they did not view fgm / c as a religious requirement and some that they did not think that uncircumcised women were promiscuous. there were 15 qualitative studies and one mixed - methods study with a qualitative component. no qualitative studies examined only men 's perspectives ; thus men 's views were incorporated with women 's views. from the thematic categories, a set of eight analytic themes was produced that most parsimoniously and accurately captured the content and meaning of all the findings. first, in almost all studies, fgm / c was mentioned as a highly meaningful and valued cultural tradition. for example, this is our tradition, it 's something we should do a participant in johnsdotter (2009, p. 129) said. in almost all studies, the participants described enforcement of the norm through community mechanisms, explaining, there are several social pressures and everyone has a say with regards to circumcising, especially from family and friends and the society as a whole nonetheless, due to exposure to western thought models, migration allowed the participants to question doxic cultural models, including those of fgm / c, a reassessment that helped slow the continuation of the practice : participants in berggren and colleagues study (2006) explained, because of migration, they got rid of most of the female peer pressure to continue all forms of fgc the two closely linked analytic themes of sexual morality and marriage were crucial as facilitators of fgm / c. in almost all studies, participants reasoned that fgm / c decreased women 's sexual desires (an uncut woman will run after men and have sex with anyone, said a participant in johansen [2007, p. 248 ]), thus protecting virginity, which was in many communities seen as prerequisite for marriage : people perform fgc to reduce a girl 's sexual desire to preserve her virginity before marriage all but two of the life history interviewees cited religion as a main reason for fgm / c, viewing it as a practice honoring their muslim faith. one participant said, a girl who is not excised is considered as a bad muslim (allag, abboud, mansour, zanardi, & qureux, 2001, p. 2). conversely, religion appeared also as a factor slowing the continuation of fgm / c in that many saw it merely as a religious option, or even in violation of islam : the most important reason for the women involved in our study for being opposed to pharaonic circumcision is that they are convinced that pharaonic circumcision is contrary to basic islamic principles, johnsdotter (2003, p. 99) concluded. fgm / c was seen as ensuring the hygiene of the genitals, which in their natural form were classified as unclean. asked what they thought to be the reasons behind the practice of fgm / c, one woman replied : some say that the girl who is not circumcised has a bad odour because she is not clean down there a last analytic theme looking specifically at the continuance factors was that some perceived womanhood to be accomplished or activated through fgm / c : as long as she hasn't been through it [excision ] she hasn't become a woman (vissandje, kantibo, levine, & n'dejuru, 2003, p. 118). factors hindering the practice constituted two additional analytic themes, most prominently negative consequences of fgm / c, particularly their loss of sexual pleasure. the health consequences were wideranging, as explained by one woman in norman and colleagues (2009) : harmful effects and complications arise from circumcision, especially the pharaonic type, which has a lot of complications emotional, physical, and health problems. sexual intercourse was hard, painful, especially the first few months (khaja, 2004, p. 113). i feel i miss something explained a third (johansen, 2007, p. 268). some women who themselves had experienced complications following fgm / c did not want to expose their daughters to such risks : i do n't want my daughter to pass through all the pain and suffering that i had stated one woman in lundberg and gerezgiher (2008, p. 221). it was clear in many of the studies that most exiled communities respected western countries laws against the practice. almost all women explained how they perceived the swedish law as supporting them in their decision to protect their daughters from fgc, a few additional factors signaled beliefs perpetuating and countering fgm / c, including the following beliefs : fgm / c enhances sexual pleasure (generally for men), being cut is a sign of honor, and the clitoris is dangerous (kleitorid dangereux). integration of data. in the last analysis step, quantitative and qualitative data integration identified six key factors perpetuating and four key factors hindering the practice of fgm / c. a conceptual model summarizes the findings (figure 3), showing the most dominant factors, those with more and credible data through convergence and corroboration (marked through capitalization), perpetuating and hindering the continuation of fgm / c. the center represents any member of a practicing community presently residing in a western country, exposed to myriad influences regarding fgm / c. key drivers perpetuating fgm / c are presented in the upper half of the model, with the most influential factor, cultural tradition, most proximal to the center because there were more data for this factor than the other five key factors situated more distally. when asked why fgm / c is performed, in almost all studies, the participants considered it a meaningful cultural tradition, which functioned both as a form of social control and identity for women, as well as a feature of the ideal girl. it was deeply rooted in their social systems, and the compulsory nature of the practice reflected in community mechanisms enforcing it. extensive collective enforcement of the tradition was strongly linked with honor and avoidance of shame, not just for the girl but also the mother and sometimes the extended family. the second key factor, sexual morals, reflected the common view that fgm / c is a cornerstone of moral virtue. fgm / c, especially infibulation, was believed to reduce sexual lust, which was seen as easily aroused and difficult to control thus likely to lead the uncut woman to sexual promiscuity. together with fgm / c, premarital chastity and marital fidelity were seen to function as proof of morality, granting the woman social respect. related, the factor marriageability was frequently found in both the quantitative and qualitative data sets and converged to a significant extent with sexual morals in that premarital virginity served as a guarantee of moral standards. fgm / c in childhood or young adulthood was considered a prerequisite for good marriage later in life. quantitative results showed that men strongly favored a future wife to have fgm / c, and findings from qualitative data indicated that the partiality concerned preference for a moral, faithful wife. as a fourth important factor influencing the continuation of fgm / c, the practice was commonly expressed as a duty according to the religion of islam. respondents from countries where fgm / c is traditionally performed believed that fgm / c is sanctioned by islamic religion. individuals who did not conform to the practice were considered to be acting against their religion and the qur'an. two additional, less influential factors reported in the included studies were health benefits and male sexual enjoyment. the perception that men preferred cut women for their sexual enjoyment was mentioned both in some quantitative and qualitative studies, perpetuating the view among women that men favored women who had been subjected to fgm / c, specifically infibulation, because they gained greater sexual pleasure from a tight vagina. the lower half of the model shows the four key factors identified as hindering the practice of fgm / c. this model area offers a negated reflection of the top area in that three factors perpetuating fgm / c are similarly found to hinder fgm / c, thus demonstrating that fgm / c among exiled communities is a tradition in transition. one factor hindering fgm / c, health issues, was mentioned most prominently in the data sets. the participants were conscious of the harmful consequences following fgm / c, mentioning pain and women 's reduced sexual responsiveness in particular. there was also strong convergence between quantitative and qualitative findings concerning the second and third factors hindering fgm / c. first, most participants knew the illegal status of fgm / c in their western host countries. the law was not just a deterrent but for many was also a support in their decision to abandon fgm / c. second, many participants stated that fgm / c was not an islamic duty and put this forth as an important reason why they would not follow the tradition. the last key factor tempering fgm / c also influenced the first three factors : migration presented individuals in exile exposure to other cultural models, models that opposed fgm / c, thereby allowing sharper scrutiny of the practice. using results from this systematic review, the authors show that fgm / c is deeply rooted culturally and held in place by reciprocal expectations within practicing communities social systems. in the included studies, exiled members of practicing communities consistently argued that fgm / c was an essential cultural traditional and so must continue. kleinman (1980) has described culture as an integrated pattern of human knowledge, beliefs, and behaviors as well as a set of shared values and practices that characterize a group. from this description it follows that, as gali (1997) explains, fgm / c is embedded in many cultural systems through multiple ties to historical tradition, tribal affiliation, social status, marriageability, and religion. most of these ties were perceptible in the datasets and will be discussed below. a related way of understanding fgm / c 's continuance at the meso level through culture is by noting how it is culture congruent : according to leininger (1997), the actions and decision for fgm / c are highly meaningful and preserve the valued lifeways of people in the community, whether in the home community or a western host community. or, as related to, for example, the somalis, among whom the practice is near universal (yoder & khan, 2008), the practice has come to occupy an important place in the psyche of the society (nkrumah, 1999). the results are also largely congruent with the who 's mental map of why the practice continues. the who concluded that members of practicing communities held culturally ingrained beliefs about fgm / c, which formed a mental map and largely included psychosexual and social reasons, religion, society, and hygiene and aesthetics (who, 1999). it seems the reasons for fgm / c as viewed by individuals living in home communities are largely the same as those expressed by communities in western exile. the results suggest that factors perpetuating fgm / c form a belief set, in which its value as a cultural tradition takes precedence. it is performed out of cultural conformity and, over time, the practice has developed social significance, signaling people 's sense of identity and respectability as an ideal member of the community. today, including in the context of life in exile, fgm / c continues to be valued with strong support, socially and culturally. in fact, the results of exile communities reasons as expressed in the 1990s and 2000s show that a great deal of pressure is enforced within the communities. accepting its place as a social organizing principle in practicing communities, the socially constructed normalizing mechanisms perceived by exile members are not wholly unexpected. gilette - frenoy (1992) writes that pressure to submit to the practice themselves or subject their daughters to it came from extended family members living in the west and those still residing in their countries of origin. on the one hand, findings showed that girls who undergo fgm / c, and their family, are met with social approval, notably respectability and honor (khaja, 2004 ; vissandje., 2003). on the other hand, in response to failure to conform to fgm / c, social mechanisms included insulting an uncut girl 's mother, teasing uncut girls, denying them social acceptance, and, most significantly, rejecting them as marriage partners (e.g., ahlberg, krantz, lindmark, & warsame, 2004 ; berggren., 2006 ; gali, 1997 ; gilette - frenoy, 1992 ; khaja, 2004 ; morison., 2004 ; vissandje., these findings show the role of fgm / c as a tool in social control. refusing fgm / c would not only introduce the psychological problem of being different, but it also would shrink women 's marriage prospects in their community. it is important to note that in most societies practicing fgm / c, being a wife and mother is of utmost value (johnsdotter, 2002 ; lightfoot - klein, 1989 ; nkrumah, 1999). relatedly, the results showed that the perpetuating factor of marriageability was significantly linked with the ideology of sexual morals. many exile community members considered premarital virginity as a guarantee of moral standards, the fundamental assurance of marriageability. in the included studies and in other reports (e.g., abor, 2006 ; ebong, 1997), a belief that a woman 's sexuality is wanton and therefore must be controlled through fgm / c was expressed. fgm / c is in this respect a means to control the sexuality of women analogous to the iron chastity belts allegedly used in medieval europe. from a physiological perspective, implications of fgm / c on sexual desire and satisfaction have been substantiated (berg & denison, 2012), but the procedure does not ensure virginity, including among infibulated women deinfibulation and reinfibulation can be and are performed (e.g., levine, 1999 ; thierfelder., 2005). albeit not unquestioned (see, e.g., johansen, 2007), the perception remains in the exile environment that through fgm / c, especially infibulations, girls bear witness of moral status and virginity (johansen, 2007 ; johnsdotter, 2002 ; khaja, 2004). as with marriage, it must be recognized that the concept of virginity holds great importance in many practicing communities in that a family 's and indeed the whole wider group 's honor depends on girls chastity (johansen, 2006 ; khaja, 2004). kassamali (1998) writes that in patrilineal societies, family honor is customarily closely associated with women 's sexual behavior. as an example, in sudanese society the greatest measure of a family 's honor is the sexual purity of its women. any transgression on the part of the woman disgraces the whole family writes lightfoot - klein (1989, p. 375). the results showed that the argument of religion coexisted on two levels, reflecting the fact that a true islamic position on fgm / c is impossible to claim, given those involved argue from their own interpretation of the written sources. there are four islamic law schools, of which three regard fgm / c as recommended and one, the shafi'i law school, regards fgm / c as compulsory. each manifests differently, however, in various countries according to sociocultural practices (roald, 2001). for example, fgm / c is virtually nonexistent in several countries that adhere to the shafi'i law school (e.g., palestine, lebanon, syria), but it is almost universal in others (e.g., somalia ; roald, 2001). lightfoot - klein (1989) concluded that fgm / c is not practiced in an overwhelming majority of muslim societies. further, the genesis of fgm / c can not be attributed to islam as the practice was evident in pre - islamic arabia, the middle east, and africa (barstow, 1999 ; giladi, 1997 ; grassivaro & viviani, 1988). what is important is to draw attention to the fact that islamic scholars interpret written sources differently. additionally, while researchers believe that today 's position of the islamic scholars urges muslims practicing fgm / c to adopt the most moderate form of fgm / c, many muslims nevertheless understand clitoridectomy and infibulation to be religious duties (giladi 1997 ; kassamali, 1998). because many parents who consider whether to perform fgm / c on their daughter are illiterate or religious texts are out of reach for them, they listen to imams, who often endorse the practice (gali, 1997). grassivaro and viviani (1988) write that fgm / c to lay people is seen as an authentic way to be religious, a sign of religious devotion. further, linked to the discussion above, while some salafi islamists consider fgm / c as a means to heighten female sexual desire, others regard it as a tool to reduce sexual desire (roald, 2001). it seems that religious faith intersects with culture and sexuality in important ways. as with religion, the findings showed that another consideration of fgm / c coexisted on two levels : fgm / c was seen as conferring health benefits while simultaneous viewed as having adverse health implications. belief in purported benefits of fgm / c in general and hygiene in particular seemed to reflect that respondents found cut female genitals somehow cleaner. conversely, participants were concerned about the intrapersonal level consequences following fgm / c, especially pain and women 's reduced sexual responsiveness. according to leading health organizations, there are no known health benefits to fgm / c (who, 2008), and a recent meta - analysis confirmed that, statistically, a woman who has been subjected to fgm / c is more likely to experience pain during intercourse and reduction in sexual satisfaction and desire than a woman whose genital tissues have not been cut (berg & denison, 2012). concerning men, one of their worries was interpersonal : women 's suffering during intercourse. as one man in johansen 's study (2007) said, how can i enjoy sex when it causes pain to my wife ? in fact, men 's interest in fgm / c ostensibly center on fgm / c 's role in preserving morality and honor, not providing sexual enjoyment (e.g., johansen, 2007 ; johnsdotter, 2002), contrary to what some female respondents suggested (e.g., johnsdotter, 2002) and literature indicates (khalifa, 1994). as presented in the results section, exile communities considered the anti - fgm / c laws in western countries and host society discourse 's rejection of the practice as important macrolevel factors slowing its continuation. since the early 1980s most western countries have instituted legislation as their main fgm / c intervention tool (european parliament, 2004 ; leye & sabbe, 2009), although neither their implementation nor effectiveness have been extensively studied (johansen, bathija, & khanna, 2008 ; unicef, 2005a). while it is possible that the existence of a law in their host country may have influenced the community members responses to questions about whether they would continue the practice, the results suggest positive implications from fgm / c - related legislation. migrating to a new social, political, and cultural context with specific laws seems to have led some to question the normalized practice of fgm / c. in sum, the findings show that like other socially entrenched practices with benefits and sanctions anchored in a broad system of collective behavior, fgm / c derives from a complex belief set, in which reasons are at once ideological, material, and spiritual. as suggested above, important factors materialize at multiple levels : intrapersonal (e.g., health consequences), interpersonal (e.g., sexual enjoyment), meso (e.g., cultural tradition), and macro level (e.g., religion, legislation). despite the grouped presentation of factors, for example, as shown in the conceptual model, some factors are perceived as both fueling and slowing the continuation of the practice. it demonstrates a migrant perspective of living in two worlds, where the multiple contexts and discourses surrounding fgm / c are negotiated and there is a cultural accommodation taking place. it also illustrates that fgm / c among exile communities is a tradition in transition with, in time, a likely transfer of relative weight to discontinuing the practice. in principle, then, the results are in agreement with mackie (1996), who suggests that fgm / c as the natural he explains that fgm / c is a self - enforcing belief, in which the cost of testing it has become so high that it traps people. outside the realm of doxa, however, it seems that people are in a state of transition, which stimulates and enables them to reflect on values of home and host communities. the results suggest that anti - fgm / c laws and actual court cases showing the effects of the law can be used as a deterrent within the communities concerned. the relevance of continuously and consistently informing citizens about the fact that fgm / c is prohibited by law and a human rights violation is indicated. while laws in themselves are not enough, they signal expectations by a government regarding the practice and they can work in a complementary fashion with prevention strategies, such as awareness, and educational intervention approaches by creating enabling environments for change. findings are in agreement with unicef (2005b), suggesting that comprehensive social support mechanisms and awareness raising campaigns may be advantageous. strategies of this kind may foster greater public discussion and reflection, such that previously nondiscussed costs of fgm / c may emerge as people share their experiences. what is crucial is that information, messages, and activities are tailored to their audiences. specifically, the results show that programs can also build upon existing beliefs about detrimental consequences from fgm / c and that the practice is not a religious obligation. the findings indicate advantages in establishing an alliance with religious leaders, who often function as norm authorities (who, 2008). health promotion professionals can also aim to modify or remove continuance factors identified, such as correcting women 's misperceptions regarding male sexual pleasure and informing community members of the greater likelihood of sexual problems with fgm / c. regarding the other factors, findings showed that parents wanting their daughters to be successful in marriage and material opportunity chose the strategy of fgm / c. when migration removes pressure, and alternative options for social and economic survival other than fgm / c seem possible, parents and other community members will consider refraining from fgm / c. as one parent in upvall and colleagues study (2009) stated, if my daughter finishes school, learns how to drive a car, and gets a job, she does n't need a man whether she is circumcised or not. there would be advantages in researching the role of islam in attitudinal change, the informational needs of the various fgm / c communities in western locales, and avenues for effective dissemination of information. groups who seek to encourage communities to discontinue fgm / c need to explore ways to address the belief set that sustains the practice. although the results suggest factors perpetuating and hindering fgm / c are fairly consistent across the many exile communities in the west, to optimally inform prevention efforts research should be done locally because the factors may vary somewhat across locations and time. a strength of this systematic review is the comprehensive and systematic literature search as well as systematic process for identifying and analyzing relevant publications. this type of integrative approach, sometimes referred to as mixed studies review, is an emerging form of literature review in the health sciences (dixon - woods., 2006 ; ploye, gagnon, griffiths, & johnson - lafleur, 2009). such reviews, by consolidating often scattered literature on a defined topic, provide detailed and highly practical understanding of complex health issues (dixon - woods., 2006 ; ploye., the integrated results informed the review 's conclusions and implications for research and practice that are optimally relevant for researchers, practitioners, and policymakers trying to understand fgm / c and behavior change, as well as groups contemplating prevention activities. first, it may be subject to publication bias because it is not always possible to identify, and retrieve, all studies addressing the question of the systematic review. in contrast to effectiveness reviews, however, for synthesis of views studies this is probably a minor problem as it is unlikely that one large additional study would drastically change the results. second, some caution is warranted in interpreting the results because about half of the studies had low methodological quality. third, given the integrative nature of the synthesis it is possible that other review authors would produce a different overall model. the methods for conducting integrative syntheses are evolving, and there is no agreement about which approaches are best for particular types of data or questions (dixon - woods., 2005). last, it is important to recognize that the identified factors are as perceived by exile communities living in western countries in the 1990s and 2000s. relatedly, it was not always clear whether the respondents in the included studies referred to situations in their home community or in their current exile setting. this is likely not problematic because many factors are, as described here, important in both contexts. understandably, this review does not mean to imply that there is unanimity among fgm / c practicing communities the reasons for fgm / c are not everywhere the same what the results show are recurring and dominant factors found in the recent literature involving exile communities in western countries. a strength of this systematic review is the comprehensive and systematic literature search as well as systematic process for identifying and analyzing relevant publications. this type of integrative approach, sometimes referred to as mixed studies review, is an emerging form of literature review in the health sciences (dixon - woods., 2006 ; ploye, gagnon, griffiths, & johnson - lafleur, 2009). such reviews, by consolidating often scattered literature on a defined topic, provide detailed and highly practical understanding of complex health issues (dixon - woods. the integrated results informed the review 's conclusions and implications for research and practice that are optimally relevant for researchers, practitioners, and policymakers trying to understand fgm / c and behavior change, as well as groups contemplating prevention activities. first, it may be subject to publication bias because it is not always possible to identify, and retrieve, all studies addressing the question of the systematic review. in contrast to effectiveness reviews, however, for synthesis of views studies this is probably a minor problem as it is unlikely that one large additional study would drastically change the results. second, some caution is warranted in interpreting the results because about half of the studies had low methodological quality. third, given the integrative nature of the synthesis it is possible that other review authors would produce a different overall model. the methods for conducting integrative syntheses are evolving, and there is no agreement about which approaches are best for particular types of data or questions (dixon - woods., 2005). last, it is important to recognize that the identified factors are as perceived by exile communities living in western countries in the 1990s and 2000s. relatedly, it was not always clear whether the respondents in the included studies referred to situations in their home community or in their current exile setting. this is likely not problematic because many factors are, as described here, important in both contexts. understandably, this review does not mean to imply that there is unanimity among fgm / c practicing communities the reasons for fgm / c are not everywhere the same what the results show are recurring and dominant factors found in the recent literature involving exile communities in western countries. this systematic review summarized 21 empirical studies examining the factors perpetuating and hindering fgm / c as perceived by exile fgm / c communities living in western countries. the integrative evidence synthesis identified that the practice of fgm / c derives from a complex belief set, in which cultural tradition takes precedence within a frame of sexual moral and religious reasons that are sustained through community mechanisms. the results showed that within this intricate web of cultural, social, religious, and medical pretexts for fgm / c, conditions hindering its continuance existed, such as a legal framework and national discourse against fgm / c. illustrated in a conceptual model, the reciprocal and dynamic relationships that these factors formed indicated that among practicing communities now living in a western country, fgm / c is a tradition in transition. | understanding the forces underpinning female genital mutilation / cutting (fgm / c) is a necessary first step to prevent the continuation of a practice that is associated with health complications and human rights violations. to this end, a systematic review of 21 studies was conducted. based on this review, the authors reveal six key factors that underpin fgm / c : cultural tradition, sexual morals, marriageability, religion, health benefits, and male sexual enjoyment. there were four key factors perceived to hinder fgm / c : health consequences, it is not a religious requirement, it is illegal, and the host society discourse rejects fgm / c. the results show that fgm / c appears to be a tradition in transition. |
overweight and obesity have been defined as abnormal or excessive accumulations of fat in the body that may impair health. overweight and obesity results from an energy imbalance in the amount of calories consumed and the amount of calories expended. the rise of overweight and obesity globally has been attributed primarily to a twofold process : (1) a global shift toward a diet richer in caloric sweeteners, animal source foods, and fats, and (2) decreased physical activity patterns due to changes in the nature of work, modes of transportation, and urbanization [2, 3 ]. overweight and obesity have been found to be major risk factors for chronic diseases, including cardiovascular disease, diabetes, musculoskeletal disorders, and some forms of cancer. although higher levels of overweight and obesity and, by extension, chronic diseases have generally characterized developed countries such as the united states, the rates of overweight and obesity are increasing much faster in the developing world [4, 5 ]. undernutrition is due to food intake that is continuously insufficient or poorly absorbed and retained in the body to meet and maintain energy requirements. undernutrition is epidemic in parts of the developing world with its occurrence generally associated with high rates of infectious diseases and has been estimated to contribute to the deaths of 5 - 6 million children under 5 each year. undernutrition affects all age groups and is especially common among the poor and those lacking clean water and good sanitation. although earlier research on global undernutrition often suggested that national economic development would lead progressively to lower levels of undernutrition within developing countries, more recent research has highlighted the cooccurrence of high levels of under- and overnutrition within countries [4, 9, 10 ] this paradoxical phenomenon places tremendous strains on the health systems of developing countries since national governments must confront the increased costs associated with nutrition - related chronic diseases as well as the continued need to address undernutrition and infectious disease [11, 12 ]. assessment of the socio - economic disparities in underweight, overweight, and obesity status globally is important to addressing the problem of chronic disease. knowledge of the worldwide distribution of weight status and how weight status may vary by socio - economic factors can provide baseline information for monitoring global and national socio - economic disparities in weight status and assist policymakers in the allocation of resources. estimates of weight status may come from either direct or self - reported measures of height and weight. direct measures of height and weight tend to provide more accurate prevalence estimates of body mass index (bmi) than self - reported measures since they reduce social desirability biases in reporting. nevertheless, self - reported height and weight measures remain as equally correlated with disease biomarkers as direct measures and have been shown associated with mortality due to cardiovascular and coronary heart disease. self - reported height and weight data thus remain valid for identifying relationships in large epidemiological studies [14, 16 ]. because they are lower in cost, less intrusive to collect. the lower costs of self - reported studies of height and weight are particularly relevant for resource - poor countries where surveillance of infectious disease may have financial priority over the surveillance of population weight status. unique about the whs is that the instruments to measure health outcomes and socio - economic indicators are directly comparable across countries. using these data, this study provides the largest directly comparable cross - national report on the association between socio - economic factors and bmi status available and identifies on a country - by - country basis, the role played by income, sex, and urbanicity in underweight, overweight, and obese status. furthermore, given the whs 's focus on population health surveillance among low - income countries, this study reports data for many developing countries that have until now lacked published information. the long version of the world health survey (whs), a large cross - sectional study, was administered in 51 countries in 2002 - 2003 to assess the global prevalence of behavioral risk factors, mental health, and chronic health conditions [17, 18 ]. the long version of the whs was administered primarily in low - income countries, although several high- and middle - income countries were also included. the whs 's sampling frame covered 100 percent of a country 's eligible population. the target population included any male or female adult aged 18 or over, present in the country and residing in a private household during the survey period. the whs was the first survey designed with explicit attention to cross - national comparability in instrument development. additional information about the whs sample and survey methodology is available on the whs website. participants were asked to report their height in either meters and centimeters or feet and inches and their weight in either kilograms or pounds. heights reported in feet and inches were converted to centimeters and weights reported in pounds were converted to kilos to calculate each person 's body mass index (bmi). bmi was calculated as weight in kilograms divided by height in meters squared (kg / m). bmi values are age - independent and applicable to both sexes [20, 21 ]. the international classification of adult underweight, normal weight, overweight, and obesity was used to group individuals according to bmi : (1) underweight (30.0). to reduce potential reporting biases, observations were dropped if their reported height was less than 122 centimeters (n = 1092) or greater than 211 centimeters (n = 56) ; observations were also dropped if their reported weight was 3 standard deviations above (n = 1359) or 1.5 standard deviations below (n = 7089) the crude sample mean of 63.1 kilograms. application of these exclusion criteria was intended to eliminate the statistically outlying height and weight self reports and resulted in the exclusion of a study sample of respondents with self - reported bmi below 11.3 and above 63.2 household permanent income was measured using the asset - based approach developed by ferguson and colleagues, which has been used in previous cross - national studies of economic status and health in developing countries. this approach assumes that economic status is an unobserved latent variable and is estimated by a random - effects probit model using measures of household ownership of assets (e.g., refrigerator, radio, car, etc.), access to services (e.g., drinking water), and known predictors of income (e.g., age and education). coefficients on the asset variables from the model indicate thresholds on the latent income scale above which households are more likely to own particular assets ; that is, if a household 's estimated permanent income is greater than the asset threshold, there is a greater than.5 probability that they own the item. previous research has shown these estimates of household income to provide valid and reliable, if imperfect, estimates of permanent income. the who provided the income information used in this study. national survey units provided definitions of what constituted urban areas and interviewers recorded urbanicity status at the time of the interview. countries were grouped into world regions based on the who regional office divisions. for this report, with certain exceptions, observations were excluded if they were missing data on sex, age, income, urbanicity, and sampling units. analyses were adjusted for age, which was grouped into six categories : (1) 1829 years old, (2) 3039 years old, (3) 4049 years old, (4) 5059 years old, (5) 6069 years old, and (6) 70 years plus. if the country as a whole was missing data on a particular variable, for example, income, analyses were conducted using the available variables. these instances were as follows : turkey was missing income data ; slovenia was lacking urbanicity information ; guatemala and slovenia were missing sampling information. countrywide and sex - specific country prevalence estimates of weight status were calculated based on who world standard age standardization estimates and weighted according to the sampling design (based on selection probability, nonresponse, and poststratification) with the exception of guatemala and slovenia. crude worldwide prevalence estimates of bmi status based on self - reported height and weight by income quintile were calculated. crude prevalence rates of bmi status are reported for urban and rural residents according to the who region. to examine patterns of bmi status according to sex, income, or urbanicity in the risk of being underweight, overweight, or obese, multivariable multinomial regression analyses with a robust error variance were conducted on a country - by - country basis. normal bmi (18.524.9) was taken as the reference category. for countries with no missing data, relative risk ratios (rrrs) and 95% confidence intervals (95% cis) countries are reported as indicating significant differences with p <.05 using clustered robust standard errors with clusters consisting of the country 's primary sampling units (psus). psu information was missing for guatemala, slovenia, and zambia and, as a result, robust standard errors were used for these countries. table 4 provides a summary report of the countries in which particular relationships were shown for each factor. results of country - by - country analyses are available in supplementary material available online at doi : 10.1155/2010/514674. there were seven countries in which sparse data within certain categories of age and/or income required that the age or income variable take a modified form sample characteristics are displayed in table 1. of the 267,926 whs participants within the 53 countries in this study, 189,258 individuals reported information on their height and weight for an overall response percentage of 74.6% to height and weight items. an additional 9596 individuals were dropped from this study for not meeting height and weight inclusion criteria. in addition, 7037 observations were dropped for missing one or more observations on socio - demographic variables. in cases where the entire country did not provide information on a specific socio - demographic variable (e.g., turkey and income) individuals the study 's final sample size was 172,625 observations, or 64.4% of the original sample. the final sample size of countries as a percentage of the original sample ranged from 6.3% (mali) to 96.3% (china). the overall percentage of males in the sample was 49.4% with a range of 33.3% (slovakia) to 88.0% (mali). mean age was 40.8 years and ranged from 31.6 years (ethiopia) to 52.4 years (croatia). the overall percentage of urban - dwelling respondents was 51.6% with a range from 15.5% (malawi) to 92.5% (russian federation). of the study 's sample, 6.7% was classified as underweight, 25.7% as overweight, and 8.9% as obese. compared to women, the distribution of weight status for men leaned toward lower bmi categories. men had a higher prevalence of underweight (7.2%m versus 6.1%w), normal (60.4%m versus 57.3%w), and overweight status (25.9%m versus 25.5%w). tables 2 and 3 provide sex - specific prevalence figures for each country in the sample. women had a higher prevalence of obese status (11.1%w versus 6.5%m). despite the globally higher prevalence of underweight status among men, statistical analyses showed that they were at higher risk of being underweight in five countries and at lower risk of being underweight in 15 countries. the specific listing of countries within each category is provided in table 4. in terms of overweight status, men were at a higher risk to be overweight in 10 countries and lower risk to be overweight in 13 countries. men were at a higher risk to be obese in two countries and lower risk to be obese in 31 countries. figure 1 provides information on the prevalence of underweight, normal, overweight, and obese status by income quintile. the prevalence of underweight status was lowest (5.8%) and the prevalence of overweight (27.4%) and obese status (9.3%) was highest in the richest income quintile. in 11 countries, individuals in lower income quintiles individuals in lower - income quintiles were less likely to be overweight in 34 countries and more likely to be overweight in three countries. in terms of obese status, lower income quintiles were less likely to be obese in 24 countries and more likely to be obese in two countries. in mexico, data suggested a curvilinear association between income and being either overweight or obese. in spain, lower income individuals were more likely to be overweight or obese than those in the richest income quintile. compared to rural areas, urban areas had the lowest prevalence of underweight status (5.1%u versus 9.3%r) and the highest prevalence of overweight (29.4%u versus 21.2%r) and obesity (10.6%u versus 6.9%r). as shown in figure 2, southeast asia had the highest prevalence of underweight status in rural (18.8%) and urban areas (14.5%) ; europe had the highest prevalence of overweight (32.9%) and obese status (11.9%) in rural areas, while the americas had the highest prevalence of overweight (34.2%) and obesity status (12.9%) in urban areas. within countries, residents of urban areas were at higher risk of being underweight in brazil and at lower risk of being underweight in laos and sri lanka. residents of urban areas were at higher risk of being overweight in four countries and at lower risk in three countries. urban residents were at higher risk of being obese in eight countries and at lower risk of being obese in three countries. the prevalence of underweight, overweight, and obesity varied extensively on a country - by - country basis. underweight had a higher prevalence in southeast asian and asian countries, whereas higher prevalence of overweight and obesity tended to characterize europe and the americas. men tended toward a higher prevalence of overweight, normal, and underweight status while women had a generally higher prevalence of obesity. this finding was expected given studies showing that women generally have higher rates of obesity than men, although men may have higher rates of overweight status. despite the overall lower bmi distribution for men, men were at lower risk for underweight status in 15 countries compared to only five countries where women were at lower risk. men were at higher risk of being obese in only slovakia and spain, although they were at lower risk in 31 countries. within countries, respondents belonging to households with greater income tended to be at lower risk for undernutrition and at higher risk for being overweight or obese. this inverse socioeconomic gradient in overweight and obesity in developing countries has been reported previously [28, 29 ]. household increases in wealth and income in developing and transitional countries have been linked to dietary changes among higher income households. greater wealth has tended historically to be associated with diets richer in animal fats leading to a higher prevalence of overweight and obesity in higher - income groups. despite the overall pattern found in this study, longitudinal research looking at trends over time suggests that lower ses groups in developing countries are increasingly at greater risk for overweight and obesity. at gross national income per capita (gni / c) values of roughly us$2500, lower - income has been reported to become a risk factor for overweight and obesity rather than a protective factor. in terms of year 2000 gross domestic product per capita (gdp / c) at purchasing power parity values, this study observed a cutoff at gdp / c values less than us$8700. below us$8700, lower income groups were at lower risk for obesity and overweight with the exception of chad ; above us$8700, there tended, with the exception of spain, to be no difference between income groups. in mexico, the lowest quintile had a lower risk for overweight and obesity compared to the highest quintile, but two middle income quintiles were at a higher risk of overweight and obesity than the richest quintile. residents of lower income households in spain (the country with the highest gdp / c in this sample) had a higher risk of overweight or obesity. rapid urbanization often results in complex societal changes that can have beneficial and adverse impacts on population health. urbanization has been identified as a key factor fueling the nutrition transition [32, 33 ]. urbanization is accompanied by technological changes that influence activity patterns and levels, increased access to modern media, and greater access and exposure to foods richer in fats, caloric sweeteners, and edible oils across all seasons of the year. urban areas were found in this study to have the lowest prevalence of underweight status and the highest prevalence of overweight and obesity. comparisons of urban and rural differences within world regions showed urbanicity important in all regions except europe. however, as a risk factor associated with bmi status, the results were mixed. countries as diverse as brazil, india, ghana, and myanmar showed residents of urban areas with a higher risk of being underweight, or overweight or obese. on the other hand, countries such as slovakia, malaysia, and spain showed residents of urban areas with a lower risk of being overweight or obese. adjustment for age and income in the regression analyses may have reduced the association of urbanicity with overweight and obesity since urban areas tend to have younger age strata and wealthier households compared to their rural counterparts. direct comparability between this study 's malnutrition prevalence estimates and those of other studies is difficult due to differences in sampling design, instruments, data collection methods (e.g., self - report versus directly measured), and the time period in which the data were collected. for these reasons, there may be variability in the extent to which this study 's prevalence estimates compare to previous reports. for example, this study found that 11.7% of chinese adults reported either overweight or obese ; 18.9% of chinese adults have been previously reported as overweight or obese [20, 36 ] ; 36.6% of brazilian adults were found overweight or obese in this study, whereas brazilian ministries reported 40.6% of its residents overweight or obese. overweight or obese and 2.1% underweight compared to 49% of adults overweight or obese and 2.7% underweight previously reported [20, 37 ]. in south africa, 23.2% of respondents were obese and 5.3% underweight in this study, whereas in 1999 the south african department of health reported 21.6% of adults obese and 8.6% underweight [20, 38 ]. one limitation of the present study is the use of self - reported measures of height and weight instead of direct measures to estimate the prevalence of overweight and obesity status. social desirability may play a role in the amount of bias that self reports introduce into population estimates of bmi. for example, research in the united states and canada has shown a tendency of overreporting of height and underreporting of weight. young college - educated students in thailand were found to underreport weight and overreport height in ways similar to persons in developed countries. correction estimates available from such studies may not however have cross - cultural validity since societies and groups may not share the same norms of what is socially desirable. yet, research has shown self - reported height and weight to be equally correlated to fasting blood glucose, high - density lipoprotein - cholesterol, and systolic blood pressure as direct measures, and related longitudinally to mortality. given the lower survey costs, higher response rates, and its relevance for epidemiological studies, use of self - reported height and weight to estimate the prevalence of obesity, overweight, and underweight will continue to have a place in current research on global malnutrition. underweight, overweight, and obesity are among the leading risk factors in the global burden of disease. knowledge of the socio - economic factors associated with overweight and obesity and how these factors are embedded in specific national contexts can contribute to the development of policies that target these factors. this is important since along with fruit and vegetable consumption, smoking, and physical activity, overweight and obesity constitute one of the major risk factors for chronic diseases. yet, recognition of overweight and obesity as a global pandemic and public health problem for all countries has been slow in coming. establishing international and regional policies that recognize national variability in the factors associated with this global pandemic may contribute to reducing global inequalities in underweight and obesity. | objective. to examine the association between socioeconomic factors and weight status across 53 countries. methods. data are cross - sectional and from the long version of the world health survey (whs). there were 172,625 whs participants who provided self - reported height and weight measures and sociodemographic information. the international classification of adult weight status was used to classify participants by body mass index (bmi) : (1) underweight (30.0). multinomial regression was used in the analyses. results. globally, 6.7% was underweight, 25.7% overweight, and 8.9% obese. underweight status was least (5.8%) and obesity (9.3%) most prevalent in the richest quintile. there was variability between countries, with a tendency for lower - income quintiles to be at increased risk for underweight and reduced risk for obesity. conclusion. international policies may require flexibility in addressing cross - national differences in the socio - economic covariates of bmi status. |
proliferating cell nuclear antigen (pcna) serves as a processivity factor (sliding clamp), encircling dna at sites of replication and repair (15). the subunits come together to form a remarkable ring - like structure with characteristic pseudo 6-fold symmetry and a central hole large enough to accommodate double - stranded dna (dsdna). each domain forms a wedge shaped structure with two -helices packed in an antiparallel arrangement against the face of a -sheet wedge. in total the -sheets forming the outer surface of the ring impart structural integrity to the protein assembly. adjacent domains are packed in a head to tail arrangement, which necessitates a long inter - domain linker passing over the central -sheet. this overall architecture (1,2,4) is remarkably well preserved across bacterial and eukaryotic clamps despite the lack of sequence similarity. pcna is known to interact with many components of the cell 's replication and signaling machinery (5) and in this context could facilitate exchange of dna repair enzymes that recognize a common dna intermediate (6). since the association of pcna and dna is topological in nature, it has not yet been possible to determine a structure of pcna loaded onto dsdna by crystallographic means. therefore, structural knowledge regarding the overall orientation as well as the precise contacts formed between a sliding clamp and the nucleic acid it encircles has been limited. furthermore, the static pcna structures determined to date give little information about possible motions of the clamp that may assist in its translocation along dna. much of the focus of recent structural work has been on determining interactions between pcna surface loops and peptides derived from known pcna binding proteins, such as flap endonuclease-1 (fen-1), dna polymerase and and cyclin - dependent protein kinase inhibitor p21(cip / waf1) (512). since no detailed computational modeling studies have been reported for this system, it would be of general interest to examine the overall conformational flexibility of the sliding clamp, the detailed interactions of residues lining the pcna hole with dna and the motions of the pcna ring, which may promote translocation along dsdna. to integrate existing structural understanding of pcna and to help address key questions regarding structural implications for pcna interactions, we here apply computational analysis to (i) map out the flexible versus rigid regions of pcna and establish their relationship to known binding motifs ; (ii) examine the overall motions of the pcna ring and (iii) establish if dynamical coupling exists between the three identical subunits and if such coupling could be affected by sequential binding. this paper is divided into three major parts plus conclusions and materials and methods sections. first, we trace the time evolution of key interactions between dsdna and basic residues on the inner surface of the sliding clamp and comment on the overall structural characteristics of the dna pcna complex as well as on the observed competition between the equivalent subunits for binding to the nucleic acid phosphodiester backbone. second, we quantify the motions of the pcna ring by carrying out covariance matrix analysis, which exposes the existence of long - range correlations concomitant with changes in dna binding. third, we present results from principal component analysis that complement the previous observations and provide more rigorous identification of distinct states for the pcna two systems were set up for classical molecular dynamics simulation : (i) archaeoglobus fulgidus pcna and (ii) human pcna. the initial structures were taken from the protein data bank (accession nos 1rwz and 1vym, respectively). a sequence of dsdna (5-acgttgactaccgtcttgaggcagagtc-3) in the canonical b - form was generated and inserted vertically through the central hole of the pcna trimer. the models were completed by adding hydrogen atoms, counterions and solvent [pre - equilibrated tip3p (13) water molecules ] with the xleap module of the amber 7.0 program. both cl and na ions were used to neutralize the overall negative charge of the pcna dna complex and mimic physiological conditions with salt concentration of 120 mm. the models were initially minimized for 1000 steps to remove unfavorable contacts, carefully brought to a temperature of 300 k by velocity rescaling and equilibrated for 0.5 ns with the dsdna atoms kept fixed. the production runs were carried out for 25.5 ns for system (i) and 24.5 ns for system (ii) in the isothermal - isobaric ensemble (npt) with pressure and temperature maintained constant at 1 atm and 300 k, respectively. the long - range electrostatic interactions were treated using the smooth particle mesh ewald (spme) algorithm (14). for the non - bonded short - range interactions, we employed a cutoff of 12 with a switching function between 10 and 12. constraining the bond lengths between hydrogen and heavy atoms to their equilibrium values with the shake algorithm (15) eliminated the fastest vibrational motions and thus, permitted an increased integration time step of 2 fs, while maintaining control over the conserved quantities. the r - respa multiple time step method (16) was employed with a 2 fs time step for bonded, 2 fs for short - range, non - bonded interactions, and 4 fs for long - range electrostatic interactions. the simulation system (i) contained 156 293 atoms in total whereas the simulation system (ii) had 125 056 atoms. all simulations were performed with the program namd 2.5 (17) using the amber parm99 forcefield parameters (18) on 160 processors of the ibm datastar system at the san diego supercomputing center. to examine in detail the relation between existing pcna structures and their biologically critical functional interactions with dsdna, we carried out extensive molecular dynamics simulations. the simulation of hpcna in complex with dsdna reveals an unexpected and striking tilt of the pcna ring relative to the dna axis, as shown by a snapshot taken from the end of the simulation trajectory for the hpcna / dsdna system (figure 1). furthermore, human pcna is evidently prototypical for pcna in general, as analogous results are obtained for archaeal pcna. the most prominent feature of the structure resulting from the simulation is the pronounced tilt (of 20) in the axis of dsdna with respect to the plane of the pcna ring. the right - hand view of the structure shows that the dsdna helix substantially departs from its initial position at the center of the sliding clamp to form contacts with basic protein residues (arginine and lysine) lining the inner surface of the hpcna ring. analogous observations are equally valid for a.fulgidus pcna, where the tilt angle approaches 20 towards the end of the simulation. the observed changes in orientation of the dna helix with respect to pcna stem from two major intrinsic structural factors. first, the relatively rigid structure of the sliding clamp preserves the diameter of the central hole of pcna (35), which is significantly larger that the lateral extent of the dsdna helix in the canonical b - form (19) (24). second, the need to maximize the interactions between the positively charged protein residues located on the inner surface -helices of pcna and the negatively charged dna phosphodiester backbone leads to an energetically more favorable tilted orientation for pcna on dsdna, as displayed in figure 2 for hpcna. the red surface, representing charged residues within a 6 cutoff from the dna phosphate groups, tracks along part of the dna backbone on both strands of the dna minor groove and is almost exactly perpendicular to the axes of the -helices forming the inner surface of hpcna (figure 2). it has been suggested that association perpendicular to the minor groove, as opposed to binding parallel to the major or minor groove seen in many dna binding proteins, promotes the formation of non - specific protein dna interactions and prevents significant distortions in the structure of the nucleic acid (4). furthermore, the inner diameter of the pcna ring is large enough to allow the solvent to pass through freely (as observed in the simulations) and compete with the protein residues in making contacts with dna. thus, we find that the process of contact breaking and formation along the inner surface of pcna is very dynamic, with competition between different protein residues for the phosphodiester groups of dna. to analyze the dynamics of contact formation in our simulations, we calculated along the entire trajectory the number of nitrogen atoms belonging to the side chains of positively charged protein residues that were located within 7 of a dna phosphate group. it is informative to examine the distribution of such contacts as a function of the distance between the phosphorus and nitrogen atoms (figure 3). the histogram data are bimodal and allow us to distinguish between close interactions with p - n distances varying from 3.2 to 4.6 and more distant interactions above 5. although weaker than the close interactions, the distant interactions are nevertheless expected to contribute to the electrostatic stabilization of dna inside the pcna central hole due to their larger number and the long - range nature of the coulomb effect. figure 4a and b depicts the time evolution of the number of contacts during our simulations. besides the total number of contacts found within a 7 cutoff, we also calculated a more complicated function, reflecting the larger contribution of close interactions. this was accomplished by weighting the number of p - n contacts by a fermi - like function f(r)=1/{exp[k(rrc)]+1}. the values of rc and k were selected to be 5 and 0.333 so that contacts below the minimum in the distributions (at 4.6) are fully weighted whereas contacts above 6 have negligible contribution. in determining the number of contacts we take into account the fact that a phosphate group can certainly interact with more than one charged amino acid side chain within the cutoff and vice versa. therefore, we count the total number of such pairs. for both systems, we observe an overall increase in the number of contacts, which is paralleled closely by an increase in the tilt angle between the axis of the dna helix and a vector normal to the plane of the pcna ring (figure 4c and d). the results indicate that the number of close interactions varies between 5 and 20, and these strong contacts are broken and reformed on a sub - nanosecond timescale (figure 4). visual inspection of the trajectory for the hpcna system revealed that the tilting of the pcna ring is a relatively simple, gradual transition. in contrast, in the case of apcna, we observe initial gradual motion with several excursions back and forth followed by a transition into a more stable tilted orientation at 7 ns ; then the ring of pcna straightens out and moves with respect to dna before transitioning into another tilted conformation for the protein nucleic acid assembly. the time scale of our simulations is insufficient to sample exhaustively the global motions of clamp with respect to dna or the process of. however, the results are stunningly suggestive as to why the clamp is capable of sliding freely despite the fact that it forms extensive contacts with the nucleic acid backbone. the answer lies in the competition between charged residues on the inner surface of pcna for forming interaction with the two strands of dna along a stretch of the minor groove coupled to the ability of the ring to tilt and partially rotate around dna. this competition is illustrated on figure 5, which presents the share of p - n contacts with the dna phosphodiester groups made by each of the pcna subunits. the numbers vary substantially between the subunits and during the different stages of the simulations. often an increase in the number of contacts made by one subunit is compensated by a decrease in the interactions with the other two subunits. we also note that for apcna the observed overall increase in the total number of close contacts arises primarily from closer association with dsdna and subunit c in the latter half of the simulation. we examined the covariance and cross - correlation (normalized covariance) matrices for the atoms in our pcna system with dsdna (20,21). the covariance matrix elements can be expressed as : where tave is the averaging time, t is the time step, the positions atoms i and j at time t are ri(t) and rj(t), respectively, and the angular brackets denote time averaging. the diagonal elements of the covariance matrix correspond to the atomic mean square fluctuations and are, in turn, related to the computed isotropic temperature factors (b - factors) by a constant multiplicative factor 8/3. the elements of the cross - correlation matrix are defined as : cij = cijcii1/2cjj1/2. these take values from 1 to 1 and, in contrast to the covariance coefficients, do not contain information about the magnitude of the correlated motions. the computed and experimental b - factors on the c atoms for our systems both serve as a measure of the inherent flexibility of the protein residues within a particular stretch of sequence (figure 6). the overall agreement between theory and experiment is reasonable especially in the case of hpcna with both experimental and computed b - factors identifying the same mobile regions. that said, the computed b - factors produce sharper peaks and the differences become particularly pronounced at both ends of the sequence. such differences are not surprising, given that the simulations are supposed to reflect a situation closer to solution phase dynamics whereas the experiment is done under different conditions and on a vastly different timescale. the calculated covariance and cross - correlation matrices between the c atoms for both hpcna and apcna reveal a clear pattern of correlated and anticorrelated motions extending both within and between the pcna subunits (figure 7). these motions are represented by off diagonal peaks, indicating correlation and valleys, indicating anticorrelation. in general, values in the range above 0.25 and below 0.25 in the cross - correlation matrices can be considered significant. the fact that strong coupling between the motions of the subunits exists opens up the possibility that affecting the dynamics of pcna in one of the subunits (e.g through binding) may have consequences for the dynamics of the other two subunits. on the other hand, such correlation may simply reflect the relatively rigid structure of the clamp and the tight coupling between the individual subunits. future work is needed to address this issue possibly by carrying out simulations of peptides bound to pcna. a key aspect of the cross - correlation maps is that they reveal a non - equivalence of the three pcna subunits in terms dynamics, with such differences arising from dsdna binding. the disparity in size between the inner diameter of pcna and the lateral extent of dsdna makes it impossible to form equally extensive contacts with all three subunits. for instance, the dna minor groove associates primarily with subunits a and b of human pcna, leaving subunit c as the most mobile of the three hpcna monomers. this is clearly represented in the covariance map (figure 7a and b), showing the largest covariance values to be associated with subunit c and extensive cross - correlation terms extending into subunit b. for apcna the second subunit (chain b in our model structure) displays a pattern of correlation and anticorrelation distinct from the patterns of the other two subunits, which appear to be similar and symmetrically related. the second subunit also exhibits motions of greater magnitude compared to the other two, especially in a region at the pcna c - terminus (residues 240244 of the pcna monomer). these results are potentially informative as the apcna c - terminus is known to form part of a hydrophobic pocket on the surface of pcna responsible for association with pcna binding enzymes, such as fen-1 (6). examination of the trajectory immediately reveals that the b subunit directly associates with the minor groove of dsdna making contacts with the phosphodiester backbone throughout the latter part of the simulation. comparison to figure 5 shows that the subunit also displays the largest variation in the number of contacts formed during the trajectory confirming its increased mobility. although the limited length of our simulations precludes a more definitive computational test of contacts and mobility at this time, the results are completely consistent with the proposal that closer association of dsdna to a given subunit can influence dynamics on the outer surface of the clamp. to identify and visualize correlated motions, including global collective motions of pcna on dna with potential functional significance, we employed principal component analysis (2224) (pca) on both the hpcna and apcna systems. the matrix is symmetric and can be diagonalized using an orthonormal transformation matrix r, with eigenvectors denoted as principal modes. the eigenvalue corresponding to a principal component represents the mean square fluctuation along this component. we applied a least squares fitting procedure to the average protein structure from the second half of the simulation to eliminate the overall rotational and translational motion of pcna. we found pca to be a versatile tool for analyzing protein dynamics for these systems and useful in confirming observations made during visual inspection of the trajectories. as pca reduces the dimensionality of the trajectory data, it highlights important dynamical features with the first few principal modes describing collective, global motions of the system. for the two pcna systems, these involved both the c - terminal region of the pcna subunits and the central part of the linker strand connecting the two domains of the pcna molecule. significantly, these structural elements on the surface of the sliding clamp are largely responsible for binding to dna - editing enzymes. for example, the first principal component eigenvector mapped onto the c carbon atoms of hpcna reveals that the largest motions involve the linker domain and the three flexible loops (guides) of hpcna (figure 8). this eigenvector accounts for 32% of the overall motion seen in the hpcna dynamics trajectory. (for comparison, the first five eigenvectors of the covariance matrix are responsible for 50% of the observed motion because the corresponding eigenvalues decrease rapidly). the magnitude of these motions appears to be largest toward the tips of the wedge shaped domains, but due to the rigid structure of the clamp, the motion also gets propagated to the inner surface alpha helices that make contact with dna. such collective motions are interesting in that they appear appropriate to facilitate the translocation of pcna along dsdna. figure 9 depicts two - dimensional projections of the trajectories for the two systems onto the first and second principal components. the analysis was again carried out using the c carbon atoms of the protein residues and, therefore, reflects the interactions with dsdna only indirectly. nevertheless, it is interesting to observe several clusters of states along the trajectory especially pronounced in the apcna case. as noted above, the hpcna system seems to undergo a more gradual transition and this fact is reflected on figure 9a, which shows a smooth transition between two broad basins encompassing many microstates. the first basin is rather broad reflecting the exploration of conformational space occurring during the initial stage of the simulation. the other two basins are much more compact and are especially pronounced in terms of density. the region between these basins is shallow and possibly serves as an intermediate state in the structural transition described earlier. thus, the global motions of the sliding clamp with respect to dsdna appear to be well correlated to the internal motions within the pcna ring. as more structures are determined the value of computational analyses that help clarify their functional interactions becomes increasingly important, especially for the reversible associations linked with many of the major decision points for cells, i.e. cell growth versus death and dna replication versus repair in the face of damage. pcna is a recognized master key for reversible associations that determine pathways since it interacts with many replication and repair proteins as well as cell cycle checkpoint inhibitors. for this reason, it has been suggested that pcna may coordinate its protein partner activities either by forming ternary complexes with two or three replication factors or by handing off dna intermediates from one enzyme to the next in the replication or repair pathway (5,2527). both mechanisms are based on current models for pcna encircling dna, which imply that pcna is perpendicular to the axis of dna. in this orientation, the three binding sites, each located on the face of one subunit of the pcna homotrimer, appear to be equivalent. in contrast, our simulations show that pcna binds to dna asymmetrically, mainly through positively charged residues on the alpha helices that line the inside of the pcna ring. this binding mode causes the plane of the pcna ring to tilt by 20 with respect to the helical axis of dna, breaking the symmetry of the pcna trimer. this result has broad implications for the mechanism in which pcna interacts uniquely with multiple enzymes. most efforts to characterize pcna interactions with replication and repair enzymes have focused on regions of the trimer surface identified by structural analyses of peptides and enzymes bound to pcna (5,26). although, it is widely recognized that interactions between pcna and dna can influence the nature of pcna interactions with bound enzymes, these interactions are difficult to characterize using biophysical techniques. for example, pcna stimulates the activity of enzymes, such as dna pol and fen-1, but only when pcna encircles dna (25,28,29). structural and biochemical studies suggest that residues from fen-1 and the c - terminus of pcna form a -sheet, which positions fen-1 for catalysis (6,29). notably, our simulations indicate that the loop and c - terminal regions of each pcna subunit, which are involved in partner enzyme interactions, may exhibit differences in flexibility depending on their association with dna. such changes in flexibility appear suitable to aid handoffs between different enzymes or facilitate unique interactions with simultaneously bound enzymes based on contacts between pcna and dna. our simulation results identified several positively charged arg and lys residues that potentially interact with the phosphate backbone of dna (figure 2) : lys13, lys14, lys20, lys77, lys217, arg146 and arg149. all of these residues are located on the inner surface of the pcna ring and are conserved in both human and yeast pcna. previous mutational analyses showed that mutating lys20 or lys77 to glu interfered with mismatch repair in saccharomyces cerevisiae, resulting in a mutator phenotype (30). biochemical analyses using purified human pcna and pol showed that mutating these pcna residues to ala prevented pcna from stimulating pol polymerization, but did not affect the processivity of the pol :pcna complex (31). these pcna mutants were originally thought to interfere with initiation of dna synthesis by pol (31). importantly, our results support this interpretation and suggest that the tilted orientation of pcna relative to the helical axis of dna, facilitated by charged interactions with the dna backbone, could uniquely position pol for efficient initiation of dna synthesis. if the asymmetric orientation of pcna is critical for stimulating pol initiation, then this orientation might also be required to stimulate fen-1 activity (6,32,33). this hypothesis can now be experimentally tested using an established biochemical assay for measuring the activity of fen-1 in the presence of pcna that has been loaded onto dna by rfc (29). mutations of the basic amino acids lining the center of the pcna do not affect rfc - dependent loading or activity (31). thus, these mutations, which disrupt interactions with dna backbone along the minor groove, could be used to test whether the asymmetric orientation of pcna is required to stimulate fen-1 activity. in addition, several key charged residues are also conserved in the heterotrimeric pcna homolog from sulfolobus solfataricus (34). in this organism, fen-1 and dna polymerase thus, mutational analysis in this pcna homolog could be used to determine how dna interactions with a single subunit affect interactions with a bound enzyme. the dna binding mode presented here also suggests a way to biophysically characterize the interaction between pcna and dna. because pcna associates with dna in a topological manner, structural and biophysical characterization of this interface has been challenging due to the difficultly in producing a homogeneous population of pcna : dna complex for analysis. similar hurdles exist for determining the structure of a ternary complex including pcna and fen-1 or dna polymerase both bound to dna. however, using the binding model presented here as a guide, it might be possible to covalently trap a complex of pcna bound to dna. the model of pcna bound to dna shows that some of the lys residues located at the center of the pcna ring (lys14, lys20, lys80 and lys217) interact close to the center of the minor groove. to trap a complex, it may be possible to use a strategy in which each of the target lys residues are systematically mutated to cys, and mixed with dna containing g or t base with a commercially available alkanethiol modification (midland certified reagent company). these modifications would allow covalent trapping via formation of a disulfide bond (35). a similar strategy was used to trap hiv - rt bound to an rna - dna hybrid substrate (36), based on computational modeling of the interaction (37). this covalent trapping strategy would facilitate structural analysis of a pcna : dna complex by small angle x - ray scattering or x - ray crystallography. thus, the results and concepts presented here provide a framework for a variety of experimental and computational approaches to test the functional implications of the asymmetric pcna subunit interactions with dna. to examine in detail the relation between existing pcna structures and their biologically critical functional interactions with dsdna, we carried out extensive molecular dynamics simulations. the simulation of hpcna in complex with dsdna reveals an unexpected and striking tilt of the pcna ring relative to the dna axis, as shown by a snapshot taken from the end of the simulation trajectory for the hpcna / dsdna system (figure 1). furthermore, human pcna is evidently prototypical for pcna in general, as analogous results are obtained for archaeal pcna. the most prominent feature of the structure resulting from the simulation is the pronounced tilt (of 20) in the axis of dsdna with respect to the plane of the pcna ring. the right - hand view of the structure shows that the dsdna helix substantially departs from its initial position at the center of the sliding clamp to form contacts with basic protein residues (arginine and lysine) lining the inner surface of the hpcna ring. analogous observations are equally valid for a.fulgidus pcna, where the tilt angle approaches 20 towards the end of the simulation. the observed changes in orientation of the dna helix with respect to pcna stem from two major intrinsic structural factors. first, the relatively rigid structure of the sliding clamp preserves the diameter of the central hole of pcna (35), which is significantly larger that the lateral extent of the dsdna helix in the canonical b - form (19) (24). second, the need to maximize the interactions between the positively charged protein residues located on the inner surface -helices of pcna and the negatively charged dna phosphodiester backbone leads to an energetically more favorable tilted orientation for pcna on dsdna, as displayed in figure 2 for hpcna. the red surface, representing charged residues within a 6 cutoff from the dna phosphate groups, tracks along part of the dna backbone on both strands of the dna minor groove and is almost exactly perpendicular to the axes of the -helices forming the inner surface of hpcna (figure 2). it has been suggested that association perpendicular to the minor groove, as opposed to binding parallel to the major or minor groove seen in many dna binding proteins, promotes the formation of non - specific protein dna interactions and prevents significant distortions in the structure of the nucleic acid (4). furthermore, the inner diameter of the pcna ring is large enough to allow the solvent to pass through freely (as observed in the simulations) and compete with the protein residues in making contacts with dna. thus, we find that the process of contact breaking and formation along the inner surface of pcna is very dynamic, with competition between different protein residues for the phosphodiester groups of dna. to analyze the dynamics of contact formation in our simulations, we calculated along the entire trajectory the number of nitrogen atoms belonging to the side chains of positively charged protein residues that were located within 7 of a dna phosphate group. it is informative to examine the distribution of such contacts as a function of the distance between the phosphorus and nitrogen atoms (figure 3). the histogram data are bimodal and allow us to distinguish between close interactions with p - n distances varying from 3.2 to 4.6 and more distant interactions above 5. although weaker than the close interactions, the distant interactions are nevertheless expected to contribute to the electrostatic stabilization of dna inside the pcna central hole due to their larger number and the long - range nature of the coulomb effect. figure 4a and b depicts the time evolution of the number of contacts during our simulations. besides the total number of contacts found within a 7 cutoff, we also calculated a more complicated function, reflecting the larger contribution of close interactions. this was accomplished by weighting the number of p - n contacts by a fermi - like function f(r)=1/{exp[k(rrc)]+1}. the values of rc and k were selected to be 5 and 0.333 so that contacts below the minimum in the distributions (at 4.6) are fully weighted whereas contacts above 6 have negligible contribution. in determining the number of contacts we take into account the fact that a phosphate group can certainly interact with more than one charged amino acid side chain within the cutoff and vice versa. therefore, we count the total number of such pairs. for both systems, we observe an overall increase in the number of contacts, which is paralleled closely by an increase in the tilt angle between the axis of the dna helix and a vector normal to the plane of the pcna ring (figure 4c and d). the results indicate that the number of close interactions varies between 5 and 20, and these strong contacts are broken and reformed on a sub - nanosecond timescale (figure 4). visual inspection of the trajectory for the hpcna system revealed that the tilting of the pcna ring is a relatively simple, gradual transition. in contrast, in the case of apcna, we observe initial gradual motion with several excursions back and forth followed by a transition into a more stable tilted orientation at 7 ns ; then the ring of pcna straightens out and moves with respect to dna before transitioning into another tilted conformation for the protein nucleic acid assembly. the time scale of our simulations is insufficient to sample exhaustively the global motions of clamp with respect to dna or the process of. however, the results are stunningly suggestive as to why the clamp is capable of sliding freely despite the fact that it forms extensive contacts with the nucleic acid backbone. the answer lies in the competition between charged residues on the inner surface of pcna for forming interaction with the two strands of dna along a stretch of the minor groove coupled to the ability of the ring to tilt and partially rotate around dna. this competition is illustrated on figure 5, which presents the share of p - n contacts with the dna phosphodiester groups made by each of the pcna subunits. the numbers vary substantially between the subunits and during the different stages of the simulations. often an increase in the number of contacts made by one subunit is compensated by a decrease in the interactions with the other two subunits. we also note that for apcna the observed overall increase in the total number of close contacts arises primarily from closer association with dsdna and subunit c in the latter half of the simulation. we examined the covariance and cross - correlation (normalized covariance) matrices for the atoms in our pcna system with dsdna (20,21). the covariance matrix elements can be expressed as : where tave is the averaging time, t is the time step, the positions atoms i and j at time t are ri(t) and rj(t), respectively, and the angular brackets denote time averaging. the diagonal elements of the covariance matrix correspond to the atomic mean square fluctuations and are, in turn, related to the computed isotropic temperature factors (b - factors) by a constant multiplicative factor 8/3. the elements of the cross - correlation matrix are defined as : cij = cijcii1/2cjj1/2. these take values from 1 to 1 and, in contrast to the covariance coefficients, do not contain information about the magnitude of the correlated motions. the computed and experimental b - factors on the c atoms for our systems both serve as a measure of the inherent flexibility of the protein residues within a particular stretch of sequence (figure 6). the overall agreement between theory and experiment is reasonable especially in the case of hpcna with both experimental and computed b - factors identifying the same mobile regions. that said, the computed b - factors produce sharper peaks and the differences become particularly pronounced at both ends of the sequence. such differences are not surprising, given that the simulations are supposed to reflect a situation closer to solution phase dynamics whereas the experiment is done under different conditions and on a vastly different timescale. the calculated covariance and cross - correlation matrices between the c atoms for both hpcna and apcna reveal a clear pattern of correlated and anticorrelated motions extending both within and between the pcna subunits (figure 7). these motions are represented by off diagonal peaks, indicating correlation and valleys, indicating anticorrelation. in general, values in the range above 0.25 and below 0.25 in the cross - correlation matrices can be considered significant. the fact that strong coupling between the motions of the subunits exists opens up the possibility that affecting the dynamics of pcna in one of the subunits (e.g through binding) may have consequences for the dynamics of the other two subunits. on the other hand, such correlation may simply reflect the relatively rigid structure of the clamp and the tight coupling between the individual subunits. future work is needed to address this issue possibly by carrying out simulations of peptides bound to pcna. a key aspect of the cross - correlation maps is that they reveal a non - equivalence of the three pcna subunits in terms dynamics, with such differences arising from dsdna binding. the disparity in size between the inner diameter of pcna and the lateral extent of dsdna makes it impossible to form equally extensive contacts with all three subunits. for instance, the dna minor groove associates primarily with subunits a and b of human pcna, leaving subunit c as the most mobile of the three hpcna monomers. this is clearly represented in the covariance map (figure 7a and b), showing the largest covariance values to be associated with subunit c and extensive cross - correlation terms extending into subunit b. for apcna the second subunit (chain b in our model structure) displays a pattern of correlation and anticorrelation distinct from the patterns of the other two subunits, which appear to be similar and symmetrically related. the second subunit also exhibits motions of greater magnitude compared to the other two, especially in a region at the pcna c - terminus (residues 240244 of the pcna monomer). these results are potentially informative as the apcna c - terminus is known to form part of a hydrophobic pocket on the surface of pcna responsible for association with pcna binding enzymes, such as fen-1 (6). examination of the trajectory immediately reveals that the b subunit directly associates with the minor groove of dsdna making contacts with the phosphodiester backbone throughout the latter part of the simulation. comparison to figure 5 shows that the subunit also displays the largest variation in the number of contacts formed during the trajectory confirming its increased mobility. although the limited length of our simulations precludes a more definitive computational test of contacts and mobility at this time, the results are completely consistent with the proposal that closer association of dsdna to a given subunit can influence dynamics on the outer surface of the clamp. to identify and visualize correlated motions, including global collective motions of pcna on dna with potential functional significance, we employed principal component analysis (2224) (pca) on both the hpcna and apcna systems. the matrix is symmetric and can be diagonalized using an orthonormal transformation matrix r, with eigenvectors denoted as principal modes. the eigenvalue corresponding to a principal component represents the mean square fluctuation along this component. we applied a least squares fitting procedure to the average protein structure from the second half of the simulation to eliminate the overall rotational and translational motion of pcna. we found pca to be a versatile tool for analyzing protein dynamics for these systems and useful in confirming observations made during visual inspection of the trajectories. as pca reduces the dimensionality of the trajectory data, it highlights important dynamical features with the first few principal modes describing collective, global motions of the system. for the two pcna systems, these involved both the c - terminal region of the pcna subunits and the central part of the linker strand connecting the two domains of the pcna molecule. significantly, these structural elements on the surface of the sliding clamp are largely responsible for binding to dna - editing enzymes. for example, the first principal component eigenvector mapped onto the c carbon atoms of hpcna reveals that the largest motions involve the linker domain and the three flexible loops (guides) of hpcna (figure 8). this eigenvector accounts for 32% of the overall motion seen in the hpcna dynamics trajectory. (for comparison, the first five eigenvectors of the covariance matrix are responsible for 50% of the observed motion because the corresponding eigenvalues decrease rapidly). the magnitude of these motions appears to be largest toward the tips of the wedge shaped domains, but due to the rigid structure of the clamp, the motion also gets propagated to the inner surface alpha helices that make contact with dna. such collective motions are interesting in that they appear appropriate to facilitate the translocation of pcna along dsdna. figure 9 depicts two - dimensional projections of the trajectories for the two systems onto the first and second principal components. the analysis was again carried out using the c carbon atoms of the protein residues and, therefore, reflects the interactions with dsdna only indirectly. nevertheless, it is interesting to observe several clusters of states along the trajectory especially pronounced in the apcna case. as noted above, the hpcna system seems to undergo a more gradual transition and this fact is reflected on figure 9a, which shows a smooth transition between two broad basins encompassing many microstates. the first basin is rather broad reflecting the exploration of conformational space occurring during the initial stage of the simulation. the other two basins are much more compact and are especially pronounced in terms of density. the region between these basins is shallow and possibly serves as an intermediate state in the structural transition described earlier. thus, the global motions of the sliding clamp with respect to dsdna appear to be well correlated to the internal motions within the pcna ring. as more structures are determined the value of computational analyses that help clarify their functional interactions becomes increasingly important, especially for the reversible associations linked with many of the major decision points for cells, i.e. cell growth versus death and dna replication versus repair in the face of damage. pcna is a recognized master key for reversible associations that determine pathways since it interacts with many replication and repair proteins as well as cell cycle checkpoint inhibitors. for this reason, it has been suggested that pcna may coordinate its protein partner activities either by forming ternary complexes with two or three replication factors or by handing off dna intermediates from one enzyme to the next in the replication or repair pathway (5,2527). both mechanisms are based on current models for pcna encircling dna, which imply that pcna is perpendicular to the axis of dna. in this orientation, the three binding sites, each located on the face of one subunit of the pcna homotrimer, appear to be equivalent. in contrast, our simulations show that pcna binds to dna asymmetrically, mainly through positively charged residues on the alpha helices that line the inside of the pcna ring. this binding mode causes the plane of the pcna ring to tilt by 20 with respect to the helical axis of dna, breaking the symmetry of the pcna trimer. this result has broad implications for the mechanism in which pcna interacts uniquely with multiple enzymes. most efforts to characterize pcna interactions with replication and repair enzymes have focused on regions of the trimer surface identified by structural analyses of peptides and enzymes bound to pcna (5,26). although, it is widely recognized that interactions between pcna and dna can influence the nature of pcna interactions with bound enzymes, these interactions are difficult to characterize using biophysical techniques. for example, pcna stimulates the activity of enzymes, such as dna pol and fen-1, but only when pcna encircles dna (25,28,29). structural and biochemical studies suggest that residues from fen-1 and the c - terminus of pcna form a -sheet, which positions fen-1 for catalysis (6,29). notably, our simulations indicate that the loop and c - terminal regions of each pcna subunit, which are involved in partner enzyme interactions, may exhibit differences in flexibility depending on their association with dna. such changes in flexibility appear suitable to aid handoffs between different enzymes or facilitate unique interactions with simultaneously bound enzymes based on contacts between pcna and dna. our simulation results identified several positively charged arg and lys residues that potentially interact with the phosphate backbone of dna (figure 2) : lys13, lys14, lys20, lys77, lys217, arg146 and arg149. all of these residues are located on the inner surface of the pcna ring and are conserved in both human and yeast pcna. previous mutational analyses showed that mutating lys20 or lys77 to glu interfered with mismatch repair in saccharomyces cerevisiae, resulting in a mutator phenotype (30). biochemical analyses using purified human pcna and pol showed that mutating these pcna residues to ala prevented pcna from stimulating pol polymerization, but did not affect the processivity of the pol :pcna complex (31). these pcna mutants were originally thought to interfere with initiation of dna synthesis by pol (31). importantly, our results support this interpretation and suggest that the tilted orientation of pcna relative to the helical axis of dna, facilitated by charged interactions with the dna backbone, could uniquely position pol for efficient initiation of dna synthesis. if the asymmetric orientation of pcna is critical for stimulating pol initiation, then this orientation might also be required to stimulate fen-1 activity (6,32,33). this hypothesis can now be experimentally tested using an established biochemical assay for measuring the activity of fen-1 in the presence of pcna that has been loaded onto dna by rfc (29). mutations of the basic amino acids lining the center of the pcna do not affect rfc - dependent loading or activity (31). thus, these mutations, which disrupt interactions with dna backbone along the minor groove, could be used to test whether the asymmetric orientation of pcna is required to stimulate fen-1 activity. in addition, several key charged residues are also conserved in the heterotrimeric pcna homolog from sulfolobus solfataricus (34). in this organism, fen-1 and dna polymerase thus, mutational analysis in this pcna homolog could be used to determine how dna interactions with a single subunit affect interactions with a bound enzyme. the dna binding mode presented here also suggests a way to biophysically characterize the interaction between pcna and dna. because pcna associates with dna in a topological manner, structural and biophysical characterization of this interface has been challenging due to the difficultly in producing a homogeneous population of pcna : dna complex for analysis. similar hurdles exist for determining the structure of a ternary complex including pcna and fen-1 or dna polymerase both bound to dna. however, using the binding model presented here as a guide, it might be possible to covalently trap a complex of pcna bound to dna. the model of pcna bound to dna shows that some of the lys residues located at the center of the pcna ring (lys14, lys20, lys80 and lys217) interact close to the center of the minor groove. to trap a complex, it may be possible to use a strategy in which each of the target lys residues are systematically mutated to cys, and mixed with dna containing g or t base with a commercially available alkanethiol modification (midland certified reagent company). these modifications would allow covalent trapping via formation of a disulfide bond (35). a similar strategy was used to trap hiv - rt bound to an rna - dna hybrid substrate (36), based on computational modeling of the interaction (37). this covalent trapping strategy would facilitate structural analysis of a pcna : dna complex by small angle x - ray scattering or x - ray crystallography. thus, the results and concepts presented here provide a framework for a variety of experimental and computational approaches to test the functional implications of the asymmetric pcna subunit interactions with dna. pcna interactions with dsdna : a) side view and b) upper view of the hpcna dsdna complex reveal the relationship between the tilting of the hpcna ring with respect to dsdna and the interactions between the nucleic acid backbone and the inner positively charged surface of the sliding clamp. the dsdna phosphodiester groups (orange spheres) plus basic (arginine and lysine) groups of hpcna within 6 of the phosphates (red surfaces) are shown. histogram of the interactions between the side chain n atoms of basic (arginine and lysine) groups of pcna located within 7 of the nucleic acid backbone p atoms. the distribution of nitrogen phosphorus contacts as a function of distance reveals two peaks with a minimum at 4.6. p contacts can be naturally subdivided into close (below the minimum) and distant (above the minimum) interactions. the red curve represents the weighting function that was used to select close interactions. time evolution of the interactions between the side chain n atoms of basic (arginine and lysine) groups of hpcna located within 7 of the nucleic acid backbone p atoms. data for apcna is shown on (a) and for hpcna on (b). (c and d) display the time evolution of the tilt angle between the dna axis and the plane of the pcna ring for apcna and hpcna, respectively. the observed increase in the number of n p contacts over the trajectory is closely paralleled by an increase in the tilt angle, which approaches 20 toward the end of the simulations. close nitrogen phosphorus contacts (within a 4.6 cutoff) between each of the pcna subunits and the nucleic acid. data for apcna is shown on (a) and for hpcna on (b), respectively. n contacts with the dna phosphodiester backbone made by each of the pcna subunits during the simulation, highlighting the dynamic nature of the observed asymmetric binding between the pcna subunits and dna. computed versus experimental b - factors identifying the mobile regions of the pcna trimer as a function of residue number (with subunits a, b and c shown sequentially) : (a) for hpcna and (b) for apcna. correlated and anti - correlated pcna motions revealed by cross - correlation and covariance maps for hpcna and apcna. a) covariance (a) and cross - correlation matrix (b) for hpcna ; b) covariance (c) and cross - correlation matrix (d) for apcna. the cross - correlation and covariance maps identify regions of the protein that may be distant in the sequence but, nevertheless, move in a concerted way. positive values (red) denote correlated motions ; negative values (blue) indicated anti - correlated motions. front (a) and side (b) representations of the first principal component vector mapped onto the c carbon atoms of hpcna. the three subunits are shown in trace representation along the c carbon atoms in red, green and blue, respectively. histograms obtained from two - dimensional projections of the trajectories onto the first two principal eigenvectors. data for hpcna and apcna is shown in (a) and (b), respectively. | proliferating cell nuclear antigen (pcna) acts as a biologically essential processivity factor that encircles dna and provides binding sites for polymerase, flap endonuclease-1 (fen-1) and ligase during dna replication and repair. we have computationally characterized the interactions of human and archaeoglobus fulgidus pcna trimer with double - stranded dna (ds dna) using multi - nanosecond classical molecular dynamics simulations. the results reveal the interactions of dna passing through the pcna trimeric ring including the contacts formed, overall orientation and motion with respect to the sliding clamp. notably, we observe pronounced tilting of the axis of dsdna with respect to the pcna ring plane reflecting interactions between the dna phosphodiester backbone and positively charged arginine and lysine residues lining the pcna inner surface. covariance matrix analysis revealed a pattern of correlated motions within and between the three equivalent subunits involving the pcna c - terminal region and linker strand associated with partner protein binding sites. additionally, principal component analysis identified low frequency global pcna subunit motions suitable for translocation along duplex dna. the pcna motions and interactions with the dna minor groove, identified here computationally, provide an unexpected basis for pcna to act in the coordinated handoff of intermediates from polymerase to fen-1 to ligase during dna replication and repair. |
patients admitted to kitunan hospital in suzenji, japan from may 1999 and april 2007 with acute psychomotor excitation were studied retrospectively. the inclusion criteria were : (i) patients diagnosed with schizophrenia according to the diagnostic and statistical manual of mental disorders, 4th edition (dsm - iv) ; (ii) achieving a score of at least 20 on the excited component of the positive and negative syndrome scale (panss - ec) scale at hospital admission ; and (iii) with laboratory values recorded at hospital admission and 35 weeks later. the panss - ec comprises five items from the panss:2 poor impulse control, tension, hostility, uncooperativeness, and excitement, each rated on a 7-point likert scale ranging from 1 (not present) to 7 (extremely severe). possible scores range from 5 to 35 ; mean scores 20 indicate severe agitation.3 patients with alcoholism and/or drug abuse were excluded. values outside the reference range were defined as abnormal values, and the frequency of abnormal values was calculated for each parameter during the acute phase and during the recovery phase on medication, between 3 and 5 weeks later. panss - ec scores between the acute and recovery phases were compared using paired t tests. for comparison between drugs for age, dose, and panss - ec score, unpaired t tests were used. for comparison of the frequency of abnormal values in the acute and recovery phases, as well as a comparison between drugs in the frequency of abnormal values, a total of 68 participants were included in the study (32 male, 36 female). mean age standard deviation was 47.0 11.9 years, and the mean panss - ec score standard deviation at hospital admission was 21.4 1.5. of the 37 participants with detailed records of previous hospital visits, 31 (83.8%) had interrupted their prescribed medication., treatment groups were defined according to the antipsychotic drug that had been used : risperidone group (14 participants), olanzapine group (16 participants), haloperidol group (12 participants), and a polypharmacy group defined as patients that had taken either risperidone and haloperidole, or olanapine and haloperidole. the medication prescribed was decided by the physician in charge at the time of their hospitalization. no information was available in the patient files as to the reason for the choice of a particular drug. table 1 shows age, sex, and panss - ec score during the acute and recovery phase by drug. no significant differences in age or panss - ec score were found between the treatment groups during the acute phase. conversion of doses of the antipsychotic drugs into chlorpromazine equivalent doses showed that the polypharmacy group received significantly higher doses compared to the risperidone and olanzapine groups (p < 0.01). as shown in figure 1, all treatment groups had a significantly improved panss - ec score by weeks 35 (p < 0.001), although there was significantly less reduction of panss - ec score in the polypharmacy group compared to the risperidone and olanzapine groups (p < 0.01). the reference values for 26 laboratory parameters and number of participants with values outside the reference range in the acute and recovery phases are summarized in table 2. laboratory parameters were classified into three groups depending on the frequency of abnormal values during the acute and recovery phases : (i) parameters with a high frequency (25%) of abnormal values during the acute phase and a low frequency (25%) during the recovery phase white blood cells (wbc), platelets, creative protein (crp), fetal bovine serum (fbs), lactate dehydrogenase (ldh), blood urea nitrogen (bun), uric acid (ua), k, na, and cl) ; (ii) parameters with a low frequency of abnormal values during the acute phase and a high frequency during the recovery phase (hba1c, tg, ldl - c, amy, and ck) ; and (iii) parameters with a low frequency of abnormal values during both the acute and recovery phases (rbc, hb, t.cho, hdl - c, tp, alb, ast, alt, gtp, t.bil, and cre). parameters with a high frequency of abnormal values during the acute phase are shown in figure 2 and those with a high frequency of abnormal values during the recovery phase in figure 3. as shown in figure 4, the frequency of abnormal values was significantly different between the acute and recovery phases for seven laboratory parameters (k, wbc, tg, ldh, ua, fbs, and bun). of these, 6 parameters normalized during the recovery phase, while the frequency of abnormal triglyceride levels was greater during the recovery phase. the frequency of abnormal values during the recovery phase in metabolism - related parameters is summarized in figure 5. abnormal triglyceride levels were significantly more frequent in the olanzapine group compared with the risperidone group (p = 0.031). a total of 68 participants were included in the study (32 male, 36 female). mean age standard deviation was 47.0 11.9 years, and the mean panss - ec score standard deviation at hospital admission was 21.4 1.5. of the 37 participants with detailed records of previous hospital visits, 31 (83.8%) had interrupted their prescribed medication., treatment groups were defined according to the antipsychotic drug that had been used : risperidone group (14 participants), olanzapine group (16 participants), haloperidol group (12 participants), and a polypharmacy group defined as patients that had taken either risperidone and haloperidole, or olanapine and haloperidole. the medication prescribed was decided by the physician in charge at the time of their hospitalization. no information was available in the patient files as to the reason for the choice of a particular drug. table 1 shows age, sex, and panss - ec score during the acute and recovery phase by drug. no significant differences in age or panss - ec score were found between the treatment groups during the acute phase. conversion of doses of the antipsychotic drugs into chlorpromazine equivalent doses showed that the polypharmacy group received significantly higher doses compared to the risperidone and olanzapine groups (p < 0.01). as shown in figure 1, all treatment groups had a significantly improved panss - ec score by weeks 35 (p < 0.001), although there was significantly less reduction of panss - ec score in the polypharmacy group compared to the risperidone and olanzapine groups (p < 0.01). the reference values for 26 laboratory parameters and number of participants with values outside the reference range in the acute and recovery phases are summarized in table 2. laboratory parameters were classified into three groups depending on the frequency of abnormal values during the acute and recovery phases : (i) parameters with a high frequency (25%) of abnormal values during the acute phase and a low frequency (25%) during the recovery phase white blood cells (wbc), platelets, creative protein (crp), fetal bovine serum (fbs), lactate dehydrogenase (ldh), blood urea nitrogen (bun), uric acid (ua), k, na, and cl) ; (ii) parameters with a low frequency of abnormal values during the acute phase and a high frequency during the recovery phase (hba1c, tg, ldl - c, amy, and ck) ; and (iii) parameters with a low frequency of abnormal values during both the acute and recovery phases (rbc, hb, t.cho, hdl - c, tp, alb, ast, alt, gtp, t.bil, and cre). parameters with a high frequency of abnormal values during the acute phase are shown in figure 2 and those with a high frequency of abnormal values during the recovery phase in figure 3. as shown in figure 4, the frequency of abnormal values was significantly different between the acute and recovery phases for seven laboratory parameters (k, wbc, tg, ldh, ua, fbs, and bun). of these, 6 parameters normalized during the recovery phase, while the frequency of abnormal triglyceride levels was greater during the recovery phase. the frequency of abnormal values during the recovery phase in metabolism - related parameters is summarized in figure 5. abnormal triglyceride levels were significantly more frequent in the olanzapine group compared with the risperidone group (p = 0.031). the frequency of abnormal laboratory values during acute agitation and one month later during the recovery phase was investigated. since it is often impossible to obtain informed consent from acutely agitated patients a retrospective nonintervention analysis was undertaken. although this investigation suffers from the typical limitations of a retrospective study, it does provide data relevant to treatment in a typical clinical situation. treatment with antipsychotic drugs resulted in a significant improvement in panss - ec score in all cases. however, multidrug combination therapy tended to be associated with higher doses and less improvement in panss - es scores after 35 weeks compared with monotherapy with risperidone or olanzapine. another interpretation, however, is that these patients were more difficult to treat, hence the decision by the clinician to give combination therapy. of the 26 laboratory parameters examined, there were only 11 parameters for which abnormal values were infrequently reported during both the acute and recovery phases, indicating the importance of systematically performing laboratory tests during the acute and recovery phases of schizophrenia. a high frequency of abnormal values was reported for 10 laboratory parameters during the acute agitation phase. of these five were reported at a frequency 35% ; wbc, k, fbs, ldh, and ua. for all of these the frequency of abnormal values was significantly lower after 35 weeks of treatment. in addition, the frequency of abnormal values of bun was also significantly lower during the recovery phases. thus, there appears to be a total of six laboratory parameters for which abnormal values are likely to occur during the acute excitation phase. in the present study abnormal values were defined as those outside the reference range and thus could be higher or lower values. all participants with abnormal wbc had values higher than the reference range, implying leukocytosis. all participants with abnormal k levels had values lower than the reference range, indicating hypokalemia. of the 28 participants with abnormal fbs levels, 26 had hyperglycemia and the remaining two had hypoglycemia. all participants with abnormal ldh, ua, and bun levels had values higher than the reference range. thus, during acute agitation there were signs of leukocytosis, hypokalemia, hyperglycemia, and elevations in ldh, ua, and bun. leukocytosis and hypokalemia seen during acute agitation were likely to be caused by the increased sympathetic tone which accompanies psychomotor excitation. the mechanism causing these abnormal laboratory values is thought to be the restraint of circulation of leukocytes by sympathetic tone, and the inflow of serum k into muscle cells during catecholaminergic stimulation.4 other findings that support this hypothesis include : (i) the low frequency of abnormal crp levels despite an increase in wbc ; (ii) the lower frequency of abnormal levels of na and ci than of hypokalemia ; and (iii) the lower frequency of abnormal cre levels related to renal function. finally when an improvement in panss - ec score was achieved, the frequency of these abnormal values was decreased, supporting a mechanism involving increased sympathetic tone via increased endogenous catecholamine activity. hyperglycemia may also be induced by catecholamines.5 ua and bun levels higher than the reference range may result from blood concentration due to excitation and dehydration.4 during the acute agitation patients are under considerable psychological and physical stress and treatment with an antipsychotic drug is immediately required. at the same time, it is necessary to perform adequate physical monitoring, including laboratory parameters. abnormal laboratory values should not, however, be regarded simply as manifestations of specific organ dysfunction and the effects of psychological symptoms (agitation, excitation) should taken into account. although in general there were less laboratory parameters with abnormal values in the recovery period, four parameters had a high frequency (25%) of abnormal values during the recovery phase : tg, amy, ck, and ldl - c. tg, however, was the only parameter for which the frequency of abnormal values was significantly increased during the recovery phase compared to the acute phase. in general the recovery phase was characterized by an increased frequency of abnormal values for parameters related to metabolism. the improvement in psychological symptoms and adverse effects of antipsychotic drugs are both potential explanations for the increase in abnormal values for parameters related to metabolism. the calming of the agitation and psychomotor excitation results in a shift from catabolism during the excitation to anabolism during the recovery phase. in addition the well characterized adverse effects of antipsychotic drugs on the metabolic system are also likely to induce abnormal values.6 the frequency of abnormal values for fbs, hba1c, tg, and ldl - c were all highest in the olanzapine group. the frequency of abnormal values for tg, in particular, was significantly higher in the olanzapine group compared with the risperidone group (p = 0.031). in an earlier study in which the laboratory data during the chronic phase during treatment with olanzapine and risperidone were compared, values for triglycerides and its intermediate metabolite, remnant - like cholesterol (rlp) were significantly higher with olanzapine compared with risperidone.7 results of the clinical antipsychotic trials of intervention effectiveness (catie) trials have shown that the dropout rate due to metabolic abnormalities was significantly higher with olanzapine and that increased triglyceride levels were frequently reported with this drug.8 increased triglyceride levels after meals are thought to constitute one of the important factors leading to cardiovascular events.9 results of a study analyzing triglyceride levels after meals in catie showed that olanzapine caused significant increases.10 thus, laboratory tests should be performed on a regular basis, especially in patients taking olanzapine. the polypharmacy group had the second highest frequency of abnormal values after the olanzapine group. multidrug combination therapy is thought to be associated with higher frequencies of metabolic syndrome and tg / hdl - c values greater than 3.5 are indicative of insulin resistance.11 as explained above, olanzapine and polypharmacy appear to be associated with risk for metabolic abnormalities. in the present study, the frequency of abnormal values for parameters related to metabolic effects during the recovery phase was increased in the olanzapine and polypharmacy groups, and the incidence of metabolic abnormalities appeared to vary depending on the type and number of antipsychotic drugs. the frequency of abnormal values during the recovery phase may depend on the type of antipsychotic drug. eleven participants had increased ck levels ; 11.1% (1 of 9) in the risperidone group, 50% (5 of 10) in the olanzapine group, 14.3% (1 of 7) in the haloperidol group, and 40% (4 of 10) in the polypharmacy group. this finding also shows that, as reported for metabolic abnormalities, increased ck levels tended to be more frequent in the olanzapine and polypharmacy groups. results of a study determining ck levels in patients with chronic - phase schizophrenia indicated that ck levels increased when olanzapine was used relative to typical antipsychotics, suggesting a probable metabolic problem in muscle cells.12 out of a total of nine participants in whom amylase (amy) levels were determined, the three who had abnormal values were all treated with olanzapine. it is interesting that the frequency of abnormal values for ck and amy by drug was similar to that of metabolic abnormalities. although in this research it is proposed that increased ck and amy levels may be related to metabolic abnormalities, this needs to be further investigated because of the small sample size used. there were certain specific limitations to the present study : (i) it was a nonrandomized retrospective study ; (ii) abnormal values were determined using standard reference values, with no adjustment for age or sex ; and (iii) the severity of abnormal values (the extent to which they exceeded the standard range) was not to taken into account. the frequencies of abnormal laboratory values during the acute and recovery phases of schizophrenia were investigated. during the acute agitation phase, the frequency of abnormal values for wbc, k, fbs, ldh, ua, and bun was high. the reason for this appeared to be the physical stress associated with agitation and excitation, such as increased sympathetic tone and dehydration. during the recovery phase olanzapine or polypharmacy appear to be associated with more abnormal values compared to risperidone and haloperidol. | during treatment of acute - phase schizophrenia, attention needs to be given to physical as well as psychological symptoms. it is often difficult, however, to obtain information on physical symptoms from patients with psychomotor excitation, and only laboratory examinations can provide objective data. the results of laboratory parameters measured in 68 patients with schizophrenia during psychomotor excitation and approximately 1 month later during the medicated recovery phase have been analyzed retrospectively. abnormal laboratory values during psychomotor excitation were frequent. the most frequent (35% of patients) were increased white blood cell count, low serum potassium levels, high levels of fasting blood sugar, lactate dehydrogenase and uric acid. there were fewer abnormal values during the medicated recovery phase. the most frequent (25% of patients) were high serum levels of triglycerides, amylase, creatinine kinase, and low - density lipoprotein cholesterol. abnormal triglyceride levels were found significantly more frequently in patients receiving olanzapine than those receiving risperidone. abnormal values during the acute phase may be the result of excitation such as increased sympathetic tone and dehydration. abnormal values during the recovery phase appeared to be related to the adverse metabolic effects of antipsychotic drugs. the frequency of these abnormal values was particularly high in patients receiving olanzapine alone or in combination with other medications. |
however many people refuse it because of fear of needle and back pain.1 many techniques have been used to obtund pain of needle insertion including infiltration analgesia and emla patch. local anesthetics themselves may produce pain on injection and many anesthetists are unsure that infiltration analgesia at the site of spinal puncture has any advantage over a straightforward puncture without analgesia. 2, 3, 4 the pain experienced during spinal puncture has both somatic and psychological components. pharmacological measures, such as the application of local anesthetics, treat only the somatic component of pain, whereas attention - diverting measures (pressing ball) address only the psychological component of pain. 4,5,6 a literature search revealed laboratory studies showing that baroreceptor activation induces nociception but there were few clinical studies exploring the effect of the valsalva maneuver on pain.6 - 10 in a study by agrawal, valsalva maneuver performed before venous canulation could decrease the incidence and severity of pain associated with venipuncture in adult patients.7 in another study by gupta the efficacy of balloon inflation were evaluated on venipuncture pain in children aged 6 - 12 year and there was a significant reduction of pain in balloon group compared with distraction and control groups.8 this study evaluated the efficacy of valsalva maneuver on needle projection pain and hemodynamic responses during spinal puncture. this randomized clinical trial was performed in dr.shariati hospital of tehran university of medical sciences from january to march 2010. the study protocol conformed to the ethical guidelines of the 1989 declaration of helsinki. to evaluate the effect of valsalva maneuver on pain, first, we searched medline, isi, and other databases. this trial was then registered with and approved by the research ethics committee of tehran university of medical sciences and iranian registry of clinical trials. ninety consecutive adults ' patients, either sex with asa physical status i and ii, scheduled for elective surgeries under spinal anesthesia, were included. patients having problems in communication, any contraindications to spinal anesthesia and patients who could not hold the mercury column up to 30 mm hg for a period of at least 20s and whose spinal puncture could not be performed in the first attempt were excluded. using a computer - generated randomization list, group i (c) : control ; group ii (b) : ball ; pressed a rubber ball (attention - diverting method) ; group iii (v) : valsalva ; blew into sphygmomanometer tubing and hold the mercury column up to 30 mm hg for a period of at least 20s. spinal needle projection pain was graded using numeric rating scale (nrs) : 1 - 10, where scales of 1 -3 were rated as mild, 4 - 6 as moderate, and > 6 as severe pain. in a pilot study of 20 patients having spinal anesthesia by 25-guage quincke needle without introducer and any local infiltration, 90% of them had moderate to severe pain using nrs and nobody had nrs=0 (unpublished observation). according to this pilot study the nrs was graded from 1 to 10. before the surgery, patients were instructed about the numeric rating scale (nrs) and how to blow into sphygmomanometer tubing. all patients were premedicated with 10 mg diazepam given orally on the morning of surgery. on arrival in the operating room, ecg electrodes and non - invasive blood pressure (nibp) monitor were applied and oxygen saturation was monitored by pulse oxymeter. the puncture pain was assessed by the patients, immediately after being placed supine for surgery using numeric rating scale (nrs). blood pressure and heart rate five minutes before the procedure, during spinal puncture and first and third minutes after that were also recorded. in a pilot study of 20 patients having spinal anesthesia by 25-guage quincke needle, 90% of them had moderate to severe pain using nrs (unpublished observation). presuming that valsalva maneuver during spinal needle projection would reduce pain to 50% ; one would need to enroll 30 patients in each group for the results to be statistically significant at a power of 95% with a level of confidence of 5%. one - way anova, kruskal wallis, chi - square and fishers exact tests were used when appropriate. repeated measures anova and post hoc tukey tests were used for comparing hemodynamic responses between the study groups. to evaluate the effect of valsalva maneuver on pain, first, we searched medline, isi, and other databases. this trial was then registered with and approved by the research ethics committee of tehran university of medical sciences and iranian registry of clinical trials. ninety consecutive adults ' patients, either sex with asa physical status i and ii, scheduled for elective surgeries under spinal anesthesia, were included. patients having problems in communication, any contraindications to spinal anesthesia and patients who could not hold the mercury column up to 30 mm hg for a period of at least 20s and whose spinal puncture could not be performed in the first attempt were excluded. using a computer - generated randomization list, group i (c) : control ; group ii (b) : ball ; pressed a rubber ball (attention - diverting method) ; group iii (v) : valsalva ; blew into sphygmomanometer tubing and hold the mercury column up to 30 mm hg for a period of at least 20s. spinal needle projection pain was graded using numeric rating scale (nrs) : 1 - 10, where scales of 1 -3 were rated as mild, 4 - 6 as moderate, and > 6 as severe pain. in a pilot study of 20 patients having spinal anesthesia by 25-guage quincke needle without introducer and any local infiltration, 90% of them had moderate to severe pain using nrs and nobody had nrs=0 (unpublished observation). according to this pilot study the nrs was graded from 1 to 10. before the surgery, patients were instructed about the numeric rating scale (nrs) and how to blow into sphygmomanometer tubing. all patients were premedicated with 10 mg diazepam given orally on the morning of surgery. on arrival in the operating room, ecg electrodes and non - invasive blood pressure (nibp) monitor were applied and oxygen saturation was monitored by pulse oxymeter. the puncture pain was assessed by the patients, immediately after being placed supine for surgery using numeric rating scale (nrs). blood pressure and heart rate five minutes before the procedure, during spinal puncture and first and third minutes after that were also recorded. in a pilot study of 20 patients having spinal anesthesia by 25-guage quincke needle, 90% of them had moderate to severe pain using nrs (unpublished observation). presuming that valsalva maneuver during spinal needle projection would reduce pain to 50% ; one would need to enroll 30 patients in each group for the results to be statistically significant at a power of 95% with a level of confidence of 5%. one - way anova, kruskal wallis, chi - square and fishers exact tests were used when appropriate. repeated measures anova and post hoc tukey tests were used for comparing hemodynamic responses between the study groups. there were no statistical differences in the demographic data between the study groups (p>0.05) (table 1). a significant reduction in nrs was observed in the valsalva group compared with the control and the ball groups (p=0.001) (table 2). the mean arterial pressure (map) throughout the time intervals of prespinal procedure to the third minutes after that were statistically different between the study groups (p= 0.008). post hoc tukey test showed there was statistical difference in map between the ball and the control groups at third minutes after spinal anesthesia (p=0.007) (fig 1). the mean heart rate (hr) throughout the time intervals of prespinal to the third minutes after that were statistically different between the study groups (p= 0.016). post hoc tukey test showed there was statistical difference in hr between the ball and the control groups at third minutes after spinal anesthesia (p=0.003) (fig 2). this study suggests that performing valsalva maneuver during spinal needle projection reduces the severity of spinal needle puncture pain. during valsalva maneuver intrathoracic pressure increases. this increase results in compression of the vessels within the chest and in turn results in baroreceptor activation. activation of either the cardiopulmonary baroreceptor reflex arc or the sinoaortic baroreceptor reflex arc induces antinociception.9,10 there were few studies for evaluating the efficacy of balloon inflation on venipuncture pain in pediatric and adult patients. in a study by gupta on seventy - five pediatric patients aged 6 - 12 yr, the efficacy of balloon inflation for attenuating venipuncture pain was evaluated. pain was self - reported by a pain face scale with a 10-cm visual analog scale (vas) placed at its back, where 0=no pain and 10 = worst imaginable pain. vas scores of 1- 3 were rated as mild, 4 - 6 as moderate, and 6 as severe. median (interquartile range) vas score in the balloon group was 1 (3), which was reduced as compared with 2 (2) and 4 (2) observed in the distraction and control groups respectively (p= 0.000). significant reduction in the incidence and severity of venipuncture pain was also observed in the balloon group compared with the other 2 groups (p<0.05).these findings were correlated with our study. in another study by agrawal, the efficacy of the valsalva maneuver on pain associated with venous canulation were evaluated on seventy - five adults patients undergoing elective surgeries. group i (c) : control ; group ii (v) : blew into sphygmomanometer tubing and raised the mercury column up to 30mmhgfor 20 s ; group iii (b) : pressed a rubber ball. venous canulation pain was graded using a 4-point scale : 0 - 3, where 0= no pain, 1= mild pain, 2= moderate pain and 3= severe pain, and visual analog scale of 0 - 10, where 0=no pain and 10= worst imaginable pain. agrawal used both, 4-point scale and vas, in which the first scaling was used during the canulation and vas was used after the canulation. they found a significant reduction in the incidence of pain in the valsalva group : 18 of 25 (72%) patients, whereas 25 of 25 (100%) experienced pain in the other two groups (p<0.001). these findings were also correlated with our study. in our study, only 9(30%) of patients had moderate and severe pain in valsalva group, where 24 (80%) in ball group and 21 (70%) in control group had moderate and severe pain. since valsalva maneuver may induce bradycardia and hypotension that are important during spinal needle projection, we also recorded hemodynamic responses that were not studied in previously mentioned studies. there were statistical differences in map and hr at third minutes after the spinal puncture between the ball and the control groups, these differences were not related to valsalva maneuver. in conclusion we suggest that valsalva maneuver can decreases the skin puncture pain associated with spinal needle projection while observing hemodynamic changes. | this study evaluated the efficacy of the valsalva maneuver that can induce baroreceptor activation and nociception, on needle projection pain and hemodynamic responses associated with spinal puncture. ninety adults, asa physical status i and ii undergoing elective surgeries were included. patients were randomized into three equal groups. group i (c) : control ; group ii (b) : ball ; pressed a rubber ball (attention - diverting method) ; group iii (v) : valsalva ; blew into sphygmomanometer tubing and hold the mercury column up to 30 mm hg for a period of at least 20s. spinal needle projection pain was graded using numeric rating scale (nrs) : 1 - 10, where scales of 1 - 3 were rated as mild, 4 - 6 as moderate, and > 6 as severe. blood pressure and heart rate, five minutes before the procedure, during the spinal puncture and first and third minutes after that, were also recorded. significant reduction in nrs was observed in the valsalva group compared with the control and the ball groups (p=0.001). there were statistical but no significant clinical differences in mean arterial blood pressure and heart rates between the study groups (p=0.008 and p=0.016 respectively). in conclusion valsalva maneuver can decrease the skin puncture pain associated with spinal needle projection while observing hemodynamic changes. |
the essential function of a classification system is as a navigation tool that helps direct research, evaluate outcomes, establish guidelines for best practices for prevention and care, and educate on all of the above. diabetes mellitus (dm) subtypes as currently categorized, however, do not fit into our contemporary understanding of the phenotypes of diabetes (16). the inherent challenges of the current system, together with the limited knowledge that existed at the time of the crafting of the current system, yielded definitions for type 1 dm, type 2 dm, and latent autoimmune diabetes in adults (lada) that are not distinct and are ambiguous and imprecise. discovery of the role played by autoimmunity in the pathogenesis of type 1 dm created the assumption that type 1 dm and type 2 dm possess unique etiologies, disease courses, and, consequently, treatment approaches. there exists, however, overlap among even the most typical patient cases. patients presenting with otherwise classic insulin resistance (ir)-associated type 2 dm may display hallmarks of type 1 dm. similarly, obesity - related ir may be observed in patients presenting with textbook the late presentation of type 1 dm provides a particular challenge for the current classification system, in which this subtype of dm is generally termed lada. leading diabetes organizations have not arrived at a common definition for lada (5). there has been little consensus as to whether this phenotype constitutes a form of type 2 dm with early or fast destruction of -cells, a late manifestation of type 1 dm (8), or a distinct entity with its own genetic footprint (5). indeed, current parameters are inadequate to clearly distinguish any of the subforms of dm (fig. 1). discussions and critiques of the current dm classification system are found in the literature (16). qualitative illustration of the spectrum of factors associated with different forms of dm, including the variable age at onset, lack of obesity, metabolic syndrome, genetic associations, different forms of immune changes, c - peptide secretion, and the need for insulin therapy. t1 dm, type 1 dm ; t2 dm, type 2 diabetes. adapted with permission from leslie. the fact that many obese patients with ir do not develop dm indicates that ir is insufficient to cause type 2 dm without predisposing factors that affect -cell function (9). patients diagnosed with lada who retain endogenous insulin production may receive default insulin therapy as treatment of choice. this decision is guided largely by the categorization of lada within type 1 dm, despite the capacity for endogenous insulin production. treatment options that do not pose the risks of hypoglycemia or weight gain might be both useful and preferable for lada but are typically not considered beyond use in type 2 dm (10). incretins and sodium glucose cotransporter 2 (sglt-2) inhibitors are examples of newer agents that have demonstrated potential and are being rigorously evaluated in the treatment of type 1 dm and lada (1017). the categorization of lada within type 1 dm also leads to myopia on the part of insurers. medications that could be logical choices as adjunctive or alternative therapies to insulin for candidate patients with lada are not designated as approved processes of care under the current classification system and accordingly are not covered by insurers. we believe that there is little rationale for limiting choice of therapy solely on the current definitions of type 1 dm, type 2 dm, and lada. we propose that choice of therapy should be based on the particular mediating pathway(s) of hyperglycemia present in each individual patient, as will be discussed. only large clinical trials can fully validate the best use of various agents across the spectrum of dm. in the interim, however, an evidence - based practice approach can allow for broader utility in routine care. metformin and pioglitazone may be safe and efficacious adjunctive therapies regardless of the current diagnostic category, as may be incretins (11,15,1723) and sglt-2 inhibitors (14,2426). it is reasonable that broader use of existing agents would extend to the management of maturity - onset diabetes of the young (23,27), as well as stress - related and steroid - induced dm. given the above discussion, the issue is not what is lada or any clinical presentation of dm under the current system. the issue is the mechanisms and rate of destruction of -cells at work in all dm. we present a model that provides a more logical approach to classifying dm : the -cell centric classification of dm. in this schema, the -cell centric classification system recognizes the interplay of genetics, ir, environmental factors, and inflammation / immune system on the function and mass of -cells (fig. the -cell centric concept can be applied to dm arising in genetically predisposed -cells, as well as in strongly genetic ir syndromes, such as the rabson - mendenhall syndrome (28), which may exhaust nongenetically predisposed -cells. finally, the -cell centric classification of all dm supports best practices in the management of dm by identifying mediating pathways of hyperglycemia that are operative in each patient and directing treatment to those specific dysfunctions. genetic determinants influence ir (whether centrally or peripherally induced), loss of -cell function and mass, environmental triggers (such as viruses, endocrine disruptors, food advanced glycosylation end products, gut biome), and immune modulation and inflammation. singly or, more commonly, in various combinations, these factors converge on the genetically susceptible -cell, impinge on -cell function and biology, and orchestrate the shift from normoglycemia to hyperglycemia. as this process takes place regardless of subtype of dm, the dysfunctional -cell is the final common denominator in all dm. the -cell centric construct suggests a more logical rationale to the eight core defects described by the ominous octet (29). our model recognizes a total of 11 interlocking pathways that contribute to hyperglycemia (fig. these mediating pathways of hyperglycemia are induced by the translation of genetic predispositions to ir, susceptibility to environmental influences, or immune dysregulation and inflammation to genetically predisposed, dysfunctional -cells. the -cell construct can incorporate newly discovered pathways to dysglycemia as these evolve, such as emerging research linking osteocalcin levels to a1c and homa of -cell function status (30). a : eleven currently known mediating pathways of hyperglycemia are shown. many of these contribute to -cell dysfunction (liver, muscle, adipose tissue [shown in red to depict additional association with ir ], brain, colon / biome, and immune dysregulation / inflammation [shown in blue ]), and others result from -cell dysfunction through downstream effects (reduced insulin, decreased incretin effect, -cell defect, stomach / small intestine via reduced amylin, and kidney [shown in green ]). the mediating pathways of hyperglycemia that contribute to -cell dysfunction include liver, muscle, and adipose tissue (organs associated with ir) and brain, colon, and immune dysregulation. this damage results in downstream hyperglycemia arising from increased glucagon secretion, as well as a reduction in insulin production, incretin effect, and amylin levels. even mild hyperglycemia resulting from -cell dysfunction can upregulate sglt-2 protein in the kidney, which further contributes to hyperglycemia (31). hyperglycemia, regardless of its source, leads to glucotoxicity, which further impairs -cell function. in a given patient, the specific mediating pathways of hyperglycemia at work are variable, though likely to involve multiple pathways (fig. three additional mediating pathways of hyperglycemia to those of the ominous octet (29) have been identified. systemic low - grade inflammation is observed in type 2 dm, type 1 dm, and lada (32,33) and has been shown to accompany the endoplasmic stress imposed by increased metabolic demand for insulin (34). early studies show incretins exert anti - inflammatory effects (35,36), which may account in part for their benefit. it would be of interest if a recent 1-year trial reporting the ability of a dipeptidyl peptidase 4 inhibitor to delay the progression of disease in lada patients (17) proved durable and reproducible. gut microbiota has been shown to be associated with type 1 dm, type 2 dm, and obesity and has been proposed to help explain the observation that only a portion of overweight individuals develop frank dm (38,39). reductions in amylin production in the diabetic state are a consequence of -cell dysfunction. decreased amylin levels lead to accelerated gastric emptying and increased glucose absorption in the small intestine, with corresponding increases in postprandial glucose levels. this pathway of hyperglycemia could theoretically be addressed, at least in part, by the ability of incretins to slow gastric emptying. a key premise is that the mediating pathways of hyperglycemia are common across prediabetes, type 1 dm, type 2 dm, and other currently defined forms of dm. accordingly, we believe that the current antidiabetes armamentarium has broader applicability across the spectrum of dm than is currently utilized. the ideal treatment paradigm would be one that uses the least number of agents possible to target the greatest number of mediating pathways of hyperglycemia operative in the given patient. it is prudent to use agents that will help patients reach target a1c levels without introducing drug - related hypoglycemia or weight gain. despite the capacity of insulin therapy to manage glucotoxicity, there is a concern for -cell damage due to ir that has been exacerbated by exogenous insulin - induced hyperinsulinemia and weight gain (41). in contrast, early data on some newer agents are suggestive of -cell sparing abilities. an improvement of early and late -cell response to glucose load has been reported with dipeptidyl peptidase 4 inhibitor treatment (18,21). incretins have been shown, in preclinical evaluations, to halt apoptosis, stimulate proliferation of -cells, increase insulin availability, improve -cell response to insulin (4446), and, in animal studies, preserve -cells (47). the -cell centric model recognizes that the final common denominator of dm is the genetically predisposed, dysfunctional -cell, which ultimately leads to compromised -cell function, loss in -cell mass, or depleted insulin content in the face of ir. these may include monogenic or polygenic defects that predispose to hyperinsulinemia, ir, more recently understood mechanisms such as inflammation by the immune system (4851), susceptibility to environmental factors (37,51,52), or other physiological factors that increase demand on or otherwise damage -cells such as elevated circulating lipids (37,5355) (fig. as not all carriers of genes associated with dm develop dm, susceptibility likely relies on combinations of genetic abnormalities, environment, and lifestyle factors to exacerbate underlying genetic predispositions. though research is nascent, implicated environmental factors have included endocrine disruptors (56), food additives (52), abnormal gut biome (38,39,57), and ingested advanced glycation end products (58). there is also evidence that certain environmental factors may epigenetically alter the genotype in reproductive cells, producing inheritable dm factors in future generations (59,60) (fig. 2). clinically evident dm ensues at or after the juncture when the combined gene environment trigger reaches a tipping point for sufficient -cell compromise to be expressed as phenotypic hyperglycemia. this fundamental concept applies to all forms of dm, substantiating that the final common denominator in dm is at the level of the -cell. in our model, as typical in obesity, ir is a monogenic or, more commonly, a polygenic disorder (59). additional contributing factors to ir may include inflammation (4851), changes in the gut microbiota (3740), and brain - modulated changes in metabolism (51,61,62). resulting hyperinsulinemia feeds back to the hypothalamus to further exacerbate peripheral ir (61,62). downstream effects of ir cause detriment to -cell function by mechanisms that may include inflammatory cytokines, adipocytokines, lipotoxicity, and decreased adiponectin, potentially representing a physiological scenario similar to that induced by hyperinsulinemia (63,64). the -cell centric construct suggests a more logical rationale to the eight core defects described by the ominous octet (29). our model recognizes a total of 11 interlocking pathways that contribute to hyperglycemia (fig. these mediating pathways of hyperglycemia are induced by the translation of genetic predispositions to ir, susceptibility to environmental influences, or immune dysregulation and inflammation to genetically predisposed, dysfunctional -cells. the -cell construct can incorporate newly discovered pathways to dysglycemia as these evolve, such as emerging research linking osteocalcin levels to a1c and homa of -cell function status (30). a : eleven currently known mediating pathways of hyperglycemia are shown. many of these contribute to -cell dysfunction (liver, muscle, adipose tissue [shown in red to depict additional association with ir ], brain, colon / biome, and immune dysregulation / inflammation [shown in blue ]), and others result from -cell dysfunction through downstream effects (reduced insulin, decreased incretin effect, -cell defect, stomach / small intestine via reduced amylin, and kidney [shown in green ]). the mediating pathways of hyperglycemia that contribute to -cell dysfunction include liver, muscle, and adipose tissue (organs associated with ir) and brain, colon, and immune dysregulation. this damage results in downstream hyperglycemia arising from increased glucagon secretion, as well as a reduction in insulin production, incretin effect, and amylin levels. even mild hyperglycemia resulting from -cell dysfunction can upregulate sglt-2 protein in the kidney, which further contributes to hyperglycemia (31). hyperglycemia, regardless of its source, leads to glucotoxicity, which further impairs -cell function. in a given patient, the specific mediating pathways of hyperglycemia at work are variable, though likely to involve multiple pathways (fig. three additional mediating pathways of hyperglycemia to those of the ominous octet (29) have been identified. systemic low - grade inflammation is observed in type 2 dm, type 1 dm, and lada (32,33) and has been shown to accompany the endoplasmic stress imposed by increased metabolic demand for insulin (34). early studies show incretins exert anti - inflammatory effects (35,36), which may account in part for their benefit. it would be of interest if a recent 1-year trial reporting the ability of a dipeptidyl peptidase 4 inhibitor to delay the progression of disease in lada patients (17) proved durable and reproducible. gut microbiota has been shown to be associated with type 1 dm, type 2 dm, and obesity and has been proposed to help explain the observation that only a portion of overweight individuals develop frank dm (38,39). probiotics and prebiotics may address this mediator of hyperglycemia. reductions in amylin production in the diabetic state are a consequence of -cell dysfunction. decreased amylin levels lead to accelerated gastric emptying and increased glucose absorption in the small intestine, with corresponding increases in postprandial glucose levels. this pathway of hyperglycemia could theoretically be addressed, at least in part, by the ability of incretins to slow gastric emptying. a key premise is that the mediating pathways of hyperglycemia are common across prediabetes, type 1 dm, type 2 dm, and other currently defined forms of dm. accordingly, we believe that the current antidiabetes armamentarium has broader applicability across the spectrum of dm than is currently utilized. the ideal treatment paradigm would be one that uses the least number of agents possible to target the greatest number of mediating pathways of hyperglycemia operative in the given patient. it is prudent to use agents that will help patients reach target a1c levels without introducing drug - related hypoglycemia or weight gain. despite the capacity of insulin therapy to manage glucotoxicity, there is a concern for -cell damage due to ir that has been exacerbated by exogenous insulin - induced hyperinsulinemia and weight gain (41). in contrast, early data on some newer agents are suggestive of -cell sparing abilities. an improvement of early and late -cell response to glucose load has been reported with dipeptidyl peptidase 4 inhibitor treatment (18,21). incretins have been shown, in preclinical evaluations, to halt apoptosis, stimulate proliferation of -cells, increase insulin availability, improve -cell response to insulin (4446), and, in animal studies, preserve -cells (47). the -cell centric model recognizes that the final common denominator of dm is the genetically predisposed, dysfunctional -cell, which ultimately leads to compromised -cell function, loss in -cell mass, or depleted insulin content in the face of ir. these may include monogenic or polygenic defects that predispose to hyperinsulinemia, ir, more recently understood mechanisms such as inflammation by the immune system (4851), susceptibility to environmental factors (37,51,52), or other physiological factors that increase demand on or otherwise damage -cells such as elevated circulating lipids (37,5355) (fig. as not all carriers of genes associated with dm develop dm, susceptibility likely relies on combinations of genetic abnormalities, environment, and lifestyle factors to exacerbate underlying genetic predispositions. though research is nascent, implicated environmental factors have included endocrine disruptors (56), food additives (52), abnormal gut biome (38,39,57), and ingested advanced glycation end products (58). there is also evidence that certain environmental factors may epigenetically alter the genotype in reproductive cells, producing inheritable dm factors in future generations (59,60) (fig. 2). clinically evident dm ensues at or after the juncture when the combined gene environment trigger reaches a tipping point for sufficient -cell compromise to be expressed as phenotypic hyperglycemia. this fundamental concept applies to all forms of dm, substantiating that the final common denominator in dm is at the level of the -cell. in our model, as typical in obesity, ir is a monogenic or, more commonly, a polygenic disorder (59). additional contributing factors to ir may include inflammation (4851), changes in the gut microbiota (3740), and brain - modulated changes in metabolism (51,61,62). resulting hyperinsulinemia feeds back to the hypothalamus to further exacerbate peripheral ir (61,62). downstream effects of ir cause detriment to -cell function by mechanisms that may include inflammatory cytokines, adipocytokines, lipotoxicity, and decreased adiponectin, potentially representing a physiological scenario similar to that induced by hyperinsulinemia (63,64). we propose that the -cell centric model is a conceptual framework that could help optimize processes of care for dm. a1c, fasting blood glucose, and postprandial glucose testing remain the basis of dm diagnosis and monitoring. precision medicine in the treatment of dm could be realized by additional diagnostic testing that could include c - peptide (1), islet cell antibodies or other markers of inflammation (1,65), measures of ir, improved assays for -cell mass, and markers of environmental damage and by the development of markers for the various mediating pathways of hyperglycemia. we uphold that there is, and will increasingly be, a place for genotyping in dm standard of care. pharmacogenomics could help direct patient - level care (6669) and holds the potential to spur on research through the development of dm gene banks for analyzing genetic distinctions between type 1 dm, lada, type 2 dm, and maturity - onset diabetes of the young. lifestyle modification is the starting point for intervention in prediabetes and dm as is normalization of dyslipidemia, given the links of prolonged lipid exposure with -cell dysfunction (9,5355). it is intuitively obvious that the constellation of mediating pathways of hyperglycemia in frank dm is likely the same as those in prediabetes. preferential use of agents with proven or strong evidence for -cell preservation is logical (70). the optimal strategy is to use the least number of agents to target the greatest number of mediating pathways of hyperglycemia operative in the given patient. it would use regimens that stabilize hyperglycemia across multiple causes, act synergistically to reduce cardiovascular and other risk factors, and preserve -cells. figure 3b illustrates the mediating pathways of hyperglycemia addressed by various available agents and provides a logic for the selection of complementary modes of action in combination therapy. our approach for using combination therapy is consistent with the recommendations within the 2015 american diabetes association (71) and 2015 american association of clinical endocrinologists (72) guidelines. we advocate the introduction of combination therapy early in the pharmacological management of the disease. critically, we avoid stratifying first-, second-, and third - line treatment sequencing. this stratification establishes undue competition between classes, which should more rightly be viewed as complementary options rather than salvage therapy after inevitable treatment failure (19,73). the ideal treatment regimens should not be potentially detrimental to the long - term integrity of the -cells. any benefits associated with sulfonylureas and glinides (including low cost) are not enduring and are far outweighed by their attendant risks (and associated treatment costs) of hypoglycemia and weight gain, high rate of treatment failure and subsequent enhanced requirements for antihyperglycemic management, potential for -cell exhaustion (42), increased risk of cardiovascular events (74), and potential for increased risk of mortality (75,76). fortunately, there are a large number of classes now available that do not pose these risks. empagliflozin has been recently shown to reduce cardiovascular outcomes and mortality in type 2 dm, while reducing weight and posing a low risk for hypoglycemia (24). insulin therapy induces hypoglycemia, weight gain, and a range of adverse consequences of hyperinsulinemia with both short- and long - term outcomes (7785). newer antidiabetes classes may be used to delay insulin therapy in candidate patients with endogenous insulin production (19). in patients requiring basal insulin, clinical research on novel combinations of classes, such as pramlintide (86) and incretins (19,22), bolus insulin accounts for most of the hypoglycemia seen with basal bolus insulin therapy (87). when insulin therapy is needed, we suggest it be incorporated as add - on therapy rather than as substitution for noninsulin antidiabetes agents. outcomes research is needed to fully evaluate various combination therapeutic approaches, as well as the potential of newer agents to address drivers of -cell dysfunction and loss. the principles of the -cell centric model provide a rationale for adjunctive therapy with noninsulin regimens in patients with type 1 dm (7,1216). thiazolidinedione (tzd) therapy in patients with type 1 dm presenting with ir, for example, is appropriate and can be beneficial (17). clinical trials in type 1 dm show that incretins (20) or sglt-2 inhibitors (25,88) as adjunctive therapy to exogenous insulin appear to reduce plasma glucose variability. this article highlights the need to replot the classification of dm, recognizing the -cell as the final common denominator of glucose dysregulation and the mediating pathways of hyperglycemia surrounding the -cell as the basis for treatment decisions. it can provide sage advice for preferential use of pharmacological interventions that address the mechanisms of hyperglycemia operative in an individual patient, avoid hypoglycemia and weight gain, and appear to be -cell sparing. novel anti - inflammation agents currently in phase 2 and 3 clinical development should be evaluated for safety and efficacy, and we should further explore suggestions that this approach could effectively treat, reverse, or even prevent dm with an inflammatory component (89). the -cell centric classification schema was envisioned as a stimulus to guide basic research, as well as clinical and translational research. it is hoped to help direct research on the genes involved in dm, the functions that these genes serve, the mechanisms that lead to -cell damage, the downstream effects of reduced -cell function, and any novel mechanisms of -cell pathophysiology. also needed the -cell centric model can be readily retrofitted into the terminology of the existing classification system. however, we submit that an entirely new nomenclature may likely best fulfill the imperative of bringing the classification in line with the known etiology and disease course. for all the above - stated reasons, we urge that the time is right to convene a committee of diabetes community leaders and researchers to reevaluate the current outmoded dm classification system. members of the american diabetes association, american association of clinical endocrinologists, european association for the study of diabetes, international diabetes federation, and world health organization should come together to address this immense, but vital, task toward delivering state - of - the - art, optimal patient care and directing future research. | the current classification system presents challenges to the diagnosis and treatment of patients with diabetes mellitus (dm), in part due to its conflicting and confounding definitions of type 1 dm, type 2 dm, and latent autoimmune diabetes of adults (lada). the current schema also lacks a foundation that readily incorporates advances in our understanding of the disease and its treatment. for appropriate and coherent therapy, we propose an alternate classification system. the -cell centric classification of dm is a new approach that obviates the inherent and unintended confusions of the current system. the -cell centric model presupposes that all dm originates from a final common denominator the abnormal pancreatic -cell. it recognizes that interactions between genetically predisposed -cells with a number of factors, including insulin resistance (ir), susceptibility to environmental influences, and immune dysregulation / inflammation, lead to the range of hyperglycemic phenotypes within the spectrum of dm. individually or in concert, and often self - perpetuating, these factors contribute to -cell stress, dysfunction, or loss through at least 11 distinct pathways. available, yet underutilized, treatments provide rational choices for personalized therapies that target the individual mediating pathways of hyperglycemia at work in any given patient, without the risk of drug - related hypoglycemia or weight gain or imposing further burden on the -cells. this article issues an urgent call for the review of the current dm classification system toward the consensus on a new, more useful system. |
considering the rapidly burgeoning older population, increased attention is being given to an accurate assessment of older adults ' cognitive and neuropsychological functioning. part of this process involves obtaining a viable estimate of their premorbid cognitive ability or their expected performance prior to any injury or relative decline in cognitive functioning. these premorbid estimates are critical toward determining the nature, type, and severity of cognitive impairment. it is vital when estimating the level of cognitive decline to account for variations in premorbid ability. for example, an older adult might be performing in the average range relative to his or her peers, but this could be a potential decline if his or her previous premorbid abilities were in the high average or superior range. it is also important to obtain a premorbid indicator in addition to age - based norms to account for other factors, such as formal education and occupation, that can contribute to one 's intellectual abilities. to this end one approach to assessing premorbid functioning among older adults involves the use of demographic variables, such as education, sex, handedness, and occupation. this approach can be useful because the data are gained without lengthy or invasive testing and independent of the patient 's current cognitive functioning and therefore remain constant throughout the patient 's adult life without being affected by any cognitive decline that may occur. the use of demographic variables has been shown in some studies to be a good estimate of premorbid intelligence among healthy controls and has been recommended over other premorbid estimates for those with alzheimer 's disease. demographic variables have been found in some cases to improve the accuracy of alternative approaches. however, other studies have found that demographic indices involving education in particular are not always the most accurate estimates of premorbid intelligence in normal aging and alzheimer 's disease [68 ], perhaps reflecting the intellectual development that may occur beyond formal education and continue throughout one 's life. to address some of the inadequacies in relying solely upon demographic variables, other methods of estimating premorbid intelligence have been suggested, such as the wechsler adult intelligence scale (wais) verbal iq, information, and vocabulary subtest scores [2, 9 ]. the most common approach is the use of word reading tests, which require the participant to verbally pronounce orthographically irregular words (e.g., it is assumed that correct pronunciation of these words, which do not follow common english grammatical rules, implies prior knowledge of them and therefore a higher premorbid intelligence. a variety of different word reading tests have been developed, all of which have their own particular strengths and weaknesses. one of the most common word reading tests is the national adult reading test (nart) [10, 11 ], which requires participants to read aloud a list of 50 irregular words. the nart appears to be a good estimate for healthy older adults and has been shown to be more resistant to the effects of age than the wais vocabulary subtest [1315 ]. although some researchers have found the nart to be a good premorbid estimate among those with dementia [12, 1517 ], others have found that it actually declines in dementia, therefore implying that it is not impervious to the effects of cognitive impairment [4, 1820 ]. similarly, while some researchers recommend that the nart should not be used among all adult populations, particularly those with organic conditions such as schizophrenia, korsakoff psychosis, or huntington 's disease [13, 21 ], others have not found any declines related to these conditions. alexander and colleagues found that the reading subtest of the wide range achievement test (wrat) correlated better than demographic estimates as a cerebral metabolic measure of premorbid cognitive functioning. the north american adult reading test (naart) was also found to be better than education as an estimate of both premorbid intelligence and overall cognitive functioning [7, 9 ].. found that both the wrat and the naart were equally accurate premorbid estimates for those in the average range of intelligence, but they were significant overestimates for those with lower intelligence and underestimates for those with higher intelligence. the spot - the - word test, which allows participants to choose the correct low - frequency english word from a pair of words that includes a nonword, has been found to be a good estimate for older adults with normal aging and even mild forms of dementia, but it appears to significantly decline in moderate - to - severe forms of dementia [26, 27 ]. the cambridge contextual reading test, which places nart words within a semantic context, seems to be a better estimate than the nart for those who have dementia or lower reading ability [2630 ]. another commonly used word reading test, the american version of the nart (amnart), was developed for american english speakers in the usa. depending on the version, this test consists of either 45 or 50 orthographically irregular english words, with about half identical to nart items. during administration, participants are instructed to pronounce each word out loud, beginning at the top of the list and continuing through the end. some researchers have suggested that the amnart is a good estimate of premorbid ability for older adults [3133 ].. discovered that it was a better premorbid estimate than demographic variables, which do not account for intellectual development occurring after the completion of one 's formal education. however, gladsjo and colleagues found that the amnart 's predictive strength was improved when it was used in conjunction with demographic estimates. one limitation of the amnart and similar word reading tests (e.g., nart, naart) is that they were developed as premorbid estimates in comparison with the wais - revised (wais - r). amnart - estimated iq, as calculated by using grober and sliwinski 's regression equation, therefore predicts premorbid intelligence in comparison with wais - r normative values. updated regression equations have not been published to convert amnart - estimated iq to the newer normative samples of the wais - third edition (wais - iii) or wais - fourth edition (wais - iv). despite the slightly outdated regression equation, the amnart remains a commonly used premorbid estimate, even in conjunction with the wais - iii [3842 ]. in fact, even after the publication of the wais - iii, schinka and vanderploeg still recommend using the amnart or a similar reading test along with demographic information (e.g., education level) and select wais - iii subtests (e.g., information, vocabulary) when attempting to predict premorbid performance. in the absence of regression equations that are updated for wais - iii or wais - iv normative values, in addition, there is insufficient amount of data regarding the utility of the amnart as a premorbid estimate for older adults. some researchers have argued that the amnart is not an equally valid measure for all populations and that it should be used with caution.. found that the amnart is an overestimate of premorbid functioning for those with lower intelligence, which may reflect a floor effect in the regression equation or a third, mediating variable. in addition, researchers have indicated that the amnart significantly declines in dementia and have recommended against its use among those with cognitive impairment [4446 ]. amnart scores have even been found to decline before the diagnosis of dementia is ever made, perhaps suggesting a link to the depletion of one 's cognitive reserve, which is the ability to employ compensatory cognitive strategies and utilize a variety of neural networks for problem solving. cognitive reserve is developed by factors such as education, occupation, and leisure activities and has been shown to act as a buffer against cognitive decline [4851 ]. the larger one 's cognitive reserve, the greater the degree of cognitive deterioration that must occur before symptoms of dementia or other forms of cognitive impairment can be detected. therefore, a decline in amnart - estimated iq, even before any formal diagnosis of cognitive impairment, may indicate an insidious depletion of one 's cognitive reserve. in light of the inconclusive data regarding the amnart, this study examined the utility of amnart - estimated intelligence scores as a measure of premorbid cognitive ability in older adults. this included examining how it compared to other commonly used measures of premorbid intelligence, namely, education, wais - iii verbal iq, and the wais - iii information subtest. to specifically examine the amnart 's utility among adults with varying levels of education similarly, the amnart was compared to other premorbid estimates among different aging - related diagnostic groups to consider its utility as cognitive functioning declines. in light of the limitations highlighted by other researchers, it was hypothesized that amnart - estimated premorbid ability would be significantly higher than all other premorbid measures and that it would be an overestimate of premorbid functioning for those with dementia and lower levels of education. the final results can help clarify and illuminate the most accurate assessment of older adults ' premorbid cognitive and intellectual abilities. one hundred and thirty community - dwelling older adults (69% female, 95% caucasian) between the ages of 56 and 104 voluntarily completed a comprehensive neuropsychological battery (see table 1 for demographic information). all data were gathered in compliance with the institutional review board affiliated with the authors ' institution. for purposes of examining measures of premorbid ability among older adults at various levels of educational attainment, the sample was divided into three educational groups : those with 012 years of education (i.e., high school or lower ; n = 17), those with 1316 years of education (i.e., college ; n = 68), and those with 17 or more years of education (i.e., graduate school ; n = 45). participants were classified as having normal aging (n = 35), age - associated memory impairment (aami ; n = 21), mild cognitive impairment (mci ; n = 59), or dementia (n = 15). aami was diagnosed according to crook and colleagues ' criteria, mci was classified according to petersen and colleagues ' criteria, and dementia was assessed using the diagnostic and statistical manual of mental disorders (4th ed. mean scores on the mini - mental status examination (mmse) were 29.23 (sd = 0.81) for the normal aging group, 28.81 (sd = 1.44) for the aami group, and 27.98 (sd = 1.85) for the mci group. the dementia group had a mean mmse score of 22.93 (sd = 3.83). the 45-item amnart was administered as the primary estimate of premorbid ability as part of a larger neuropsychological battery that included the mmse and eight subtests of the wais - iii. on the amnart, amnart - estimated iq score was calculated using grober and sliwinski 's formula, which also accounts for years of education. other premorbid estimates included wais - iii verbal iq (viq) and wais - iii information subtest (information) scores [34, 56 ]. all scores were then converted to z - scores for standardization and ease in statistical analyses. information z - scores were based on the normative data provided in the administration manual. viq and amnart - estimated iq scores were converted to z - scores based on a mean iq score of 100 and standard deviation of 15. they were notified that they would have the option for free feedback at a later time. a brief interview was conducted to gather demographic information, as well as to ascertain the presence of subjective memory complaints and difficulties with activities of daily living. participants were then administered a comprehensive neuropsychological battery that took approximately three hours to be completed. correlations were conducted between the amnart and demographic variables, as well as between the amnart and all other estimates of premorbid functioning (i.e., wais - iii viq and information). to explore differences in means between the amnart and other premorbid estimates, a one - way, repeated measures anova was conducted, with the premorbid estimates entered as different levels of the within - subject variable. to specifically compare premorbid estimates between those with different levels of education, a one - way manova was conducted, which examined performance on tests of premorbid intelligence among different educational groups (i.e., 012, 1316, and 17 or more years of education). to account for an interaction between the variables, a mixed - model anova was run with educational group entered as the between - group independent variable and premorbid estimate entered as the within - subject dependent variable. similarly, to assess differences among those with varying degrees of cognitive impairment, a one - way manova was conducted that examined premorbid estimates between diagnostic groups. to consider an interaction, a mixed - model anova was run with diagnostic group entered as the between - groups independent variable and premorbid estimate entered as the within - subjects dependent variable. amnart - estimated iq was significantly correlated with education, mmse, viq, and information (see table 1 for descriptive and inferential statistics). amnart performance was not correlated with age, and a t - test did not reveal significant gender differences in amnart scores, ps = ns. a one - way, repeated measures anova revealed significant differences between premorbid estimates, (3,369) = 35.61, p <.001. a scheffe 's post hoc test indicated that amnart - estimated iq was significantly higher than all other premorbid estimates, ps <.01. a one - way manova indicated significant differences between educational groups for amnart - estimated iq, f(2,127) = 11.95 ; viq, f(2,123) = 6.17 ; and information scores, f(2,124) = 5.69, ps <.01 (see table 2 for means). scheffe post hoc analyses revealed that the participants with 012 years of education had significantly lower scores on all premorbid measures than those with 17 or more years of education, ps <.02. amnart scores for those with 012 years of education were significantly lower than scores for those with 1316 years of education, which in turn were significantly lower than scores for those with 17 or more years of education, ps <.03. a mixed - model anova did not reveal a significant interaction between educational group and premorbid estimate scores, f(6,363) = 0.60, p = ns. a one - way manova indicated significant differences among diagnostic groups for amnart - estimated iq, f(3,126) = 8.60 ; viq, f(3,122) = 15.85 ; information scores, f(3,123) = 6.58, ps <.001 (see table 2 for means). scheffe post hoc analyses revealed that amnart - estimated iq and viq scores were significantly lower for those in the dementia group than for those in all other diagnostic groups, ps <.05. viq scores were significantly lower in the mci group than in the normal aging and aami groups, ps <.01, and there was a trend toward significance for amnart scores to be higher in the mci group than in the normal aging group, p <.06. in addition, information scores were significantly lower in the dementia group than in the normal aging and aami groups, ps <.01. a mixed - model anova revealed a significant interaction between diagnostic group and premorbid estimate, f(9,360) = 8.39, p <.001, such that the discrepancy between amnart scores and other premorbid estimates increased with greater cognitive impairment (see figure 1). this study was designed to investigate the amnart 's utility as an estimate of premorbid functioning for older adults. consistent with the original hypotheses and the intimations of other research [4446 ], the results suggest that the amnart may overestimate premorbid ability relative to other tests of premorbid intelligence. in particular, it may overinflate premorbid estimates among those with greater cognitive impairment and lower levels of education. overall, amnart - estimated iq was found to be significantly higher than scores on other indices of premorbid ability, namely, wais - iii viq and information. in fact, the mean amnart score was an average of one - half standard deviation above the other premorbid estimates. this suggests that the amnart may be an overestimation of premorbid ability in comparison with other established premorbid measures. while it is important not to underestimate premorbid iq, it is also equally crucial to avoid overestimation of premorbid ability. for instance, if an older adult was premorbidly performing in the average range but is estimated to have high average premorbid intelligence on the amnart, this would alter the threshold for determining the level or extent of cognitive impairment. for this individual, a clinician using the amnart might classify cognitive decline as any score in the lower end of the average range, when in fact these scores may be within normal limits and consistent with that older adult 's premorbid functioning. thus, an overestimation of premorbid intelligence is linked to an increased false positive rate for diagnosing the presence and severity of cognitive impairment. patients may be diagnosed with a more severe form of cognitive impairment than is objectively present. in addition, clinicians will have difficulty providing the best treatment to a patient without a clear and accurate assessment of his or her premorbid functioning. these are potentially detrimental and misleading errors. when educational groupings were examined, there were significant differences among all groups for all premorbid estimates. it would be expected that those who have completed more years of education would have a higher level of premorbid functioning, which is consistent with our findings. however, a qualitative analysis of the premorbid estimate means for those with 012 years of education yields interesting results. wais - iii viq information scores for those with 012 years of education were in the average range (57th and 64th percentiles, resp.), whereas mean amnart - estimated iq was in the high average range (81st percentile). this raises an important question of whether the premorbid ability of those with 012 years of education, whose highest educational attainment would be a high school diploma, should be estimated in the high average range based on the amnart. rather, those who have a high school education should generally cluster around average premorbid skills. it was also discovered that there was a significant decline in amnart - estimated iq scores among those with dementia, implying that the amnart does not remain unaffected by increased cognitive impairment. this is consistent with previous research [4447 ] and is expected considering that as a performance - based cognitive measure, the amnart should be somewhat affected by progressive dementia. this finding could also suggest that the amnart is increasingly less valid as individuals develop a greater degree of cognitive decline. this is supported by the finding that the discrepancy between amnart - estimated iq and all other premorbid estimates increased with greater cognitive impairment. amnart - estimated iq was particularly inflated relative to other premorbid estimates among older adults with mci and dementia. it may be that the amnart is most accurate as a premorbid estimate among those with normal aging, but the evidence from this study suggests that it even overinflates premorbid functioning in the normal aging group, as well as other aging - related diagnostic groups. another possibility is that amnart - estimated iq may be less affected by cognitive impairment and may be a better premorbid estimate among those with more severe forms of cognitive impairment. however, it still does not interact with cognitive decline in the same way as other commonly used premorbid measures, and therefore the amnart 's criterion validity decreases in the presence of greater cognitive impairment. it seems that the amnart overestimates premorbid ability across diagnostic groups, particularly among those with the greatest cognitive impairment. this overestimation of premorbid iq among those with greater cognitive impairment is particularly troubling, since premorbid measures are often of greater importance when working with individuals whose current level of cognitive functioning no longer matches their premorbid abilities, such as older adults with dementia. when interpreting and generalizing these results, one should keep in mind particular limitations. amnart - estimated iq was calculated using a regression equation that was developed in conjunction with the wais - r. therefore, any comparison between amnart - estimated iq and wais - iii scores should be held with a degree of tentativeness. considering the present study 's participants, a convenience sample was used ; therefore, those who participated may be more concerned about their memory or interested in scientific research than those who chose not to participate. in addition, the sample was highly educated, with the average level of education just under 16 years (i.e., a bachelor 's degree). there were a limited number of participants with a high school education, so the present findings regarding educational groups should be interpreted with some degree of caution. similarly, the diagnostic group sizes were not equivalent. finally, the sample was predominantly caucasian. future research should continue to explore these issues with a sample that is more evenly distributed and representative of the population. the implementation of a longitudinal design may contribute important information as to how premorbid estimates change with time and the development of cognitive decline. finally, this study should be replicated to compare amnart - estimated iq with wais - iv premorbid estimates. overall, the amnart appears to be an overestimation of premorbid ability in older adults. comparison with other premorbid measures indicates that the amnart seems to be most appropriate for use with those with higher levels of education, as well as with those experiencing normal aging. however, for all groups, the amnart appears to yield an inflated estimate of premorbid ability. these overestimates are clinically relevant, since such discrepancies between actual premorbid ability and amnart - estimated iq may lead to misdiagnosis of cognitive impairment or to the overestimation of the severity of cognitive decline. collectively, these results suggest that the amnart should be used cautiously with older adults, especially those with cognitive impairment or lower levels of education. future research should further examine the utility and accuracy of the amnart as a premorbid measure among older adults, and clinicians using this test should interpret amnart - estimated iq scores in conjunction with other premorbid estimates to guard against questionable estimates of premorbid functioning. | this study examines the utility of the american version of the national adult reading test (amnart) as a measure of premorbid intelligence for older adults. in a sample of 130 older adults, aged 56 to 104, the amnart was compared to other tests of premorbid intelligence. the results revealed that amnart - estimated iq was significantly higher than other premorbid estimates. across specific educational groups (i.e., 012, 1316, and 17 or more years of education), amnart - estimated iq was inflated relative to all other premorbid estimates. the amnart also declined as cognitive impairment increased, and there was a significant interaction between aging - related diagnostic group and premorbid estimate. the amnart may therefore overestimate premorbid ability relative to other premorbid measures, particularly among those with greater cognitive impairment and lower levels of education. these results suggest that the amnart should be used cautiously among older adults and in conjunction with other estimates of premorbid ability. |
reaching these niches is sometimes a challenging feat as pathogens have to travel long distances across organs, cross several cellular barriers, invade cells and travel inside the cell itself. the talk by robert mnard (pasteur institute, paris) followed one of these journeys. it travels the blood stream and reaches the liver where it divides, forms merozoites, infects other liver cells and finally enter the host 's bloodstream to invade erythrocytes and initiate disease. using intravital imaging, robert 's lab tracked the parasite from its injection site to the liver and came across several surprises. first, a large number of parasites persist in the skin, multiply and even form merozoites. a second surprise was that the exit step from the liver sinusoids occurs by endothelial cell traversal, a process that does not appear to require kpffer cells as was generally thought. arriving in proximity with host cells, pathogens can exploit the specific morphology of host cells to enter them. ari helenius (eth, zurich) described how vaccinia viruses use cellular extensions known as filopodia to reach the cellular body before entering host cells. exposed phosphatidylserine groups in the vaccinia virus viral membrane mimic apoptotic bodies and are used by the virus to enter the host cell by macropinocytosis in an epidermal growth factor receptor (egfr)-dependent manner. bacterial pathogens on the other hand have at their disposal an arsenal of secretion systems to invade cells such as the needle - like type iii secretion system (t3ss). salmonella enterica, for example is an intracellular pathogen that replicates within host - cell vacuoles and delivers virulence factors (effector proteins) through a t3ss. the process involves assembling the needle and shuttling the effector proteins across two bacterial membranes and the host vacuolar membrane in a highly coordinated sequential set of events. david holden (imperial college, london) showed recent data from his laboratory that address one aspect of this spectacular coordination. as the t3ss is assembled and crosses the vacuolar membrane, the intravacuolar salmonella can sense the neutral cytosolic ph, trigger degradation of a regulatory complex and finally allow effector translocation. legionella pneumophila is another intracellular pathogen that thrives inside macrophages in a vacuole that evades fusion with lysosomes. to control endosomal traffic and guarantee its intravacuolar survival legionella secretes an unexpectedly high number of effectors in the host cytoplasm through a type iv secretion system. (yale university, new haven) identified and functionally characterized a family of such effectors displaying an ankyrin repeat homology domain. one of these, ankx fragments the golgi and affects several endocytic rab proteins possibly interfering with the endocytic pathway in this manner. john boothroyd (stanford university) and isabelle tardieux (institut cochin, paris) both discussed toxoplasma gondii invasion and the role of rhoptries and their components in this process. isabelle showed how, toxofilin, one such component secreted into the host cell can mimic cellular cofilin by locally increasing actin flow and turn - over to disassemble the cortical actin meshwork at the site of invasion and facilitate vacuole folding. john focussed on different rhoptry components that enable the formation of a so - called moving junction that allows the parasite to slide into host cells in an intracellular vacuole. he highlighted the importance of rop16, a polymorphic kinase that mimics cellular jak2 and is injected into the host cell to activate stat6 and alter the host immune response. matt welch (university of california, berkeley) described how rickettsia express an actin modulator, sca2 that acts as a cellular mimic of formins that binds profilin to assemble the long parallel actin filaments that rickettsiae use for their intracellular movement. this molecular mimicry leads not only to the movement of the bacterium in the host cell cytoplasm but it also allows bacteria to move to neighboring cells. herpes viruses are another striking example as they travel from the epithelium to the nervous system. lynn enquist (princeton university) described how, once inside neurons, pseudorabies virus take advantage of the microtubule network to travel large distances from the cell body to the tip of the axon and back. lynn also discussed a viral strain, bartha that is impaired in its anterograde movement. surprisingly, it can instead jump to the axon of a neighbouring neuron and then move in a retrograde fashion to the cell body. after efficient replication and the full exploitation of the local resources pathogens may need to move to a different site inside the same host or to another host. guillaume dumnil (paris cardiovascular research center) explored this understudied step of the infection process during neisseria meningitidis infections. guillaume showed that after proliferating on the epithelial cell surface in the throat, neisseria trigger a cascade of events involving post - translational modifications of its type iv pili leading to the detachment of individual bacteria from the microcolonies. this detachment step allows the propagation of bacteria to new host but also the dissemination of bacteria across the epithelium, a prerequisite for invasive infections. from the experimental point of view, observation of some of these events requires real - time imaging techniques that are now a requirement in the field. spinning disc confocal microscopy and two - photon microscopy are necessary to efficiently describe how pathogens travel across tissues. at the cellular level, observation of single cells reveals a wealth of information but novel assays are needed to describe specific steps. for instance, jost enninja (institut pasteur, paris) reported the design of a fluorescence resonance energy transfer (fret)-based assay to visualize the exit of single shigella flexneri from the intracellular vacuole following its internalization. in another elegant analysis, this revealed a temporal link between the induction of interleukin 8 (il-8) with specific steps of the invasion process. in an attempt to maximize the impact of the limited number of proteins coded by the small genomes of the majority of the organisms that were discussed at the meeting, these pathogens have developed strategies to take advantage of the cell machinery for their own benefit or to target it as broadly as possible with their limited resources. one example common to many pathogens is exploiting regulators of post - translational modifications such as phosphorylation, ubiquitination or sumoylation. these modifications can affect the stability, localization or activity of the proteins that are targeted and pathogen effector molecules can use several of these components for different purposes. indeed, ari helenius (eth, zurich) presented data showing that vaccinia virus proteins are highly ubiquitinated and that the virus depends on a cluster of proteasome and ubiquitin related genes for disassembling its core and for its replication. ari 's approach consists of high throughput cellular small interfering (sirna) screens to identify what he calls the infectome, i.e. the cellular proteins and pathways involved in the entry of the viruses studied. jorge galan (yale university) also reported on salmonella typhimurium and the ubiquitination of its type iii effector phosphoinositide phosphatase sopb. sopb is ubiquitinated by the cellular machinery and its localization and functions depend on this ubiquitination. eric oswald (universite de toulouse) showed that also the cycle inhibiting factors (cif), produced by the enteropathogenic escherichia coli (epec) usurps another ubiquitin ligase complex. cifs are known to stabilize the cell cycle inhibitors p21 and p27, hence inducing cell cycle arrest. eric showed that epec cif (and also other cifs) induces accumulation of nedd8-conjugated cullins and binds components of the cullin - ring e3 ligase complex. by modulating the ligase activity of this complex, cifs could target proteins involved in processes such as control of the cell cycle progression, but also dynamic of actin cytoskeleton and host immune response. like some of the salmonella effector proteins discussed by jorge galan, chihiro sasakawa (university of tokyo) reported that this shigella e3 ligase promotes aberrant polyubiquitylation of nemo and tempers the nf - kappab - mediated inflammatory response to bacterial infection. clearly, the ubiquitin and proteasome machinery are part of the infectome of many pathogens. david ribet (institute pasteur, paris), from pascale cossart 's laboratory gave yet another example whereby the pathogen appears to be dampening the host response by affecting sumoylation. small ubiquitin - like modifier (sumo) are ubiquitin - like proteins that can also be covalently linked to protein substrates. david showed that listeria monocytogenes infection, and in particular the expression of its virulent pore - forming listeriolysin o (llo), induces degradation of the sumo e2 ligase ubc9 both in cell culture and in infected mice. the result is a decrease in the levels of cellular sumo - conjugated proteins, which is likely to be beneficial for the bacteria as sumo overexpression leads to decreased growth of intracellular listeria. among other broad regulatory molecules that pathogens elegantly the never - ending tale of antiviral rna silencing in plant pathogens was the topic of the talk of olivier voinnet (ibmp, strasbourg). although rnai is thought to act as a host defense mechanism, oliver described how plant viruses produce counter defense proteins that supress rnai pathways, and the host in turn produces resistance proteins that can counteract the pathogen 's counter defenses. bryan cullen (duke university) focused on human viruses and micro - rnas coded not only by the host genome but also by the virus itself. bryan detailed the role of the oncogenic kaposi 's sarcoma - associated herpesvirus (kshv) mir - k1 that appears to directly down - regulate p21 and overcome the cell cycle blocking activities of p53. this in turn will promote proliferation, hence increasing the carcinogenic potential of the virus. kshv mir - k11 is another interesting example of mirna - based interactions between viruses and humans. mir - k11 is an ortholog of the cellular mir-155 and consequently can affect expression of the same mir-155 targets. while in this case, the relevance of this regulation for viral infectivity, pathology or immune response is not yet clear the fact that epstein barr viruses (ebv) also affect the same mirna155this time by promoting its expression in b - cells and possibly affecting the nf - kappab pathway stabilizing latent persistence indicates an important function for this molecule in the context of infections. the apicomplexan t. gondii also expresses several small rnas, including heterochromatic sirna typical of plants and metazoan like mirnas that are likely to act as translational regulators in this system. mohamed ali hakimi (universit joseph fourier, grenoble) has been studying the small rnaome of this parasite and showed how varied it is and how dynamic its expression is. the fact that expression levels of several parasite mirnas differ greatly in freshly egressed or intracellular growing parasites indicates that they too are important regulators of toxoplasma 's life cycle. the first line of defense against pathogens is the presence of cellular barriers such as epithelia but pathogens use these barriers as docking sites to invade the host. in the case of hiv, it has been known that semen prevents the adhesion of the virus to epithelial cells. sebastian amigorena (curie institute, paris) reported that the soluble protein clusterin is the active component in semen that blocks hiv capture. although this protein is expressed ubiquitously, its glycosylation state is key for the effect and specific for semen. despite this early blockage the tripartite motif 5 (trim5) is a host e3 ubiquitin ligase which plays an important role in restricting hiv-1 infections in several primates. trim5 directly binds to the hiv-1 capsid in the cytoplasm and prevents reverse transcription of the virus by mechanisms that remain unclear. studying downstream signaling cascades and transcription regulation jeremy luban suggested that trim5 acts as a pattern - recognition receptor to detect hiv and trigger an intracellular antiviral response. mounting an efficient acquired immune response requires the coordination of several different cell types such as dendritic cells, b cells and t cells. one key challenge is understanding how rare populations of cells, for example antigen specific t cells and ag bearing dendritic cells encounter each other in infected animals. using intravital two - photon microscopy, ron germain demonstrated that t cells move along a fibroblastic reticular cell (frc) network in the lymph nodes. this movement favours t - cell contacts with dendritic cells that are sitting on the same network and deposit chemoattractants on the fibres. in his talk, of particular interest was the correlation observed between antigen - induced arrest of effector t - cell migration and interferon (ifn)-gamma production, consistent with the immune system focusing effector cytokine production locally where and when it is needed. salmonella can also cause long - term chronic infections and denise monack (stanford university) reported that ssei, one effector of the t3ss expressed by salmonella contributes to persistent infection in mice. the protein is able to reduce the motility of infected dendritic cells towards the t - cell and can thus slow down the immune response and favour persistence. ssei appears to exert its inhibitory function by interacting with the iqgap1 protein, an actin binding protein involved in cellular morphogenesis. arthur scherf (pasteur institute, paris) detailed another evasion strategy, this time by the parasite plasmodium. plasmodium repeatedly changes the expression of variant molecules at the surface of infected erythrocytes to avoid immune clearance and arthur described the epigenetic regulation of the expression of the var genes as well as their particular spatial organization and tethering to the transcriptionally repressive nuclear periphery and the role that actin appears to have in these phenomena. both innate and acquired immune responses need to be tempered before their activation causes unnecessary damage. arturo zychlinsky (max planck institute for infection biology, berlin) provided a striking example of this necessity by describing the role of neutrophils and neutrophil extracellular traps (nets) in systemic lupus erythematosus (sle). nets are extracellular structures composed of deoxyribonucleic acid (dna), histones and neutrophil proteins that capture and kill pathogens. serum dnase1 is essential for disassembly of nets after an infection and arturo reported that a considerable number of sle patients ' sera do not degrade nets efficiently. these patients either lack an active dnase i or the nets produced are resistant to dnase i activity because of the large amounts of autoantibodies accumulated in these structures. the net result (so to speak) is the accumulation of nets and the inflammatory syndrome that results. a workshop on infection biology would not be complete without mentioning the well - established role of several infections in cancer development. harald zur hausen (dkfz, heidelberg) dedicated most of his career to proving the involvement of high - risk human papilloma viruses in cervical cancer and his closing keynote lecture highlighted many other instances where an association between a particular infection and cancer exists or is to be expected. zur hausen put forward the hypothesis that an infectious agent may be associated with the higher incidence of colorectal, lung and breast cancer in red meat consumers. the field of infection biology is replete with incredible examples of strategically planed and well - executed pathogen interventions. the host response is not always effective and indeed the outcome of an infection is often deleterious to the host. however, the quality of the science, the nature of the discoveries and the imaging and technical advances presented at this meeting made it clear that we are well on the way to knowing our enemies and ourselves. | the first embo workshop on emerging themes in infection biology was held last june in the south of france. it gathered scientists working on various pathogens from viruses and bacteria to larger eukaryotic fungi and parasites. topics included not only the crosstalk between pathogens and their hosts but also the tools researchers are using to study and image such cellular and molecular conversations.so it is said that if you know your enemies and know yourself, you can win a hundred battles without a single loss.the art of war, sun tzu |
intussusception of the appendix is an uncommon type of intussusception with an incidence of 0.01% (1). although it may clinically mimic more common acute and chronic abdominal conditions, it is an important entity to recognize since it could be discovered as cecal mass and mistaken for a neoplasm. in korea, two cases of intussusception of the appendix were reported ; these were diagnosed by barium enema (2, 3). a 17-yr - old female admitted with repeated periumbilical pain, nausea, vomiting and febrile sensation. physical examination revealed mild periumbilical tenderness without rebound tenderness. in laboratory data, white blood cell count was 12,800/l and the other findings were normal. simple abdominal radiography and abdominal computerized tomography revealed no abnormality except mild paralytic ileus. the following day a colonoscopy was performed, which revealed a polypoid lesion in the cecum (fig. 1). an exploratory laparotomy was performed and it was seen dimple at cecum, the appendix invaginated into the cecum and an appendectomy was performed. the appendix was 2.5 cm in length, and it showed a chronic inflammation pathologically (fig. intussusception of the appendix is a rare entity that is difficult to diagnose preoperatively. in 1858 mckid (4) first described a complete invagination of the appendix into the cecum of a 7 yr - old boy. in 1890, the first operation for appendiceal intussusception in a 13-month - old child was reported (5). wright, renshaws, pitts and mcgraw reported successful operations for appendical intussusception (6). in 1964, collins (1) concluded a 40-yr study on 71,000 appendices obtained from surgical and autopsy material and reported prevalence of 0.01% for intussusception of the appendix ; the prevalence of the endometriosis and adenocarcinoma of the appendix were 0.05% and 0.08%, respectively. intussusception of the appendix may occur at any age, however, the majority of the reported cases reviewed in the earlier literature occurred in the pediatric aged group, with the average of 16 yr (5, 6, 8). the condition is more common four to five times in males than in females (9). the signs and symptoms of the intussusception of the appendix are vague, recurrent, cramping abdominal pain, intermittent rectal bleeding and mucus in the stools (6, 9, 10). the condition assumes a greater importance in the differential diagnosis of right lower quadrant abnormalities, particularly if the symptoms are subacute or recurring (11). (12) described five possible clinical features : 1) acute appendicitis, 2) intussusception, 3) recurrent right iliac fossa pain, 4) intermittent painless rectal bleeding, and 5) asymptomatic findings at laparotomy, barium enema, or colonoscopy. intussusception of the appendix probably occurs by the same mechanism and pathogenesis as intussusception elsewhere and has been reviewed extensively (5, 7, 13). many pathological conditions have been considered responsible for its presentation, including abnormal appendiceal peristalsis, irritation of the appendix secondary to fecalith, foreign body, polyp, parasites, and lymphoid hyperplasia, but the appendix may be normal (6, 9). other causes such as mucocele, appendiceal adenocarcinoma, carcinoid tumor and endometrial implants have not been reported in the pediatric age group. however, intussusception may occur in an appendix even without an underlying abnormality (6). anatomically, various types of the appendiceal intussusception have been reported (5). in 1941, mcswain (8) modified the original classification of the appendiceal intussusception by moschcowitz (5), which in turn was simplified by langsam. the tip of the appendix is the intussusceptum and its more proximal portion is the intussuscipiens., the proximal portion of the appendix forms the intussusceptum and is received into the distal portion. in type 4, there is a complete inversion of the appendix into the cecum from progression of types 1 and 2. either type 1 or type 2 can result in a complete inversion of the appendix into the cecum thereby resulting in the inside - out appendix as demonstrated in 2 cases (14). intussusception of the appendix has been noted during barium enema, sometimes in asymptomatic patients, and the intussusception has been reduced during the study (15, 16). levin. (15) reported 11 cases with the characteristic coiled - spring sign in the cecum and nonfilling of the appendix on double - contrast barium enemas. therefore, this finding defect has to be differentiated from other causes such as polyps, carcinoma, and lymphoma by colonoscopy (12). radiological findings may be : 1) no abnormality seen in cecal region without any mention of the appendix ; 2) oval or round bosselated intraluminal filling defects, usually in the medial wall of the cecum, with no visualization of the appendix ; 3) intraluminal finger - like filling defects within the cecum, usually arising from the medial wall of the cecum ; or 4) reduction of the filling defect out of the cecum during fluoroscopy (16, 17). sonogram has a proven sensitivity for the diagnosis of intussusception, and the typical sonographic appearances are well documented (6, 11). it enables a physician to identify a lead point within a complex target sign, but there are few publications about the sonographic pattern of intussusception of the appendix (6, 18). axial sonogram, intussusception of the appendix may present with a permanent multiple concentric sign. intussusception of the appendix may produce a target like appearance or concentric ring sign on ultrasound examination (19). colonoscopic view of the appendiceal intussusception has often been misdiagnosed as a cecal polyp or a neoplasm (20, 21). it is important for the endoscopist to remember that the appendiceal intussusception may mimic a cecal polyp, especially if the appendiceal lumen is not identified. therefore, it is important to make an accurate preoperative diagnosis in order to avoid endoscopic removal and the potential hazards. pathologically intussusception of the appendix has the following characteristics : 1) lesions of the wall - fecaliths, foreign bodies, and peristalsis ; 2) lesions of the wall - hypertrophic lymphoid follicles and adenovirus infections affecting the younger age groups ; mucosal polyps, mucoceles, adenocarcinoma, carcinoid, endometriosis, and tuberculosis (8, 9, 23). | intussusception of the appendix is an uncommon condition and the diagnosis is rarely made preoperatively. intussusception of the appendix may mimic a neoplastic lesion. colonoscopy is a valuable tool for diagnosis of the appendiceal intussusception. a 17-yr - old female admitted with repeated abdominal pain, nausea, vomiting and febrile sensation. we diagnosed as appendiceal intussusception by colonoscopy, which showed a polypoid tumor (about 1.5 cm) in the cecum. this sessile polypoid mass looks like foreskin or glans. we present colonoscopic finding of appendiceal intussusception and review the literature. |
an obvious example of a current debate within hoarding research is the question of where hoarding belongs within our diagnostic nosology. the uncertainty regarding the most appropriate classification of compulsive hoarding syndrome has had important consequences for our understanding of hoarding, and in some ways has constituted an obstacle to hoarding research. the lack of clear placement within dsm has led to an underestimation of the significance of the burden of disease associated with compulsive hoarding, inconsistencies with respect to an appropriate clinical comparison group in hoarding research, difficulties comparing findings across hoarding studies, and misconceptions regarding which assessment and treatment models are most relevant to hoarding. in the dsm - iv - tr, hoarding is described as difficulty discarding items, and is listed as one of the eight diagnostic criteria for obsessive - compulsive personality disorder (ocpd). accumulating evidence, however, suggests that it is misleading and invalid to classify hoarding as part of ocpd. when studies examining the prevalence of ocpd in hoarding samples exclude the criterion describing difficulty discarding, most studies suggest that hoarding is no more associated with ocpd than it is with other axis ii disorders. in addition, hoarding has been found to have the lowest specificity and predictive criteria of all eight of the diagnostic criteria for ocpd based on these findings, saxena argued convincingly that hoarding should be removed from the diagnostic criteria for ocpd. nevertheless, there is some evidence to suggest a link between hoarding and ocpd. a recent study of hoarding within a collaborative ocpd genetics study found that hoarders had a greater prevalence of certain ocpd traits, particularly miserliness and preoccupation with details. in addition, thus while the consensus appears to be that hoarding is inappropriately classified as a criterion of ocpd, the broader issue of the relation of hoarding to ocpd, as well as to other axis ii disorders, remains unresolved. despite its placement in the diagnostic and statistical manual of mental disorders (dsm)-iv, clinicians and researchers typically consider hoarding a symptom or subtype of obsessive - compulsive disorder (ocd). for example, the y - bocs checklist lists hoarding obsessions and compulsions, and many investigations into hoarding have involved comparing ocd individuals with and without hoarding. this view of hoarding as part of ocd derived from early findings that approximately one third of individuals with ocd have hoarding symptoms. more recent studies, however, have found ample evidence that hoarding should not be conceptualized only as an ocd symptom. for example, wu and watson found that hoarding correlated more weakly with other symptoms of ocd than these other symptoms correlated with each other. moreover, saxena found that patients who hoard, compared with other ocd patients, had different functional neuroimaging findings, response to treatment, and clinical profiles. in a large study of hoarding among ocd patients, individuals with hoarding were more likely to have symmetry obsessions and counting, ordering, and repeating compulsions. they also were more likely to have greater illness severity, more difficulty initiating and completing tasks, and problems with indecision. a recent study by abramowitz and colleagues provided further evidence that although some individuals with ocd may show hoarding behavior, hoarding is most likely distinct from ocd. abramowitz and colleagues compared ocd patients, patients with other anxiety disorders, and unscreened undergraduate students. ocd patients scored higher on all ocd symptoms except hoarding, in which the student group scored slightly, but significantly higher than both clinical groups. similarly to wu and watson, abramowitz and colleagues found that the magnitude of the correlations between hoarding and other ocd symptoms was significantly weaker than the magnitude of the correlations amongst all other ocd symptoms. in addition, the hoarding items loaded weakly on a unitary oci - r factor. in a second study, abramowitz found that hoarding was correlated weakly with depression, but not with anxiety. other ocd symptoms showed at least a moderate association with anxiety. due to these recent findings, there is a growing consensus that hoarding should not be considered as a symptom of ocpd or ocd, but as a separate clinical syndrome. several researchers have also examined whether there are important differences between hoarding behavior seen in the context of ocd and hoarding that occurs without any other ocd symptoms. a recent study conducted by petrusa compared individuals with severe compulsive hoarding who met criteria for ocd (ocd plus hoarding group) with individuals with severe hoarding who did not meet criteria for ocd (monosymptomatic hoarding). individuals in the ocd plus hoarding group differed from the monosymptomatic hoarding group in several important ways. for example, ocd plus hoarding participants were more likely to hoard bizarre items and more likely to report other obsessions and compulsions related to their hoarding than those in the monosymptomatic hoarding group. in addition, the ocd plus hoarding group endorsed more cluster c personality traits than the monsymptomatic hoarding group. given that hoarding can occur in the absence of ocd and that it shares some similarity to impulse control disorders (icds) such as pathological gambling, pyromania, and kleptomania, it may have a place within behavioral addiction. although hoarding behavior is sometimes motivated by a desire to reduce anxiety, it also sometimes appears to be driven by anticipation of pleasure and impaired self - regulation. since both anxiety and approach behaviors may play a role in compulsive hoarding, samuels reported a greater frequency of trichotillomania and skin picking among hoarding compared with nonhoarding individuals with ocd. in addition, frost found that pathological gamblers reported significantly more hoarding symptoms than light gamblers. although grant found a low prevalence of icds overall among individuals with obsessive - compulsive disorder, obsessive - compulsive disorder participants with a lifetime and current impulse control disorder were more likely to report hoarding symptoms. in a recent study, hayward and coles examined the relation of hoarding to ocd and icds in an undergraduate sample, and found that hoarding behaviors were related moderately to symptoms of compulsive buying, and more weakly related to pathological gambling, trichotillomania, and kleptomania. the possible association between hoarding and icds is consistent with mcelroy and colleagues ' conceptualization of a compulsive - impulsive spectrum, but requires further exploration. the nosological issues surrounding hoarding will influence its placement in the next edition of the dsm. one position is that compulsive hoarding should be included in our diagnostic system as an independent syndrome, which is sometimes comorbid with ocd. including hoarding as a separate syndrome has a number of important practical advantages, well - summarized by rachman and colleagues. for example, it would expand the boundaries of the hoarding population to be consistent with the data showing a high incidence of hoarding not associated with ocd. it would also encourage clinicians and researchers to use hoarding - specific assessment tools rather than measures designed for ocd, and facilitate the development of new treatment methods for hoarding. another possibility is that hoarding may be listed in dsm-5 as both a separate syndrome and as an ocd symptom. hoarding researchers also have made substantial progress in understanding the prevalence and manifestation of compulsive hoarding in the population. until very recently, researchers estimated the prevalence of hoarding as a subportion of individuals with ocd in the community. similarly, information regarding the burden of hoarding was based on anecdotal evidence and small samples. recent epidemiological studies, however, suggest that compulsive hoarding may be far more prevalent and burdensome in the community than previously thought. data from the baltimore epidemiologic catchment area follow - up survey suggest that 5% of the general population experiences clinically significant hoarding, while data from the national comorbidity survey replication indicate that the lifetime prevalence of compulsive hoarding may be as high as 14%. these studies estimated hoarding based upon reports of difficulty discarding, and did not specifically target clutter and excessive acquisition, and thus it is unknown whether cases met criteria for compulsive hoarding as defined by frost and hart. a recent twin study that utilized a self -report instrument to assess the broad hoarding phenotype found that 2% of its sample reported clinically significant hoarding symptoms. as symptom severity obtained by self - report tends to be lower than clinician - rated severity, the current prevalence of clinically significant compulsive hoarding may be somewhere between 2% and 5%. importantly, a large proportion of individuals who hoard report having at least one first - degree relative who experiences hoarding problems. in a sample of individuals with ocd, samuels and colleagues reported that probands of individuals with hoarding symptoms were four times more likely to experience hoarding symptoms than probands of individuals who did not report hoarding symptoms. genetic factors and unshared environmental factors may explain this familial connection. in a large sample of female twins, genetic factors accounted for approximately 50% of the variance in compulsive hoarding, while shared environmental factors encountered by twins growing up in the same household did not substantially contribute to the other half. samuels and colleagues reported that hoarding was almost three times more prevalent in individuals over the age of 54 than it was in individuals aged 34 to 44. this finding most likely is due to compulsive hoarding being a chronic and progressive disorder. hoarding symptoms often develop during childhood or adolescence, and become clinically significant during middle age. having the means to acquire and accumulate objects as a child may be substantially restricted ; therefore, it may take a decade or more for symptoms tobecome clinically significant. in other cases, hoarding may have a sudden onset in adulthood, such as after a traumatic life event or brain injury fifty - five percent of grisham and colleagues ' sample reported experiencing a stressful life event at the onset of hoarding symptoms, and these individuals had a significantly later age of onset than individuals who did not experience a stressful life event. clinical studies have demonstrated that hoarding often co - occurs with other psychological disorders. in a large clinical sample, almost all individuals with a hoarding diagnosis met criteria for another axis i disorder, and these individuals had significantly more co - occurring disorders than nonhoarding individuals with ocd. compared with nonhoarding individuals with ocd, hoarders are consistently more likely to meet criteria for social anxiety disorder, bipolar disorder, and pathological grooming behavior. hoarders also appear more likely to experience an alcohol - use disorder at some point in their lives. a community study has found that the prevalence of co - occurring disorders differs for men and women. in men, hoarding is associated with generalized anxiety disorder and tics, while among women, hoarding is associated with social phobia, post - traumatic stress disorder, body dysmorphic disorder, nail biting, and skin picking. women and men also may not be affected equally by hoarding symptoms. while clinical samples tend to be predominantly female, the identification of a significant prevalence of men who compulsively hoard, and genderspecific comorbidity differences, presents a significant challenge for developing and engaging all individuals in effective treatment. a growing body of research suggests that hoarding is associated with a lower quality of life. although longitudinal studies are needed to determine if hoarding is a cause or consequence of financial insecurity, a recent internet study indicated that hoarding may at least contribute to financial insecurity. five percent of the web sample reported they had been fired because of hoarding, and on average, employed individuals reported seven psychiatric work impairment days per month. individuals who hoard are very likely to be overweight or obese and suffer from a severe medical condition. third, several clinical and community studies have reported a low rate of marriage among compulsive hoarders. those who are married or cohabitating tend to have a lower degree of hoarding severity fourth, hoarding is associated with high rates of family frustration. family members who cohabit with hoarders report being embarrassed about the condition of their home, arguing about the clutter, and feeling rejection and hostility toward the hoarder. in summary, emergent research suggests that the prevalence of compulsive hoarding ranges from 2% to 5%, and men may be more likely to hoard than women. in most cases, hoarding is a chronic disorder. although some people may experience a gradual rise in symptoms throughout their lifetime, others may develop hoarding symptoms quite quickly after a stressful life event. men and women who hoard may experience different cooccurring disorders, yet both genders are likely to experience a substantial amount of burden associated with their hoarding. the initial clues that hoarding was related to frontal - lobe dysfunction came from case reports of pathological collecting and saving that began after a brain injury, typically along with other changes in personality and social functioning. in the last decade, two papers presented findings suggesting that hoarding is the result of frontal - lobe lesions. in the first report, hahm and colleagues described the case of a 46-year - old korean man who began unusual collecting behavior after he suffered an injury to his left ventromedial prefrontal cortex and caudate. this man had difficulty with social decisionmaking and judgment processes. in the second report, anderson examined compulsive hoarding behavior within a sample of 86 patients with focal lesions, and found that 13 of these participants exhibited abnormal collecting behavior. magnetic resonance imaging (mri) showed that all 13 individuals with hoarding symptoms had damage to the mesial frontal region of the brain, including the right polar sector and anterior cingulate. if excessive collecting and saving behaviors can begin after brain injury, individuals who hoard in the absence of lesions may possess similar deficits in neuropsychological functioning or impaired self - regulation that contribute to compulsive hoarding symptoms. self - report and laboratory studies of neuropsychological functioning in hoarding have highlighted potential areas of subtle impairment. in a study by hartl, hoarding patients reported increased symptoms of attention deficit - hyperactivity disorder (adhd). they also have been found to perform worse on certain neuropsychological tasks, including measures of attention and nonverbal intelligence, memory, and decisionmaking. deficits in executive function marked by inhibition, planning, and decision - making difficulties may limit hoarders ' ability to discard and organize their possessions. although this is an intriguing and rapidly advancing area within hoarding research, there has been some inconsistency with respect to the specific pattern of deficits associated with hoarding. they tend to believe a disproportionate number of their possessions are very important, and feel paralyzed by seemingly commonplace decisions about what items to discard and what items to keep, which items are valuable, and how to organize the items they decide to keep. these decision - making problems have been associated with hoarding in several studies using self - report measures. with respect to laboratory studies, however, research has provided mixed results regarding decision - making deficits. grisham found that hoarders displayed relatively intact decision making on the iowa gambling task relative to a clinical and community control groups. a recent study in our laboratory has replicated this finding, showing that individuals with compulsive hoarding did not demonstrate decision - making problems on the computerized cambridge gambling task. however, lawrence found that hoarding symptoms were associated with specific decision making impairments on the same gambling task and that these deficits were related to the severity of the hoarding symptoms. lawrence suggested that hoarders have difficulty deciding whether to save or discard their possession due to general decision - making difficulties. one important difference between the grisham and lawrence studies was the composition of the hoarding group. in the grisham study, the hoarding group comprised participants who met criteria for compulsive hoarding, regardless of whether they had ocd, while the hoarding group in the lawrence study consisted of ocd patients who displayed hoarding behaviors. this difference in the samples may explain the discrepancy on the decision - making task in the two studies. future studies may compare hoarding patients with and without other ocd symptoms to nonhoarding ocd patients and community controls in order to clarify the source of the decision - making difficulties. another area that remains unresolved is the role of proposed categorization problems in hoarding patients. compulsive hoarding patients appear to exhibit problems grouping their possessions into categories, which contributes to the disorganization and clutter that are hallmark features of this disorder. wincze contrasted hoarding participants, obsessive -compulsive nonhoarding participants and healthy control participants on categorization tasks. the results of this study suggested that categorization problems occur only when compulsive hoarders sort their own possessions. in contrast, luchian found that nonclinical hoarders also created more categories when categorizing nonpersonal objects. they also took almost twice as long to sort objects, and found sorting to be more difficult and stressful than did nonhoarding participants. inconsistencies between this study and wincze may be due to differences between nonclinical and clinical hoarding participants or because of methodological differences between the two studies. thus, the circumstances under which hoarders have categorization difficulties remains unknown due to the lack of systematic comparisons between personal and nonpersonal objects. despite recent advances in the study of cognitive functioning among individuals who hoard, while there is some indication of deficits in hoarding patients, it is unclear how reliably these deficits can be identified. it is also uncertain whether these deficits are present to varying degrees in all hoarding patients, or a subset of patients. future research also should provide greater understanding regarding the specific nature of information processing difficulties and/or cognitive impairment. finally, it will be important as we gain greater understanding of cognitive difficulties to examine whether these difficulties may be remediated in order to improve treatment outcome. several earlier studies found that hoarding symptoms are negative treatment predictors for therapies that have demonstrated effectiveness for ocd. in serotonergic medication trials for ocd, individuals with hoarding symptoms typically have poorer outcomes. only one that has examined the effectiveness of selective serotonin reuptake inhibitors in reducing obsessive - compulsive symptoms has demonstrated equivalent outcomes for individuals with and without hoarding symptoms. the authors only measured obsessive - compulsive symptoms, symptom response was poor in both groups (23% to 24% symptom reduction), and individuals with hoarding symptoms took paroxetine for significantly more days. as with pharmacological approaches, the presence of hoarding symptoms is a negative predictor of cognitive - behavioral treatment outcome for ocd only one third of hoarders with ocd demonstrate clinically significant improvement in response to exposure and response prevention, while one half to two thirds of nonhoarders with ocd demonstrate such improvement. in response to these disappointing outcomes, researchers have developed psychological treatments for compulsive hoarding that are based on frost and hartl 's cognitive - behavioral model. treatments outcomes based on frost and hartl 's model are encouraging, but suggest that many sessions are required to produce change and that clutter is slow to improve. the first case study reported that approximately 45 sessions were needed to completely reduce clutter. after 20 weeks of treatment, steketee demonstrated a 16% reduction in y - bocs scores, while saxena demonstrated a 35% reduction in y - bocs scores after 6 weeks of daily intensive treatment. utilizing steketee and frost 's cognitive - behavioral treatment manual for compulsive hoarding, tolin offered 26 individual sessions (in - office sessions and at least one home visit) over a 7- to 12-month period to 14 individuals. on average, treatment completers (n=10) demonstrated 25% improvement in their clutter and difficulty discarding, and 35% reduction in acquiring. following this open trial, steketee made minor modifications to the treatment and examined its efficacy in a randomized controlled trial. findings from this trial indicated that improvements in hoarding symptoms were greater after receiving 12 sessions of cognitive behavioral therapy (cbt) than after waiting for a comparable period. after 26 sessions of cbt, 68% to 76% of patients were rated as improved by their therapists or themselves, respectively, and 41% of patients met criteria for clinically significant improvement. given that changes are slow to occur during the treatment of compulsive hoarding, researchers have been examining alternative delivery models in hopes of increasing the cost - effectiveness of treatment. using a multiple cohort pretest - post - test design, muroff and colleagues examined the effectiveness of group cbt using steketee and frost 's treatment manual. after 16 to 20 sessions and two home visits, patients evidenced a mean reduction of 8.6 points on the saving inventory - revised (si - r), which is less than that produced from individual treatment using the same manual (18.7 or 16.9). after these investigators modified their research procedures to more thoroughly screen group members and utilized a more detailed and structured manual for the group, the mean si - r reduction in the final group was 14.25. as access to clinicians trained in cbt for compulsive hoarding is limited, a web - based self - help group has also been examined for its effectiveness. this web - based treatment was also based on steketee and frost 's manual and required individuals to take active steps to reducing their hoarding behavior within 2 months of membership. after 6 months of memberships, si - r scores decreased by an average of 6 points. these two group studies suggest that highly structured, in - person groups may lead to greater improvements in hoarding outcomes than less - structured groups. internet treatment approaches are important because they have the potential to expand significantly the number of individuals with hoarding who receive treatment, and thus, ways to improve outcomes achieved from internet - delivered therapy are much needed. novel pharmacotherapies, such as cognitive enhancers and stimulants, should be evaluated for their utility with hoarding patients. cognitive enhancers may improve memory, attention, and overall cognitive functioning, while stimulants may improve attention, alertness, and information - processing speed. only one case report has been published describing the effects of a stimulant in an individual with compulsive hoarding. in this case, a combined treatment of fluvoxamine, risperidone, amphetamine salts, and behavior therapy was used to treat a 56-year old man diagnosed with ocd, compulsive hoarding, adhd, and schizotypal personality disorder. although the patient reported that after treatment he procrastinated less, kept appointments better, and was less upset when throwing things away, the patient 's clutter did not significantly decrease. in order to determine if stimulants or cognitive enhancers are effective adjuncts for the treatment of compulsive hoarding, systematic, overall, research findings indicate that compulsive hoarders do respond to cbt, although improvements are moderate in comparison with gains observed in nonhoarders with ocd. a number of methodological limitations, however, curtail these findings. first, there is a lack of properly controlled treatment studies that involve random allocation to treatment (cbt or medication) and a placebo group. also, the lack of specificity of the measures used to index symptoms makes it difficult to determine whether improvements are due to changes in hoarding symptoms or to reductions in nonhoarding ocd symptoms. despite the increased research on compulsive hoarding in recent years, several avenues still require exploration. researchers must continue to unravel the complex story of hoarding 's etiology and pathogenesis through additional laboratory studies examining the cognitive, emotional, neural, and behavioral features of the disorder. future research may also help to establish the relation of hoarding symptoms to ocd, anxiety, adhd, and icds. finally, further treatment studies investigating the efficacy of cognitive rehabilitation, behavioral interventions, internet applications, and novel medication treatments are essential for improving clinical outcomes. | compulsive hoarding is a disabling psychological disorder characterized by excessive collecting and saving behavior. this article reviews four key areas of recent advances in hoarding research. first, we provide an overview of the evolving controversy regarding the diagnostic status of hoarding, highlighting accumulating evidence that it may be best conceptualized as a separate syndrome. second, we describe advances in our understanding of the epidemiology, course, and demographic features of compulsive hoarding. third, we review the latest findings regarding possible neuropsychological correlates of the disorder. finally, we discuss ongoing progress and future directions related to the clinical management of compulsive hoarding. |
in middle - aged populations, differences between males and females are described in terms of various aspects of coronary artery diseases. males, for example, have a higher rate of previous myocardial infarction (mi).1,2 females, at least in some studies, have been reported to have a longer delay to reach hospital and less frequently to receive invasive therapy for acute myocardial infarction (ami) than males.3,4 in addition, females develop symptomatic cardiovascular disease about a decade later in life than males do.5 age is a demographic predictor of short- and long - term mortality in ami.6 an analysis from the american national registry of myocardial infarction7 noted an interaction between sex and age with regard to 30-day mortality. there was a progressive decrease in the difference (higher mortality in females) with increasing age until the age of 75 years. study results for sex differences in the prevalence of and outcome for ami among patients over 75 years of age have not been described as well as they have been described for middle - aged patients. both female sex and increasing age have been reported to be risk factors for atypical symptoms and a less aggressive treatment in ami.8 therefore, it is of interest to compare females and males when both criteria are fulfilled, that is, to look at sex differences among the elderly patients. this study analyzed age - adjusted sex differences among ami patients aged 75 years and above with regard to 7-year mortality (primary end point) and the frequency of angiograms and admission to the coronary care unit (ccu) as well as 1-year mortality (secondary end points). the hospital has facilities for emergency percutaneous coronary intervention at all hours of the week. gothenburg is the second largest city in sweden and has 550,000 inhabitants, 7% of whom are aged 75 years and above.9 we collected data during two distinct periods, 2001/2002 and 2007, to evaluate possible changes over time. we have previously described these periods,10 and the first period was furthermore described in a comparison between patients living in minneapolis / st paul and gothenburg.11 all patients admitted to the sahlgrenska university hospital with a final diagnosis of ami during the two periods (from july 1, 2001 to june 30, 2002 and from january 1, 2007 to december 31, 2007), regardless of whether or not they were admitted to the ccu, were registered. patients who were residents of gothenburg and aged 75 years and above were included in this analysis. it was recommended that the diagnosis should be based on a dynamic serial elevation of isoenzyme creatine kinase - mb > 5 g / l or troponin t > 0.05 g / l, or an equivalent increase in troponin i plus at least one further criterion (eg, typical chest pain or electrocardiographic signs of acute myocardial ischemia).1214 a 12-lead electrocardiogram (ecg) was recorded on admission to the hospital. the ecg definition of acute myocardial ischemia in the absence of left bundle branch block (lbbb), right bundle branch block, and pacemaker rhythm was st - segment elevation at the j point in two contiguous leads with the cutoff points of 1 mm (0.1 mv) in all leads other than leads v2v3, where the cutoff points of 2 mm (0.2 mv) in males and 1.5 mm (0.15 mv) in females applied. in cases of st - segment depression, the changes regarded as manifestations of acute myocardial ischemia were a downsloping st segment with a j - point depression of 0.5 mm (0.05 mv) in at least two contiguous leads. contiguous leads refer to anterior leads (v1v6), inferior leads (ii, iii, avf), or lateral / apical leads (i, avl).14 when the v4r lead was recorded instead of the v3 lead, an st - segment elevation of 0.5 mm (0.05 mv) or greater was regarded as a sufficient ecg criterion for acute myocardial ischemia.14 in the presence of lbbb, a new lbbb in comparison with the previous ecgs, such as a prehospital ecg, would fulfill the criterion for acute myocardial ischemia. similarly, if a new st - segment elevation or a new q - wave is found in the presence of right bundle branch block, the ecg criteria for acute myocardial ischemia would be fulfilled.14 in the event of suspected mi, other investigative cardiac procedures, such as an exercise bicycle test, echocardiography test, and coronary angiography were considered to obtain a final diagnosis. a recurrent mi was considered to be present if the features of ami started from the 28th day or later following the first incident of ami and the criteria for acute myocardial ischemia were fulfilled.14 a reinfarction was considered to be present if an ami occurred within 28 days of the first incident of ami or within 28 days of a recurrent mi.14 previous history, smoking habits, ecg records, cardiac treatment procedures, cardiovascular complications, medical treatment, and events after discharge within 1 year of admission to hospital were extracted from the database of the health care system by specially trained monitoring nurses. the date of admission to hospital and the date of discharge from hospital were registered. the date of death was obtained from the swedish national population register, together with survival confirmation. all percentages are presented as crude results (ie, not adjusted for age). for comparisons of age between sexes, the mann whitney u - test was used ; otherwise p - values, including the calculation of odds ratios (ors) for two of the secondary objectives (admission to the ccu and performance of coronary angiography), were age adjusted using multiple logistic regression. multiple logistic regression analysis was also used to analyze the interaction between sex and time period. the kaplan meier method was used to estimate mortality, as well as for cardiovascular event rates after hospital discharge. the cox proportional hazards model was used for age - adjusted p - values and time period by sex interaction in terms of mortality and event rates and to calculate the hazard ratio (hr) with corresponding 95% confidence intervals (cis). for the analysis of mortality and event rates after discharge, all tests were two - sided, and a p - value of 5 g / l or troponin t > 0.05 g / l, or an equivalent increase in troponin i plus at least one further criterion (eg, typical chest pain or electrocardiographic signs of acute myocardial ischemia).1214 a 12-lead electrocardiogram (ecg) was recorded on admission to the hospital. the ecg definition of acute myocardial ischemia in the absence of left bundle branch block (lbbb), right bundle branch block, and pacemaker rhythm was st - segment elevation at the j point in two contiguous leads with the cutoff points of 1 mm (0.1 mv) in all leads other than leads v2v3, where the cutoff points of 2 mm (0.2 mv) in males and 1.5 mm (0.15 mv) in females applied. in cases of st - segment depression, the changes regarded as manifestations of acute myocardial ischemia were a downsloping st segment with a j - point depression of 0.5 mm (0.05 mv) in at least two contiguous leads. contiguous leads refer to anterior leads (v1v6), inferior leads (ii, iii, avf), or lateral / apical leads (i, avl).14 when the v4r lead was recorded instead of the v3 lead, an st - segment elevation of 0.5 mm (0.05 mv) or greater was regarded as a sufficient ecg criterion for acute myocardial ischemia.14 in the presence of lbbb, a new lbbb in comparison with the previous ecgs, such as a prehospital ecg, would fulfill the criterion for acute myocardial ischemia. similarly, if a new st - segment elevation or a new q - wave is found in the presence of right bundle branch block, the ecg criteria for acute myocardial ischemia would be fulfilled.14 in the event of suspected mi, other investigative cardiac procedures, such as an exercise bicycle test, echocardiography test, and coronary angiography were considered to obtain a final diagnosis. a recurrent mi was considered to be present if the features of ami started from the 28th day or later following the first incident of ami and the criteria for acute myocardial ischemia were fulfilled.14 a reinfarction was considered to be present if an ami occurred within 28 days of the first incident of ami or within 28 days of a recurrent mi.14 previous history, smoking habits, ecg records, cardiac treatment procedures, cardiovascular complications, medical treatment, and events after discharge within 1 year of admission to hospital were extracted from the database of the health care system by specially trained monitoring nurses. the date of admission to hospital and the date of discharge from hospital were registered. the date of death was obtained from the swedish national population register, together with survival confirmation. all percentages are presented as crude results (ie, not adjusted for age). for comparisons of age between sexes, the mann whitney u - test was used ; otherwise p - values, including the calculation of odds ratios (ors) for two of the secondary objectives (admission to the ccu and performance of coronary angiography), were age adjusted using multiple logistic regression. multiple logistic regression analysis was also used to analyze the interaction between sex and time period. the kaplan meier method was used to estimate mortality, as well as for cardiovascular event rates after hospital discharge. the cox proportional hazards model was used for age - adjusted p - values and time period by sex interaction in terms of mortality and event rates and to calculate the hazard ratio (hr) with corresponding 95% confidence intervals (cis). for the analysis of mortality and event rates after discharge, all tests were two - sided, and a p - value of < 0.05 was considered statistically significant for 7-year mortality (primary objective), otherwise a p - value of < 0.01 was required for statistical significance. approval for this study was given by the research ethics committee at gothenburg university, guldhedsgatan 5a, se-413 20 gothenburg, sweden. this study was retrospective and the research ethics committee at gothenburg university deemed informed consent not necessary. in total, there were 1,414 elderly patients with a final diagnosis of ami in both periods (2001/2002 and 2007), including 748 (53%) females and 666 (47%) males. females were older and had a lower prevalence of previous ami and coronary artery bypass grafting (cabg) but a higher prevalence of diagnosed hypertension. a history of renal disease was more common among males, more so in 2001/2002 than in 2007. males were admitted to the ccu (secondary end point) more frequently than females, age - adjusted or for males vs females was 1.38 (95% ci 1.11, 1.72, p=0.004). the duration of hospital stay was similar in both females and males (table 1). the proportion of patients with symptoms of chest pain, cardiogenic shock, and congestive heart failure on admission to hospital did not differ significantly between females and males (table 2). sixty - five percent of the patients had an ecg recorded on admission to the hospital. among them, st - segment elevation, especially with a location in the anterior leads, occurred more frequently in females, while lbbb was more frequent in males. there was no statistically significant difference for the secondary end point of coronary angiography, and age - adjusted or for males vs females was 1.34 (95% ci 1.00, 1.79, p=0.05) (table 3). the proportions of females and males who underwent other cardiac procedures, including reperfusion therapy, did not differ significantly, except for cabg, which was performed significantly more frequently in males (table 3). table 3 also shows that there was no significant difference in the frequency of various complications in relation to ami between males and females. there was no statistically significant difference between females and males with regard to medication at discharge from hospital (table 4). of all the patients, 18% died during hospitalization in both sex groups (table 3). among the patients who were alive at discharge from hospital, 29% of females and 33% of males died within 1 year of their index admission (table 5). there were no statistically significant differences regarding ami or stroke after hospital discharge between males and females during 1 year after index admission. the 7-year mortality rate was above 80% for both males and females during both time periods (figure 1), although there was a statistically significant difference between the sexes for this parameter (the primary end point) when adjusting for age, with an hr (males vs females) of 1.16 (95% ci 1.03, 1.31, p=0.02). for the two separate time periods, the hr was 1.20 (95% ci 1.02, 1.41, p=0.02) in 2001/2002 and 1.12 (95% ci 0.93, 1.35, p=0.23) in 2007. for death during the first year following index ami admission including in - hospital mortality (secondary objective), there was a nonsignificant trend toward higher mortality among males than females, with an age - adjusted hr of 1.18 (95% ci 0.99, 1.39, p=0.06). when adjusting also for all previous history variables (table 1), hr for 7-year mortality was 1.13 (95% ci 0.99, 1.30, p=0.07), and hr for 1-year mortality was 1.11 (95% ci 0.92, 1.34, p=0.28). females were older and had a lower prevalence of previous ami and coronary artery bypass grafting (cabg) but a higher prevalence of diagnosed hypertension. a history of renal disease was more common among males, more so in 2001/2002 than in 2007. males were admitted to the ccu (secondary end point) more frequently than females, age - adjusted or for males vs females was 1.38 (95% ci 1.11, 1.72, p=0.004). the duration of hospital stay was similar in both females and males (table 1). the proportion of patients with symptoms of chest pain, cardiogenic shock, and congestive heart failure on admission to hospital did not differ significantly between females and males (table 2). sixty - five percent of the patients had an ecg recorded on admission to the hospital. among them, st - segment elevation, especially with a location in the anterior leads, occurred more frequently in females, while lbbb was more frequent in males. there was no statistically significant difference for the secondary end point of coronary angiography, and age - adjusted or for males vs females was 1.34 (95% ci 1.00, 1.79, p=0.05) (table 3). the proportions of females and males who underwent other cardiac procedures, including reperfusion therapy, did not differ significantly, except for cabg, which was performed significantly more frequently in males (table 3). table 3 also shows that there was no significant difference in the frequency of various complications in relation to ami between males and females. there was no statistically significant difference between females and males with regard to medication at discharge from hospital (table 4). of all the patients, 18% died during hospitalization in both sex groups (table 3). among the patients who were alive at discharge from hospital, 29% of females and 33% of males died within 1 year of their index admission (table 5). there were no statistically significant differences regarding ami or stroke after hospital discharge between males and females during 1 year after index admission. the 7-year mortality rate was above 80% for both males and females during both time periods (figure 1), although there was a statistically significant difference between the sexes for this parameter (the primary end point) when adjusting for age, with an hr (males vs females) of 1.16 (95% ci 1.03, 1.31, p=0.02). for the two separate time periods, the hr was 1.20 (95% ci 1.02, 1.41, p=0.02) in 2001/2002 and 1.12 (95% ci 0.93, 1.35, p=0.23) in 2007. for death during the first year following index ami admission including in - hospital mortality (secondary objective), there was a nonsignificant trend toward higher mortality among males than females, with an age - adjusted hr of 1.18 (95% ci 0.99, 1.39, p=0.06). when adjusting also for all previous history variables (table 1), hr for 7-year mortality was 1.13 (95% ci 0.99, 1.30, p=0.07), and hr for 1-year mortality was 1.11 (95% ci 0.92, 1.34, p=0.28). this study is a comparison between females and males over the age of 75 years with regard to the diagnosis of ami and survival follow - up data of all patients for more than 7 years. the primary end point was mortality after 7 years, and during that period, the mortality was as high as above 80% for both sexes. males had higher age - adjusted 7-year mortality than females. for one of the secondary end points, that is, mortality (including in - hospital mortality) during 1 year previous surveys have shown higher mortality in females in both short- and long - term outcomes,15,16 which differs from our results in this respect and the explanation is not clear. similar findings were reported in previous studies.17 the reason behind this finding is not known, but it has even been discussed in terms of inappropriate sex bias. age, previous mi and/or cabg, and clinical symptoms seem to be the factors predicting the initial degree of suspicion of ami.18 in our study, when adjusting for age and previous mi and/or cabg, rate of ccu admission was still significantly higher in males than females (or 1.38 [95% ci 1.10, 1.72 ]). we speculated that a higher prevalence of clinical severity in males than in females might be the main reason. some of the previous studies showed a lower rate of invasive therapy for ami among females when compared with males,19 but others reported the opposite results.20 in our study, cabg was performed more frequently in males than in females. the reason for this finding might be that extensive coronary artery disease is more frequent in males than in females.21 a higher rate of previous mi or cabg and the more frequent presence of lbbb might also indicate more extensive coronary artery disease in males. the use of coronary angiography was one of our secondary end points for which there was no statistically significant difference between the sexes (age - adjusted or 1.34 [95% ci 1.00, 1.79 ]). other aspects of therapy, such as medication at discharge from hospital, did not differ between the sexes, in line with previous reporting.22 other studies have reported that beta - blockers and angiotensin - converting enzyme inhibitors were used more frequently in males than in females.23 both these medicines have been shown to reduce mortality in ami patients.24 in this study population, females were older than males and had a lower prevalence of previous mi. our findings are in agreement with those reported in the literature.23,25 in a population of young patients with previous ami, a higher prevalence of hypertension was found in males. however, in those over 40 years of age, opposite results were found.15 in our study, females had a higher prevalence of previous hypertension, which is in agreement with that reported by other studies.15,23,25 previous studies have suggested that, for anatomic reasons, st - elevation myocardial infarction (stemi) is less frequent in females than in males.26 in our region, the vstra gtaland region of sweden, a study in 2008 of unselected patients with ami found a similar degree of st deviation in both sexes.25 in the present study, stemi was more frequent in females than in males. we speculate that amis in elderly patients might differ from those in younger patients.27 however, a lack of availability of ecgs recorded on admission to hospital in both study periods might have influenced this finding. there was a lower prevalence of lbbb in females, which is in agreement with the previous observations.23,25 the main strength of this study is that all elderly patients hospitalized with an ami in a certain catchment area were included and followed for a long period, with 7-year mortality data for all patients. furthermore, the study population is unselected and has a high degree of representativeness, as all patients with a final diagnosis of ami were included, regardless of whether or not they were admitted to the ccu. the date of death was obtained from the swedish national population register, which has a high degree of quality control and high validity. the availability of ecgs registered in the emergency room on admission was low in the first study period. this might influence the ecg findings and limit our opportunity to draw the correct conclusions. we presume that prehospital ecgs and ecgs monitored during ambulance transport were used instead of ecgs recorded in the emergency department.28 unfortunately, these registrations were not available to us. another limitation was that the ami diagnosis was based on the official clinical judgment and not on study - specific criteria. finally, patients were recruited some years ago, and treatment routines for ami differ in some respects today as compared with the time of the surveys. in this study population, males had higher age - adjusted 7-year mortality and higher rate of admission to the ccu than females. one - year mortality did not differ significantly between females and males, nor did the frequency of performed coronary angiograms. the mortality was high for both sexes, both in the first year and during the 7-year follow - up. from our results, there seems to be room for improvement at all levels of care for these elderly patients. this includes providing initial ccu treatment, invasive investigations, revascularization procedures, and medication. basically, these elderly patients should be treated according to the guidelines2931 to a maximum extent possible, while keeping in mind their individual needs. | objectivesthis study analyzed age - adjusted sex differences among acute myocardial infarction (ami) patients aged 75 years and above with regard to 7-year mortality (primary end point) and the frequency of angiograms and admission to the coronary care unit (ccu) as well as 1-year mortality (secondary end points).methodsa retrospective cohort study comprised 1,414 ami patients (748 females and 666 males) aged at least 75 years, who were admitted to sahlgrenska university hospital in gothenburg, sweden, during two periods (2001/2002 and 2007). all comparisons between female and male patients were age adjusted.resultsfemales were older and their previous history included fewer amis, coronary artery bypass grafting procedures, and renal diseases, but more frequent incidence of hypertension. on the contrary, males had higher age - adjusted 7-year mortality in relation to females (hazard ratio [hr ] 1.16 with corresponding 95% confidence interval [95% ci 1.03, 1.31 ], p=0.02). admission to the ccu was more frequent among males than females (odds ratio [or ] 1.38 [95% ci 1.11, 1.72 ], p=0.004). there was a nonsignificant trend toward more coronary angiographies performed among males (or 1.34 [95% ci 1.00, 1.79 ], p=0.05), as well as a nonsignificant trend toward higher 1-year mortality (hr 1.18 [95% ci 0.99, 1.39 ], p=0.06).conclusionin an ami population aged 75 years and above, males had higher age - adjusted 7-year mortality and higher rate of admission to the ccu than females. one - year mortality did not differ significantly between the sexes, nor did the frequency of performed coronary angiograms. |
until recently, influenza a virus infection in indonesia did not cause much concern for the public due to the mild, self - limiting nature of seasonal influenza. however, since the outbreaks of avian influenza a (h5n1) in poultry in 2003 and the subsequent fatal human infections in 2005, the risk of influenza caused alarm worldwide. until the beginning of 2012, indonesia has 189 human h5n1 cases with a mortality rate at more than 80%. therefore, there is a growing concern that h5n1 became the pandemic.1,2 the big influenza pandemics that caused huge death tolls in 1918, 1957, 1968 high morbidity in 2009 are still fresh in memory.3 laboratory assessment of influenza virus infection is an essential component for prompt and accurate diagnosis. however, this task remains difficult to accomplish in indonesia, the largest archipelago in the world with a population of more than 235 million people. there was a strong need to establish a reliable laboratory network throughout the country to provide prompt and accurate diagnosis of influenza in suspected sufferers. here, we review our efforts and learnings in establishing a reliable laboratory network for influenza a diagnosis during the avian flu (h5n1) outbreaks. the republic of indonesia s territory extends 5120 kilometers from east to west, a similar distance as from san francisco to new york in the united states. the archipelago lies from 6 north latitude to 11 south latitude and from 04 to 141 east longitude and consists of more than 17,500 islands. the inclusion of indonesia s exclusive economic zone brings the total area to about 7.9 million square kilometers of sea area four times larger than the land area (1,992,579 km). based on the 2010 census, the total population of indonesia is 237,556,363 people, which are unevenly distributed among five major islands (java, sumatra, kalimantan, sulawesi, and papua) plus the islands of moluccas, of bali, of west and east nusa tenggara, and thousands of smaller islands. administratively, the republic of indonesia is divided into 33 provinces with jakarta as the capital city.4 laboratory confirmation is required to establish the etiology of clinical influenza and also allows the genetic characterization of viruses and identification of genetic changes favorable to the emergence of a pandemic strain.5 to monitor the circulation of influenza a virus in indonesia, the indonesian ministry of health (moh) initiated a surveillance program in 1975 when the national institute of health research and development (nihrd) of the moh became the national influenza center (nic). this program was supported by wide collaboration among international parties such as usa centers for disease control and prevention (us cdc) and national institute of infectious diseases (niid), japan which are the world health organization (who) collaborating center under the global influenza surveillance network coordinated by the who. since then, all evaluation specimens of suspected influenza cases from the nic and us naval medical research unit 2 (namru-2) sentinels were sent to nic and namru-2 laboratories, respectively. since the first h5n1 human case until 2007, the nic laboratory collaborated with namru-2 laboratory to diagnose specimens from suspected h5n1 cases. us cdc (as a who collaborating center) and hong kong university (as h5n1 reference laboratory) have helped with confirmation and characterization of h5n1 viruses isolated from humans in indonesia from 2005 to 2007. since human h5n1 infections started being reported in mid-2005, specimens of suspected cases from all over indonesia that were requested to be confirmed by laboratory diagnostic test substantially increased. with indonesia s geographical and demographic characteristics, throat and nose swab specimens sent to jakarta needed 1 to 3 days to arrive. the limitation on a number of nic laboratory staff who handle many specimens from all over indonesia has also become a concern. additional laboratories were required to provide timely and accurate diagnosis of influenza a (h5n1) cases throughout indonesia. it was a real challenge for both central and province / district governments to build and develop an influenza laboratory in every province in indonesia. to increase timely diagnostic confirmation of influenza a (h5n1) virus infection, the moh decided to establish a laboratory network to facilitate early detection of influenza a (h5n1).5 in 2007, the minister of health issued a written policy which assigned 44 laboratories as the diagnostic laboratories for influenza a (h5n1) in indonesia. these 44 laboratories came not only from the general hospitals under the moh but also from medical schools among universities and other research institutions. these laboratories were organized as follows : two referral laboratories in jakarta (nihrd moh / nic and eijkman institute) ; eight regional labs in eight provinces and 34 subregional labs in 13 provinces. the two referral laboratories had all the necessary laboratory equipment and had more human resource capabilities than the other laboratories. however, insufficient equipment and reagents stocks in other laboratories were other challenges faced by the network. in 2009, now the organization of the 44 laboratories is simpler since there are only two referral laboratories and 42 designated avian influenza laboratories. although initially all specimens were sent to the nic laboratory, diagnostic tests can now be performed in many more laboratories within the network.68 the moh collaborated with several donor agencies through who in indonesia to provide diagnostic equipment for all the laboratories within the network in 2006. equipment was provided based on the who recommendation for influenza virus detection, ie, real - time polymerase chain reaction (pcr) and gel - based pcr.8 the referral and regional laboratories should have both real - time and gel - based pcr equipment, while the subregional laboratories have only gel - based pcr equipment. in addition, a biosafety cabinet type iia was provided in each laboratory. in 2006, laboratory staff in nihrd received further training for pcr and serology tests from more experienced staff at institutions such as the niid japan and the us cdc in atlanta, georgia, as the who collaborating center for influenza for the region. expert staff from nihrd then trained staff in regional and subregional laboratories in pcr techniques based on the protocol developed by who.8 since 2008, nihrd has routinely given pcr training to all laboratories. additional training to accommodate the use of different pcr primers was given when the novel influenza a (h1 n1) pandemic occurred in 2009. therefore, specimen collecting and delivery are important. in addition, all health personnel should adopt safe behavior when handling and preparing samples. regarding this safety issue, the nihrd also held periodic training in each laboratory on specimen collection, packaging, and delivery based on who guidelines.9 the training is not only provided to the laboratory staff, but also to the staff of hospitals, public health centers and the provincial / district health office. to ensure quality control for specimens, the nihrd regularly carries out external quality control (qc) scheme in each laboratory within the network. the nihrd has developed specific test panels containing h1, h3, h5, h1n1 (2009), and influenza b strain, which are sent annually to all laboratories for this purpose. the nihrd also checks the ribonucleoprotein (rnp) for influenza specimens serving as controls for the qc of specimen collection by the health personnel in the laboratory. lastly, the nihrd made efforts to increase the biosafety and biosecurity of the influenza test by performing assessments of all laboratories. recommendations were then given to the laboratories that performed to the laboratory standards of biosafety level 2 (bsl2). biosafety and biosecurity training of the laboratory personnel is done periodically by the nihrd biosafety and biosecurity team in order to maintain the safety and security of all laboratory personnel. firstly, the public ignorance of the impact of clinical influenza may cause a low demand for laboratory confirmation of disease etiology.7,10 as we mentioned earlier, influenza a, which commonly manifests as seasonal flu in indonesia, was perceived as a mild condition which does not require laboratory diagnosis. this situation contributes to the small number of specimens sent to laboratories, which causes underutilization of diagnostic equipment and reagents. secondly, rotation among health personnel is high so there is a decrease in skilled laboratory staff in many laboratories. in addition, the low demand for diagnostic confirmation in turn lowers the capability of the laboratory staff to perform pcr screening tests. thirdly, the need to continuously provide reagent stocks, maintain laboratory equipment, and calibrate the laboratory network is costly and challenging for the moh.11,12 to overcome these problems and improve utilization of network capacity, the nihrd issued a policy to start a stepwise approach to expand some laboratories to include a role as sentinel influenza surveillance laboratories. in 2008, three laboratories were converted to sentinel laboratories, ie, the microbiology laboratory of hasanuddin university (makassar, south sulawesi), north sumatra islamic university (medan, north sumatra), and diponegoro university (semarang, central java). the results from these laboratories are considered good and in 2009, another two laboratories were added as sentinel laboratories, ie, the microbiology laboratory of udayana university (denpasar, bali) and the university of indonesia (jakarta). each laboratory routinely received specimens of influenza - like illness from neighboring areas of the sentinel surveillance laboratories (figure 1). in this way, the nihrd, together with the help from us cdc, is responsible for maintaining reagent stocks by providing updated primers and probes and calibration of equipment in every sentinel laboratory. the establishment of a laboratory network for influenza diagnosis in indonesia was initially a response to accelerate influenza a (h5n1) confirmation throughout the archipelago. maintenance of such networks was partly supported by international partners and was combined with the organization of a sentinel influenza surveillance laboratory network. diagnosis of other emerging infectious diseases in addition to influenza could be a potential future development of these laboratory networks. | indonesia has been part of the global influenza surveillance since the establishment of a national influenza center (nic) at the national institute of health research and development (nihrd) by the indonesian ministry of health in 1975. when the outbreak of avian influenza a (h5n1) occurred, the nic and us naval medical research unit 2 were the only diagnostic laboratories equipped for etiology confirmation. the large geographical area of the republic of indonesia poses a real challenge to provide prompt and accurate diagnosis nationally. this was the main reason to establish a laboratory network for h5n1 diagnosis in indonesia. currently, 44 laboratories have been included in the network capable of performing polymerase chain reaction testing for influenza a. diagnostic equipment and standard procedures of biosafety and biosecurity of handling specimens have been adopted largely from world health organization recommendations. |
an estimated 569,490 americans are expected to die of cancer in 2010, accounting for approximately 25% of the overall mortality 1. bone metastases are a common manifestation of advanced disease with autopsy studies showing an incidence of 33 - 36% in patients with lung cancer 2, 3, 68% in prostate cancer 3, and 73% in breast cancer 2, 3. while many patients receive therapy at major cancer centers, numerous other patients choose local or regional hospitals, and most imaging studies include the skeleton secondarily if not primarily (e.g. chest radiography, body computed tomography [ct ]). thus, the appearance and behavior of bone metastases can be detected on a wide variety of imaging studies that are performed for many different indications. response criteria represent the standard by which the efficacy of new therapeutic agents is determined in cancer treatment trials. the most commonly used set of criteria is the response evaluation criteria in solid tumors (recist). these and similar anatomic criteria focus predominantly on the physical measurement of solid tumors. disease that is not easily measurable with a ruler or calipers, such as most bone metastases, is designated as unmeasurable. cancer patients with no measurable disease (e.g. individuals with bone - only metastases following the resection of a primary tumor) are often ineligible for clinical trials, which may be the only available source of therapy. this article reviews anatomic (recist 1.1), bone (md anderson [mda ]), and metabolic (positron emission tomography response criteria in solid tumors [percist ]) cancer response criteria, with a focus on the developing role of bone metastases and the interpretation of the treatment response of bone metastases seen on imaging studies. change in tumor size following therapy, also known as objective response 4, 5, is a robust indicator of outcome in the treatment of numerous solid tumors 6 - 9 and forms the basis for anatomic response criteria. recist 10, updated to recist 1.1 in 2009 11, was designed to standardize the assessment of therapeutic response to allow meaningful comparison of drug efficacy among individuals in the same study and across different studies 12, 13. recist 1.1 specifies that up to 5 target lesions, representing all affected organ systems but with no more than 2 target lesions per organ, be selected for measurement throughout the course of a therapeutic trial. to be considered as target lesions, at baseline nodules must measure 10 mm on ct (or twice the slice thickness if the interval is > 5 mm), the short axes of lymph nodes must measure 15 mm on ct (recommended slice thickness is 5 mm), palpable masses must be 10 mm as measured with calipers ; and lung lesions must be 20 mm, clearly delineated, and surrounded by lung parenchyma on chest radiographs. lesions may be measured using ct or magnetic resonance imaging (mri), but ct is preferred in most situations because of the variability of mri scan parameters. measurements made using ultrasonography are not acceptable because of operator dependency and lack of objective reproducibility. according to recist 1.1, drug efficacy is primarily determined by the sum of the measurements of the greatest longitudinal dimension of each target lesion. one of the differences between recist and recist 1.1 is that bone metastases with soft tissue masses measuring 10 mm are now accepted as target lesions. the soft tissue component is to be measured in an identical manner to that used for other target lesions (fig. 1). measurements are to be made in the plane of acquisition (typically axial for ct unless isotropic reconstructions are performed). the largest lesions are preferred if they are clearly and reproducibly measurable (e.g. the largest well - defined lesion is preferred over larger, ill - defined lesions), and no previously irradiated lesion is eligible as a target lesion unless it demonstrates progression after irradiation. therefore, a careful search of the medical record for previous therapeutic radiation exposure is indicated prior to the selection of a bone metastasis as a target lesion. recist 1.1 states that ct is the best currently available and reproducible method to measure lesions selected for response assessment 11. however, mri has been shown superior to ct in delineating the extent of primary bone tumors (which are similar to target bone lesions because they typically produce large soft tissue masses) and their relationship to adjacent structures 14, 15. the value of the high soft tissue contrast resolution of mri was shown in a prospective study comparing mri and ct for the detection of locally recurrent tumors in 49 patients following the resection of musculoskeletal malignancies 16. in the 33 biopsy - proven locally recurrent tumor nodules, mri demonstrated sensitivity, specificity, and accuracy of 82.5%, 96.3%, and 92.6%, respectively ; ct values for sensitivity, specificity, and accuracy were 57.5%, 96.3%, and 85.0%, respectively. mri scans with and without the use of intravenous gadolinium contrast can be considered for the follow - up of measurable bone lesions. the 4 response categories included in recist 1.1 are complete response (cr), partial response (pr), progressive disease (pd), and stable disease (sd) (table 1). cr is defined as the disappearance of all target lesions and reduction of the short axes of target lymph nodes to 30% in sul peak with at least a 0.8-unit decline (fig. progressive metabolic disease includes an increase of > 30% in sul peak with at least a 0.8-unit increase, a visible increase in the extent of fdg uptake (increase in the color field representing fdg uptake), or the development of new lesions. in the absence of clear evidence of disease progression on the fused ct image, new fdg - avid foci are to be verified on a follow - up scan 1 month after discovery. stable metabolic disease is the absence of change or mild changes that do not meet the minimum qualifications of the other categories. anatomic change in tumor size remains an important factor under percist and is to be measured according to recist 1.1. if lesions increase or decrease in size without a corresponding change in metabolic activity, disease progression or response is to be verified on a follow - up scan. when evaluating the potential role of functional imaging modalities such as pet, the recist working group decided that there was not sufficient standardization or evidence to abandon anatomical assessment of tumor burden 11. considering the numerous areas of potential variability that must be overcome in the acquisition and interpretation of pet / ct scans, this hesitation is understandable. nevertheless, if the attempt at standardization represented by percist is successful, fdg pet / ct may be considered as an alternative source of disease measurement in future revisions of the recist criteria. functional imaging criteria can also be considered for use in conjunction with anatomic criteria such as recist or mda (table 4). the mda criteria can allow more bone lesions to be considered measurable disease than does the recist 1.1 system by allowing physical measurement of well - defined bone lesions regardless of soft tissue extension, by allowing regimented subjective assessment of ill - defined lesions, and by taking into account characteristic behaviors such as the development of healing sclerosis. metabolic imaging criteria can allow bone metastases to be measured in the absence of anatomic change by assessing tumor metabolism. response criteria are of crucial importance to the care of many cancer patients, and the tumor response assessment of bone metastases is assuming a greater role in therapeutic management. knowledge of the fundamental concepts of tumor response criteria (anatomic, bone, and functional) and the appearance of bone metastases as they respond to treatment or progress can aid in the interpretation of studies in a manner that will render them of optimal value to the patient and clinician. | response criteria represent the standard by which the efficacy of therapeutic agents is determined in cancer trials. the most widely used criteria are based on the anatomic measurement of solid tumors. because bone metastases are typically located in irregularly shaped bones and are difficult to measure with rulers, they have been previously considered unmeasurable disease. new developments in cancer response criteria have increased awareness of the importance of the response of bone metastases to therapy. the recently updated response evaluation criteria in solid tumors (recist 1.1) now consider bone metastases with soft tissue masses > 10 mm to be measurable disease. response criteria specific to bone metastases have been developed at the university of texas md anderson cancer center (mda criteria) and can be used to assess therapeutic response in numerous types of bone metastases. functional imaging criteria, such as the recently developed positron emission tomography response criteria in solid tumors (percist) allow response to be measured in the absence of anatomic change through assessment of metabolic activity. as monitoring tumor response of bone metastases becomes more important in the management of cancer, so does the demand on radiologists and nuclear medicine physicians for accurate interpretation of the behavior of these lesions. this article reviews anatomic, bone, and metabolic response criteria, providing illustrations for the interpretation of therapy - induced change in bone metastases. |
most cellular functions are carried out by a complex network of genes, proteins, and metabolites that interact through biochemical and physical interactions (gerstein, 2002 ; barabsi and oltvai, 2004 ; albert, 2005 ; basso, 2005 ; almaas, 2007 ; alon, 2007 ; yildrim, 2007). therefore, disease - causing defects may initiate cascades of failures that trigger the co - emergence of multiple diseases in a patient, such as diabetes and obesity. yet, given the environmental, lifestyle, or treatment - related factors that all contribute to comorbidity, it is not obvious whether these cellular network - based interdependencies manifest themselves at the individual or at the population level. discovering such systematic correlations between cellular networks and disease patterns could potentially open new avenues for understanding the human interactome, and may help uncover hitherto unknown disease mechanisms (ergun, 2007 ; loscalzo, 2007 ; braun, 2008). the possibility that there may be systematic links between hereditary diseases, thanks to their common genetic origins, was postulated recently by goh, who created a human disease network (hdn) by connecting all hereditary diseases that share a disease - causing gene according to the online mendelian inheritance in man (omim) database (goh, 2007 ; feldman, 2008). although some of the diseases connected in the hdn captured well - known comorbidity patterns, the functional relevance of the links in the network remains to be demonstrated, leaving open the question whether most diseases connected in the hdn exhibit significant comorbidity. interestingly, the most disconnected disease class in the hdn is that of metabolic diseases. however, lee recently showed that metabolic diseases can be also organized in a metabolic disease network if the enzymes and their associated diseases are linked through metabolic pathways (lee, 2008). most importantly, the study found that metabolic diseases connected through shared pathways tend to show significant comorbidity, suggesting that information encoded in the structure of the metabolic network is amplified, becoming discernible at the population level as comorbidity patterns. metabolic networks represent only one of the several networks functionally relevant to our understanding of cellular activity. indeed, when it comes to cellular interactions of potential importance to human diseases, we need to consider protein protein interaction (ppi) and coexpression networks as well as the links between diseases generated by shared genes. therefore, earlier research raises an important question : are the cellular - level relationships encoded by ppis, coexpression, and shared genes amplified at the population level ? that is, should we expect statistically significant comorbidity patterns for disease pairs that share a gene, whose proteins interact, or whose genes show high coexpression patterns ? to answer these questions, we analyzed the large - scale comorbidity pattern extracted from the us medicare claims database and the gene -- disease association network from omim (mccusick, 1998). we find that cellular interaction links indeed manifest themselves at the population level, resulting in statistically significant comorbidity patterns. we quantify the relative magnitude of these correlations and discuss the current difficulties in mapping population- and cellular - level data into each other, as well as the benefits of such an approach toward elucidating disease mechanisms. the starting point of our study is the medicare claims database containing the diagnoses that cij led to the hospitalization of n=13 039 018 elderly patients, each disease or condition identified by an icd-9-cm code. we denote the incidence of disease i with ii, and the number of patients who were simultaneously diagnosed with diseases i and j with cij. the comorbid tendency between the two diseases can be quantified using either the relative risk, rr = cij / cij, where cij=iiij / n is the expectation value of cij when the two diseases are independent, or the -correlation defined as. when two diseases co - occur more frequently than expected by chance, we have rr>1 and >0. note, however, that although rr and are not independent of each other, each carries unique biases that are complementary. therefore, we use both measures of comorbidity to ensure the robustness of our findings (see supplementary information (si) for further details). the disease -- gene associations used in the study were obtained from the omim database, which contains > 4900 such associations as of october 2008. although the disease -- gene record is far from complete, omim is currently the most complete repository of all known disease genes and their associated disorders. it is important to note that disease names used in the medicare database by the medical and the insurance communities (the icd-9-cm scheme) and those used in the omim database by geneticists are not identical. therefore, we enlisted a professional icd-9-cm coder to manually map the omim disease names into icd-9-cm codes and established connections between the genetic associations and the comorbidity measures (see box 1 and si sections s1 and s2 for more detail). on account of the discrepancies in disease names and the complex, hierarchical nature of the icd-9-cm scheme, we recognize that the mapping is not perfect, and may contain debatable and occasionally erroneous icd-9-cm - to - omim correspondence. therefore, we are providing the mapping used by us in the si, offering a chance for the community to improve on it in future studies. as omim comprises the set of hereditary or complex diseases with validated gene -- disease associations, it is anticipated that only a subset of icd-9-cm codes would correspond to the diseases in the omim. indeed, we find that, of the > 12 000 available icd-9-cm codes, 763 unique icd-9-cm codes can be mapped to omim diseases. the fact that our analysis is limited to 5% of possible diagnosis codes contained in the medicare database, could limit our population (patient) coverage. we find, however, that this is not the case : as figure 1a shows, 90% of patients in the medicare database are diagnosed with at least one disease whose icd-9-cm code is contained in our mapping to the omim database. we use the following three quantities to capture the cellular network - level relationship between diseases i and j, as illustrated in figure 1b for the case of breast cancer (icd-9-cm 174) and cancer of bone and cartilage (icd-9-cm 170.9, see also si) : (i) nij, the number of shared genes associated with both diseases i and j, which quantifies the potential common genetic origin of the two diseases (goh, 2007) ; (ii) nij, the number of ppis between the proteins of diseases i and j capturing the ppi network - level relationships between them (rual, 2005 ; stelzl, 2005). (iii), the average pearson correlation of coexpression between pairs of genes from each disease, capturing the degree to which the genes associated with the two diseases are coexpressed (ge, 2005). the main question can be formulated as follows : does the existence of these cellular - level links (i.e., nij>0, nij>0, ij>0) between the two diseases increase the likelihood that individuals simultaneously develop both conditions ? we start our investigation by measuring the pearson correlation between the cellular variables (nij, nij, ij) and comorbidities (rr and) for 83 924 disease pairs. of these, 2239 pairs are linked through either shared genes (nij1) or ppis (nij1 ; 658 with shared genes, and 1873 with ppis). in figure 2a and table i we present the pearson correlation coefficients (pccs) between the comorbidity measures and the genetic variables. although n, in general, has the highest correlation with comorbidity, we do observe positive pcc with all three variables. there are numerous factors that determine whether two diseases co - occur in a patient, some of which are environmental, lifestyle - related or treatment - induced. the small magnitude of the correlations observed by us suggests that the cellular network offers only a small contribution to the observed comorbidity. note, however, that placing significant emphasis on the magnitude of these correlations is premature, as two known effects limit the correlations observed by us. first, the magnitude of the correlation is limited by the predictive power of specific genetic mutations catalogued in the omim database, and the likelihood of a patient developing a particular disease. indeed, it is known that genetic mutations result in an increase of at most a few percentage points in the likelihood of an individual developing a specific complex disease (loscalzo, 2007) and the correlations observed by us can not exceed the known disease -- gene correlations. second, the correlations are further limited by the noise in the mapping between the omim diseases and the icd-9-cm codes. as we noted, there is inherent ambiguity both in the mapping as well as in the process of assigning a particular diagnosis to specific icd-9-cm codes in hospitals. each instance of such misdiagnosis or mapping ambiguity decreases the magnitude of the observed correlations. therefore, at this point it is not the magnitude, but the statistical significance of the correlations that we can rely on. as summarized in table i, the observed correlations are statistically significant. to quantify the degree of comorbidity caused by the observed correlations, we measured the average comorbidities rr and for disease pairs that are connected at the cellular network level. compared with the entire set of 83 924 pairs of hereditary diseases considered in our study, we find (see figure 2b ; table ii) a two- to four - fold increase in the average comorbidity in disease pairs that share genes (nij1), indicating that if a patient develops a particular disease associated with a gene or multiple genes in the hdn, then they have a two - fold higher chance of developing another disease mapped to one or more common genes in the hdn, compared with diseases that are not. an increased comorbidity is also observed for disease pairs linked through ppis (nij1) and high coexpression (ij geqslant 0.5). the observed correlations between the cellular links and comorbidities raise a related question : would disease pairs that are more interconnected than others (i.e., have larger nij, nij, or ij) show higher comorbidity ? to address this, in figure 2c we show that comorbidity increases rapidly with the number of shared genes : sharing two or more genes (nij2) results in nearly a five - fold increase in comorbidity compared with hereditary disease pairs that do not share genes. an increase in comorbidity is observed with increasing nij and ij as well (figure 2d and e), although the effect is weaker than that observed for nij, which is not unexpected given the smaller impact that nij and ij have on comorbidity in comparison with nij. note that the average comorbidity measured between all pairs of diseases is > 1 (figure 2b), indicating that many patients develop multiple disorders, whether or not the specific diseases are linked at the cellular level. such correlations have been observed in other studies focused on comorbidity patterns (rzhetsky, 2007 ; hidalgo, 2009). these overall comorbidity patterns are not particularly surprising considering that the medicare population is 65 years of age or older, the age at which individuals do develop multiple disorders. thus, the overall comorbidity represents the baseline against which we can assess the impact of the genetic and cellular networks. it is reassuring, therefore, that hereditary diseases that are linked in the hdn (and thus at the cellular level) show comorbidity higher than the baseline rr=1.920.01 and =(1.840.02) 10 observed for the set of all disease pairs. despite the significant increase in rr and, there are many disease pairs that share genes yet fail to show significant comorbidity. we hypothesize that this is, in part, because of pleiotropy, which in this context means that different mutations on the same gene can have different pathological effects on a protein (dudley, 2005), thereby predisposing an individual to different disorders. in general, we expect that disease pairs associated with mutations on the same functional domain of the shared protein show higher comorbidity than disease pairs whose mutations occur in different functional domains. to test this hypothesis, we identified the functional domains of disease - causing mutations on shared genes using the pfam database (finn, 2006). in agreement with our hypothesis, we find higher rr and, for disease pairs whose mutations are on the same domain of the shared gene, compared with disease pairs whose mutations are in distinct functional domains (figure 2b). the observed correlations suggest that a combination of disease data and cellular network information may assist us in identifying new comorbidity patterns alongside their potential genetic origin. indeed, upon inspection of the 2239 disease pairs that are genetically linked (i.e., nij1 or nij1), we find several disease pairs whose comorbidity patterns are already well known to the medical community, such as diabetes and obesity (evans, 2002), or breast cancer and osteosarcoma (knowling and basco, 1986). at the same time, due to the aforementioned mismatch between disease names used by clinicians (within the icd-9 coding scheme) and by geneticists (within the omim tabulation), several highly comorbid disease pairs are readily anticipated (such as diabetes and hypoglycemia, as hypoglycemia is a common side effect of the treatment of diabetes) or cases in which one disease is a broader version of the other (such as mononeuritis and hereditary peripheral neuropathy). such mapping limitations notwithstanding, we find several interesting disease pairs that are linked at the cellular level and also show significant comorbidity. for example, consider alzheimer 's disease (icd-9-cm 331) and myocardial infarction (icd-9-cm 410.9), for which earlier comorbidity studies were either inconclusive or contradictory (bursi, 2006). as figure 3a shows, we not only find statistically significant comorbidity (p10) between the two, but the figure suggests that the shared ace and apoe genes may contribute to the observed effect. similarly, we observe significant comorbidity (p10) between autonomic nervous system disorder (icd-9-cm 337.9) and carpal tunnel syndrome (icd-9-cm 354, figure 3b). a known mechanism is l - chain amyloidosis, which may affect the autonomic nervous system and causes carpal tunnel syndrome when the amyloid infiltrates the flexor retinaculum of the patient 's wrist (haan and peters, 1994). figure 3b, however, suggests that a ppi between the associated genes of each disorder may also play a role in the observed effect. although there may be additional possible physiological or social explanations for some of the observed comorbidities (see si), the method described above has the potential to offer new, testable hypotheses about the biological basis of disease interrelationships. these examples were selected only to demonstrate the potential of the combined investigation of the network and population - level data in identifying potentially interesting disease pairs worthy of further study. a more detailed description of these disease pairs, along with the complete list of the 2239 genetically linked disease pairs and their genetic associations are provided in the si. the main finding of this paper is that health care and treatment data on a large number of individuals offer information useful to systems biology that can complement the information from the well - established genomic studies. indeed, medicare and insurance databases already collect the health care history of millions of individuals, allowing us to uncover the correlations in the occurrence of various diseases. in parallel, increasing knowledge about the molecular origin of disease indicates that many disorders are rooted in defects in gene products that are part of the same cellular network, raising the possibility that these diseases should co - occur in the same individual. admittedly, much of the currently available network data are incomplete and probably noisy. we may be approaching a tipping point, however, where we have acquired sufficient knowledge of human cellular networks to begin understanding the way a disturbance in the networks may contribute to the development of a disease and suggest potential disease - modifying factors. to test the validity of this hypothesis, here we correlated cellular level information for human cells, namely data on shared genes, ppis, and coexpression patterns, with comorbidity data obtained from the medicare database. despite the aforementioned limitations of the mapping and the data collection process we also found that disease pairs with higher correlations tend to be linked more strongly in the cellular network. although our work was mainly driven by the desire to uncover evidence that cellular information is amplified in the human population and thus can be detected from patient data, our results point to the potential usefulness of our approach in uncovering disease mechanisms. indeed, we discuss two disease pairs in which the network - based information offers a plausible mechanism for statistically significant comorbidity patterns. these results suggest that medicare and other insurance databases could play an increasing role in future studies of the systems biology of human cells and diseases. we used the hdn from goh for disorder -- gene associations (goh, 2007), updated based on the version of the morbid map from omim at the time of the study. the ppi data were taken from rual (2005) and stelzl (2005). the genetic coexpression levels were calculated on the basis of an affeymetrix microarray data (ge, 2005) (see www.affymetrix.com). protein domain information is available on unitprot (http://www.uniprot.org/) and pfam (http://www.sanger.ac.uk/software/pfam/). the p - values for the pccs shown in figure 2a and table i were calculated by a monte carlo sampling method. we generate a randomized sequence of the genetic variables and calculate its pcc with comorbidity. after 2 million randomizations, the p - value is the fraction of the total trials that resulted in a pcc that is larger than what was observed. as rr and are monotonically increasing functions of cij, their one - sided p - value is equal to the sum of probabilities that the co - occurrence cij is larger than the actual value. it can be obtained using standard computational software such as mathematica (www.wolfram.com) by approximating the binomial distribution generated from the number of patients n and cij = np = iiij / n as a poisson distribution, and therefore the errors on the comorbidity values (figure 2) were calculated using the bootstrap method based on resampling (efron, 1979 ; newman and barkema, 1998). we used the hdn from goh for disorder -- gene associations (goh, 2007), updated based on the version of the morbid map from omim at the time of the study. the ppi data were taken from rual (2005) and stelzl (2005). the genetic coexpression levels were calculated on the basis of an affeymetrix microarray data (ge, 2005) (see www.affymetrix.com). protein domain information is available on unitprot (http://www.uniprot.org/) and pfam (http://www.sanger.ac.uk/software/pfam/). the p - values for the pccs shown in figure 2a and table i were calculated by a monte carlo sampling method. we generate a randomized sequence of the genetic variables and calculate its pcc with comorbidity. after 2 million randomizations, the p - value is the fraction of the total trials that resulted in a pcc that is larger than what was observed. as rr and are monotonically increasing functions of cij, their one - sided p - value is equal to the sum of probabilities that the co - occurrence cij is larger than the actual value. it can be obtained using standard computational software such as mathematica (www.wolfram.com) by approximating the binomial distribution generated from the number of patients n and cij = np = iiij / n as a poisson distribution, and therefore the errors on the comorbidity values (figure 2) were calculated using the bootstrap method based on resampling (efron, 1979 ; newman and barkema, 1998). schematic description of the procedure used to connect comorbidity (calculated in the medicare layer, top) and genetic associations (given in the omim layer, bottom) between a pair of diseases. breast cancer and bone and cartilage cancer are treated as the example here, also presented in figure 1b. in the medicare layer (top), each disease is represented by an icd-9-cm code, a widely used hierarchical disease diagnosis code system. the incidence ii of each disease (represented by a blue line) is found by counting patients in the medicare database diagnosed with the corresponding icd-9-cm code and its sub - level codes (i.e. 174.1 is also counted as an incidence of 174 for breast cancer), while the co - occurrence cij (red line) of a disease pair is found by counting patients diagnosed with both codes. the comorbidity measures rr and can be calculated from these quantities and the total number of patients in the medicare database (approximately 13 million). the associated genes of each disease are provided in the omim layer (bottom, green lines). because of differences in the disease - labeling schemes in the medicare (icd-9-cm) and the omim databases (the codes are as given in goh, 2007), we manually constructed a mapping between the two (grey lines). supplementary information and supplementary figure s1-s2 mapping between omim and icd-9-cm comorbidity between genetically connected diseases disorder - genetic associations | the impact of disease - causing defects is often not limited to the products of a mutated gene but, thanks to interactions between the molecular components, may also affect other cellular functions, resulting in potential comorbidity effects. by combining information on cellular interactions, disease -- gene associations, and population - level disease patterns extracted from medicare data, we find statistically significant correlations between the underlying structure of cellular networks and disease comorbidity patterns in the human population. our results indicate that such a combination of population - level data and cellular network information could help build novel hypotheses about disease mechanisms. |
bilateral sagittal split osteotomy (bsso) is one of the most common orthognathic surgical procedures for the correction of mandibular deformities. the obwegeser - dal pont osteotomy and hunsuck modifications are widely used1 - 3. when performing bsso, it is crucial to control the lingual split because it may result in an unfavorable fracture or inferior alveolar nerve (ian) damage. conventional radiographs, such as panoramic views and cephalograms, have limitations in regard to evaluation of the lingual split pattern. using cone - beam computed tomography (cbct) and three - dimensional (3d) reconstruction plooij.4 and muto.5 evaluated the mandibular ramus split pattern in a symmetric mandible using 3d - ct. in an asymmetric mandible, the morphology and anatomy of each side differ. the length of the ramus and body, the inclination of the ramus, and the ramal volume are significantly different between the deviated side and the contralateral side6,7. therefore, the aim of this study was to evaluate the lingual split line when performing bsso in asymmetric mandibular prognathism ; we used cbct and a 3d software program. the study group comprised 40 patients with asymmetric mandibular prognathism (20 males and 20 females) who had undergone bsso from january 2012 through june 2013 in pusan national university dental hospital. patients ' age ranged from 18 to 31 years (mean age : 23.2 years). the inclusion criterion comprised chin deviation > 3 mm compared to the facial midline (mean deviation : 5.7 mm ; right / left : 17/23). this study protocol was approved by institutional review board of pusan national university dental hospital, yangsan, korea (pnudh-2013 - 036). preoperative and postoperative cbct images were acquired using dct pro (vatech co., hwaseong, korea). the images were post - processed to a digital imaging and communications in medicine (dicom) 3.0 file (simplant ; materialise inc., leuven, belgium). the mandible was digitally isolated from the maxilla and skull and split up the midline. the deviated and contralateral sides of the mandible were rotated along the vertical axis to visualize the lingual surface of the ramus. the frankfort horizontal plane was set as the horizontal reference plane, and the mid - sagittal plane was set as the sagittal plane.(fig. b) measurements were performed at the level of the mandibular lingula which is the reference point for horizontal osteotomy.(fig. c) to evaluate contributing factors, we observed the position of the lateral cortical bone cut end using 3d reconstructed images. type a was positioned lingually, type b was inferior, and type c was located on the buccal side of the mandibular inferior border. all statistical analyses were conducted with pasw statistics 18.0 (ibm co., armonk, ny, usa). to evaluate intraobserver reliability, the kappa - coefficient was used for the lingual split pattern and the vertical osteotomy end. following the measurement of the ramal thickness, a paired t - test was performed to identify any significant differences between the deviated side and the contralateral side. all parameters were measured twice by one examiner 48 hours apart using the paired t - test for intraobserver reliability. preoperative and postoperative cbct images were acquired using dct pro (vatech co., hwaseong, korea). the images were post - processed to a digital imaging and communications in medicine (dicom) 3.0 file (simplant ; materialise inc., leuven, belgium). a 3d image analysis program was used to reconstruct the 3d images. the mandible was digitally isolated from the maxilla and skull and split up the midline. the deviated and contralateral sides of the mandible were rotated along the vertical axis to visualize the lingual surface of the ramus. the frankfort horizontal plane was set as the horizontal reference plane, and the mid - sagittal plane was set as the sagittal plane.(fig. b) measurements were performed at the level of the mandibular lingula which is the reference point for horizontal osteotomy.(fig. to evaluate contributing factors, we observed the position of the lateral cortical bone cut end using 3d reconstructed images. type a was positioned lingually, type b was inferior, and type c was located on the buccal side of the mandibular inferior border. all statistical analyses were conducted with pasw statistics 18.0 (ibm co., armonk, ny, usa). to evaluate intraobserver reliability, the kappa - coefficient was used for the lingual split pattern and the vertical osteotomy end. following the measurement of the ramal thickness, a paired t - test was performed to identify any significant differences between the deviated side and the contralateral side. all parameters were measured twice by one examiner 48 hours apart using the paired t - test for intraobserver reliability. the lingual split line pattern was categorized into five groups according to the path of the fracture line on the lingual surface of the ramus.(table 1, fig. 2) the kappa - coefficient of the intraobserver reliability was 0.91 (p=0.000). intended split pattern (type i) comprised 60.00% (n=48). type ii, iii, and iv accounted for 11.25% (n=9), 16.25% (n=13), and 5.00% (n=4), respectively, and a bad split (type v) occurred in 7.50% (n=6). there was no split pattern difference between the deviation and contralateral sides.(table 2) the ramal thickness at the level of the mandibular lingula was measured. on the deviated side, no significant difference was found between thicknesses on the deviated and the contralateral sides (p<0.864) ; however, males had a statistically significant difference in ramal thickness.(table 3) the position of the lateral bone cut end was related to the lingual split pattern. in type a, type i was predominant (41/48), and type v was absent. however, in type c, the buccal side end was observed in types iii, iv, and v.(table 4, fig. bsso is a widely used orthognathic surgical procedure for the correction of mandibular deformities. since its initial description by obwegeser, modified procedures proposed by dal pont and hunsuck are currently in common usage1 - 3. the important aspect of this technique is that it safely separates the proximal and distal segments in an intended direction ; it is difficult to identify the separation pattern precisely using a conventional cephalogram or a panoramic view. cbct and 3d reconstruction software provide effective means for evaluation of the facial skeleton and are currently used in large - scale studies of the maxillofacial region8,9. due to its 3d mandibular bone reconstruction capability, cbct enables accurate assessment of the lingual surface of the ramus, which is hard to evaluate with traditional radiography. the 3d evaluation of the lingual split line pattern in a bsso procedure was first reported by plooij.4 they categorized the lingual split line pattern of 40 consecutive patients with symmetric mandibular hypoplasia who underwent advanced bsso into four groups. only 51% of the splits coursed as described by hunsuck3 ; 13% extended to the posterior border, 33% coursed along the outer side of the mandibular canal, and 2.5% had an unfavorable split pattern. he noted the length and position of the medial bone cut during horizontal osteotomy and reported that the likelihood of splitting according to hunsuck 's description increases when the bone cut end lies behind the mandibular foramen ; however, it decreases if the bone cut end extends through the mandibular canal. a study of mandibular prognathism performed by muto.5 reported a relatively high prevalence among asian populations. thirty patients were categorized into five types of lingual split patterns, and 33% were in the range of hunsuck 's description ; however, 15% suffered a buccal fracture. the most important factor influencing such a split tendency is the location of the lateral bone cut end during vertical osteotomy ; the lateral bone cut was on the buccal side of all incidences of buccal fracture. the deviated side appears to have a shorter ramal and body length than the contralateral side ; in addition, it has a smaller degree of ramal inclination, measured in the sagittal plane, and a smaller ramal volume6,7,12,13. these measurements are based on orthodontic reference points or landmarks ; however, information regarding the thickness of the ramus or its relationship with the ian, which should be considered for bsso, can not be obtained. the aim of this study was to evaluate whether the lingual split line pattern during bsso in patients with asymmetric mandibular prognathism is affected by anatomic differences. we observed that there was no differences in the split pattern between the deviated and the contralateral sides ; 60% of split lines were type i, in agreement with hunsuck 's description, and 7.5% had an unfavorable fracture. to verify contributing factors, ramal thickness and the location of the lateral bone cut end were investigated. ramal thickness was not significantly different between the deviated side (mean : 7.10 mm) and the contralateral side (mean : 7.00 mm). yamamoto.14 measured the distance from the mandibular canal to the buccal cortex and the distance from the mandibular lingula to the inferior border of the ramus. in cases where the distance was < 0.8 mm, the incidence of neurosensory disturbance increased significantly. for a medial osteotomy, we measured the ramal thickness only at the level of lingula ; however, the need to evaluate the entire ramus following the split requires further investigation. the lateral cortical bone cut end had a correlation with the split pattern. in case of type i, type a was the most common (41/48). in addition, six cases of type v (either type b or type c) were observed ; these results concurred with those of muto. lee.15 studied mandibular body anatomy in patients with asymmetric prognathism. in that study, the distance from the mandibular canal to the buccal cortex was not significantly different between deviated and contralateral sides. wolford and davis16 reported the use of a reciprocating saw to cut the inferior border of the mandible, without using a mallet, to achieve mandible splitting. with preoperative cbct and a 3d program to locate the lateral bone cut end lingually, it is important to analyze the cross - sectional view of the mandible, the pathway of the ian, and the distance between the mandibular canal and buccal cortex. other studies17 - 19 have reported an incidence of bad splits during bsso ranging from 0.9% to 20% ; the incidence in this study was 7.5% (6 splits). the risk factors for bad splits were old age, the presence of a third molar, a thin mandibular ramus, a high mandibular lingula, and an incomplete split of the inferior border of mandible20 - 23. in this study, in six cases of unfavorable fracture, the factors were presence of a third molar (2 cases), a high mandibular lingula (2 cases), and a buccaly - positioned lateral bone cut end (2 cases). this study was conducted to evaluate whether there is any difference in the split pattern that occurs with bsso in an asymmetric mandible. pre- and post - operative cbct data and a 3d reconstruction program were used to analyze the lingual split pattern, lateral bone cut end, and to measure the thickness of the ramus. 1. we categorized the lingual split pattern of the asymmetric prognathic mandible into five types, and there were no differences in split pattern. the ramal thickness, which was measured at the level of the lingula was not significantly different between the deviated and contralateral sides. the lateral bone cut end was categorized into three types ; a correlation with ramal thickness and split pattern was found. | objectivesthe aim of this study was to evaluate the pattern of lingual split line when performing a bilateral sagittal split osteotomy (bsso) for asymmetric prognathism. this was accomplished with the use of cone - beam computed tomography (cbct) and three - dimensional (3d) software program.materials and methodsthe study group was comprised of 40 patients (20 males and 20 females) with asymmetric prognathism, who underwent bsso (80 splits ; n=80) from january 2012 through june 2013. we observed the pattern of lingual split line using cbct data and image analysis program. the deviated side was compared to the contralateral side in each patient. to analyze the contributing factors to the split pattern, we observed the position of the lateral cortical bone cut end and measured the thickness of the ramus that surrounds the mandibular lingula.resultsthe lingual split patterns were classified into. the true " hunsuck " line was 60.00% (n=48), and the bad split was 7.50% (n=6). ramal thickness surrounding the lingual was 5.551.07 mm (deviated) and 5.661.34 mm (contralateral) (p=0.409). the position of the lateral cortical bone cut end was classified into three types : a, lingual ; b, inferior ; c, buccal. type a comprised 66.25% (n=53), type b comprised 22.50% (n=18), and type c comprised 11.25% (n=9).conclusionin asymmetric prognathism patients, there were no differences in the ramal thickness between the deviated side and the contralateral side. furthermore, no differences were found in the lingual split pattern. the lingual split pattern correlated with the position of the lateral cortical bone cut end. in addition, the 3d - ct reformation was a useful tool for evaluating the surgical results of bsso of the mandible. |
despite the marked advances in the perioperative management of the liver transplant recipient, an assessment of clinically significant graft injury following preservation and reperfusion remains difficult. the lack of a sensitive clinical marker of acute liver injury has a profound impact on clinical practice and the success of translational research in liver transplantation. such a clinical marker could aid the management of graft dysfunction in early identification of recipients who will require retransplantation. currently, posttransplant peak aspartate aminotransferase (ast) is a widely accepted clinical marker for graft damage in liver transplant practice [26 ]. in fact, the united network for organ sharing (unos) suggests relisting criteria for the recipients with primary graft dysfunction (pnf) on the basis of the post - transplant peak ast. ast is an enzyme that is involved in amino acid metabolism, primarily existing in hepatocytes. during transplantation graft hepatocytes are inevitably injured, and intracellular enzymes are subsequently released into the systemic circulation of the recipient. we theorized that the recipient ast serum concentration is a function of the total amount of ast released by the graft liver diluted by the total circulating blood volume at the time of blood sampling. therefore, the ast value could be misleading or inaccurate, depending on the size of the graft (total mass of injured hepatocytes) and the volume of blood in the recipient 's circulation. we have previously reported on the use of the ratio of donor to recipient body surface area (bsai) to predict donor - recipient graft size mismatch [8, 9 ]. we showed that size mismatch has a bidirectional impact on graft survival with a progressive increase in the risk of graft failure towards both ends of the size - mismatch spectrum, small - for - size to large - for - size 5 by generalized additive model. bsai has an important value in deceased liver transplant to quickly and reliably predict size - mismatch - related comorbidities and graft outcome. in this study, we hypothesized that serum ast when adjusted by the ratio of the donor - to - recipient body surface area (asti) could more accurately approximate the true mass of injured hepatocytes than the uncorrected or absolute peak ast, subsequently providing a more precise assessment of graft fate. in order to test this hypothesis, we conducted a study to investigate the sensitivity and specificity of asti versus absolute peak ast to predict pnf and graft survival in a large cohort of postliver transplant patients. after institutional review board approval, primary retrospective data collection was from patient records who underwent whole orthotopic liver transplantation (lt) from january 2002 to march 2008 at our institution (jackson memorial hospital, university of miami, leonard miller school of medicine, miami, fl, usa). we were interested in the effect of size - matched peak ast (asti) on postoperative outcomes. therefore, among all transplant recipients, we excluded those who underwent split or partial lt as well as those who underwent simultaneous other organ transplantation, such as liver - kidney, liver - heart, liver - intestine, or multivisceral transplantation or live donor lt. body surface area index (bsai) to investigate donor - recipient size mismatch on postoperative primary graft dysfunction [8, 9 ]. the bsai was calculated by the following equation : (1)bsa = weight (kg)0.425height (cm)0.7250.007184bsai = donor bsarecipient bsa. size - adjusted peak ast (asti) was calculated by dividing peak ast by bsai. peak ast was defined as maximum ast within 7days after lt. statistical analysis. primary nonfunction (pnf) of liver graft is the most detrimental condition after transplantation. unos suggests the listing criteria for recipients with pnf as an ast 3,000 and a pt - inr 2.5 or acidosis ph 7.30 (arterial), ph 7.25 (venous), and/or lactate 4 mmol / l or anhepatic candidate within 7 days after implantation of a graft. of these criteria, the combination of ast and pt - inr was used for comparative purposes in this study. scatter plotting was used to show the statistical association between ast and asti with subsequent development of pnf. the area under the receiver - operating - characteristic curve (auc) was used to evaluate diagnostic accuracy of asti relative to that of conventional ast. the predictive value of current unos criteria for pnf (by combination of ast and pt - inr) was assessed by sensitivity and specificity in our study cohort. the logistic regression model was used to describe the association between seven study groups differentiated by ranges of asti and the incidence of pnf. all recipient and donor variables were included in the multivariable model. to assess the independence of asti effect on pnf, a backward conditional elimination procedure was then undertaken using the donor and recipient demographic factors. cox proportional - hazards models were used to examine the association between seven study groups differentiated by ranges of asti and graft survival ; data were censored at the time of the last visit or patient death. one - year graft survival was used to assess the short- and mid - term effect of asti. covariates were analyzed by using a backward conditional method with the donor and recipient demographic factors. kaplan - meier survival analysis with generalized wilcoxon analysis was used to assess the difference in graft survival between asti 3,500 and asti 3,500 iu / l group and 83.7% in the asti 3,500 iu / l group (p = 0.004). despite the significant advances in the management of the liver transplant (lt) recipient, the quantitative assessment of graft damage remains uncertain. acute injury and inflammation of transplanted organs during the immediate postoperative period may be linked to early organ dysfunction resulting in higher graft failure rates and rejection in the recipient. therefore, development of appropriate clinical markers to identify organ injury in the early postoperative period will have a profound impact on patient outcomes as well as on the success of clinical and translational research [11, 12 ]. such clinical markers of liver damage can be of assistance in identifying the mechanisms of liver inflammation and injury and predisposing factors of donor organ injury, making it possible to implement injury - specific treatment strategies to promote organ resuscitation. in response to the transplant community 's urgent need for a useful clinical marker of injury, ongoing nih clinical trials in organ transplantation have attempted to investigate posttransplant genomic expression patterns, which may correlate with a high probability of graft dysfunction or failure. currently, in lt practice posttransplant peak ast is the most widely accepted clinical markers of graft damage. ast is an enzyme that is involved in amino acid metabolism, primarily existing in liver as well as heart, skeletal muscles, kidney, and brain. when hepatic parenchymal cells are injured, ast is released into the systemic circulation, causing an elevation of serum ast. however, the serum ast level depends on the mass of the donor liver graft (the degree and extent of injured and dying hepatocytes) and on the recipient circulatory blood volume. the working hypothesis that guided this study is that the peak ast corrected by the ratio of donor - to - recipient body surface area could be determined to be more accurately associated with the degree of graft injury, thus better predicting graft outcome in lt. indeed, the results of this study prove that the size - adjusted peak ast (asti) has higher sensitivity and specificity to predict pnf compared to size - unadjusted ast. iu / l and either (a) inr 2.5 or arterial ph 7.30 or venous ph 7.25 and/or lactate 4mmol / l, (b) anhepatic patient within 7 days after transplantation. in our study cohort, when we applied unos criteria, we found a relatively low sensitivity of 75.0% and specificity of 97.7%. this relatively high false positive result with conventional unos criteria may be due to the fact that some cases of pnf occurred in small - for - size donors as shown in figure 1, where ast is more likely to underestimate the severity of graft damage based on the reasoning previously described. also, we found that the severity of graft damage in large - for - size donors can be overestimated by size - unadjusted ast. in fact, one case was underestimated and three cases were overestimated by conventional criteria and this error was corrected by asti criteria as shown in table 2. when we used size - adjusted ast (asti), the combination of size - adjusted asti and pt - inr predicted pnf more accurately, and the sensitivity increased to 87.5% with a specificity of 98.0% at cutoff of asti 3,500 iu / l and pt - inr 2.3. in addition, asti correlated well with the incidence of pnf and short - term graft survival. increases in asti were significantly associated with increasing incidence of pnf. also asti was highly correlated with the incidence of 1-year graft failure, if asti exceeded 3,500 iu / l. a more sensitive prediction of pnf with asti will enable clinicians to more accurately identify the recipient who requires retransplantation, perhaps, at an earlier stage in the postoperative period, resulting in better patient outcome. one - year graft survival (shown in figure 4) showed that asti > 3,500 iu / l group was associated with a steep decrease in graft survival in the early postoperative period compared to asti < 3,500 iu / l group. these findings suggest that in lt markers of postoperative graft injury should be interpreted with caution by taking into account graft size mismatches. bsai appears to be a powerful tool to detect donor - recipient size mismatch in cadaveric lt but awaits further validation. small - for - size or large - for - size grafts not only have been attributed to decreased graft survival, increased additional graft usage (retransplantation), and increased incidence of other complications but also have made an assessment of posttransplant graft injury difficult. this study has demonstrated that size mismatch may have an even greater negative impact on graft outcome in the presence of other donor risk factors, such as prolonged cold ischemia. furthermore, the lack of a suitable clinical measure to reliably predict size mismatch has probably hindered our ability to judge its clinical importance in cadaveric lt. bsai provides a simple, reproducible, and sensitive way of detecting size mismatch in cadaveric lt. second, fluid excess (ascites and edema) is universal in end - stage liver disease. fluid excess causes weight - to - height parameters such as percentage ideal body weight, bmi, and bsa to be underestimated, and the bsa of the recipient may not accurately reflect the liver volume in these situations. alternative ways to more accurately detect size disparity may be (i) measuring the donor graft weight at back table or (ii) using the recipient 's dry weight instead of recipient bsa. however, we believe that asti is a simple, reproducible, and sensitive predictor of graft outcome associated with size mismatch. the liver has the excellent regenerative capacity even in the face of significant loss of hepatocyte mass. this has been demonstrated after liver resection or traumatic hepatic injury [14, 15 ]. once mass (volume) or functional compensation starts, bsai may no longer accurately reflect the size mismatch between donor and recipient. in experiments with rodents, regeneration of liver mass occurred by 3 days after hepatectomy and restoration is complete by 57 days. recent clinical observations showed that recipients of partial grafts have a rapid proliferation of liver mass, with a majority reaching a calculated standard liver volume by one month [17, 18 ]. although several factors have been proposed to explain the delay in the regeneration process, such as small - for - size donor, severity of ischemic injury, immunosuppressive medications, presence of steatosis, and older age, for these reasons the use of bsai - adjusted liver enzymes as more sensitive and specific markers of liver injury should be limited to the immediate postoperative period. lastly, although asti seems to be a useful index to predict outcome in adult olt, it remains unclear whether asti is also useful in pediatric olt. since slv calculated with bsa can be applied to pediatric populations, asti should also be applied to pediatric patients and should provide useful information for preoperative estimation of liver grafts regardless of age. however, predicting outcome by bsai for pediatric population warrants further investigation. we conclude that the asti appears to be a sensitive predictor of postoperative liver graft injury in the early postoperative period. despite some shortcomings, we believe our analysis shows that the asti may provide significantly better prediction of size mismatch - related graft dysfunction in lt. finally, we believe that asti should be considered in the early identification of graft dysfunction or pnf and should be validated in a prospective study to see whether it accurately and consistently predicts pnf or graft function and reduces posttransplant mortality. | background. despite the marked advances in the perioperative management of the liver transplant recipient, an assessment of clinically significant graft injury following preservation and reperfusion remains difficult. in this study, we hypothesized that size - adjusted ast could better approximate real ast values and consequently provide a better reflection of the extent of graft damage, with better sensitivity and specificity than current criteria. methods. we reviewed data on 930 orthotopic liver transplant recipients. size - adjusted ast (asti) was calculated by dividing peak ast by our previously reported index for donor - recipient size mismatch, the bsai. the predictive value of asti of primary nonfunction (pnf) and graft survival was assessed by receiver operating characteristic curve, logistic regression, kaplan - meier survival, and cox proportional hazard model. results. size - adjusted peak ast (asti) was significantly associated with subsequent occurrence of pnf and graft failure. in our study cohort, the prediction of pnf by the combination of asti and pt - inr had a higher sensitivity and specificity compared to current unos criteria. conclusions. we conclude that size - adjusted ast (asti) is a simple, reproducible, and sensitive marker of clinically significant graft damage. |
heart function fails when the organ is unable to pump blood at a rate proportional to the body s need for oxygen or when this function leads to elevated cardiac chamber filling pressures (cardiogenic pulmonary edema). despite our sophisticated knowledge of heart failure, even the so - called ejection fraction up 175% over the past 25 years and costs of nearly $ 15.4 billion dollars, acute heart failure is a critically important health concern. furthermore, half of the patients discharged from the hospital are readmitted within half - a - year. in - hospital mortality remains high between 4% and 7%.1,2 heart failure is a significant problem as the population ages. the prevalence is 2.5% of the us population or 5 million patients (from the national health and nutrition education survey).2 common etiological mechanisms of heart failure include coronary ischemia, valvular disease, hypertension, and diastolic dysfunction. yet, other causes include : postpartum cardiomyopathy ; postinfectious, chronic tachycardia ; metabolic dysregulation ; adverse medication side effects (particularly adriamycin chemotherapy) ; orphan disease duchenne s muscular dystrophy ; infiltrative diseases (such as amyloidosis) ; and inflammatory / connective tissue diseases (such as systemic lupus erythematosus). when known causes of heart failure less often studied versus chronic heart failure, acute decompensated heart failure is associated with abrupt - onset symptoms associated with hospitalization. nearly half of the admitted patients with heart failure have preserved ejection fraction.2,3 in order to appreciate the physiology of heart failure, a review of fundamental principles is on order. the frank starling law states that the stroke volume of the heart increases in response to an increase in the volume of blood filling the heart (the end diastolic volume) when all other factors remain constant. the increased volume of blood stretches the ventricular wall, causing cardiac muscle to contract more forcefully (the so - called frank starling mechanism). the stroke volume may also increase as a result of greater contractility of the cardiac muscle during exercise, independent of the end - diastolic volume. the frank starling mechanism appears to make its greatest contribution to increasing stroke volume at lower work rates, and contractility has its greatest influence at higher work rates.4 while the frank starling mechanism may contribute at lower work rates, cardiac injury at lower work rates may activate additional mechanisms. cardiac injury in heart failure leads to not only reduced left ventricular (lv) dysfunction but also activation of the renin angiotensin system and sympathetic nervous system, ultimately causing lv thickening, remodeling, vasoconstriction (with pulmonary hypertension and cell dysregulation), and, possibly, apoptosis or programmed cell death. severity is graded by the new york heart association class, with increasing severity of classes i to iv. another grading system is the american heart association staging system with stage c, classified as the symptomatic stage. this stage c is a result of known structural heart diseases and is associated with shortness of breath, fatigue, and reduced exercise tolerance. the obvious goal is to prevent progressive lv dysfunction to end - stage d, resulting in lv assist device implantation, cardiac transplant, or palliative care.5 medication management of acute heart failure often entails diuretics to reduce preload, vasodilators to reduce preload and/or afterload, and inotropes to augment contractility. angiotensin - converting enzyme (ace) inhibitors and angiotensin receptor blockers (arbs) and aldosterone antagonists also aim to reduce preload and afterload.6 one adverse side effect (usually nonallergic antibody, non - ige mediated) of ace inhibitors is angioedema (risk up to 15.5%)., this angioedema results from increased bradykinin production, which can occur at any time while taking these medications due to interruption of the conversion of angiotensin i to angiotensin ii. angioedema from ace inhibitors and arbs may lead to life - threatening swelling of the throat, requiring critical interventions, ie, intubation or tracheostomy. for arbs, the mechanism of action of angioedema is not due to direct production of bradykinin and is unclear. some evidence suggests that treatment with a bradykinin b2 receptor blocker (icatibant) may attenuate angioedema attacks early.79 patients with hereditary angioedema (deficiency of c1 esterase inhibitor) and spontaneous attacks of swelling should not take ace inhibitors. clearly, it would be prudent to design a heart failure drug that does not incur risk of angioedema. other medications used for heart failure include inotropes such as dopamine and dobutamine. while inotropes increase cardiac output, they often do so at the expense of increasing myocardial oxygen demand, predisposition to arrhythmias, and neurohormonal activation there is 50% survival at 6 months and nearly 100% mortality at 1 year for those heart failure patients who are dependent on inotropes.10 long - term survival among heart failure patients may be improved with -blockers,11 ace inhibitors,12 aldosterone antagonists,13 electrophysiology devices such as automatic implantable cardiovascular defibrillators, and vasodilators. another aspect of heart failure is diastolic dysfunction with preserved lv ejection fraction, accounting for half of the hospitalizations. pathophysiologically, there is concentric remodeling and increased lv end diastolic pressure from a stiff left ventricle, thereby preventing relaxation. medications for diastolic dysfunction are similar to systolic dysfunction : ace inhibitors, arbs, diuretics, and -blockers.14 in august of 2014, the first new drug for heart failure in many years, called lcz696, was shown to prevent deaths and hospitalizations in heart failure patients better than the gold standard enalipril, which is an ace inhibitor. in the paradign - hf study, lcz696, which is comprised of two drugs an arb and a neprilysin inhibitor called sacubitril reduced the risk of cardiovascular - related death to 13.3%, versus the ace inhibitor cohort rate of 16.5%. this use of sacubitril is an example of utilizing new or different pathways to treat heart failure.15 sacubitril is a prodrug that is activated to lbq657 by de - ethylation via esterases.16 lbq657 inhibits the enzyme neprilysin,17 which is responsible for the degradation of atrial and brain natriuretic peptide, two blood pressure lowering peptides that work mainly by reducing blood volume. as a result of reduced degradation of helpful peptides, these heart failure patients on sacubitril have less circulating blood volume and have done well in terms of prevention of deaths and hospitalizations (supra vide). on april 15, 2015, the food and drug administration approved corlanor (ivabradine) to reduce hospitalizations from worsening heart failure. the indications for ivabradine are specific : symptoms of heart failure that are stable and a normal heartbeat with a resting heart rate of at least 70 beats per minute and patients must already be taking -blockers at the highest dose they can tolerate. ivabradine is thought to work by decreasing heart rate and represents the first approved product in this drug class. it works via the if (funny channel) and does not affect other cardiac ionic currents (not beta receptor mediated). ventricular contractility is preserved.18 in clinical trials of 6,505 participants, hospitalizations for heart failure were reduced compared to placebo. the most notable side effects were bradycardia, hypertension, atrial fibrillation, and temporary visual disturbance entailing seeing flashes of light. the bradycardia may be severe.19 other novel therapeutic agents for heart failure may consequently have plausible efficacy on the heels of sacubitril and ivabradine. for example, vasoactive intestinal peptide (vip), discovered by drs sami i. said and victor mutt in the 1970s, is a 28amino acid peptide found in nerve terminals and in mast cells.20 vip increases cyclic adenosine monophosphate (camp) intracellularly to cause cardiac contraction in an analogous fashion to glucagon.21 vip also is a smooth muscle relaxant, which may be beneficial in pulmonary hypertension associated with heart failure22 and vip inhibits the proliferation of smooth muscle cells, attenuating vascular remodeling.23 inasmuch as sometimes vascular remodeling is a compensatory mechanism for dysfunction in heart failure, vip has promise to treat this deleterious component. enhanced peripheral vascular tone is a major factor in determining deterioration of clinical heart failure. nakamura has reviewed peripheral circulatory failure in heart failure and notes that it is caused not only by simple arterial muscle constriction, but also by structural and functional changes, including receptor and post - receptor levels in the vasculature. thus, vascular remodeling potentially may be an important mechanism underlying vasodilatory failure in both limb conduit and intraskeletal muscle vessels, contributing significantly to lv dysfunction and exercise intolerance in patients with heart failure.24 what is compelling about vip as a novel therapeutic agent for heart failure is evident in mice lacking the gene for vip. gene programs are overexpressed in vip knockout mice.25 so, the plausibility of vip as both a treatment for heart failure in patients and a modifier of one s biology in a pharmacogenomic fashion may have traction. four categories of genes triggering heart failure are increased in mice without heart - protective vip : (1) force generation and propagation ; (2) energy production and regulation ; (3) ca cycling ; and (4) transcriptional regulators. force generation gene mutations encoding proteins of the muscle sarcomere have been associated with both hypertrophic and dilated cardiomyopathy. overexpression of sarcomere protein expression in the setting of a dilated cardiomyopathy26 may suggest a compensatory mechanism. upregulated expression of other force generation genes such as sarcoglycan and caveolin27 in vip knockout mice supports the concept of increased force to the extracellular matrix, with ventricular hypertrophy, for apparent release of protein products of these genes in heart failure. regarding energy production and regulation genes, it has been increasingly recognized that nuclear - encoded metabolic gene mutations are key regulators of hypertrophic cardiac remodeling and heart failure. for example, mutations in genes encoding the lysosome - associated membrane protein are associated with heart failure in patients.28 in the absence of vip in vip knockout (ko) mice, upregulation of prkag2, for example,29 would support a compensatory response to thinning of the cardiac wall. ca cycling is critically important, since protein and rna levels of key calcium modulators are altered in acquired and inherited forms of heart failure. overexpression of fkbp30 in mice supports the concept that these mice would have aberrant excessive release of calcium during the relaxation / diastolic phase of the cardiac cycle, enhancing the opportunity to generate hypertrophy. cardiac hypertrophy is attenuated in murine models with overexpression of caveolin-3, a potent upstream inhibitor of t - type ca current,18 and rats treated with calhex231, an inhibitor of calcium - sensing receptor.31 similarly, deleterious lv hypertrophy may therefore plausibly be suppressed with exogenous vip administration. mechanisms which activate or repress cardiac gene transcription may induce key molecules, which directly or indirectly lead to cardiac remodeling. lack of vip in vip ko mice with the phenotype of biventricular dilated cardiomyopathy and primary pulmonary hypertension is concurrent with strong overexpression of cardiac muscle genes, supporting the concept of vip in vivo as a maestro conductor maintaining homeostasis of the heart.26 table 1 shows on the next page multiple heart failure gene programs which are overexpressed in vip ko mice. these physiological abnormalities are associated with premature death compared to the wild type (fig. 3). compensatory and pathogenic gene programs are displayed in figure 4, with overexpression of leptin as a compensatory reaction for cardiac cachexia and low body weight along with proinflammatory interleukin-6 (il-6) and il-1. vip increases intracellular camp and acts via protein kinase a to activate transcriptional promoters.32,33 vip upregulates il-10, which is anti - inflammatory.34 kupari found that vip levels in serum from healthy subjects and patients with aortic stenosis and heart failure were released into the coronary sinus. these authors concluded that, although vip was marginally elevated systemically in heart failure, it is the major neuroendocrine contributor to heart failure.35 smitherman infused vip into the left coronary artery of four other men at four levels. the maximum decline in coronary vascular resistance was 46% and was not associated with an increase in myocardial oxygen uptake. he concluded that 1) intravenous administration of low to intermediate doses of vip in humans is associated with substantial coronary vasodilation, 2) the coronary bed appears to be at least as responsive as other vascular beds, 3) the coronary vasodilation is due to both direct and indirect effects, and 4) the coronary vasodilation does not appear to be mediated by prostaglandins.36,37 so, the potential of an exogenous, sustainable vip drug would be to help relax coronary arteries. lucia also found that vip plasma levels were higher in heart failure patients with dilated cardiomyopathy compared to healthy subjects and was higher in older subjects. heart failure patients with higher new york heart association severity had lower levels of vip. these data also suggest that heart failure patients with worse disease have an inability to produce vip. the capacity of the elderly to produce vip is heartening, so that low levels are not due to aging itself.38 phasebio corporation recently conducted a phase 1, single ascending dose (sad) study in patients with essential hypertension. pb1046 was administered subcutaneously and was found to be well - tolerated and demonstrated a prolonged, dose - dependent effect on blood pressure, which was used as a biomarker for activity in this study. (see http://clinicaltrials.gov/ct2/show/nct01523067 for further details.) with this demonstration of safety and tolerability at efficacious doses, phasebio conducted a second phase 1 sad study in patients with essential hypertension, with the product administered intravenously to support the use of pb1046 in an acute setting (http://clinicaltrials.gov/show/nct01873885). subsequent phase 2 clinical studies will examine the efficacy of multiple ascending doses of pb1046 in patients with pulmonary arterial hypertension and heart failure. in august of 2014, the first new drug for heart failure in many years, called lcz696, was shown to prevent deaths and hospitalizations in heart failure patients better than the gold standard enalipril, which is an ace inhibitor. in the paradign - hf study, lcz696, which is comprised of two drugs an arb and a neprilysin inhibitor called sacubitril reduced the risk of cardiovascular - related death to 13.3%, versus the ace inhibitor cohort rate of 16.5%. this use of sacubitril is an example of utilizing new or different pathways to treat heart failure.15 sacubitril is a prodrug that is activated to lbq657 by de - ethylation via esterases.16 lbq657 inhibits the enzyme neprilysin,17 which is responsible for the degradation of atrial and brain natriuretic peptide, two blood pressure lowering peptides that work mainly by reducing blood volume. as a result of reduced degradation of helpful peptides, these heart failure patients on sacubitril have less circulating blood volume and have done well in terms of prevention of deaths and hospitalizations (supra vide). on april 15, 2015, the food and drug administration approved corlanor (ivabradine) to reduce hospitalizations from worsening heart failure. the indications for ivabradine are specific : symptoms of heart failure that are stable and a normal heartbeat with a resting heart rate of at least 70 beats per minute and patients must already be taking -blockers at the highest dose they can tolerate. ivabradine is thought to work by decreasing heart rate and represents the first approved product in this drug class. it works via the if (funny channel) and does not affect other cardiac ionic currents (not beta receptor mediated). ventricular contractility is preserved.18 in clinical trials of 6,505 participants, hospitalizations for heart failure were reduced compared to placebo. the most notable side effects were bradycardia, hypertension, atrial fibrillation, and temporary visual disturbance entailing seeing flashes of light other novel therapeutic agents for heart failure may consequently have plausible efficacy on the heels of sacubitril and ivabradine. for example, vasoactive intestinal peptide (vip), discovered by drs sami i. said and victor mutt in the 1970s, is a 28amino acid peptide found in nerve terminals and in mast cells.20 vip increases cyclic adenosine monophosphate (camp) intracellularly to cause cardiac contraction in an analogous fashion to glucagon.21 vip also is a smooth muscle relaxant, which may be beneficial in pulmonary hypertension associated with heart failure22 and vip inhibits the proliferation of smooth muscle cells, attenuating vascular remodeling.23 inasmuch as sometimes vascular remodeling is a compensatory mechanism for dysfunction in heart failure, vip has promise to treat this deleterious component. enhanced peripheral vascular tone is a major factor in determining deterioration of clinical heart failure. nakamura has reviewed peripheral circulatory failure in heart failure and notes that it is caused not only by simple arterial muscle constriction, but also by structural and functional changes, including receptor and post - receptor levels in the vasculature. thus, vascular remodeling potentially may be an important mechanism underlying vasodilatory failure in both limb conduit and intraskeletal muscle vessels, contributing significantly to lv dysfunction and exercise intolerance in patients with heart failure.24 what is compelling about vip as a novel therapeutic agent for heart failure is evident in mice lacking the gene for vip. human heart failure gene programs are overexpressed in vip knockout mice.25 so, the plausibility of vip as both a treatment for heart failure in patients and a modifier of one s biology in a pharmacogenomic fashion may have traction. four categories of genes triggering heart failure are increased in mice without heart - protective vip : (1) force generation and propagation ; (2) energy production and regulation ; (3) ca cycling ; and (4) transcriptional regulators. force generation gene mutations encoding proteins of the muscle sarcomere have been associated with both hypertrophic and dilated cardiomyopathy. overexpression of sarcomere protein expression in the setting of a dilated cardiomyopathy26 may suggest a compensatory mechanism. upregulated expression of other force generation genes such as sarcoglycan and caveolin27 in vip knockout mice supports the concept of increased force to the extracellular matrix, with ventricular hypertrophy, for apparent release of protein products of these genes in heart failure. regarding energy production and regulation genes, it has been increasingly recognized that nuclear - encoded metabolic gene mutations are key regulators of hypertrophic cardiac remodeling and heart failure. for example, mutations in genes encoding the lysosome - associated membrane protein are associated with heart failure in patients.28 in the absence of vip in vip knockout (ko) mice, upregulation of prkag2, for example,29 would support a compensatory response to thinning of the cardiac wall. ca cycling is critically important, since protein and rna levels of key calcium modulators are altered in acquired and inherited forms of heart failure. overexpression of fkbp30 in mice supports the concept that these mice would have aberrant excessive release of calcium during the relaxation / diastolic phase of the cardiac cycle, enhancing the opportunity to generate hypertrophy. cardiac hypertrophy is attenuated in murine models with overexpression of caveolin-3, a potent upstream inhibitor of t - type ca current,18 and rats treated with calhex231, an inhibitor of calcium - sensing receptor.31 similarly, deleterious lv hypertrophy may therefore plausibly be suppressed with exogenous vip administration. mechanisms which activate or repress cardiac gene transcription may induce key molecules, which directly or indirectly lead to cardiac remodeling. lack of vip in vip ko mice with the phenotype of biventricular dilated cardiomyopathy and primary pulmonary hypertension is concurrent with strong overexpression of cardiac muscle genes, supporting the concept of vip in vivo as a maestro conductor maintaining homeostasis of the heart.26 table 1 shows on the next page multiple heart failure gene programs which are overexpressed in vip ko mice. these physiological abnormalities are associated with premature death compared to the wild type (fig. 3). compensatory and pathogenic gene programs are displayed in figure 4, with overexpression of leptin as a compensatory reaction for cardiac cachexia and low body weight along with proinflammatory interleukin-6 (il-6) and il-1. vip increases intracellular camp and acts via protein kinase a to activate transcriptional promoters.32,33 vip upregulates il-10, which is anti - inflammatory.34 kupari found that vip levels in serum from healthy subjects and patients with aortic stenosis and heart failure were released into the coronary sinus. these authors concluded that, although vip was marginally elevated systemically in heart failure, it is the major neuroendocrine contributor to heart failure.35 smitherman infused vip into the left coronary artery of four other men at four levels. the maximum decline in coronary vascular resistance was 46% and was not associated with an increase in myocardial oxygen uptake. he concluded that 1) intravenous administration of low to intermediate doses of vip in humans is associated with substantial coronary vasodilation, 2) the coronary bed appears to be at least as responsive as other vascular beds, 3) the coronary vasodilation is due to both direct and indirect effects, and 4) the coronary vasodilation does not appear to be mediated by prostaglandins.36,37 so, the potential of an exogenous, sustainable vip drug would be to help relax coronary arteries. lucia also found that vip plasma levels were higher in heart failure patients with dilated cardiomyopathy compared to healthy subjects and was higher in older subjects. heart failure patients with higher new york heart association severity had lower levels of vip. these data also suggest that heart failure patients with worse disease have an inability to produce vip. the capacity of the elderly to produce vip is heartening, so that low levels are not due to aging itself.38 phasebio corporation recently conducted a phase 1, single ascending dose (sad) study in patients with essential hypertension. pb1046 was administered subcutaneously and was found to be well - tolerated and demonstrated a prolonged, dose - dependent effect on blood pressure, which was used as a biomarker for activity in this study. (see http://clinicaltrials.gov/ct2/show/nct01523067 for further details.) with this demonstration of safety and tolerability at efficacious doses, phasebio conducted a second phase 1 sad study in patients with essential hypertension, with the product administered intravenously to support the use of pb1046 in an acute setting (http://clinicaltrials.gov/show/nct01873885). subsequent phase 2 clinical studies will examine the efficacy of multiple ascending doses of pb1046 in patients with pulmonary arterial hypertension and heart failure. current therapies do not control progression of heart failure well and must often be used in combinations to optimize results. lcz696 utilizes a novel pathway to treat heart failure and improves mortality in comparison to the current standard of care, enalipril. it is important to remember that as a combination therapy utilizing an arb, it carries a risk of inducing life - threatening angioedema. ivabradine reduces rates of heart failure related hospitalization, but the indications for use are strict and this medication carries undesirable side effects that potentiate etiologic mechanisms of heart failure. vip, an endogenous peptide, shows widespread cardiovascular benefits as a vasodilator, an attenuator of smooth muscle proliferation, and a homeostatic regulator of heart failure gene programs thereby preventing hypertrophy of cardiac myocytes. endogenous levels of vip have been shown to be elevated in certain stages of heart failure. however, later stages of the disease show significantly diminished levels. administration of exogenous vip in early and late stages of heart failure may provide the most benefit for patients. use of an elastin polymer to protect and stabilize vip as a therapeutic agent shows the interface between engineering and medicine, and it bodes well for the influence of technology on society, particularly since heart failure will continue to grow as a prevalent problem as the population ages. vip may therefore have a chance to satisfy the unmet need as a treatment for acute heart failure, thus fulfilling discoverer dr sami i. said s dream to promote human health. | heart function fails when the organ is unable to pump blood at a rate proportional to the body s need for oxygen or when this function leads to elevated cardiac chamber filling pressures (cardiogenic pulmonary edema). despite our sophisticated knowledge of heart failure, even so - called ejection fraction - preserved heart failure has high rates of mortality and morbidity. so, novel therapies are sorely needed. this review discusses current standard therapies for heart failure and launches an exploration into emerging novel treatments on the heels of recently - approved sacubitril and ivbradine. for example, vasoactive intestinal peptide (vip) is protective of the heart, so in the absence of vip, vip knockout mice have dysregulation in key heart failure genes : 1) force generation and propagation ; 2) energy production and regulation ; 3) ca+2 cycling ; 4) transcriptional regulators. vip administration leads to coronary dilation in human subjects. in heart failure patients, vip levels are elevated as a plausible endogenous protective effect. with the development of elastin polymers to stabilize vip and prevent its degradation, vip may therefore have a chance to satisfy the unmet need as a potential treatment for acute heart failure. |
mutations of the gjb2 gene, encoding gap junction protein connexin 26 (cx26), are the most common cause of hereditary congenital hearing loss in many countries (2). cx26 is a member of the connexin family of gap junction proteins, which facilitate intercellular communication by encoding channels that directly link the cytoplasm of adjacent cells (3, 4). in spite of contribution of several different genes as causative agents of deafness, mutations in one gene encoding connexin 26 (genbank m86849, mim 121011) with chromosomal location 13q11 - 12 known as dfnb1 (omim 220290) responsible for half of severe to profound autosomal recessive non syndromic deafness in many populations (58). gjb2 is a small gene about 5500-bp length with two exons, of which only one (exon 2) contains the coding region. the coding region consists of 681 bp that encodes a gap - junction protein with 226 amino acids (6). in the present study, we report a novel dominant mutation (c202r) in an iranian patient with bilateral non - syndromic sensorineural hearing loss. a 40-year - old woman with bilateral hearing loss, for molecular analysis of deafness, was referred to welfare organization of marand, iran in august 2012. for molecular analysis, genomic dna was extracted from 1 ml of edta anticoagulated peripheral blood by rapid genomic dna extraction (rgde) method (9). polymerase chain reaction of coding region of cx26 gene was performed using cx26f : 5-tct ttt cca gag caa acc gc-3 as a forward primer and cx26r : 5-tgg gca atg cgt taa act ggc-3 as a reverse primer. pcr reactions were carried out in 25 l reaction mixture containing 1 pcr buffer, 1.5 mm mgcl2, 0.2 mm dntp, 10 pmoles of each primer, 0.5 u of taq dna polymerase and about 1g of genomic dna on a sensoquest (labcycler / germany) thermal cycler. pcr was performed under the following conditions : initial denaturation at 95c for 5 min, 34 cycles of denaturation at 95c for 45 sec, annealing at 60c for 1 min, extension at 72c for 1 min, followed by 8 min of final extension at 72c. the sequencing results were analyzed by sequencing - analysis chromas lite 2.1 software and were compared with the wild type. direct sequencing of pcr products in both directions revealed a novel heterozygous -t to -c transition at nucleotide 604 in codon 202 (tgccgc) of the gjb2 gene which replaces a cysteine with an arginine residue (c202r) (fig. 1). dna sequence of the gjb2 coding exon for the heterozygous -t to -c transition mutation. for molecular analysis, genomic dna was extracted from 1 ml of edta anticoagulated peripheral blood by rapid genomic dna extraction (rgde) method (9). polymerase chain reaction of coding region of cx26 gene was performed using cx26f : 5-tct ttt cca gag caa acc gc-3 as a forward primer and cx26r : 5-tgg gca atg cgt taa act ggc-3 as a reverse primer. pcr reactions were carried out in 25 l reaction mixture containing 1 pcr buffer, 1.5 mm mgcl2, 0.2 mm dntp, 10 pmoles of each primer, 0.5 u of taq dna polymerase and about 1g of genomic dna on a sensoquest (labcycler / germany) thermal cycler. pcr was performed under the following conditions : initial denaturation at 95c for 5 min, 34 cycles of denaturation at 95c for 45 sec, annealing at 60c for 1 min, extension at 72c for 1 min, followed by 8 min of final extension at 72c. the sequencing results were analyzed by sequencing - analysis chromas lite 2.1 software and were compared with the wild type. direct sequencing of pcr products in both directions revealed a novel heterozygous -t to -c transition at nucleotide 604 in codon 202 (tgccgc) of the gjb2 gene which replaces a cysteine with an arginine residue (c202r) (fig. 1). dna sequence of the gjb2 coding exon for the heterozygous -t to -c transition mutation. in the present study, we report very rare, novel and dominant mutation c202r in gjb2 gene that has not previously been reported in the cx26-deafness database. gap junctional intercellular communication (gjic) fulfills a multitude of different functions, tailored to meet the specific needs of organs, tissues or groups of cells in which cx are expressed. in the auditory system, intercellular channels formed predominantly by cx26 but also cx30 and cx31 seem crucial for maintaining a high extracellular electrical potential in the cochlea by facilitating the local circulation of k ions (2). the identification of mutations in the connexin 26 gene (gjb2 : mim # 121011) as a cause for profound sensorineural hearing impairment prompted a series of studies on gjb2 in affected families and subjects from europe, north america, the near east, north africa, and japan (10, 11). mutations in the gjb2 gene are the cause of an important number of cases of non - syndromic recessive deafness but are not as common in non - syndromic dominant deafness cases (2). a few dfnb1 mutations have been related to dominantly inherited hearing impairment, both nonsyndromic [r184q (11), w44c (12), c202f (1) and r143q (13) ] and syndromic with accompanying skin disease [g12r (14), g59a (15), dele42 (16) ]. to date, more than 14 different types of connexins have been identified, and each one contains four transmembrane domains (tm1tm4), two extracellular domains (ec1ec2), one cytoplasmatic loop (cl), and n and c - cytoplasmatic termini (nt ct). the n - terminal domain is involved in the insertion of the nascent polypeptide chain into the endoplasmic reticulum and, along with the first transmembrane domain, determines voltage gating. the extracellular loops regulate the connexon - connexon interactions, including heterotypic channel formation ; each loop contains three cysteine residues, conserved across all connexins that form essential intramolecular - disulphide bonds. c202r substitution, which lies in the fourth (tm4) transmembrane domain of cx26, may impair connexin - oligomerisation. alignments of the residue conservation of the mutated area of the gjb2 gene among the species (a) and different gap junction genes (b) dominant mutation, c202r, which associated with non - syndromic sensorineural hearing loss, has not previously been described in affected or control samples from other populations, indicating that it is a rare substitution. probably this novel mutation has prevalence in hearing loss patients in marand region and it seems that study of causative mutations in gjb2 gene and other genes related to deafness is necessary in this region. this work was helpful in providing genetic counseling to the affected family and helps in confirming the clinical diagnosis. ethical issues (including plagiarism, informed consent, misconduct, data fabrication and/or falsification, double publication and/or submission, redundancy, etc.) have been completely observed by the author. | mutations in the gjb2 gene are the most common known cause of hereditary congenital hearing loss. rapid genomic dna extraction (rgde) method was used for genomic dna extraction. after amplification of coding region of cx26 gene with specific primers, expected pcr products with 724bp length were subjected to direct sequencing in both directions. we describe here a novel heterozygous -t to -c transition at codon 202 (tgccgc) of the gjb2 gene in a patient, 40-year - old iranian woman, which replaces a cysteine with an arginine residue (c202r). the dominant mutation c202r associated with non - syndromic sensorineural hearing loss. this mutation has not previously been described in affected or control samples from other populations investigated for gjb2 mutations, indicating that it is a rare substitution. this dominant mutation was recorded in ncbi genbank with accession number kf 638275. |
linear growth retardation, or stunting, which is a manifestation of chronic malnutrition, is highly prevalent in under - developed and developing countries.[14 ] according to the world health organization, 215 million children were stunted in 2000 throughout the world. the national data from iranian national study of malnutrition prevalence in 2005 has indicated that 4.7% of iranian children are affected by stunting. earlier studies have shown that child 's stunting is associated with concurrent, and possibly later, delayed mental and motor development. the adverse effects of stunting could affect the adulthood by limiting work capacity owing to reduced muscle mass and increasing obstetric risks in women. although, several studies have identified determinants of stunting in different populations,[51015 ] limited data are available indicating its associates among iranian children. earlier studies have identified low birth weight, inadequate child care, feeding practice and birth order as a probable correlates of stunting # 94 ; wamani, 2007 # 82). it also has been shown that birth weight, breast feeding status, mothers age, family size, socio - economic status and birth order are associated with a higher prevalence of stunting among iranian children.[51822 ] previous studies have been limited in sample size and their sampling procedure ; such that their findings can not be easily extrapolated to other parts of the country. it remains unknown if the determinants of stunting in a metropolitan area like tehran are different from those in deprived regions of the country. this study was, therefore, conducted to investigate determinants of stunting in first grade primary school children of tehran. in this case - control study, participants were selected by multistage cluster random sampling method from 5 districts of tehran in 2007. first, 42 public schools (19 schools for girls and 23 for boys) were randomly selected based on proportional to size method in each district. in each selected school, all first grade pupils aged 7 years old were invited to participate in the study (n = 3147, 1343 boys and 1804 girls). stunting was defined as height for age less than the 5 percentile of cdc2000 cut - off points. based on this definition, 86 stunted children were identified and considered as case group in the current study. after matching for age, sex and residence area, 308 non - stunted children (height for age more than the 5 percentile of cdc2000 cut - off points) were randomly selected as control group. the study was approved ethically by research ethics committee of national nutrition and food technology research institute, tehran, iran and all participants provided informed written consent forms. required data about children 's birth weight and height, duration of breastfeeding and family demographic characteristics (including family size, parity, parents job, parents; education, marital status, mothers age, and home ownership) were collected by trained nutritionists using questionnaires. anthropometric measurements were done in all participants by two trained nutritionists and were randomly re - checked by a senior investigator (fe). weight was measured by means of a portable calibrated digital scale (breuer, germany) to the nearest 0.1 kg in light clothing without shoes. height was measured according to a standard protocol to the nearest 0.1 cm using height measure (seca 214, germany). mothers weight and height were measured by standard methods and fathers weight and height were collected through self - reporting. data were analyzed using statistical package for social science (spss inc, version 14). to ensure normal distribution of variables, kolmogrov - smirnov test was applied. continuous variables were compared using independent samples t - test between stunted and non - stunted children. distribution of participants in terms of categorical variables was examined through the use of chi - square test. multivariate logistic regression analysis was applied to identify the determinants of stunting. in these analyses, first the continuous variables were entered into the models and then these variables were treated as categorical variables. in this case - control study, participants were selected by multistage cluster random sampling method from 5 districts of tehran in 2007. first, 42 public schools (19 schools for girls and 23 for boys) were randomly selected based on proportional to size method in each district. in each selected school, all first grade pupils aged 7 years old were invited to participate in the study (n = 3147, 1343 boys and 1804 girls). stunting was defined as height for age less than the 5 percentile of cdc2000 cut - off points. based on this definition, 86 stunted children were identified and considered as case group in the current study. after matching for age, sex and residence area, 308 non - stunted children (height for age more than the 5 percentile of cdc2000 cut - off points) were randomly selected as control group. the study was approved ethically by research ethics committee of national nutrition and food technology research institute, tehran, iran and all participants provided informed written consent forms. required data about children 's birth weight and height, duration of breastfeeding and family demographic characteristics (including family size, parity, parents job, parents; education, marital status, mothers age, and home ownership) were collected by trained nutritionists using questionnaires. anthropometric measurements were done in all participants by two trained nutritionists and were randomly re - checked by a senior investigator (fe). weight was measured by means of a portable calibrated digital scale (breuer, germany) to the nearest 0.1 kg in light clothing without shoes. height was measured according to a standard protocol to the nearest 0.1 cm using height measure (seca 214, germany). mothers weight and height were measured by standard methods and fathers weight and height were collected through self - reporting. data were analyzed using statistical package for social science (spss inc, version 14). to ensure normal distribution of variables, kolmogrov - smirnov test was applied. continuous variables were compared using independent samples t - test between stunted and non - stunted children. distribution of participants in terms of categorical variables was examined through the use of chi - square test. multivariate logistic regression analysis was applied to identify the determinants of stunting. in these analyses, first the continuous variables were entered into the models and then these variables were treated as categorical variables. it was more prevalent in girls (4.4%) than boys (2.8%, p 3000 g were less likely to be stunted (or : 0.25 : 95% ci : 0.11 - 0.54) compared with those with a birth weight of 35 years) was associated with greater odds of being stunted (3.01 ; 1.19 - 7.60) compared with being born to younger mothers (160 cm were less likely to be stunted (0.04 ; 0.005 - 0.37) than those whose fathers height was less than 160 cm. birth weight, fathers height and maternal age were the main contributing factors to stunting in the studied population. the prevalence of stunting in this study was in agreement with the one reported by national study of malnutrition prevalence in iranian children. it seems that the prevalence of stunting has been decreased in the past decade in the country. the decreasing trend of stunting has been reported in south east asia, where the stunting has reduced in elementary school children from 52.4%in 1998 to 32.8% in 2000. worldwide stunting prevalence this can be explained by improved primary health care during pregnancy and early childhood, breastfeeding and growth monitoring. in this study, the prevalence of low birth weight and short duration of breastfeeding (< 6 mo) was higher in cases than that in controls. increased requirements in these infants, additional amounts of nutrients are needed for these babies to reach normal height growth. findings of the current study about breastfeeding are in agreement with those of marquis., in peru and semba., in indonesia and bangladesh which indicated that short duration of breastfeeding was positively related to prevalence of stunting. short duration of breastfeeding would prohibit adequate dietary intakes of energy, protein and micronutrients required for normal height growth. this finding is in line with earlier studies among brazilian, mexican and pakistani children. birth weight has also been reported as a strong predictor of child size in later life among children with intra - uterine growth retardation (iugr). low birth weight and subsequent stunting could be induced by maternal under - nutrition during pregnancy. in the current study adequate nutrition during pregnancy would result in increased birth weight and in turn would improve child growth in later life. this finding is in agreement with previously reported ones. in egypt, children born to mothers aged 35 years showed a higher prevalence of stunting than children born to mothers in other age groups. maternal age has also been reported as a determinant factor of stunting in south african children. the effect of the age of the mother on child health outcomes has also been explored in several studies. in the case of india, raj., showed that children born to mothers who were married below the age of 18 were at a higher risk of stunting and underweight compared to children of women who had married at age 18 or older. although, several studies have reported the importance of mothers young age at pregnancy on health outcomes of children, limited data have focused on older age. the current findings highlight the importance of old age of mothers as a contributing factor to stunting. as we all know socioeconomic status and dietary intakes we tried to collect data on economical status of the households ; however, after completion of the study, we found that these data are not reliable. one might suggest the use of living district as a proxy of socioeconomic status. in the current study, when we explored the frequency of stunting in different districts of tehran (5 districts were selected in this study), we failed to find a significant difference. due to the inclusion of children from different socioeconomic status in the current study, the findings can easily be extrapolated to the tehrani children, however, more caution are required about the generalizability of the findings to the whole country. we found that birth weight, maternal age and fathers height are the major contributing factors to stunting in this group of iranian children. furthermore, taking into account the determinants of stunting might help policy makers designing appropriate interventions. | background : limited data are available indicating associates of stunting among iranian children. this study was conducted to investigate determinants of stunting in first grade primary school children of tehran in 2007.method:in this case - control study, 3147 school children were selected by multistage cluster random sampling method from 5 districts of tehran. anthropometric measurements were done and stunting was defined as height for age less than the 5th percentile of cdc2000 cut - off points. eighty six stunted children were identified and considered as case group. after matching for age, sex and residence area, 308 non - stunted children were randomly selected as control group. required data were collected by trained nutritionists using questionnaires.results:stunting was prevalent among 3.7% of the study population (girls 4.4% vs. boys 2.8%, p 3000 g were less likely to be stunted (or : 0.25 : 95% ci : 0.11 - 0.54) compared with those with a birth weight of 35 years) was associated with greater odds of being stunted (3.01 ; 1.19 - 7.60) compared with being born to younger mothers (160 cm were less likely to be stunted (0.04 ; 0.005 - 0.37) than those whose fathers height was less than 160 cm.conclusions:we found that birth weight, maternal age and fathers height are the major contributing factors to stunting in this group of iranian children. taking into account the determinants of stunting might help policy makers designing appropriate interventions. |
already back in 1850, ignatz semmelweis was able to demonstrate in a spectacular manner that hand hygiene could prevent infections, but since then very few studies have been able to attest to the importance of hand hygiene in everyday practice. it is therefore all the more remarkable that in two studies of simple design, it has been possible to furnish proof of the influence of hand hygiene prior to insertion of peripheral venous catheters on the incidence of complications. both studies were conducted as practical exercises within the framework of the training course for infection control nurses in austria. using a questionnaire, hans hirschmann had the persons placing peripheral venous catheters record anonymously the following data : patient data, data and time of insertion, localization of insertion site, professional status of person placing the catheter (physician, nurse, medical student), place catheter fitted (or, ward, outpatient department), type of skin antisepsis (exposure time to an alcohol - based skin antiseptic 15 s), type of hand hygiene practiced before placement of the catheter (no hygienic measure, handwashing, hygienic hand disinfection with an alcohol - based product, donning of non - sterile disposable gloves) and, finally, the reason for catheter placement. on removing the peripheral venous catheter the person doing so now entered the following information on the same questionnaire : date and time of removal, reason for removal as well as complications (redness, swelling, pain, fever of unknown origin, pus at insertion site, other reason). for the first study, 1148 questionnaires were completed in an austrian hospital. on the basis of the average consumption of peripheral venous catheters (approx. 600 per week) a data registration quota of approx. each patient received on average 1.4 peripheral venous catheters during their hospital stay, calculated on the basis of the number of patients and consumption of venous catheters. evaluation of the questionnaires revealed that there was a sharp increase in the incidence of complications in line with an increasing duration of the indwelling period of between 24 and 96 hours. the unexpected but conspicuous finding of this study was that in the case of catheters with an indwelling period of more than two days there were highly significantly lower complication rates in those cases in which hand disinfection was carried out or gloves donned before catheter placement. conversely, there was a higher rate of complications in those cases in which the hands were washed or hand hygiene measures dispensed without before placement. the findings of the first study gave rise, under the aegis of euridiki (european interdisciplinary committee for infection prophylaxis), to the initiation of a further analogous study, so as to verify these remarkable findings regarding the influence exerted by hand hygiene prior to insertion of peripheral venous catheters on the incidence of complications. this study was also carried out as part of the training course for infection control nurses in parallel at three austrian hospitals, with the aforementioned questionnaires being completed and evaluated for a total of 1132 peripheral venous catheters. overall, 379 separate complications were recorded, with more than one complication being seen per catheter (table 1 (tab. 1)). the proportion of peripheral venous catheters implicated in complications was 24%. using uni- and multivariate analyses, various factors implicated in the complication rates 2)). there was a sharp rise in the relative risk of complications the longer the catheter was in place. for the period between 49 - 72 h and more than 72 h, the complication rates were significantly higher than those in the 0 - 24 h period. this finding lends credence to the demand that the duration of the indwelling period be limited in general to 2 - 3 days. catheters placed in the or during the patient s hospital stay showed a significantly lower risk of complications than those inserted on wards, or even in the outpatient department. the likely explanation for this is that already when entering the or sluice, hand hygiene measures are taken that have a positive effect on the complication rates. univariate analysis showed a significantly higher relative risk for venous catheter complications in patients older than 70 years compared with younger patients, but this was not the case for the multivariate analysis. is all probability this was due to the fact that in the case of patients younger than 70 years, only in 5% of cases did the duration of the catheter indwelling period exceed 72 h, whereas this was longer than 72 h for 13.8% of the catheters in the case of patients older than 70 years. as regards the role of hand hygienic measures prior to catheter insertion, it was possible to fully confirm the results of the first study. conductance of hygienic hand disinfection or donning of disposable gloves, as compared with normal hand washing or omission of hand hygienic measures, led to markedly lower complication rates, which proved to be significant in the uni- as well as the multivariate analysis. the most plausible explanation for this positive effect exerted by the wearing of disposable gloves or conductance of hygienic hand disinfection prior to catheter placement is that there was no recontamination during repalpation, as often seen, of the prepared venepuncture site. the study reported here was one of the chief reasons why the guideline drafted by the robert koch institute (rki) prevention of vascular - associated infections assigned hygienic hand disinfection prior to insertion of peripheral venous catheters to category i a (recommendation based on well - designed experimental or epidemiological studies). specialist for hygiene ; head of austrian agency for health and nutrition safety, institute for microbiology and hygiene. gnther wewalka studied medicine at the university of vienna and received his doctoral degree there in 1971. first he became assistant doctor at the institute for hygiene, and then changed to the professorship for chemotherapy of the 1. medicinal university clinic until he went back to the institute of hygiene where he habilitated in 1981. in 1987 he was appointed head of the federal bacteriological - serological analysis institute (today : austrian agency for health and nutrition safety, institute for medicinal microbiology and hygiene) in vienna. apart from this he is hospital hygiene consultant for the city of vienna, as an internationally respected hospital hygienist he is advisor for several hospitals, also a board member of the austrian society for hygiene, microbiology and preventive medicine (ghmp from 2002 - 2004 chairman thereof), member of the task force influenza pandemieplan, member of the national committee for the eradication of polio and much more. the european task force on legionella - infections (ewgli) as well as all national associations and committees are highly benefiting from his special knowledge in this area. specialist for hygiene ; head of austrian agency for health and nutrition safety, institute for microbiology and hygiene. gnther wewalka studied medicine at the university of vienna and received his doctoral degree there in 1971. first he became assistant doctor at the institute for hygiene, and then changed to the professorship for chemotherapy of the 1. medicinal university clinic until he went back to the institute of hygiene where he habilitated in 1981. in 1987 he was appointed head of the federal bacteriological - serological analysis institute (today : austrian agency for health and nutrition safety, institute for medicinal microbiology and hygiene) in vienna. apart from this he is hospital hygiene consultant for the city of vienna, as an internationally respected hospital hygienist he is advisor for several hospitals, also a board member of the austrian society for hygiene, microbiology and preventive medicine (ghmp from 2002 - 2004 chairman thereof), member of the task force influenza pandemieplan, member of the national committee for the eradication of polio and much more. the european task force on legionella - infections (ewgli) as well as all national associations and committees are highly benefiting from his special knowledge in this area. | using two studies of a simple design it has been possible to furnish proof of the influence of hand hygiene prior to insertion of peripheral venous catheters on the incidence of complications. in the first study detailed data were collected anonymously for each patient on the procedure used for catheter insertion or on any complications. data were collected for around 64% of patients in one hospital. evaluation of the questionnaires revealed that there was a significant increase in the incidence of complications in line with an increasing duration of the indwelling period of between 24 and 96 hours. the unexpected finding of this evaluation study was that in the case of catheters with an indwelling period of more than two days there were highly significantly lower complication rates in those cases in which hand disinfection was carried out or gloves donned before catheter placement. the second study, based on the former, documented the cases giving rise to complications. the proportion of peripheral venous catheters implicated in complications was 24%. here, too, there was a sharp rise in the risk of complications in line with the duration of the indwelling period. catheters placed in the or during the patient s hospital stay showed a significantly lower risk of complications than those inserted on wards, or even in the outpatient department. conductance of hygienic hand disinfection or the wearing of disposable gloves resulted in significantly lower complication rates compared to normal handwashing or omission of a hand hygiene measure. the most plausible explanation for this positive effect exerted by the wearing of disposable gloves or conductance of hygienic hand disinfection prior to catheter placement is that there was no recontamination during repalpation, as often seen, of the prepared venepuncture site. |
important factors determining the rate of tooth alignment include the bracket slot dimension, the associated inter bracket span, the choice of arch wire, and frictional forces generated between bracket and arch wire. friction has been defined as the resistance to motion that occurs when two objects move tangentially to each other. friction, which impedes sliding movements, is determined by multiplying the coefficient of friction of the materials in contact by the normal force, which is the force of ligation in orthodontic appliance. an ideal ligation system for alleviation of crowding in orthodontic treatment primarily should ensure full bracket engagement of the arch wire and should exhibit low friction between brackets and arch wire. currently, two main types of brackets are available for practicing orthodontists - conventional (cl) and self - ligating (sl) brackets. in a conventional tie wing bracket, wire ligatures and elastomeric ligatures have been documented with higher frictional forces, which reach undesirable levels relative to those that are ideal for tooth movement. sl brackets are considered a ligature - less bracket system with an inbuilt metal labial face, which provides the combination of low friction and full arch wire engagement.. the claimed advantages of both types of sl bracket systems include increased patient comfort due to the absence of ligatures, improved oral hygiene, less chair time, and shorter overall treatment time. however, there are also certain disadvantages, including difficulty with the full expression of torque, frequent failure of the clips, and brackets that are bulkier and more expensive than conventional brackets. to address some deficiencies of sl brackets, novel ligatures such as the leone slide ligature (ll) leone slide ligatures (ll) have been developed with a polyurethane fourth wall, which allows the arch wire to slide freely in the slot while transmitting its full force to the tooth. several retrospective and prospective in - vivo studies have compared the efficiency of sl and cl brackets during various stages of treatment, reporting no significant differences during initial alignment. did not find any significant differences when they prospectively compared damon sl, smart clip (3 m unitek) sl with victory series cl brackets during initial alignment. pandis. also found no difference when comparing time to alignment in the mandibular arch between damon 2 (ormco) sl and microarch (gac international) cl brackets. a retrospective study by hamilton comparing in - ovation (gac) sl brackets and victory series cl brackets found no measurable difference in initial alignment time. the above - said studies were based on non - extraction treatment plan and lack the evidence about the benefits of sl brackets for extraction patients during initial alignment. scott. conducted a randomized controlled trial of patients having mandibular first premolar extractions and concluded damon 3mx sl brackets were no more efficient than synthesis (ormco) cl brackets during mandibular alignment. ong. compared efficiency of damon 3mx sl brackets and victory series (3 m unitek) cl brackets for anterior arch alignment and passive space closure and found no significant difference between sl and cl groups. early retrospective studies include the possibility of outcome bias and subsequent prospective clinical trials had the effect of confounding factors, such as various malocclusions treated with many modalities and methods, lack of testing for the equivalence pre - treatment characteristics of the sample, ill - defined selection criteria and variability with respect to arch wire sequences, and bracket types among the groups. to date, there is little robust clinical evidence in the form of a well - designed prospective randomized clinical trial with adequate control over clinical variables : (bracket composition, dimension, arch wire type and sequence, inter - appointment interval and treatment plan), comparing the efficacy of the 5 available modes of ligation over treatment time to alignment, passive space closure, and changes in mandibular incisal inclination at the end of first phase of fixed appliance therapy. hence, the purpose of this prospective randomized clinical trial was to compare the efficacy of 5 ligation modes, elastomeric ligation (el), stainless steel ligation (ssl), leone slide ligation (ll), passive self - ligation (psl), and active self - ligation (asl). the primary outcome measure was rapidity of tooth alignment, and secondary outcome measures were passive space closure and changes in the mandibular incisal inclination. the participants and their parents or guardians were informed of the study, its implications, and written consent was received from all. a randomized prospective clinical trial design was opted. the subjects were recruited from consecutive patients attending the orthodontic department between january 2011 and february 2012 and satisfied the inclusion criteria. the subjects were randomly allocated for treatment with elastomeric ligation (ell), stainless steel ligation (ssl), leone slide ligatures (ll), passive self - ligation (psl), and active self - ligation (asl) brackets using unstratified subject allocation sequence made using a computer - generated randomization program done by a statistician independent from the study (http://www.randomization.com). numbered opaque sealed envelopes containing the treatment allocation card were prepared by the statistician before the trial commencement. after the satisfaction of inclusion criteria and consent obtained from the subjects, the successive subjects were handed over the sealed envelopes by the secretary of the department, in order to conceal the assignments from the clinician until the appointment at which the appliance was to be placed. demographic and clinical characteristics of sample a post - hoc sample size calculation with = 0.05 and power = 80% reveals, a sample of 10 subjects per ligation group could be justified as a median sample size. the bracket systems used were 0.022 slot mbt (gemini series, 3 m unitek) for el, ssl, ll and 0.022 slot mbt (smart clip, 3 m unitek) for psl, and 0.022 slot mbt (in - ovation euro, dentsply gac, usa) for asl. two operators treated the patients in all the 5 groups, and the bonding method was standardized between the groups using conventional etching and transbond (3 m unitek) bracket adhesive. copper - ni - ti (cu - ni - ti) 35c and 0.016 0.022-inch stainless steel (ormco ; true - arch form) working wire in place for 1 month. the patients were reviewed every 6 weeks, and the first arch wire was left in place until the teeth were passively engaged in all the bracket slots before proceeding to the second arch wire. in bracket systems ligated with elastomeric modules anchorage augmentation procedures like tpa in the upper arch and lower lingual arch were placed in all the patients at the initial appointment of appliance placement and were carried on to the retraction phase of mechanotherapy. passive lace backs were placed in both the quadrants from the power arm of first molar to the canines in a figure 8 fashion in all the groups. the distal ends of all the wires were cinched distal to the first molar tube. pre - treatment characteristics were recorded including patient age, gender, sex, lower incisal irregularity index, lower first premolar extraction space, and lower incisal inclination. pre - treatment study models were evaluated by using little 's irregularity index to quantify the alignment of 6 mandibular anterior teeth. mandibular dental casts and lateral cephalograms were taken at appliance placement (t1) and on the alignment date (t2) when the patient was considered complete after 1 month of working 0.016 0.022 stainless steel archwires and upon visual inspection of the correction of proximal contacts. the time to alignment (t2 - t1) was calculated for each patient in days. extraction space (t1) was measured from the closest points on the adjacent teeth before extraction. the mesiodistal widths of the teeth to be extracted were not used because they were often displaced from the arch form ; this decreased the extraction space to be closed. similarly, extraction spaces at (t2) were measured from the closest points on the crowns of the teeth on either side of the extraction space. coded lateral cephalograms were traced, and mandibular incisal inclination was assessed for all patients at t1 and t2 using the angular measurement : mandibular incisor to mandibular plane. therefore, the researcher was blinded to patient 's name, the ligation type during data collection to minimize systematic error. the study models were measured with digital calipers (digimatic ntd 12 - 6c, mitutoyo corp, tokyo, japan). all model and cephalometric measurements were measured by the researcher. to assess the intra - examiner reliability, 15 plaster models and 15 cephalometric radiographs were randomly selected from the records. the cephalometric radiographs were retraced, and the measurements of the cephalometric variables were repeated. in the dental casts, the reproducibility of the measurements was investigated with intra - class correlation coefficient (icc). the analysis showed no significant difference between the first and second measurements (icc = 0.96). demographic and clinical characteristics were investigated with conventional descriptive statistics. before performing the test of significance, normality assumptions were tested by kolmogorov - smirnov and shapiro - wilk test. anova was performed for normally distributed variables, and non - parametric tests (chi - square and kruskal - wallis) were used for non - normally distributed variables. comparison between 5 ligation groups with respect to the treatment duration, passive space closure, and change in the mandibular incisal inclination was investigated by anova followed by post - hoc bonferroni analysis. correlations as well as partial correlations for time to alignment as the dependent variable and other independent variables were calculated followed by a stepwise regression analysis. the statistical analyses were performed with spss software (release, 11.0, spss for windows, spss chicago, iii). demographic and clinical characteristics were investigated with conventional descriptive statistics. before performing the test of significance, normality assumptions were tested by kolmogorov - smirnov and shapiro - wilk test. anova was performed for normally distributed variables, and non - parametric tests (chi - square and kruskal - wallis) were used for non - normally distributed variables. comparison between 5 ligation groups with respect to the treatment duration, passive space closure, and change in the mandibular incisal inclination was investigated by anova followed by post - hoc bonferroni analysis. correlations as well as partial correlations for time to alignment as the dependent variable and other independent variables were calculated followed by a stepwise regression analysis. the statistical analyses were performed with spss software (release, 11.0, spss for windows, spss chicago, iii). fifty subjects were recruited for the study and of them, 48 subjects completed the study and those who failed to complete the treatment were omitted from the statistical analysis (per - protocol analysis). baseline demographic and clinical characteristics for the 5 groups showed no discrimination with respect to age, gender, irregularity index, thus validating the random assignment of ligation methods to the 5 groups [table 1 ]. the mean duration of treatment time to alignment was 161.19 days overall : 176.30 days in the ell group, 175.56 days in the ssl group, 159.10 days in the ll group, 152.67 days in the psl group, and 142.90 days in the asl group [table 2 ]. anova was used to compare the mean treatment time to alignment between 5 ligation systems followed by post - hoc analysis. a statistically significant difference was found between all the groups, except for ell vs.. ssl, ll vs.. psl, and psl vs.. asl [table 3 ]. table 4 shows the mean passive space closure in each ligation group with 2.90 mm overall : 2.00 mm in ell, 2.09 mm in ssl, 3.06 mm in ll, 3.50 mm in psl, and 3.85 mm in asl. inferential statistical investigation showed a significant difference between all the groups, except for ell vs.. ssl, ll vs.. psl, and psl vs.. asl [table 5 ]. mean treatment time to alignment by ligation group comparison of mean treatment time to alignment between ligation systems mean change in the passive space closure by ligation group in millimeters comparison of the passive space closure between ligation groups table 6 shows the mean change in mandibular incisal inclination in each ligation group with 3.17 overall : 4.40 in ell, 4.39 in ssl, 2.50 in ll, 2.78 in psl, and 1.85 in asl. inferential statistical investigation showed a significant difference between all the groups, except for ell vs.. ssl and ll vs.. psl [table 7 ]. mean change in the mandibular incisal inclination by ligation group in degrees comparison of change in the mandibular incisal inclination between ligation groups pearson correlation coefficient for time to alignment, change in mandibular incisal inclination, passive space closure and irregularity index are 0.72, -0.839, and 0.457, respectively. all these correlation coefficients are significant (p < 0.01) [table 8 ]. correlation matrix indicates that the time to alignment is positively correlated with change in mandibular incisal inclination and irregularity index, while passive space closure and time to alignment are negatively correlated. partial correlation of time to alignment with change in mandibular incisal inclination, passive space closure is significant (p < 0.05), and the partial correlation of time to alignment with age and change in mandibular incisal inclination is not significant [table 9 ]. pearson correlation coefficients partial correlation coefficients multiple regression analysis was done to alignment as dependent variable, age, irregularity index, change in mandibular incisal inclination, and passive space closure as independent variables. r square value of 0.925, indicating that this linear model explained 93% of variability in time to alignment as influenced by the independent variables. the anova indicated that the dependent and independent variables are having a linear relation and the linearity is significant. the final linear model included passive space closure, irregularity index, and mode of ligation as significantly contributing variables in deciding the time taken for alignment while age and change in mandibular incisal inclination were identified as non - significant variables, hence were excluded from the regression model. the final regression equation is : time to alignment = 134.07 - (7.38 passive space closure) + (11.68 irregularity index) - (5.28 ligation group). the model shows that 1 mm of passive space closure contributes to 7.38 days reduction in time to alignment when irregularity index and ligation group are fixed variables, 1 mm increase in the irregularity index increases the time to alignment by 11.67 days by keeping the passive space and ligation groups as fixed variables. there will be a reduction of 5.28 days in time to alignment if the ligation groups are changed in the order from ell to asl per each group when passive space closure and irregularity are held constant [table 10 ]. our study is the first to simultaneously quantify, relate, and compare time to alignment, passive space closure, and change in the mandibular incisal inclination between 5 different ligation systems. the results of this study suggest that sl groups (psl, asl) were more efficient than cl groups (el, ssl) in terms of treatment time to alignment. the results of this study emphasize the clinical relevance of the in - vitro assessment of frictional protocols in many studies during the past decade, which has often demonstrated a significant decrease in friction with sl systems compared with cl brackets designs, which is highly desirable during the initial alignment phase. this observation could be attributed to the substantial greater free play of the sl appliances in contrast to the conventional ligatures that act as obstacles because of the stress they exert on the arch wire adjacent to the bracket sides, precluding the free sliding of the wire into the slot walls, thus reducing the rate of tooth movement. our findings have demonstrated asl performing better than the psl system with respect to the treatment time to alignment, but the difference was statistically insignificant. the possible explanation behind this observation could be due to the storage of some of the force in the active clip as well as in the wire with asl brackets, which means a given wire will have its range of labiolingual action increased and, therefore, produce more alignment than would a passive slide with the same wire. in sufficiently lingually placed tooth with an active spring clip in relation to its neighboring teeth, a higher total force will usually be applied to the tooth in comparison to a passive clip. but, such force is unlikely to be detrimental with modern low modulus wires since several studies have shown that only large deflections are likely to enable a superelastic wire to show a plateau of force for a range of deflection, which is unlikely to happen with passive self - ligation. ll group was significantly more efficient from the cl group but was comparably inferior to the asl group in terms of treatment time to alignment. the results are in accordance with an in - vitro study in the past, which states that sl systems and leone slide ligatures on conventional brackets produce significantly lower frictional forces compared with conventional ligatures. passive space closure, incisal proclination, and dental arch expansion have been considered as contributing factors for alleviation of crowding during orthodontic arch alignment. however, studies have reported intentional development of the arch width, with appliance systems has been found to be highly unpredictable. hence, we, in our study, have abstained from evaluating the transverse arch dimensions. the present study reflected a significant advantage of sl systems over cl systems with respect to passive space closure with less extraction space to close actively in sl category, which could eventually reduce the overall treatment time. the results demonstrate an identical performance between ll and psl groups, psl and asl groups, while the asl groups had significantly increased passive space closure in comparison to ll groups. the results concur with a previous study, which suggested that sl brackets might encourage passive space closure during initial alignment. however, ong., in their study found no difference in the amount of passive space closure during initial alignment between the bracket types. mandibular incisal inclination during initial alignment was significantly influenced by the bracket system with sl brackets resulting in significantly lesser incisal proclination. the results suggest that cl appliances resolve crowding more through an incisal proclination while sl brackets relieve crowding by facilitating more passive space closure. this phenomenon with sl brackets would help in preventing the torque loss during the initial alignment, prevent round tripping of the anterior teeth, thus minimizing root resorption and would greatly minimize the net effective anchorage loss during the overall treatment time of an individual. to summarize, sl systems (psl, asl) had a significant measurable advantage over cl systems (ell, ssl) with respect to the three variables : (a) reduction in treatment time to alignment, (b) increased passive space closure, and (c) decrease in incisal proclination. ll system was comparatively beneficial over cl systems but inferior to asl systems and with an insignificant difference from psl systems. the study showed an insignificant difference in the clinical performance between ell, ssl in conventional, while asl performed significantly superior to psl system in sl groups. the results of our study, however, differ with previous studies in the rapidity of alignment, amount of passive space closure, and mandibular incisal inclination. the difference could be explained by the variability of bracket designs utilized in sl and cl appliances with their characteristic prescriptions, variability in arch form, arch wire sequence, and final working arch wires. certain studies have compared damon2, damon3 psl brackets with a 0.019 0.025-inch working stainless steel arch wire to cl systems with mbt prescription (victory series, 3 m unitek). a thicker wire of 0.019 0.025-inch is likely to completely fill the slot contributing to an increase in friction as well as torque effectiveness, thereby increasing the time to alignment with sl systems when the arch wire sequence is carried on till the 0.019 0.025-inch stainless steel working arch wire. in studies that had 0.014 0.025-inch copper - nickel - titanium damon arch form wherein measurements are taken, the lack of significant difference could be attributed to the high torque version of damon mx that increases the frictional resistance at the bracket slot arch wire interface. in addition, narrower bracket design of damon 3mx can increase the contact angle and contribute to elastic binding and notching with wires of higher dimension and stiffness. in our study, we have standardized mbt prescription, arch forms, and arch sequence to all the 5 ligation groups with measurements taken at the end of a 0.016 0.022-inch stainless steel working arch wire. a 0.016 0.022-inch wire would create a slop of 0.005-inch, which reduces the frictional resistance in all the ligation groups and thereby could enhance the alignment efficiency of active spring clip and passive sl systems through a more passive space closure and lesser incisal inclination. for majority of bracket arch wire combinations, the friction values obtained were significantly higher than with only the terminal brackets ligated, confirming that ligation plays an important role in generating friction. the strength of the study was standardization of clinical variables such as bracket composition, dimension, prescription, arch wire type, arch wire sequence, inter - appointment intervals, category of dento - alveolar malocclusion and extraction pattern. consecutive eligible patients were included to minimize confounding variables with method of ligation being the critical difference between treatment groups. despite the strict inclusion, exclusion criteria, and standardization of the clinical variables, a major limitation with the study is the sampling bias with a low initial sample size recruited into the study although lost to follow up was minimal. the aim of a randomized clinical trial is to provide an unbiased assessment of the effects of a treatment intervention. a small sample size could eventually lead to a random error that can arise when the results of the clinical trial could be different from the truth. in small trials, the probability of reporting a clinically important effect due to chance is relatively higher and the effects of random error on the estimates decreases as the trial becomes larger. hence, a similar study with a larger sample size is needed in future to minimize the random error that could be associated with the present study. the results of this rct suggest that clinicians can appreciate the effectiveness of sl systems when used with undersized wires. this situation, however, could be negated as treatment progresses and wires of higher dimension and stiffness are engaged in the brackets. treatment efficiency is the product of many mechanical and biological factors. along with individual biological variation, bracket type can significantly influence the treatment efficiency and overall treatment span in orthodontic mechanotherapy. self - ligating brackets were more efficient than conventional brackets in anterior alignment, passive space closure, and mandibular incisal inclination during initial phase of orthodontic treatment. | objectives : to conduct a prospective randomized study comparing the efficiency of 5 different ligation systems (ell ; elastomeric ligature, ssl ; stainless steel ligature, ll ; leone slide ligature, psl ; passive self - ligation and asl ; active self - ligation) over the duration of mandibular crowding alleviation.materials and methods : fifty consecutive patients (54.2% male, 45.8% female ; mean age : 16.69 years) satisfying the inclusion criteria were randomly allocated to 5 ligation groups with an equal sample size of 10 per group. the 5 groups received treatment with 0.022-inch mbt pre - adjusted edge - wise technique (ell : gemini 3 m unitek, ssl : gemini 3 m unitek, ll : gemini 3 m unitek, psl : smartclip 3 m unitek and asl : in - ovation r euro gac international). the models and cephalograms were evaluated for anterior arch alignment, extraction space closure, and lower incisal inclinations at pre - treatment t1 and at the end of initial alignment t2. analysis of variance (anova) and post - hoc tests were used for data analysis.results:forty-eight participants completed the study, and sl systems showed a significant difference over cl groups in time to alignment, passive space closure, and incisal inclination. multiple regression showed a reduction of 5.28 days in time to alignment by changing the ligation group in the order of ell to asl group and 1 mm increase in initial irregularity index increases time to alignment by 11.68 days.conclusion:self-ligation brackets were more efficient than conventional ligation brackets during initial leveling and alignment. |
mucinous carcinoma of the breast is uncommon, the reported incidence of pure mucinous carcinoma being 2% of all breast cancers. pure mucinous carcinoma has a far better prognosis than the mixed variety noted in several studies. we report a case diagnosed as pure mucinous carcinoma breast on fine needle aspiration cytology (fnac) in a female of reproductive age group. a 30-year - old female presented with a large firm lump in upper quadrant of right breast for 4 years, which was gradually increasing in size and was associated with pain. there were numerous moderately pleomorphic epithelial cells lying either discretely forming loose clusters or entrapped within stromal material [figure 1 ]. at places, fnac smear showing abundant pink mucoid material (arrow) with entrapped tumor cells (arrow head) (h and e, 400) right mastectomy specimen with axillary tail measuring 20 cm14 cm4 cm was received. overlying skin measured 20 cm11 cm, and the axillary tail measured 9 cm6 cm2 cm. cut surface revealed circumscribed mass with variegated appearance, grey brown in color, and solid to cystic with areas of necrosis and hemorrhage. tumor mass measured 7 cm6 cm5 cm, and extended from subepidermal tissue to posterior resected margin. four lymph nodes were identified in axillary tail, with the largest measuring 3 cm2 cm1 cm and the smallest measuring 1 cm0.5 cm0.5 cm. cut surface of the largest was gelatinous and necrotic. three axillary lymph nodes with fibrofatty tissue received separately ranged from 0.5 to 1.5 cm, and had unremarkable cut surface. sections from different areas of specimen were studied. sections from tumor mass revealed large areas of necrosis, myxoid degeneration and lakes of mucoid material [figure 2 ]. the tumor cells were round to polyhedral, exhibiting mild pleomorphism but frequent mitotic figures. these cells were mostly arranged in thin trabeculae, single rows and few solid cellular areas. tumor was reaching close to posterior resected margin but was separated by a thin rim of uninvolved fibroadipose tissue. microscopically, seven lymph nodes were identified of which the largest was almost completely replaced by tumor metastasis. tumor tissue was negative for estrogen receptor (er) and progesterone receptor (pr). section showing lakes of mucoid material (arrow) with enmeshed column of tumor cells (arrow head) (h and e, 400) there were numerous moderately pleomorphic epithelial cells lying either discretely forming loose clusters or entrapped within stromal material [figure 1 ]. at places, fnac smear showing abundant pink mucoid material (arrow) with entrapped tumor cells (arrow head) (h and e, 400) right mastectomy specimen with axillary tail measuring 20 cm14 cm4 cm was received. overlying skin measured 20 cm11 cm, and the axillary tail measured 9 cm6 cm2 cm. cut surface revealed circumscribed mass with variegated appearance, grey brown in color, and solid to cystic with areas of necrosis and hemorrhage. tumor mass measured 7 cm6 cm5 cm, and extended from subepidermal tissue to posterior resected margin. four lymph nodes were identified in axillary tail, with the largest measuring 3 cm2 cm1 cm and the smallest measuring 1 cm0.5 cm0.5 cm. cut surface three axillary lymph nodes with fibrofatty tissue received separately ranged from 0.5 to 1.5 cm, and had unremarkable cut surface. sections from tumor mass revealed large areas of necrosis, myxoid degeneration and lakes of mucoid material [figure 2 ]. the tumor cells were round to polyhedral, exhibiting mild pleomorphism but frequent mitotic figures. these cells were mostly arranged in thin trabeculae, single rows and few solid cellular areas. tumor was reaching close to posterior resected margin but was separated by a thin rim of uninvolved fibroadipose tissue. microscopically, seven lymph nodes were identified of which the largest was almost completely replaced by tumor metastasis. tumor tissue was negative for estrogen receptor (er) and progesterone receptor (pr). section showing lakes of mucoid material (arrow) with enmeshed column of tumor cells (arrow head) (h and e, 400) mucinous carcinoma of the breast is an uncommon entity seen in postmenopausal females, accounting for only 2% of all breast carcinomas. however, we are reporting a case of mucinous carcinoma in a young female aged 30 years. the behavior of the tumor tends to be less aggressive, so it has a better prognosis than other breast malignancies. as soon as another pattern becomes evident as a component of tumor mass, the lesion qualifies as a mixed tumor. two lesions most likely to be confused with mucinous carcinoma are mucoid fibroadenoma and mucocele like lesion. mucinous carcinoma is er positive, and in less than 70% cases, it is pr positive. nearly all pure mucinous carcinomas are diploid, while over 50% of mixed variety is aneuploid. only 315% of pure variety shows axillary node metastasis compared to 3346% of the mixed type. in the present case, the present case did not show any distant metastasis either at the time of diagnosis or during the 1 year of follow - up. adjuvant chemotherapy containing cyclophosphamide, adriamycin and 5 fluorouracil (5 fu) was given at every 3 weeks interval. histochemically, the mucins secreted by this tumor are distinct o - acylated forms of sialomucin. immunohistochemically, there is strong muc (mucin) 2 cytoplasmic immunoreactivity and decreased muc i immunoreactivity compared with ductal carcinoma not otherwise specified (nos). we have reported a case of pure mucinous carcinoma in a young female emphasizing the role of fnac in its early diagnosis. | mucinous carcinoma of the breast is a relatively rare, pure form accounting for 2% of all breast cancers. pure mucinous carcinoma of the breast has a favorable prognosis. the common age is postmenopausal group. here, we report a 30-year - old female patient diagnosed on cytology as mucinous carcinoma of the breast with lymph node metastasis and subsequently confirmed by histopathology. in 1 year follow - up, the patient did not show pulmonary or distant metastasis and received adjuvant chemotherapy at every 3 weeks interval. |
for the pharmacometrician and the system pharmacologist, computer software is, quite simply, the equivalent of the laboratory for the molecular biologist, or of the research clinic for the practicing clinician. it is the environment where tools are developed, observations and conjectures are made, hypotheses are formulated and tested, multiple scenarios are run, and discoveries are made for the betterment of science or patients. it is only natural that cpt : psp, in its efforts to merge pharmacometrics and systems pharmacology, wishes to be the home to cutting - edge tutorial papers that can bring practitioners of one discipline closer to the other, spreading beyond the initiated to familiarize our audience with an ever - increasing number of tools. such familiarity is expected to facilitate new findings, developments, and discoveries by indicating capabilities and potential applications of pharmacometrics and systems pharmacology software and helping with repeatability of the study, an essential element in any scientific discipline. in the words of umberto eco, thinking back to another powerful medium used to propagate information, if you want to use television to teach somebody, you must first teach them how to use television. the cpt : psp designation for a tutorial, as stated in the journal 's guide to authors, is an educational article providing practical tutorial on tools, methodologies, and approaches in pharmacometrics and systems pharmacology. this is of crucial importance for the continued health of both pharmacometrics and systems pharmacology because the pace of changes in these dynamic fields appears to outpace any concerted efforts to provide timely didactic needs by current higher education systems. an integrated technical education on modeling and simulation tools has been sporadic, carried out individually in centers scattered around the globe. as explained in the inaugural editorial and in a description of our disciplines foundational tutorials, cpt : psp strongly supports (technical) tutorials that aid the dissemination and uptake of model - based approaches in pharmacometrics and systems pharmacology, targeting a wide spectrum of audiences from novices to advanced modelers. inevitably, many of these tutorials may focus on one or more software tools, some of which may be commercial products or otherwise proprietary. the cpt : psp editorial team has been debating the content of the tutorials, their scientific scope, and also whether in any way to limit the scope of such tutorials to those where the software is available for free (whether tout court, for research purposes, or for research purposes to noncommercial organizations only), because this fits into the mission of the journal to support the dissemination of the discipline. this editorial is the result of our deliberations and defines our current position on this topic ; we are putting it forth in cpt : psp to be proactive and in the hope that it will stimulate some needed debate. an important (but often underappreciated) distinction we need to make early on is between a software tutorial and a user 's manual. we would most often consider user 's manuals not publishable by cpt : psp, as they consist of a sequence of steps and/or reference material that software users can consult as they solve real - world problems or attend a hands - on workshop to learn the mechanics of the software. cpt : psp looks to publish software tutorials that contain a real - world scientific problem (case study) and apply the method or the software to solve that problem step - by - step, much like it is done in a real - world scientific inquiry scenario. that way, the usefulness of the document is increased, as the reader now learns twice over : from the scientific basis of the solution to the case study, and the operational aspects of the specific computer software being used. a software tutorial is thus richer than a user 's manual, as it includes both a description of the scientific problems the software is meant to solve and the practicalities of the software tool itself, whereas a user 's manual most often contains only the latter. we collectively believe that cpt : psp should focus on software tutorials, whereas user 's manuals may be published separately (in a biomedical computing journal, for example) or become a useful appendix to a tutorial manuscript (e.g., available through a web link). in general, the tutorials should provide a balanced view of the field and include consideration of other, similar software. the journal requires all software to be freely available under the open source initiative for an article to be considered for publication. the best case in point would probably be that we could not publish a nonmem tutorial if we were to apply such criteria. to us, there seems to be a fundamental difference between the bioinformatics / computational biology discipline (where software has always been free and open source, since the very early days of the field) and ours (where software has very often, almost exclusively, been commercial). however, unambiguously adjudicating the scientific merit of publications requires that the study be repeatable by other researchers, and this will still be facilitated when the files and data used for any piece of research are made available to others, as opposed to the software platform itself. an analogy can be drawn when details of an analytical method to assay chemicals or genes are given, but the ability to repeat the study is still impinging on access to a liquid chromatography mass spectrometry machine or gene sequencer, which obviously can not be easily provided as an appendix to the report ! let us elaborate further on these aspects and on what this implies for the authors and readers of software tutorials published in cpt : psp. in many ways, the intent of the more traditional approach of including in research articles the model (differential) equations in plain text ensures that mathematical models can be ultimately implemented (with a bit of work !) in any computer code. however, everyone with an interest in this topic is painfully aware that what gets in the way of the process is that the information is usually incomplete and the textual reporting is error prone. publishing or making available computer code or modeling scripts is intended to streamline the process of implementing anew models described in cpt : psp research articles. curated databases are also increasingly available that are accessible to all free of charge, without significant restrictions. it is apparent that databases such as these are of value, and unrestricted access has in general to be evaluated for costs and benefits against other modes of dissemination. we certainly see availability of computer code as a very important avenue to maximize the impact of our published tutorials. a compromise that we are already striving to implement in the cpt : psp research articles would be to upload enough information to enable the reader to reproduce the model results, assuming the software platform is broadly available, perhaps at a cost. this could be as simple as a parameter set (in native format), or as detailed as the actual model code or tutorial script (e.g., control files for nonmem), workspace file, and so forth, depending on the specifics of the platform used. this would enable the reader access to the software to reproduce the simulations, even if the software is commercial or otherwise proprietary. the only requirement would be that the journal reader needs to be able to run the code in the native software format and reproduce the published simulations (for verification) with no modifications. of note, this approach is indirectly borne out in the life sciences : a dna sequence is required to be published in some journals, but this does not require the author to supply a sequencer machine as well. the tools to run a simulation can be separate from the conditions and inputs that need to be specified, i.e., all the details that permit an independent reader re - run the study. this is in line with the requirements from other journals, and it is our belief that such an approach would satisfy the great majority of readers. financial interests are often complex and hard to define, and software fees vary widely in the amount of profit they bring. some tools may require a fee which includes user support, and others may charge separately for extended service (through consulting, for example). at any rate, we feel that there may be a place in cpt : psp also for commercial software. this is provided the considerations we outlined above about reproducibility of the simulations can be met. we encourage submissions of tutorials where authors are not exclusively employed by the developer, or where not all authors have a financial interest in the software. we recognize the potential limitations of this ; however, we believe the user base who regularly applies these tools would be attracted to manuscript co - authorship with the developers, thus bringing the user community 's specific perspective to the tutorial text. finally, regardless of its potential shortcomings, it seems to us that peer review will continue to be the method of choice for propagation of scientific information on new discoveries and novel approaches to technical problems, and our journal of course wholeheartedly supports this notion. our view is that peer - reviewed software tutorials to be published in cpt : psp should include : a description of one or more scientific motivating problems that will be used to structure the tutorial flow ; instructions as to how to use the software to solve this or similar modeling analysis problems ; scripts or other computer code or pseudocode, as appropriate, to enable a user to reproduce the tutorial sequence ; and an inclusive author list, ideally comprising users without monetary interests in the software, in addition to the software developers. a line attributed to the mathematician paul halmos states that the best way to learn is to do ; the worst way to teach is to talk. after committing some of our thoughts to paper, we now prefer to let our submissions do the talking. accordingly, we hope that the software tutorials we will host on the pages of cpt : psp will contain relevant, useful, reproducible, durable, and practical examples to effectively assist our readers in learning, and in turn sharing and thus growing, our craft. | in addition to methodological tutorials,1 cpt : psp has recently started to publish software tutorials.2,3 our readership and authors may be wondering what kind of format or product is expected, and the review of submissions we have already received prompted several discussions within the psp editorial team. this editorial reflects on these discussions and summarizes their salient points. it aims at providing some details about the current vision of cpt : psp for software tutorial articles. in addition, it brings some clarity on the topic of what role commercial software tutorials can have in cpt : psp and how cpt : psp tutorials differ from publications which describe the software itself, as those which can be found in other computer science journals. finally, the discussion includes reproducibility considerations and the general use of commercial and noncommercial software in cpt : psp publications. we hope our thoughts, and especially a stated requirement to publish user input to the software to aid in reproducibility, will help in guiding our authors and will stimulate healthy debate among our readers about the evolving nature of our science, how it can be facilitated using software and associated databases as a conduit, and what role this journal can play in fostering both the best modeling and simulation practices and the best scientific approaches to computational modeling, to bring the advantages of modeling and simulation to all regular practitioners, and not to just a (self) selected few. |
the institute of medicine (iom) report to err is human revealed the number and significance of adverse events and errors that occur during hospitalization. the report was a call to action to transform healthcare systems to ensure patient safety and higher quality care. in one step toward healthcare transformation, the centers for medicare and medicaid (cms) no longer reimburses institutions for the care, or treatment, associated with certain hospital - acquired conditions. understanding what factors contribute to adverse incidents during hospitalization is essential to developing effective counter measures. in order to improve factors that are modifiable within a hospital structure or with healthcare delivery, there are a number of potential contributing factors that need to be considered such as the patient 's condition, the care the patient receives, and the environment in which they receive care [3, 4 ]. battles and lilford provide a conceptual model for patient safety that includes antecedent conditions, which would include the patient 's comorbid conditions, the primary reason the patient was admitted to the hospital, and characteristics the patient possessed before entering the hospital. their model also includes the structure, or environment, in which the patient receives care such as the hospital, or nursing unit. also acting within the structure are the processes of care (the interventions or treatments) delivered by the multidisciplinary team caring for the patient in the hospital. none of these components exist in isolation, which is why it is important to examine all of these factors and how they interact. the purpose of this study was to examine factors that contribute to adverse incidents that occur during hospitalization by creating a model that included patient characteristics, clinical conditions, nursing unit context of care variables, medical treatments, pharmaceutical treatments, and nursing treatments. the research question addressed in this study is : what patient characteristics, clinical conditions, context of nursing care variables (e.g., nursing hours per patient day, rn skill mix, number of units resided on during hospitalization), and treatments (medical, pharmaceutical, and nursing treatments) explain the occurrence of adverse incidents for hospitalized, older adults at risk for falling ? a model that has been used successfully to guide multidisciplinary effectiveness research in the hospital setting can be seen in figure 1 [5, 6 ]. data for this exploratory study came from a large, health service effectiveness grant and was approved by the institution 's human subjects review board. data from a four - year period (july 1, 1998 to june 31, 2002) were extracted for the primary study from one large midwestern academic medical center. data sources came from nine electronic data repositories, including the nursing information system that used the nursing interventions classification (nic) to electronically document nursing care delivered. detail of the nine electronic repositories and methods to assure validity and reliability are discussed elsewhere. extracted data were stored in a structured query language (sql) server and relational databases were built using a unique subject number. the inclusion criteria were hospitalizations to one midwestern tertiary care hospital over a four - year period, patients 60 years of age or older upon admission, and at risk of falling. patients were determined to be at risk of falling based on a fall risk assessment that was completed upon admission or when the patient received the nursing intervention of fall prevention as recorded in the electronic documentation system. patients at risk for falling were selected with the rationale that they would be at risk for experiencing one adverse incident (i.e., falling), and therefore interventions would be initiated to prevent the adverse incident. in addition, the hospitalizations were selected as the unit of analysis rather than individual patients and a variable was included to control for patients who had more than one hospitalization. conceptual and operational definitions for the independent variables included in the explanatory model are displayed in table 1 and organized by the conceptual model seen in figure 1 (patient characteristics, clinical conditions, context of care, and treatments). when appropriate, the source used to guide coding of variables is provided ; for example, pharmaceutical treatments, or medications, were coded using the american hospital formulary service (ahfs) codes. the dependent variable for this analysis was the first occurrence of an adverse incident during an episode of hospitalization. adverse incidents were defined as any undesired circumstance that lead to, or could have led to, personal harm. adverse incidents included falls, medication errors, procedure - related events (e.g., wrong patient, wrong procedure or test), equipment - related events (e.g., equipment malfunction, unplanned removal, improper set - up), and new conditions (e.g., skin breakdown). due to the large number of study variables, a four - step model building process using logistic regression was used to answer the research question. each independent variable included in the analysis was tested independently using a bivariate analysis and a score statistic to determine the association with occurrence of an adverse incident. in this bivariate analysis, no other variables were statistically controlled for. variables with p values 0.15 were retained for step two. a p value 0.15 was used as the criterion to guard against eliminating variables too soon in this exploratory analysis. the variables retained in step one (p values 0.15) were then analyzed within their respective conceptual variable blocks (i.e., patient characteristics, clinical conditions, context of care, medical treatments, pharmaceutical treatments, and nursing treatments) using logistic regression. a backward elimination process was used, indicating that the variable with the largest p value was eliminated and the analysis was rerun on the remaining variables within the block. this procedure was repeated until all variables within the block had a p value 0.15. a p value of 0.15 during step two was chosen to guard against eliminating variables too soon because they might yet prove to have a statistically significant effect when combined with variables from other conceptual blocks. a model integrating all of the conceptual variable blocks was built in a progressive fashion using the variables that were retained in step two. the significant variables were added to the model by their respective blocks. starting with the significant variables in block one (patient characteristics) and block 2 (clinical conditions), a model was built using the backward elimination process described in step two until the only variables remaining in the model were those with a p value 0.15. the significant variables from block three (context of care) were then added to what remained of blocks one (patient characteristics) and two (clinical conditions) in the model. once again, a backward elimination process was performed until the only variables remaining in the model were those with values 0.15. this process of adding blocks and using the backward elimination continued until the last block (nursing treatments) was added. at this point, when the significant variables from the final block were added and backward elimination was performed, the criterion for significance was decreased to a p value 0.05. this resulted in a final model containing only those variables with a p value 0.05. in step three, variables with a p value 0.05 in the logistic regression indicated that variables were significantly related to the dependent variable (occurrence of an adverse incident) after controlling for the other variables in the model. covariates used for risk adjustment included age, severity of illness, and number of hospitalizations during the study period (see table 2). step four added these covariates used for risk adjustment (severity of illness, age, and more than one hospitalization during the study period) to the model to those that were significant in step three. categorical variables with more than two categories were analyzed by comparing each level to a reference category. for example, severity of illness (four levels from minor to severe) was analyzed by comparing each of the three upper level categories to the lowest level of severity of illness (i.e., minor). the mean age was 73.7 years ; most were retired (74.4%), caucasian (93.5%), female (52.6%), and admitted from home (64.4%). this patient group, defined primarily by receiving the nursing treatment fall prevention, was medically diverse. the most common primary medical diagnoses were diseases of the circulatory system (28.5%), neoplasms (13.8%), and injury, including fractures, or poisoning (11.5%). the most commonly experienced adverse incident for this patient group included medication errors (37%), falls (27%), and equipment - related events (14%). the bivariate correlations completed in step one are not included in table 2 due to space constraints but are available from the authors upon request. the second column in table 2 illustrates variables retained from step one that were analyzed within blocks with p values 0.15 (step two of model building) and thus retained for step three. the third column includes p values from the third part of the modeling building process, prior to adding covariates used for risk adjustment to the final model (step four). five patient characteristics entered step one of the model building process but none were significant beyond step two. age, although not significant in any of the three model building steps, was entered in the final model for risk adjustment. nine primary medical diagnoses were retained from step two, four were retained from step three, and three were retained (p 0.05) in the final model (see tables 2 and 3). as the results in table 3 indicate, other nervous system disorders, other primary cancer and senility and organic mental disorders were all significant (p 0.05) in the final model. other nervous system disorders was the only primary medical diagnosis of the three inversely associated with experiencing an adverse incident (o.r. = 0.43), indicating that hospitalizations with this medical diagnosis were less likely to suffer an adverse incident compared to hospitalizations that did not have this condition. other primary cancer and senility and organic mental disorders were both positively associated with experiencing an adverse incident with odds ratios of 1.94 and 1.57, respectively. severity of illness, although not significant in step three, was entered into the final model for risk adjustment. severe and major severity of illness categories were significantly (p 0.05) and positively associated with experiencing an adverse incident compared to the lowest severity of illness category (i.e., mild) (see table 3). seven comorbid conditions were retained from step two for inclusion in step three but none were significant and thus were not retained for inclusion in the final model. past hospitalizations during the study period were significant in step two but not in step three (see table 2). however, this variable was entered into the final model to adjust for patients that had experienced more than one hospitalization during the study period. in the final model (table 3) four context of care variables, the number of units the patient resided on during hospitalization, the dip proportion (falling below the unit 's average staffing), skill mix, and the average caregiver patient ratio (cgpr), were significant in step two (see table 2) but only two variables, the dip proportion and average cgpr, were significant in step three and retained for the final model (see table 2). both were significant in the final model (step four) as illustrated in table 3 the average cgpr (rn hours per patient day (hppds)) was categorized as quartiles to enable comparison and interpretation for this nonlinear variable. the two highest average cgpr quartiles (9.5 rn hppds and 6.6 rn hppds) were significantly (p 0.05) and inversely associated with experiencing an adverse incident, indicating that when compared to the lowest quartile of staffing (4.1 rn hppds), the odds of experiencing an adverse incident decreased in the highest two quartiles of nursing hours per patient day. the odds of experiencing an adverse incident for hospitalizations with the highest average cgpr quartile (9.5 rn hppds) were 0.76 of the odds for hospitalizations that experienced the lowest average cgpr quartile (4.1 rn hppds). the odds of experiencing an adverse incident for hospitalizations with the second highest average cgpr (6.6 rn hppds) were 0.62 of the odds for hospitalizations in the lowest cgpr average quartile. the cgpr dip proportion was significantly (p = 0.011) and positively associated with experiencing an adverse incident. the results shown in table 3 are in terms of 0.2 increments of change and indicate that for each 20% fall in staffing below the average, the odds of experiencing an adverse incident increase by 15% (o.r. the number of medical treatments received during hospitalization and 20 types of medical treatment were significant in step two (see table 2) and were therefore included in step three. in step three of the analysis, the number of medical treatments received during hospitalization and one medical treatment type, physical therapy, were significant (p 0.05) and retained for the final model. both were positively associated with experiencing an adverse incident (see tables 2 and 3). the results indicate that for each additional medical treatment received during hospitalization, the odds of experiencing an adverse incident increased by approximately 3% (o.r. = 1.52) more likely to experience an adverse incident than hospitalizations that did not receive this medical treatment. the number of unique medications received during hospitalization and 27 specific pharmaceutical treatments (i.e., medications types) were significant in step two of the analysis (p 0.15) and thus retained for step three. the number of unique medication types and four types of medications were significant in step three (see table 2) and all were significant in the final model (see table 3). the number of unique medications was positively associated (p < 0.001) with experiencing an adverse incident (o.r. receipt of succinimides, caloric agents, and eent anti - infectives during hospitalization increased the odds of an adverse incident. ammonia detoxicants were inversely associated (p = 0.021) with experiencing an adverse incident (o.r. the number of unique nursing treatments received during hospitalization was not significant but 38 types of nursing treatments were significant (p 0.15) and entered into step three (see table 2). eleven were significant at step three and ten were significant in the final model (see tables 2 and 3). the nursing treatment pressure ulcer care, received by 91.5% of the sample, was divided into thirds based on the average number of times per day it was delivered (see table 1). the results for the three categories of use are interpreted in comparison to hospitalizations that did not receive the nursing treatment. the middle and low use categories of pressure ulcer care were significantly (p 0.05) and positively associated with experiencing an adverse incident, indicating that hospitalizations that received pressure ulcer care a little less than once every other day (use rate = 0.41) or once every four days (use rate = 0.25) were more likely to experience an adverse incident than hospitalizations that did not receive pressure ulcer care. the medium (use rate = 0.34) and low (use rate = 0.10) categories were significantly (p 0.05) and positively associated with experiencing an adverse incident (see table 3). both health screening and neurologic monitoring had low use categories that were significantly (p 0.05) and positively correlated with experiencing an adverse incident. the results indicate that hospitalizations that received the low use of these two nursing treatments were more likely to experience an adverse incident than hospitalizations that did not receive the associated nursing treatment (see table 3). the medium use category of blood products administration (use rate = 0.89) was significantly (p 0.05) and positively (o.r. = 1.49) associated with experiencing an adverse incident. hospitalizations that received blood products administration a little less than once a day were almost 50% more likely to experience an adverse incident than hospitalizations that did not receive blood products administration. all three categories of use for the nursing treatment restraint were significantly (p < 0.01) and positively associated with experiencing an adverse incident (see table 3). the high use category had an average delivery of 16.47 times a day and hospitalizations that received high use of restraint had more than double the odds (o.r. = 2.16) of experiencing an adverse incident compared to hospitalizations that did not receive this nursing treatment. hospitalizations that received restraint approximately four and a half times a day (medium use category) had almost double the odds (o.r. = 1.86) of experiencing an adverse incident compared to hospitalizations that did not receive restraint. the lowest category of use was delivered an average a little more than once a day and increased the likelihood of experiencing an adverse incident by 58% (o.r. the remaining significant nursing treatments were delivered to less than 5% of the sample and were therefore operationalized as dichotomous variables so that hospitalizations that received the nursing treatment at least once are compared to hospitalizations that did not receive the treatment (see table 1 for definition). active listening received at least once by 4.8% of the sample was significantly (p < 0.001) and positively (o.r. mood management was received by only 2.5% of the sample but was delivered an average of 3.1 times per day when it was delivered. = 1.84) of experiencing an adverse incident compared to hospitalizations that did not receive mood management. cast care maintenance was another nursing treatment that was delivered frequently (more than five times a day on average) when hospitalizations required it. = 2.00) of experiencing an adverse incident compared to hospitalizations that did not receive this nursing treatment. the average use rate for hospitalizations that received this treatment was slightly more than once every ten days (use rate = 0.21). = 2.03) for hospitalizations that received this nursing treatment compared to hospitalizations that did not receive music therapy (see table 3). none of the patient characteristics were significant, indicating that patient characteristics were not explanatory variables of adverse incidents, given the other variables that entered the model. also nonsignificant were two clinical conditions : number of past hospitalizations during the study period and comorbid medical conditions. this indicates that after controlling for other variables in the model, patient characteristics of this sample of older adults were not significant for experiencing an adverse incident during hospitalization. other nervous system disorders were inversely associated with experiencing an adverse incident. this inverse relationship may be explained by considering the type of nursing unit these patients are typically admitted to. a primary medical diagnosis of nervous system disorder, which is composed of peripheral and central nervous system disorders along with more generic symptoms of a nervous system disorder, would likely warrant admission to a neurology unit in this academic medical setting where the nursing personnel are skilled in the care of these patients and may recognize the need for increased surveillance. this heightened surveillance for patients hospitalized with the primary medical diagnosis of other primary cancer are on high - risk medications, some that call for double - checks, and that may increase the number of medication errors that are discovered. the third primary medical diagnosis, senility and organic mental disorders, appears similar in nature to other nervous system disorders but is positively associated with experiencing an adverse incident, unlike other nervous system disorders. this may be because patients who have senility and organic mental disorders are less capable of using safety equipment in their environment like call lights and hand rails and are more likely to be dispersed among a variety of general medical or surgical units. the environment and specialized nursing expertise may not be readily available to meet the unique care demands of individuals with this primary medical condition. in the final model, the top two severities of illness categories (i.e., severe and major) were significantly and positively associated with experiencing an adverse incident. this is not surprising, as patients who are sicker often have complex care issues which may place them at greater risk to experience an adverse incident. related to the structure of care (context of care), the two highest categories of the average cgpr (rn hppds) were significantly and inversely associated with experiencing an adverse incident compared to the lowest quartile, indicating that when there are more nursing hours per patient day, there is a decreased likelihood of preventing an adverse incident. the more the rn staffing fell below the nursing unit average, the more likely an adverse incident was to occur during that hospitalization. this finding indicates that not only is the number of nurses, or hppds, an important predictor of adverse incidents but so is staffing below the average on a nursing unit. this may indicate that units develop effective processes dependent upon their average staffing and when the staffing is altered, the processes are impacted. staffing below the unit average places the patient at greater risk for having an adverse incident. processes of care included medical, pharmaceutical, and nursing treatments. both the number of medical treatments and the number of unique medications received during hospitalization were positively associated with experiencing an adverse incident. as the number of procedures and medications increased so did the odds of having an adverse incident (e.g., medication error, wrong site surgery, trauma, etc.). there was one medical treatment, physical therapy, and two medication types, succinimides and ammonia detoxicants, that were significantly associated with experiencing an adverse incident, which may be related to falls. the positive association between physical therapy and adverse incidents may be a reflection of patients with decreased functional status who are at greater risk for falling. similarly, succinimides are anticonvulsives and are in the same ahfs class as barbiturates and benzodiazepines, which are positively associated with falls. ammonia detoxicants was the only pharmaceutical treatment in the final model inversely associated with experiencing an adverse incident (see table 3). patients who require ammonia detoxicants often have conditions associated with liver dysfunction, which makes it more difficult for them to excrete ammonia that builds up in their body. patients that have high ammonia levels are often confused, disoriented, difficult to direct, and are at great risk for falling for these reasons. there was one nursing treatment, pressure ulcer care, that is used to treat an adverse incident (i.e., pressure ulcer). there were also a number of nursing treatments positively associated with adverse incidents where providing the treatment showed that the patient likely had greater exposure to an adverse incident than patients who did not receive the treatment. one example is the nursing treatment specimen management where a patient is more likely to have a mislabeled lab as an adverse incident than a patient who did not receive this treatment. similarly, all three categories of restraint were significantly and positively associated with experiencing an adverse incident. only 8.5% of the hospitalizations in this sample received restraint at least once but the use rates were relatively high, especially the high use category with an average delivery of 16.47 times per day. these findings also show that use of restraints does not prevent adverse incidents (e.g., falls) and in fact may contribute to them as has been demonstrated in other research [26, 27 ]. active listening, mood management, and music therapy may be used as complementary therapies for patients who are distressed, confused, or combative when other treatments have not worked. hospitalizations that require these nursing treatments may be at greater risk for falling because the patient is unable to follow commands, is impulsive or unable to communicate effectively. this study was conducted at one academic medical center and therefore further multisite research is needed. although the effectiveness research model used in this study includes many important, patient and multidisciplinary components, there were important aspects of care that impact patient safety such as the individual characteristics of the clinicians involved in care (e.g., experience, education) and how they interact with one another (e.g., teamwork, communication) that were not included in this study. this study examined a number of patient conditions, structural variables, and process of care variables to better understand what factors contribute to adverse incidents during hospitalization. this is one of the first studies to show that delivered nursing treatments help explain adverse incidents in hospitalized, older adults. this study also used a multidisciplinary model that considered medical and pharmaceutical components of treatment, which are critical when providing care of the older adult in acute care. with this more robust multidisciplinary model, rn staffing was still an important explanatory variable for adverse incidents, which is congruent with findings from other research [29, 30 ]. | the purpose of this study was to examine factors that contribute to adverse incidents by creating a model that included patient characteristics, clinical conditions, nursing unit context of care variables, medical treatments, pharmaceutical treatments, and nursing treatments. data were abstracted from electronic, administrative, and clinical data repositories. the sample included older adults hospitalized during a four - year period at one, academic medical facility in the midwestern united states who were at risk for falling. relational databases were built and a multistep, statistical model building analytic process was used. total registered nurse (rn) hours per patient day (hppd) and hppds dropping below the nursing unit average were significant explanatory variables for experiencing an adverse incident. the number of medical and pharmaceutical treatments that a patient received during hospitalization as well as many specific nursing treatments (e.g., restraint use, neurological monitoring) were also contributors to experiencing an adverse incident. |
therefore, action must be taken immediately to diagnose and treat such intracranial complications before irreversible tissue damage has occurred, and both physicians and patients at putative risk should be aware. here, a case of sinusitis originating from an odontogenic focus leading to severe subdural empyema with subsequent neurological sequelae a 45yearold man was admitted to the ophthalmological department with a periorbital swelling so severe it disabled him to open his left eye. he was overweight (bmi = 28), without any known diabetes, was a smoker, and was undergoing treatment for rheumatoid arthritis (ra) ; methotrexate tablets (25 mg / week). at the time of admission the patient was febrile (38.7c) and his state of consciousness was moderately affected. on inspection the left periorbital region showed massive swelling ; the skin was warm, red, and sore ; and the left pupil was vaguely responsive equaling a small relative afferent pupillary defect. the vital signs were as follows : pulse, 103 beats / min ; respiration, 16 breaths / min ; blood pressure, 118/71 mmhg ; hemoglobin, 7.3 gm / dl ; white blood cell count, 18.5 10/l ; neutrophils, 15.6 10/l ; creactive protein (crp), 298 meq / l ; procalcitonin, 12.1 meq / l ; potassium, 3.3 meq / l ; and sodium, 133 meq / l. after ophthalmic examination a computed tomography (ct) scan of the orbit was performed and showed left orbital inflammation interpreted as postseptal cellulitis, exophthalmos, and these findings led to the performance of an acute canthotomy in an effort to save the patient 's vision on the left eye. computed tomography scan in the axial plane showing exophthalmos and dirty fat (arrow) on the left eye. on the same day large amounts of pus were evacuated from the left ethmoidal, frontal, and maxillary sinuses, and only a limited volume was present in and removed from the periorbita. simultaneously two teeth (10, 14) were extracted due to apical periodontitis with pus under pressure. the patient was given a dose of antibiotics, cefuroxime (1500 mg) and metronidazole (500 mg), following surgery and 6 h postoperatively. an acute ct scan of the cerebrum was performed and a subdural accumulation visualized on the left side along with midline shift (fig. 2). computed tomography scan with contrast in the axial plane showing midline shift and a subdural accumulation (arrow) on the left side. under general anesthesia a burr hole surgery (bhs) was performed, and large amounts of pus were emptied under high pressure, and an external ventricular drain (evd) was placed in the anterior horn of the right lateral ventricle by neurosurgeons. postoperatively the patient was aphasic and could only answer questions with onesyllable words. on hospital day 3, a control ct still showed subdural empyema along with swelling of the left hemisphere, bleeding in the left frontal lobe, and perifocal edema (fig. 3). computed tomography scan in the axial plane showing swelling of the left hemisphere, empyema, bleeding in the left frontal lobe (arrow), and perifocal edema. day 4, the patient 's gcs was 9, and a new ct showed extensive swelling of the left hemisphere and still some empyema. the ventricles had collapsed and therefore the evd did not produce any pus or cerebrospinal fluid. the crp had not changed much despite the antibiotics and antifungal medication ; voriconazole (400 2 mg the first day, then 200 2 mg the following) now was initiated. in addition to the bhs, 510 ml pus was drained from the extracranial abscess in relation to the left eye. subsequently a sinuscopy was performed revealing an edematous mucosa and large amounts of coagulated blood, which were removed. upon ophthalmic examination on hospital day 5 the patient 's left pupil was found nonresponsive and the vision of this eye was declared lost. there were still clear signs of inflammation and staphylococcus aureus, fusobacterium nucleatum, propionibacterium species, and group c streptococcus were found in the pus ; only group c streptococcus was found in the blood. the antifungal medication was discontinued, while metronidazole (500 3 mg), meropenem (2 3 g), and linezolid (600 2 mg) treatment was initiated. by hospital 4) of the brain showed improvement ; decreasing midline shift, increasing depth of sulci, and visible basal cisterns. computed tomography scan in the axial plane showing decreased midline shift. during the next 2 days over the course of 3 weeks he regained the ability to walk and his ability to construct more advanced sentences improved. fortyfour days after admission the patient was discharged from the hospital. he still had neurological sequelae from the infection, such as loss of vision on the left eye and speech impairment as well as loss of shortterm memory. generally, most cases of sinusitis, including suppurative infections, are uncomplicated. however, in some instances like this case the suppurative infection can spread and affect the orbita as well as intracranial structures, causing subdural empyema. the sinusitis is believed to infect the subdural space either through osteomyelitis of the skull or by retrograde movement of septic thrombi via the small veins of the involved sinus 2, 3, 4. since subdural empyema is such a rare complication to sinusitis and in this case odontogenic, it can be difficult to diagnose because of a low index of suspicion. when the patient was initially admitted to the hospital no dental complaints were mentioned and the symptoms from the eye were the predominant focus of the examination. in addition, the initial examination did not reveal any dental abnormalities nor raise any suspicion to the fact that the patient was immunosuppressed. this may explain why the dental infectious focus was not detected until during the first surgery. the rapid progression of the infection in this case is probably due to immunosuppression as a result of methotrexate treatment. it is important that action is taken early on to provide effective treatment and avoid a fatal outcome 5. therefore, it is important that patients experiencing orbital symptoms undergo cranial imaging at the slightest suspicion of suppurative complications. in this case the patient underwent acute canthotomy as a result of the ct scan performed on the date of admission. despite this early intervention, the patient 's vision was lost, which may be contributed to the fact that he waited for several days before consulting a physician, while the swelling was building up. a contributing factor to the rapid progression of the infection in this particular patient may be the suppression of the patient 's immune system due to ra and methotrexate intake, allowing the infection to persist despite the first bhs. acute severe infection is a known risk of methotrexate treatment 8 ; however, the dose in this case was within the lowdose interval (25 mg / week). despite the low dose, the patient might have benefitted from being instructed by the prescribing doctor to see a physician at the representation of infectious symptoms. generally the management of a subdural empyema has three components : early localization and treatment of the primary infectious focus, drainage of the empyema itself, and administration of antibiotics 1. in this case the infection originated from an odontological focus and the two affected teeth were extracted early on. no consensus exists on the surgical intervention of choice as either bhs or craniotomy can be used. because of limited exposure when using bhs there is a higher incidence of recurring pus formation 3. this might explain why the first bhs in this case was not sufficient to eliminate the subdural empyema. as shown in this case of an aggressive purulent odontogenic and sinogenic infection with severe orbital and neurologic sequelae, early awareness, diagnosis, and treatment are essential, especially in immunosuppressed patients. it is important to recognize that purulent spread to the orbit and intracranial requires immediate medical and surgical intervention. | key clinical messagesubdural empyema is a rare but potentially lifethreatening complication to sinusitis. awareness of infection and early diagnosis is of the essence when dealing with a putatively immunosuppressed patient. furthermore, patients at increased risk of infections due to immunosuppression need to be fully informed of risks associated with their treatment. |
establishing good communication, either verbal or nonverbal, with patient is an essential and important component to develop a good doctor - patient relation. face - to - face interaction (including facial expressions and eye contact), expressive touch, body language, paralinguistics (vocal communication which is discrete from actual language), interpersonal proximity, physical appearance, and eloquent gestures all make verbal conversation more expressive and meaningful. evidence shows that physician 's nonverbal behavior leads to higher patient satisfaction, but this is affected by a number of factors, including gender of the doctor as well as of the patient [3, 4 ]. a study, from switzerland, showed both male and female doctors should display different set of nonverbal behavior to maximize patients satisfaction. nonverbal communication has been shown to be important in dealing with pediatric age groups and with those recovering from disabilities [79 ]. eye contact and physical touch are commonly used as effective tools in nonverbal communication [1, 10 ]. general practitioners may feel reluctant to use touch other than procedural touch, because of the fear of misinterpretation of such behavior however, many patients believe that, particularly in distressing situations, expressive touch is acceptable. a study conducted in canada on female patients concluded that, as compared to males, females were more tolerant of comforting touch. eye contact is another important nonverbal behavior and is especially essential for building good rapport with elderly individuals. it is mostly taken as a sign of respect, care, and attention from a doctor. however, if eye contact is coupled with attentive listening it inclines the interaction towards more patient - centered communication. nowadays, the use of computers and especially the electronic health records (her), during medical interviews, is a big obstacle in using eye contact as an effective way to communicate. we therefore directed our study to explore the expectations and perceptions of patients, in a developing asian country, regarding touch and eye contact by physicians during consultancy, as most of studies on nonverbal communication were conducted in west and we consider that responses in our part of the world would differ. this cross - sectional study was conducted at the community health centre (family practice clinic), at the aga khan university hospital, karachi, pakistan (akuh), during the months of january to march 2012. akuh is one of the major, tertiary care teaching hospitals in karachi, with state - of - the - art family practice clinics. on an average, 200 family practice patients with mostly primary and secondary care level problems were seen daily by 1216 family physicians at the clinics. an interview based questionnaire was developed by the researchers through literature search (for validated questions), input from colleagues and from experts in the field of family medicine. the questionnaire was then translated from english version to urdu language and was back translated in english to check for reliability and any discrepancies found were removed. the questionnaire was pretested on 10 participants (data not included in the results) before finalizing it. interviewers (two medical graduates) were trained for data collection and on the use of the questionnaire to ensure uniformity of application. the questions were asked in english and in urdu depending on the participant 's ease with the language. patients were chosen randomly and interviewed in the waiting area, before their consultancy with the physician, irrespective of their age, gender, educational status, and the reason for their visit. those who were willing to participate did not receive any monetary compensation and were asked to sign a written consent form and the provision of confidentiality was ensured to them. section 1 included demographics like age, gender, religion, education, marital status, and occupation. in section 2 the questions were focused on the nonverbal communication modes of touch and eye contact only, exploring the patient 's expectations about willingness, comfort, and perceptions regarding the two modalities. other questions were directed to know which part of the body they would allow the physician to touch and the duration of eye contact. the study was reviewed and approved by the dow university of health sciences, karachi. the data was entered on statistical package for social sciences (spss) version 16. proportions were calculated for all the variables of interest and chi - square test was used to assess relations of gender and educational level with nonverbal communication modalities. a p - value of < 0.05 was considered statistically significant throughout the study. a total of 133 patients were approached, out of which 120 patients agreed to participate and were interviewed. the mean age of the participants was 34.9 10.9 years, out of which 32.5% were between 2030 years of age and 39.2% were 3040 years. eighty - three respondents were married with 80.1% (96) having grade ten or more education (table 1). in the study 62.6% (62) males and 37.3% (37) females formed a total of 99 participants who stated that a physician should use touch during their visits (p - value : 0.03) (table 2) ; however there was no statistical significance in relation to their educational level (p - value : 0.28), religious beliefs (p - value : 0.63), and marital status (p - value : 0.61). about 75.8% of 120 patients said that they can be tapped at their shoulder, 38.3% on their upper back, and 14.2% on their hands ; however 59.1% do not want to be touched on either knee (figure 1). if tapped on their shoulder, 35.8% would take it as a gesture of comfort and 24.1% would take as respect followed by 21.6% as healing and 19.1% as a way to increase mutual understanding (p - value : 0.001) (table 3). the desire to have eye contact was not statistically significant to gender (p - value : 0.08), religion followed, or literacy level. however, 95.8% (115) patients felt comfortable if eye contact was established by the clinician in order to develop the patient - doctor relation (table 2). eighty - six percent of the respondents felt that this is a sign that the doctor is paying attention to their complaints. forty - six percent felt that this was done to gain confidence in communication. according to 54.1%, eye contact should be brief but regular rather than only when patient is talking about symptoms (1.7%). long stares (36.7%) and objects in the surrounding environment, like computers (19.1%) make them uncomfortable during eye contact (table 4). this is a pioneer study from the south asian region to assess patient 's attitudes towards nonverbal communication through touch and eyecontact during consultations. the outcomes from this study should reassure many medical practitioners regarding the use of comforting touch to patients, as 82.5% of respondents wanted sympathetic touch, especially in distressing situations. however, as opposed to more distal touch, this study reveals that touch on the shoulder (75.8%) or upper back (38.3%) is more acceptable, when compared to touching on the hands (14.2%). yet there exists some gender bias, as patients (of both sexes) are more comfortable to touch from a female doctor as compared to that from a male. our results are consistent with study by street and buller, who studied the nonverbal communication in patient - doctor interactions and they also found no statistically significant correlation between the level of education of patients and their perceptions regarding physician 's touch, whereas a study from john hopkins has shown contrasting results that physician 's touch can be dominating or controlling to people, but as our results highlight it is also taken as a gesture of comfort or respect and as healing, which is in agreement with osmun.. a good physician begins to care for the patient as soon as he / she looks at him. in this present study, 86.1% of the patients eagerly wanted the doctor 's attention through his / her eye contact as pointed out by marcinowicz.. even a simple gesture of frowning can have a positive impact on the patient 's satisfaction. we also found that in the opinion of 54.1% participants, a regular but brief visual interaction is more effective, rather than long durations of eye contact. the use of the computer for keeping electronic records has been a hindrance in developing a good patient - doctor relation, as in this study 19.1% people are distracted from objects in the clinical environment and feel that less attention is being directed to them. this has been acknowledged by many general practitioners (gps) as there was a drastic decrease (from 2001 to 2008) in the use of computer by gps during consultancy. observing the patient together with listening and informative responses makes a medical interview more patient centered and thus results in better therapeutic outcome. this research is a first step in exploring the importance of nonverbal communications among physicians in the south asian region, but it does have many limitations. first, the study was conducted at a single, privately governed hospital and needs a larger sample size to generalize the results to all private or to public sector hospitals. secondly, only two nonverbal modalities have been evaluated in this study, rather than exploring all nonverbal means of communication. fourth, online literature search revealed a study by stepanikova. who concluded that racial backgrounds do play a role in influencing nonverbal communication and this aspect is not covered in the present study. positive, effective, and sensitive nonverbal behavior helps to strengthen the doctor - patient bond. this study does require further clarification and elaborations, but the results do demonstrate the importance of touch and eye contact during the physician 's consultancy. patients do require, from their doctors, a comforting touch on shoulder and regular but brief eye contacts to demonstrate his / her attention towards the patients. we believe further research on this important subject, which could be multicentric, should be further explored, with larger sample populations and covering all aspects of nonverbal communication. moreover, researches investigating physician 's perspective regarding nonverbal means of communication during consultancy are warranted to enhance the importance of this ignored yet eminent topic. annex : questionnaire name (optional):age : gender : male femalereligion : education : illiterate can read and write primary secondary (grade viii) grade 10 higher secondary graduate postgraduatemarital status : married unmarriedoccupation : private job government job self - employed retired the following questions are designed to assess the preferences for touch and eye contact during consultations. you may decide not to participate or if you decide to participate, you are free to withdraw from the study at any time. results of the survey will only be reported in the aggregates ; individuals will not be identified in any way in the reports. your data will be coded under a random identifier that can not be linked to you. signature : date : (1) do you want physical touching of your body by doctor during consultation ? (a) yes (b) no (c) do n't know(2) do you approve physical touching of your body by doctor during consultation ? (a) yes (b) no (c) do n't know(3) do you feel comfortable with physical touching of your body by doctor during consultation ? (a) yes (b) no (c) do n't know(4) you would take a touch from a doctor during consultation as a gesture of ? (a) empathy (b) cure (c) respect (d) comfort (e) communication with doctor (f) other (5) which part of your body you would comfortable with ? (a) hand (b) head (c) shoulder (d) knee (e) upper back (f) other (6) which part of your body you will not be comfortable with ? (a) abdomen (b) thigh (c) others (7) do you feel comfortable with his / her eye contact ? (a) yes (b) no (c) do n't know(8) for how long should an eye contact be ? (a) should be throughout consultation (b) should be brief but regular (c) should be more at beginning less in the end (d) should be more at endless in the beginning (e) others (9) what do you feel when a doctor makes an eye contact ? (a) confidence in yourself (b) confidence in communication (c) secure (d) attended (e) much more important (f) other (10) what are the things which would make you uncomfortable during eye contact ? (a) long stare (b) smiling eyes (c) frequent blinking (d) others(11) what do you feel if a doctor does not make an eye contact ? (a) religious (b) less confident (c) short attention span (d) liar (e) lack of interest | background. nonverbal behaviors have a significant impact on patients during consultations. this study was undertaken to find out the attitudes and preferences of the patients regarding nonverbal communication during consultations with physicians, in a tertiary care hospital. methods. a questionnaire based cross - sectional study was carried out at the aga khan university hospital, karachi, pakistan, during the months of january to march 2012. all patients (> 18 years of age) coming for consultancy in the family medicine clinics were approached ; out of 133, 120 agreed to participate. the subjects were asked questions regarding physician 's comforting touch and eye contact and their responses were noted. the data were analyzed using spss and chi - square test was used to identify corelations. results. overall, 120 patients were enrolled. about 58.3% were men and 41.7% were women with a mean age of 34.9 10.9 years. 95.8% were muslims and 57.6% had more than 12 years of education. among females 74% wanted supportive touch from doctors, used to comfort the patient (45%) or to show respect (27.5%) or as healing (30%). 86.1% of the respondents believe that establishing eye contact with the patient shows that the doctor is attentive towards his / her patient. the eye contact should be brief but regular (54.1%) and prolonged staring (36.7%) makes them uncomfortable. conclusion. nonverbal communication helps to strengthen the doctor - patient relation as patients do appreciate positive touch and eye contact from their physicians. |
ischemic colitis usually develops due to arteriosclerosis, diabetes, hypovolemic shock, portal hypertension, malignant tumors, pancreatitis, or colonic obstruction. it is very common in elderly people.1 phlebosclerotic colitis (pc) is a rare form of ischemic colitis that develops as a result of mesenteric venous calcification and calcification in the invaded colonic submucosal layer.23 pc is usually observed in the colons of elderly men.4 a young woman without any underlying diseases visited our hospital owing to abdominal pain. colonoscopy revealed edematous dark - bluish colonic mucosa, a sclerotic colon wall, and multiple ulcers without clear boundaries from the ascending colon to the transverse colon. the edematous dark - bluish colonic mucosa and sclerotic colon wall were observed only in the sigmoid colon. abdominopelvic computed tomography (ct) revealed linear calcification on the colon wall from the terminal ileum to the transverse colon. arterial phase superior mesenteric angiography revealed an irregular and convoluted marginal artery and vasa recta., we report a case of pc that showed symptomatic improvement with conservative treatment only. a 36-year - old woman visited our hospital with a major complaint of acute pain in her lower right abdomen. she had no history of tobacco, alcohol, or herbal medicine use, underlying diseases such as hypertension or diabetes, and surgery. her vital signs were as follows : blood pressure, 100/70 mm hg ; pulse, 98 beats per minute ; respiratory rate, 18 breaths per minute ; and body temperature, 36.6. physical examination revealed mild oppressive pain and rebound tenderness in the lower right abdominal area. peripheral blood examination yielded the following results : white blood cell count, 16,700/mm hemoglobin level, 14.0 g / dl ; platelet count 340,000/mm ; and neutrophil proportion, 91.1%. biochemical testing revealed the following levels : aspartate aminotransferase / alanine aminotransferase, 21/14 iu / l ; total protein / albumin, 7.4/4.5 g / dl ; total / direct bilirubin, 0.29/0.28 mg / dl ; alkaline phosphatase/-glutamyl transpeptidase, 125/18 iu / l ; amylase / lipase, 97/86 iu / l ; and blood urea nitrogen / creatinine, 13.3/0.84 mg / dl. normal findings were also obtained for rheumatoid factors : antinuclear antibody (-), anti - neutrophil cytoplasmic antibody (-), antiphospholipid immunoglobulin m (igm) (-), and antiphospholipid igg (-). plain abdominal x - ray revealed a paralytic ileus and linear calcification along the colon wall in the lower right abdominal area (fig. abdominopelvic ct revealed that the colon wall from the terminal ileum to the transverse colon was thickened, and multiple calcifications were observed on the colon wall and in the ramifications of the colonic blood vessels (fig. 2). abdominopelvic ct also revealed that the portal vein size was normal in size, implying that portal pressure was within the normal range (fig. 2). colonoscopy showed edematous dark - bluish colonic mucosa, a sclerotic colon wall, and multiple ulcers without clear boundaries from the ascending colon to the transverse colon. the edematous dark - bluish colonic mucosa and sclerotic colon wall was seen only in the sigmoid colon (fig. superior mesenteric angiography arterial - phase images showed an irregular and convoluted marginal artery and vasa recta, and delay - phase images showed convoluted and expanded veins along the vasa recta (fig. 4). a biopsy conducted during colonoscopy revealed fibrosis in the blood vessel walls and lamina propria mucosae, but fibrosis in the submucosal layer, we made a diagnosis of pc, after which conservative treatment including fasting and fluid therapy was initiated. these conservative treatments were continued given the absence of any findings of inflammation along the entirety of the colon wall and the presence of accompanying peritonitis caused by necrosis and perforation. the patient 's abdominal pain was alleviated ; she no longer complained of symptoms even after starting a solid food diet, and was discharged 10 days after admission. she underwent follow - up colonoscopy 10 days after discharge, and slightly improved ulcerative findings were obtained (fig. endoscopic mucosal resection was performed to obtain submucosal tissues, but insufficient samples could be collected due to severe sclerosis in the colonic mucosa. pc is a rare type of ischemic colitis, an on - thrombotic venous colitis characterized by colonic and mesenteric venous calcification and fibrosis.1 the pathogenesis of pc is unclear, although calcinosis, raynaud phenomenon, esophageal dysmotility, sclerodactyly, and telangiectasia syndrome ; toxic chemicals ; and pressure increases in the colon lumen, which may be hereditary, have been reported ; as none of these were observed in this patient, they are not of clinical relevance.356789 kusanagi.2 suggested that secondary portal hypertension caused by liver cirrhosis or chronic liver disease may represent the etiology of pc. meanwhile, yao.1 reported two cases of pc accompanied by normal portal vein pressure, implying that no clear mechanism has yet been established. the six cases of pc reported in south korea to date included three men with underlying diseases such as renal failure, liver cirrhosis, diabetes, and hypertension (table 1). in women, there was no underlying disease in two patients, while a history of herbal medication was observed in one patient. the average age of the six patients was 65.8 years, and all patients were aged from 60 to 70 years. in south korea, there was no common cause considering the associated diseases (table 1). in the present case, a young woman had a medical history of treatment for rhinitis of unknown origin and dark blue pigmentation around her eyes for 5 years. we suspected a relationship between pc and the pigmentation around her eyes, even though it is not clear whether pc invades any organs other than the mesenteric vein. pc usually involves symptoms such as acute abdominal pain, diarrhea, bloody stool, and vomiting, but sometimes shows no symptoms. pc is found in the appendix or ascending colon by accident during colonoscopy, and appears as thread - like calcifications on plain abdominal radiography and venous congestion and hyperplasia in the colon wall, calcification in the colon wall, and mesenteric veins on abdominopelvic ct scans.145671011121314151617 among the six pc cases reported in south korea, three involved ascending to transverse colon wall calcification, one involved ileum to descending colon wall calcification, and two involved whole colonic wall calcification detected by abdominopelvic ct scans.356789 among the four reported cases studied by superior mesenteric angiography, three showed a convoluted marginal artery on arterial - phase images, and one showed expanded and convoluted veins on delay - phase images.131013 in the present case, an irregular and convoluted marginal artery and vasa recta were observed on arterial - phase images, and convoluted and expanded veins were observed along the vasa recta (fig. colonoscopy often reveals bluish violet or dark - brownish mucosa invading the colon, cylindrical ulcers, and loss of the fissures of the semilunar colon fold.456711121314 in the tissues obtained during surgery, findings of colonic hyperplasia accompanying sclerosis and edema, and loss of the fissures on the semilunar colon fold are observable. microscopic findings often include mucosal and submucosal necrosis, submucosal blood vessel calcification, and sclerosis. mucosal atrophy or treatable ulcers may also be seen.141015 pc can be treated with surgery or conservative therapy. markos.10 suggested surgical treatments such as colectomy in cases involving inflammation is present on the whole colon wall in addition to peritonitis caused by necrosis and perforation. however, yu.18 reported a case in which conservative treatments were used, which showed no specific symptoms during the 4-year follow - up period. the six cases of pc reported in south korea included one case in which a right hemicolectomy was performed, and five cases in which the patient was discharged and experienced improvement in symptoms after conservative treatment (table 1). it is necessary to differentiate pc from diseases that can cause chronic mesenteric ischemia, including churg - strauss syndrome, behcet 's disease, lymphocytic phlebitis, degos disease, wegener 's granulomatosis, and polyarteritis nodosa.1920 however, these diseases do not involve fibrosis and calcification in the veins. the etiology and mechanisms of pc are not yet fully understood, and appropriate treatment methods other than colectomy in cases with complications have not yet been established., pc was diagnosed through radiography and colonoscopy in a 36-year - old woman who visited our hospital with a major complaint of pain in her lower right abdomen. here, we report, with a literature review, a rare case of pc that developed in a young woman with no underlying diseases. | phlebosclerotic colitis is a rare disease of intestinal ischemia and differentiating it from the typical ischemic colitis. it is caused by venous obstruction due to colonic and mesenteric venous calcification. we report a 36-year - old woman presenting with intermittent abdominal pain. initial radiologic findings showed multiple tortuous thread - like calcifications in the region of the right side of the colon and transverse colon on plain abdominal radiographs and computed tomography images. in the colonoscopy, edematous dark - bluish colonic mucosa, sclerotic colon wall, and multiple ulcers without clear boundaries were observed from the ascending colon to the transverse colon. in the sigmoid colon only showed the edematous dark - bluish colonic mucosa, sclerotic colon wall. on the basis of these findings, we diagnosed the patient as having phlebosclerotic colitis. we report a rare case of phlebosclerotic colitis in healthy young woman. |
soft tissue sarcomas are rare tumors (1% of all adult malignancies) with a dismal prognosis. approximately 70% of sarcomas occur in the extremities (49% in the lower and 19% in the upper limbs), while the remaining 30% occur in the trunk, abdomen or retroperitoneum (1,2). among the most common histological types of sarcomas of the extremities, malignant fibrous histiocytoma (mfh) is the one exhibiting the highest incidence, followed by liposarcoma, fibrosarcoma and neurofibrosarcoma, with a peak incidence in the sixth decade of life. according to the world health organization 2002 classification of soft tissue tumors (3), mfh may be categorized into three morphological subtypes : i) storiform - pleomorphic mfh (undifferentiated high - grade pleomorphic sarcoma) ; ii) giant - cell mfh (undifferentiated pleomorphic sarcoma with giant cells) ; and iii) inflammatory mfh (undifferentiated pleomorphic sarcoma with prominent inflammation) (3). mfh is also considered by the american joint committee on cancer to be a high - grade sarcoma by definition and, therefore, exhibiting high local aggressiveness. the typical histological appearance of mfh is that of a hypercellular neoplasm, with marked nuclear and cytological pleomorphism, bizarre appearance of stromal cells and a fascicular or storiform model growth (4). the presence of a lesion suspicious for soft tissue sarcoma should prompt the surgeon to perform diagnostic - instrumental tests. x - ray may reveal ectopic calcifications in the surrounding tissues (520%), sclerosis or clear erosion of adjacent bone due to tumor compression. magnetic resonance imaging (mri) may reveal a lobulated mass, with intermediate signal in both t1 and t2 images (5,6). there are also frequent intracapsular areas of tumor necrosis and hemorrhage, which are occasionally so extensive that the tumor is mistakenly diagnosed as hematoma of the soft tissues (7). a correct surgical approach is crucial in several tumors, as previously demonstrated by our studies (8,9), and a survey of the literature suggests that surgery is the first choice for the treatment of mfh (10). owing to advances in surgical techniques, the advanced reconstructive methods and the development of effective adjuvant therapies, the use of widely demolitive surgery is now considered unnecessary. over the last few decades, studies have demonstrated that conservative surgery, with or without adjuvant therapy, appears to be an effective treatment for sarcomas, including high - grade sarcomas, with a local recurrence rate of 715%, with no significant differences in terms of overall survival and disease - free interval compared with amputation (1113). neoplastic infiltration of the resection margins at the end of surgery appears to be among the major factors affectng the rate of local recurrence. in fact, the local recurrence rate approximates 13% with margins of 1 cm. in light of these data, amputation may no longer be considered to be the primary approach for the treatment of sarcomas of the limbs, but rather be reserved for those cases in which adequate resection can not be achieved without severe aesthetic and functional deficits, or when the sarcoma is very proximally located in the limb. the functional compartmental resection represents the surgical evolution of amputation, although not without functional consequences. however, the literature over the last 20 years reveals that soft tissue sarcomas of the extremities and precisely localized within a muscle may be treated by excision of the entire affected muscle, with preservation of the surrounding innervated muscles, without performing total compartmental resection (14). the present study stems from a multidisciplinary management involving a team with expertise in general microsurgery, where microsurgical technique is routinely applied by the use of magnification devices such as loupes 4.5, operating microscope 440, dedicated instruments and led coaxial light. a 79-year - old caucasian man presented with a swelling in the middle third of the dorsal aspect of the left forearm. a core needle biopsy previously performed in a different hospital had not reached a definitive diagnosis. the patient also reported history of trauma 6 months earlier, with progressive volume increase, without pain or functional deficit. on clinical examination, the patient had a tense elastic 5-cm swelling in the dorsal aspect of the left forearm (fig. 1), hypomobile in the deep region, while the overlying skin appeared to be normal. the tinel 's sign and other tests assessing the functionality of the muscles and tendons of the hand and fingers were negative. the patient reported a dull pain elicited by palpation of the region in question and was subjected to instrumental diagnostic examinations. ultrasound imaging revealed a heterogeneous lesion with hyperechoic areas, perilesional hypervascularization and intralesional spots on signal - enhanced color doppler sonography. an mri revealed a neoformation sized ~84 cm within the extensor digitorum communis (edc) muscle, without invasion of the interosseous space (fig. furthermore, the patient was subjected to total - body computed tomography (ct) staging that was negative for lymph node involvement or distant metastases. finally, electromyography (emg) was performed with normal findings, thus excluding any nerve compression or infiltration. as the case history, physical examination and diagnostic tests were strongly suggestive of a tumor, we decided to perform an open biopsy according to the criteria of enneking (15). this procedure was performed under temporary ischemia of the arm, applying a tourniquet for 20 min at 220 mmhg at the root of the arm. the bioptic specimen included skin, subcutaneous tissue, fascia, muscle and a portion of the tumor that appeared to be localized inside the muscle. on histological examination, the neoformation exhibited irregular margins and consisted of spindle cells with nuclear atypia set in a dense, sclerotic, richly vascularized stroma, as evidenced by cd34 staining. on immunohistochemical staining, the tumor was positive for cd68 and negative for smooth muscle actin (sma) and s-100. the proliferative index, as assessed by ki-67, was 40% of the neoplastic elements. thus, the final diagnosis was mfh. based on the histological findings, mri imaging and in accordance with the national comprehensive cancer network (nccn) guidelines (16), we decided to remove the tumor by excising the extensor compartment of the forearm with subsequent tendon transfer. indeed, the guidelines suggest radical surgical treatment followed by radiotherapy for such soft tissue sarcomas. a wide longitudinal incision was performed that also removed the scar of the earlier biopsy (fig. surgery was performed by a microsurgical technique under magnification (loupes 4.5) and the use of dedicated instruments and bipolar coagulator, allowing the identification of the correct anatomical cleavage planes and the preservation of the sensory branches of the radial nerve. according to the mri findings, the tumor was considered to be localized within the edc muscle ; however, the surgical exploration revealed that the lesion was located at a deeper plane, within the abductor pollicis longus muscle (apl). the muscle fibres appeared to be subverted and discontinued, while the apl muscle sheath did not exhibit macroscopic discontinuity or pathological adhesions to the adjacent and overlying muscles ; the underlying bones did not appear to be infiltrated (fig. 3). therefore, we decided to change the surgical strategy and perform a wide excision of the tumor, including the apl muscle, without discontinuing the intact muscle sheath and the tumor pseudocapsule (fig. 4), thus preserving the edc muscle. subsequently, through a palmar mini - incision, we performed a transfer of the palmaris longus (pl) tendon to the distal apl metacarpal tendon insertion with a pulvertaft suture, in order to restore the abduction function of the thumb. the histological examination revealed an encapsulated, whitish neoformation, sized ~94.5 cm, with regular margins. 5) with nuclear atypia set in a dense, sclerotic, richly vascularised stroma, as evidenced by cd34 staining. on immunoistochemical staining, the proliferative index, as assessed by ki-67, was 40% of the neoplastic elements. the mfh was classified as stage iib (pt2b, pn0, pm0, g2 and r0) according to the american joint committee on cancer (7). after 2 weeks, the recovery of the strength and functional ability of the hand and thumb was satisfactory, compared to the contralateral side. after 2 months of rehabilitation, there was no functional deficit in the abduction of the thumb. in accordance with the nccn guidelines (16), we consulted the oncologist and the radiologist, who did not suggest any adjuvant therapy, also taking into consideration the patient 's comorbidities (prostate cancer, heart disease and chronic obstructive pulmonary disease). moreover, the patient was informed of the options of adjuvant radiation therapy vs. observation and he selected observation. the patient has been followed up every 6 months for 4 years with clinical and instrumental examinations and he remains alive and recurrence - free. the diagnosis of soft tissue sarcomas located in the extremities is frequently delayed, as the tumor often grows slowly and is not associated with specific symptoms, apart from tension in the movements of the limb. in the present case, the open biopsy according to the criteria of enneking (15) established the diagnosis of mfh. the biopsy usually represents a strategy and not a small surgical intervention, therefore it is essential to first perform ultrasound, mri or ct examinations to limit the risk of a biopsy procedure. a biopsy performed incorrectly may negatively affect subsequent therapy, increasing the risk of local recurrence and postoperative complications, worsening the functional outcome. according to the mri findings therefore, surgery was planned to excise the tumor along with the edc, which would mean complete detachment of digit tendons and severe functional damage. accurate surgical dissection allowed us to identify the incorrect localization diagnosed by mri, i.e., the sarcoma was unusually and unexpectedly located within the apl muscle. therefore, we performed a wide excision of the sarcoma, including the apl muscle, without discontinuing the intact muscle sheath and the tumor pseudocapsule, preserving the edc muscle. simultaneous pl tendon transfer was performed, to restore the abduction function of the thumb. the surgical excision and reconstruction were performed by the same microsurgical team through the application of a microsurgical technique and the functional outcome was excellent. the follow - up at 4 years did not reveal tumor recurrence or metastatic spread. in conclusion, when the sarcoma is entirely enclosed within a muscle, with free margins and without regional or distant metastases, a wide excision maintaining the muscle sheath intact may be considered as the optimal treatment. | soft tissue sarcomas are rare tumors with a dismal prognosis. among the most common histological types of sarcomas of the extremities, malignant fibrous histiocytoma (mfh) is the one with the highest incidence. surgery is considered to be the first choice of treatment for mfh. to the best of our knowledge, this is the first case report in the literature of a patient with mfh within the abductor pollicis longus (apl) muscle. this unusual location was also unexpected by the treating surgeons, as the preoperative magnetic resonance imaging localized the tumor inside a different muscle. a 79-year - old caucasian man presented with a swelling in the middle third of the dorsal aspect of the left forearm. mfh was diagnosed following biopsy and instrumental diagnostic examinations. surgical excision and simultaneous reconstruction was performed by the same microsurgical team, achieving an excellent functional outcome. the present case highlights the significance of microsurgical approach for improving strategic planning in oncologic surgery. accurate surgical dissection, performed by a team of microsurgeons, allowed for the identification of the unusual and unexpected tumor localization within the apl muscle. for this reason, a change of surgical strategy allowed for preservation of the extensor digitorum communis muscle, which would otherwise have to be resected, with tendon transfer and successful restoration of the thumb abduction function. |
nevus depigmentosus is a hypomelanotic condition which is usually refractory to medical modalities of treatment.a few cases of surgical repigmentation have been reported.one case report from south korea described the recurrence of hypomelanosis at the surgically treated site after eight years. nevus depigmentosus is a hypomelanotic condition which is usually refractory to medical modalities of treatment. one case report from south korea described the recurrence of hypomelanosis at the surgically treated site after eight years. nevus depigmentosus (nd) is a rare, congenital, stable hypomelanosis first described by lesser in 1884. the lesions usually present as dermatomal or quasidermatomal macules commonly on the trunk, lower abdomen, or proximal extremities. they are off - white in colour and have irregular, serrated, feathered, or geographic margins. the face, when involved, is a cause of social embarrassment for the patient. unfortunately, there is no effective treatment for this condition. here we report a patient of nd treated successfully with suction blister grafting, in whom the pigment has been well maintained even after 10 years. a 15-year - old female student presented with a hypopigmented asymptomatic macule on the left half of the upper lip [figure 1 ] slightly extending to the left cheek, since birth. examination of other areas of the skin did not reveal any similar lesions or any other type of skin lesions and the other systems particularly the skeletal and nervous system were within normal limits. her investigation revealed normal blood counts ; liver function test (lft), renal function test (rft), clotting profile, and test for hiv were negative. nevus depigmentosus distributed on the left half of the upper lip with extension to the chin in march 2002, suction blister grafting was performed on her after taking informed consent. multiple blisters were raised on the left thigh using a suction blister technique described by gupta. then, the blister roof from the thigh was removed with the help of iris scissors, transferred on to a glass slide ; the edges were trimmed to fit the shape of the recipient area and then applied on to the recipient nd site [figure 2 ]. when the dressing was removed after 48 hours the patient was put on topical puvasol (psoralen plus ultraviolet - a of solar origin) after the grafts fell off and the resultant pigmentation was satisfactory at a follow - up after 1 month. application of suction blister grafts at the recipient site the patient followed up in the clinic after a gap of 10 years in may 2012, and on examination, it was found that the pigmentation achieved with suction blister grafting was well maintained [figure 3 ], though there were a few areas of patchy hyperpigmentation toward the angle of the lip on the left side. the disease is primarily limited to the skin though there are reports of association of neurological abnormalities and limb hypertrophy the commonly used clinical diagnostic criteria for nd are as follows : leukoderma present at birth or of an early onsetno alteration in the distribution of leukoderma throughout lifeno alteration in texture or change in sensation in the affected areaabsence of hyperpigmented border leukoderma present at birth or of an early onset no alteration in the distribution of leukoderma throughout life no alteration in texture or change in sensation in the affected area absence of hyperpigmented border lee. found that the majority (92.5%) of nd present before the age of three years and have serrated irregular borders (77.4%). wood 's lamp examination shows an off - white accentuation in nd as compared to the chalky white accentuation in the case of vitiligo. described the procedure of suction blister grafting as helpful in nd but the follow - up details in that case were not known. kim. reported a case of recurrence of nd eight years after autologous epidermal grafting where he considered the newly developed hypopigmented macules to be a functional impairment of melanocytes in nd. in the present case, the follow - up was after 10 years and the pigment gained after suction blister grafting was stable. the epidermal sheets used for grafting in nd should completely cover the area of nd for a uniform repigmentation. in contrast, there have been reports of poor quality of repigmentation with cell suspension or melanocyte keratinocyte transplantation. as the resultant cosmetic discomfort for facial lesions of nd can be embarrassing to the patient and no effective therapeutic options are available, surgical repigmentation with blister grafting should be offered to all interested patients. pigmentation was well maintained at the site of suction blister grafting after 10 years, and hence, suction blister grafting should be offered to all willing patients. | nevus depigmentosus is a congenital hypomelanotic condition for which no effective treatments are available. the hypopigmentation is permanent and enlarges in proportion with growth in the person. here, i report a patient of nevus depigmentosus on whom we performed suction blister grafting and the resultant pigmentation was satisfactory even at a follow - up after 10 years. |
in bone, endogenous nucleotides, such as atp and utp, are released from many types of cells, including bone cells [1, 2 ]. nucleotides are released in response to a number of stimuli including inflammation [3, 4 ] and mechanical stimulation [57 ]. they act as autocrine and paracrine signaling molecules by activating purinergic (p2) receptors. bone cells express several types of p2 receptors, allowing them to respond differently to nucleotides, depending on the types of nucleotide present, their concentration, and the duration of exposure [8, 9 ]. the p2x7 receptor subtype is probably one of the most important p2 receptors in the regulation of bone turnover. it is most abundant in cells of haematopoetic origin, including osteoclasts [8, 11 ], but also in osteoblasts that are of mesenchymal origin [1214 ]. several roles of p2x7 have been suggested including atp - induced apoptosis [12, 15, 16 ], maturation of interleukin-1, and its subsequent release. low agonist concentrations lead to low - level activation of p2x7 receptors and might initiate osteoclast formation through activation of pathways initiating production of transcription factors [18, 19 ]. prolonged exposure to high agonist concentrations induces the formation of large pores in the membrane, permeable to hydrophilic molecules as large as 900 da [20, 21 ]. p2x7 receptors are expressed in both osteoclast precursors and active osteoclasts [8, 11, 22 ]. therefore, in addition to activating the apoptotic pathway, the p2x7 receptor could play a role in osteoclast development [23, 24 ] and activation. to further explore the role of the p2x7 receptor in the regulation of the skeletal system, the bone phenotype of two mouse models with ablation of the p2x7 receptor was examined [13, 22, 26 ].. showed reduced total bone mineral content (bmc), decreased periosteal bone formation, and increased bone resorption, which resembles the effects of disuse on the skeleton. in line with this the effects are partially due to increased osteoclast numbers which were further supported by in vitro studies showing that the p2x7 receptor is not necessary for murine osteoclast formation. shows a different bone phenotype with increased cortical thickness of the tibial shaft, but surprisingly no changes in total bmd. the contradicting observations have been attributed to the dissimilar sample sizes, methods of the gene knockout, and different genetic backgrounds of the inbred strains used to generate the mice. solle 's p2x7 mice were generated on 129/ola, c57bl/6 (b6), and dba/2 mixed genetic background but afterwards maintained on the b6xdba/2 background [13, 27 ], hereafter called dbaxb6 p2x7. the p2x7 mice generated by chessell. were maintained on b6 background but originate from a b6/129 hybrid, hereafter called b6 p2x7. interestingly, mice of the dba and b6 background have a naturally occurring mutation in the p2x7 receptor as shown by adriouch.. presence of the mutation impairs the normal function of the receptor ; hek cells transfected with constructs of both genotypes showed that atp - induced pore formation was reduced by 50% in cells carrying the mutated allele (451l). in osteoclasts from mouse strains carrying the 451l allele we found a reduction in pore - forming activity compared to osteoclasts from strains carrying the p451 allele. in murine thymocytes the p451l mutation affects the mechanism of apoptosis acting through the pore formation induced by atp. as the response of the p2x7 receptor to atp is lower in cells harbouring the 451l allele of the p2x7 gene, consequently the observed effects of the p2x7 in the two models may have been underestimated in the published studies. by introducing a different genetic background to a p2x7 model a more pronounced bone phenotype could maybe be found. in support of this, we recently showed that dba/2 and b6 mice have low bmd along with impaired bone strength, which could make it difficult to detect ovariectomy - induced bone loss in mouse models of postmenopausal osteoporosis. therefore, both the b6 and dba are less suitable as mouse models of osteoporosis. of the four inbred strains of mice that have been reported to have the p451 allelic genotype, only 129x1/svj and balb / cj proved ideal for a new p2x7 strain. both strains had high baseline bmd, relatively strong bones and high trabecular bone volume. since the 129x1/svj strain is resistant to cancellous bone loss induced by ovariectomy only the balb / cj strain is suitable as a candidate strain for the generation of a new p2x7 knock - out model. the balb / cj mice are characterized by having high bmd, high tb.th, and trabecular numbers along with strong bones. since the balb / cj mice clearly respond to ovariectomy with bone loss, it is well suited for studying the bone specific effect of p2x7 knock - out. therefore, the overall aim of this study was to generate a new p2x7 strain with a genetic background not harbouring the p451l mutation subsequent to characterizing the balb / cj p2x7 mice and comparing the bone phenotypes of this model with that of the b6 p2x7. female mice of the b6 p2x7 strain were kindly provided by glaxosmithkline and crossed into the balb / cj inbred strain from jackson laboratories (bar harbor, me) for five generations. founders were selected by pcr for the knockout of exon of the p2x7 gene. since the mice generated were only insipient congenic with the balb / cj genome (~97%), wild type littermates from heterozygote breeding couples were used as control groups in both strains, instead of inbred b6 or balb / cj. requests for balb / cj p2x7 animals should be addressed to niklas rye jrgensen ([email protected]). the authors do not currently have access to b6 p2x7 animals, so requests for these should be directed to glaxosmithkline. all animal procedures were approved by the danish animal welfare council prior to initiating the experiments (protocol : 2002/561634 and 2006/5611209). female mice (n = 15 mice per strain) were kept at a 12 : 12 hour light / dark cycle at 20 0.7c, fed the purina mouse diet formula 5k52 (purina, st. ten and two days before sacrifice, animals were injected with 25 mg / kg tetracycline i.p. or 20 mg / kg calcein i.p.. at the age of 120 days, they were starved overnight (to minimize biological variation in bone markers) and euthanized by co2. blood was collected into 2 ml syringes by cardiac puncture, and serum was collected for later measurements of bone markers. p2x7 animals of both strains (b6 and the balbc / j) were genotyped by pcr using the protocol outlined in the following. earpieces or tail parts were used for dna isolation with the qiaamp dna blood mini kit (qiagen), which was used as template in pcr reactions. the primers used were forward primer (p2x7-gt1) : 5 ggg gtg gtg aag aaa tga a 3, reverse primer (p2x7-gt2) : 5 gga tgt gct gca agg cga tt 3, reverse primer (p2x7-gt3) : 5 cca ctt gac ggt gcc ata 3. the p2x7-gt1 and p2x7-gt2 primers amplify a 2473 bp product for the p2x7 mouse, while the p2x7-gt1 and p2x7-gt3 primers amplify a 2447 bp product for the wild type mouse. because of the similarity of the two bands the two primer pairs were used in two different pcr reactions. the samples were amplified using a pcr cycler (mj research inc.) with the following program. after preheating at 95c for 15 min, 35 cycles were run, with denaturation at 94c, annealing at 56c for 1 min, extension at 72c for 1 min before final extension for 10 min at 72c. pcr products were loaded on a 1% agarose gel (ibi agarose, shelton scientific inc.) for electrophoresis. the amplicons were visualized under uv light using the genegenius gel imaging system from syngene. to investigate possible differences in bone formation markers between the genotypes, osteocalcin was measured in serum samples in duplicate using the mouse osteocalcin ria reagents from biomedical technologies, inc. bone resorption as expressed by fragments of type i collagen (ctx) in mouse serum was measured in duplicate using the ratlaps elisa assay (c - telopeptide collagen type i fragment assay) developed by nordic bioscience diagnostics (herlev, denmark) and following the procedure supplied with the kit. alkaline phosphatase activity was measured in duplicate in mouse serum using the alkaline phosphatase reagent kit (sigma). the kit measures total alkaline phosphatase activity and does not distinguish the different organ - specific subtypes. alkaline phosphatase activity was measured directly on the serum in the multiwell plates, using a slight modification of the standard clinical chemistry procedure. serum replicas were diluted with alkaline buffer solution, and substrate solution was added to each well, and the plate was incubated at 37c for 30 min. interassay cv was 5.9% and intra - assay cv was 2.4%. on the day of sacrifice the mouse femurs were collected, cleaned for soft tissue, wrapped in saline moistened gauze in a tube, and frozen at 20c for later ex vivo biomechanical measurements, as described earlier. the strength of the femoral diaphysis was determined by a 3-point bending test on a lloyd instruments compression device (lr50k, lloyd instruments, fareham, uk), after rehydration in a saline solution at room temperature. load - deformation curves were generated, and maximal load was recorded at a speed of 2 mm per minute with a 100 n load cell. to investigate histologic and morphometric changes in the p2x7 models histomorphometric analyses were performed. in short after methyl - methacrylate embedding, bones were sectioned into 7 m thick slices on a polycut e (heavy - duty microtome) and mounted on slides. five subsequent slides were stained with goldner - trichrome for determination of bone volume in percentage of total volume (bv / tv, %), cortical thickness (c.th, m), trabecular thickness (tb.th, m), and eroded surface as percentage of bone surface (es / bs). five slides from each bone were left unstained for quantification of mineralizing surfaces as percentage of bone surface (ms / bs, %) under fluorescent light. the previously mentioned indices were determined using an olympus bx51 microscope equipped with a c.a.s.t.- grid system and reported according to standard bone histomorphometry nomenclature. simple descriptives were presented as means standard error of the mean (sem). the two types of knockout animals were compared to the respective wild type animals by student 's t - test. the new p2x7 strain was made by backcrossing the b6 p2x7 mice generated by chessell. into the balb / cj inbred strain for five generations, there were no significant differences between the wild type (p2x7) mice of the two strains used for the knockout in the majority of assessed bone parameters (table 1), but the bmd, femoral strength, and concentration of bone markers were significantly lower in the b6 strain (table 1, figures 1(b) and 1(c)) than in the balb / cj. furthermore, bone resorption (levels of s - ctx and es / bs%) were lower in the wild type b6 compared to the wild type balb / cj strain (tables 1 and 2 and figure 1(d)), and the bone formation markers alkaline phosphatase and osteocalcin were decreased in the b6 (table 1). there were no significant differences in the basic characteristics such as weight, length, and body composition between the two genotypes of the b6 p2x7 animals (data not shown). the markers of bone resorption and bone formation did not alter significantly when p2x7 was ablated in the b6 mice (table 2). the increase in whole body bmd (4.5%, p = 0.011) in b6 p2x7 compared to the p2x7 animals (figure 1(a)) was accompanied by significant increased bone strength in b6 p2x7, 18.22 n compared to 16.29 n in b6 p2x7 (p = 0.018). the histomorphometric analysis of bone indices in the vertebrae showed only significant increase in tb.th in b6 p2x7 mice (p = 0.011) compared to b6 p2x7 (table 2). histomorphometric analysis of the proximal tibia revealed that in resemblance with the vertebrae tb.th was increased (32.3 m to 35.5 m) however not significant in this region (p = 0.119). as seen on figure 2, the thickness of the tibial cortex was increased to 162.6 m in b6 p2x7 compared to 127.4 m in b6 p2x7 (p < 0.001). when the histomorphometric indices of b6 p2x7 were compared to b6 p2x7, the changes were of different amplitude and direction in the two regions, tibia and vertebrae, as presented in table 2. first, we analyzed the data concerning the basic characteristics such as weight, length, and body composition (lean versus fat tissue). no significant difference between balb / cj p2x7 mice and wild type littermates was found (table 1). the total bmd in balb / cj p2x7 assessed by dxa, was higher compared to wild type animals (figure 1(a), table 1) ; however here the difference between genotypes was higher in the balb / cj strain than in the b6 strain. also differences in bmc were detected, with the higher value in the knock - outs (0.470 g versus 0.558 g, p < 0.001), as well as in bone area, again assessed by dxa analysis, from 9.19 cm to 9.90 cm (p < 0.001). bone mineral density of the femur was increased by 8.2% (p < 0.001, figure 1(b)), and bone strength was improved as much as 37.9% in balb / cj p2x7 animals (p < balb / cj p2x7 mice have low levels of s - ctx, as seen in figure 1(d) and table 1 ; interestingly, it is markedly reduced in balb / cj mice (8.76 ng telopeptide / ml serum) compared to wild type animals (19.09 ng telopeptide / ml serum) (p < 0.001). the non - bone - specific alkaline phosphatase decreased significantly from 314.2 nmol / ml to 270.6 nmol / ml in balb / cj p2x7 mice (p = 0.028). there were no significant changes in concentration of the formation markers osteocalcin (p = 0.101, table 1). no significant differences for the balb / cj strain in any of the bone histomorphometric indices analyzed were detected (table 1). the role of p2x7 in the regulation of bone turnover has already been established over the last decade. extracellular nucleotides have been shown to be involved in calcium signalling and thus intercellular communication among osteoblasts. nevertheless, the reported effects of the relatively specific p2x7 agonist bzatp on the activity of bone cells in vitro have been conflicting [12, 18, 19 ]. in the current study, we have shown that the p2x7 receptor has a distinct role in the regulation of bone mass and turnover, as bmd was increased in p2x7 mice from two strains with different genetic backgrounds. the mechanisms underlying the rise in bmd are presumably multiple but could be related to the observed changes in bone formation and resorption markers, even though no difference was seen in the histomorphometric analyses. the new p2x7 on balb / cj background (balb p2x7) showed significantly increased bmd, bmc, and bone area determined by dxa compared to the wild type genotype. bmd / bmc was also increased in the femoral region, which was accompanied by an increase in bone strength of the femoral diaphysis. the difference could be caused by reduced resorption compared to bone formation, since we found big reduction in s - ctx and a significant decrease in the bone formation marker alkaline phosphatase. morphological changes in other compartments could also explain the observed results, but these have not been investigated in this study. almost the same picture was present when the b6 p2x7 mice were examined though the effect sizes were smaller. when histomorphometric analysis of the vertebrae was compared to the tibia, site - specific differences were detected in the resorption index es / bs%, but even the effect was in the opposite direction ; it was insignificant at both sites. the fact that we observe the same effect of p2x7 ablation, but with a smaller effect size in the strain carrying the 451l allele, confirms the role of the p2x7 receptor in regulation of bone mass. serum ctx levels express the total bone resorbing activity of the organism, and interestingly we found striking differences in s - ctx between the two strains. in balb / cj, the wild type animals have a significantly higher s - ctx level than both the corresponding knock - out and the b6 wild type (figure 1(d)). in contrast, the b6 wild type and the b6 p2x7 s - ctx levels are not different, suggesting a distinct role of the cytoplasmic tail of the p2x7 receptor in osteoclastic bone resorption. in a former study we found a reduction in pore formation in osteoclasts from the b6 strains compared to osteoclasts from the balb / cj, but no difference in resorptive activity of cultures not stimulated with nucleotides. even though there are multiple genetic differences between the two strains, the major finding in this study is that the genetic background is of significant importance when determining the effect of the p2x7 ablation. as seen in figure 2, our study confirms the results of the previous study where gartland since they were not able to detect any difference in the formation and number of osteoclasts, they suggested that the apoptosis of the osteoblasts was affected. they also observed an increased number of osteocytes in cortex, suggesting that instead of undergoing apoptosis the osteoblasts were incorporated in matrix. that could be a plausible explanation for the differences found in b6 p2x7, but it can not explain the decrease in s - ctx in the balb p2x7. in the study by ke., they found an increased number of osteoclasts in pfizers dba / b6 p2x7 tibiae but could not detect any difference in osteoclastogenesis in vitro. also reported that there was no significant difference in the development of osteoclasts between b6 p2x7 and b6 wt in vitro. the question is if the unnatural culture conditions, for example, the lack of systemic hormones, accessory cells, and mechanical stimulation, could explain the lack of effect of p2x7 ablation upon osteoclasts when assessed in vitro compared to the differences seen in vivo / ex vivo. finally, the recently described p2x7-k splice variant suggested to escape knockout in the glaxo mice could be a contributing factor to the differences between the glaxo and pfizer models. however, even though normal osteoclasts express p2x7, none could be detected in osteoclasts derived from balb / cj - p2x7 mice, indicating that the p2x7-k expression splice variant was undetectable on the protein level. however, even if there are some conflicting results from the two different models of p2x7 knock - out in mice, they both point towards an important role of the receptor in regulation of bone formation. it emphasizes that even though mice are widely used as models to study the regulation of bone mass and turnover, there are important differences between murine and human bone physiology. human studies are highly warranted to investigate the role of the p2x7 receptor in human bone turnover and in conditions of altered bone metabolism. the two background strains on which the b6xdba p2x7 from pfizer and b6 p2x7 from glaxo mice were based carry the 451l allele of the naturally occurring mutation in p2x7. the contradictory effects on bone status between the two strains could be due to a number of factors, including differences in experimental parameters like sex, age, or diet [13, 22 ], which makes it difficult to compare the experiments with different background. ke. found a larger effect of the knockout in dba / b6 p2x7 males compared to the females. do not inform about the sex of the b6 p2x7 animals in their study. in our study, only females were used. of critical importance is the choice of wild type animals, whether it is wild type siblings from heterozygote breeding couples or purchased from the inbred background strain. another important parameter to consider is the diet, as it is known to influence bone mass and structure. our earlier experiments with the b6 p2x7 mice showed only insignificant differences in s - ctx. by changing the diet from altromin 1430 or 1319 to labdiet 5k52, which contains more calcium (0.8% to 1.15%), more d - vitamin (1 iu to 4 iu), and less fat (7.5% to 4%) it showed significant effects on the bmd, also in female mice. in this study the balb and b6 p2x7 mice were fed the same diet and had similar age and sex, so the only difference is the genetic background. have shown that the genetic background is extremely important for the bone status in the different strains. in this study balb / cj mice had higher bone turnover (indicated by levels of s - ctx and alkaline phosphatase) than b6. therefore it is possible that it is easier to detect the decrease in resorption in the p2x7 on the balb / cj background. similar strain - specific differences are seen in other knockout mice, like diverse effects of treatment are seen in different strains. in conclusion, we have shown that the p2x7 receptor plays an important role in the control of bone remodelling. furthermore, absence of p2x7 receptor expression seems to be associated with increased bone mass and strength in two mouse models. the effect of p2x7 ablation was underestimated in the model based on the b6 strain carrying the naturally occurring mutation, since the difference between the knock - outs and their wild type littermates is higher in the balb / c model than in the b6 model. further studies are warranted in order to understand the complex roles of p2x7 in bone turnover, where especially human studies are important in order to fully understand the role of the receptor in human bone physiology. | the purinergic p2x7 receptor is expressed by bone cells and has been shown to be important in both bone formation and bone resorption. in this study we investigated the importance of the genetic background of the mouse strains on which the p2x7 knock - out models were based by comparing bone status of a new balb / cj p2x7/ strain with a previous one based on the c57bl/6 strain. female four - month - old mice from both strains were dxa scanned on a piximus densitometer ; femurs were collected for bone strength measurements and serum for bone marker analysis. bone - related parameters that were altered only slightly in the b6 p2x7/ became significantly altered in the balb / cj p2x7/ when compared to their wild type littermates. the balb / cj p2x7/ showed reduced levels of serum c - telopeptide fragment (s - ctx), higher bone mineral density, and increased bone strength compared to the wild type littermates. in conclusion, we have shown that the genetic background of p2x7/ mice strongly influences the bone phenotype of the p2x7/ mice and that p2x7 has a more significant regulatory role in bone remodeling than found in previous studies. |
we studied cross - sectionally clinic- and hospital - based adult diabetic patients (aged > 30 years) within 1 year from diagnosis. we ascertained subjects with lada and type 2 diabetes from the lada china multicenter study (46 hospitals throughout china contributed) ; type 1 diabetes, defined by who criteria (4), from the type 1 diabetes collaboration net (18 hospitals in hunan province) ; and normal subjects were screened by 75-g oral glucose tolerance test from 5,000 chinese in a population - based study. definition of lada (57) was 1) gad antibody (gada) positive, 2) age > 30 years at diagnosis, 3) independence from insulin treatment 6 months postdiagnosis, and 4) without ketoacidosis. subjects with hs - crp > 10 mg / l or taking anti - inflammatory drugs within 3 months were excluded. metabolic syndrome was defined by national cholesterol education program - adult treatment panel iii criteria (8). this study was approved by ethics committees at second xiangya hospital of central south university and at each center and conducted according to the declaration of helsinki. height, weight, waist circumference, hip circumference, and blood pressure were recorded locally, and fasting sera samples were processed centrally. elisa determined high - sensitivity il-6 (il-6 hs ; r&d systems, minneapolis, mn), 7% intraassay and 5% interassay coefficient of variation (cv) ; lcn2 (10), 3.86.0% intraassay and 3.15.2% interassay cv ; adiponectin (11) 56% intraassay and 68% interassay cv. hs - crp was measured by immunoturbidometric assay (orine diagnostica) with 2.13.3% intraassay and 3.44.6% interassay cv. we used spss (version 13 ; spss, chicago, il) for statistical analysis. data are expressed when normally distributed as means sd and when skewed as median (25th75th percentiles). we used a general linear model of univariate analysis adjusted for age, sex, and bmi ; log - transformed cytokine value was the dependent variable, different diabetes groups were fixed, and age, sex, and bmi were covariates (fig. multivariate regression models investigated differences in log - transformed cytokine concentrations (dependent variables), with increasing number of variables (model 1 : unadjusted ; model 2 : sex and age adjusted ; model 3 : age, sex, and bmi [independent variables ] adjusted) (supplementary table 2). linear relationships were evaluated by partial correlations test adjusted age, sex, and bmi. data were not corrected for multiple comparisons and are descriptive ; p < 0.01 was considered significant. univariate of general linear model adjusted according to age, sex, and bmi between groups. p < 0.05, p < 0.01, p < 0.001. ngt, normal glucose tolerance subjects ; t1d, type 1 diabetic subjects ; t2d, type 2 diabetic subjects. we used spss (version 13 ; spss, chicago, il) for statistical analysis. data are expressed when normally distributed as means sd and when skewed as median (25th75th percentiles). we used a general linear model of univariate analysis adjusted for age, sex, and bmi ; log - transformed cytokine value was the dependent variable, different diabetes groups were fixed, and age, sex, and bmi were covariates (fig. multivariate regression models investigated differences in log - transformed cytokine concentrations (dependent variables), with increasing number of variables (model 1 : unadjusted ; model 2 : sex and age adjusted ; model 3 : age, sex, and bmi [independent variables ] adjusted) (supplementary table 2). linear relationships were evaluated by partial correlations test adjusted age, sex, and bmi. data were not corrected for multiple comparisons and are descriptive ; p < 0.01 was considered significant. univariate of general linear model adjusted according to age, sex, and bmi between groups. p < 0.05, p < 0.01, p < 0.001. ngt, normal glucose tolerance subjects ; t1d, type 1 diabetic subjects ; t2d, type 2 diabetic subjects. of 784 subjects, patients with both lada and type 2 diabetes were significantly older than subjects with normal glucose tolerance or type 1 diabetic patients. as expected, lada patients had lower insulin secretion than type 2 diabetic patients, while in type 1 diabetic patients, insulin secretion was lower than in both lada and type 2 diabetic patients. metabolic syndrome in type 1 diabetic and control subjects was less frequent than in both lada and type 2 diabetic subjects. we used univariate and multivariate regression models to adjust age, sex, and bmi differences (fig. 1 and supplementary table 2). after adjustment, all three types of diabetes (type 1 diabetes, lada, and type 2 diabetes) showed increased il-6 and lcn2. all four cytokines were increased in lada. in type 1 diabetes, adiponectin but not hs - crp was increased. in type 2 diabetes, hs - crp but not adiponectin was increased, and adiponectin was decreased after adjustment. gada titer was positively correlated with adiponectin (r = 0.14, p < 0.01) and negatively correlated with hs - crp (r = 0.15, p < 0.001) (supplementary tables 3 and 4). in lada subjects, hs - crp was strongly related to bmi (r = 0.363, p < 0.001). some inflammatory markers were increased in diabetes (i.e., il-6 and lcn2) irrespective of diabetes type, implying that cytokine changes are secondary features of the disease independent of obesity, and consistent with a proinflammatory effect in diabetes in general. nevertheless, obesity (waist - to - hip ratio) was associated with all four cytokines assayed, and the pattern of cytokine changes differed according to diabetes type. cytokines (il-6, tnf, and il-1 receptor antagonist) in europeans with these same major diabetes types were also positively associated with bmi, which was higher in diabetic patients than in control subjects and highest in type 2 diabetes but similar in lada and type 1 diabetes ; after correction, only tnf in lada and type 1 diabetes was no longer different from that in control subjects (5). in both the european study and this chinese study, il-6 showed remarkably similar increases in major diabetes types ; il-6, as well as lcn2, levels were higher than in control subjects and highest in type 2 diabetes (5). however, hs - crp was increased in both lada and type 2 diabetes, but not type 1 diabetes, while adiponectin was only increased in lada and type 1 diabetes. despite adiponectin being positively correlated with gada titer, it has no established role in autoimmune diabetes and certainly can not predict it (12). however, low - grade inflammation could be important in autoimmunity, potentially explaining any benefit of rosiglitazone (13). indeed, each cytokine (il-6, lcn2, hs - crp, and adiponectin) was increased in chinese lada, though probably for different reasons (14). given the relative lack of obesity in china, comparative chinese and european studies could delineate the relative roles of obesity, diabetes, and autoimmunity on cytokines. surprisingly, therefore, metabolic syndrome was prevalent in this large lada cohort, in contrast to european studies (5,6). these results point toward a complex relationship between diabetes types and altered cytokine levels. | objectiveto determine the relationship between selected cytokines and diabetes in chinese subjects.research design and methodsadult patients with recent - onset type 1 diabetes (n = 53), latent autoimmune diabetes in adults (lada) (n = 250), and type 2 diabetes (n = 285) from multiple centers were compared with normal subjects (n = 196). we centrally tested serum gad antibodies (gadas), interleukin-6 (il-6), lipocalin 2 (lcn2), high - sensitivity c - reactive protein (hs - crp), and adiponectin.resultsafter adjustment for age, sex, and bmi, all diabetes types had increased il-6 and lcn2 (p < 0.01), and all four cytokines were increased in lada (p < 0.01). in type 1 diabetes, adiponectin but not hs - crp was increased (p < 0.01), whereas in type 2 diabetes, hs - crp but not adiponectin was increased (p < 0.01). adiponectin was correlated positively with gada titer and negatively with hs - crp (p < 0.01 for both).conclusionsin china, inflammatory markers are increased in all three major types of diabetes, but probably for different reasons, even in autoimmune diabetes. |
azathioprine is a prodrug of the purine mimic antimetabolite, 6-mercaptopurine, widely used in various autoimmune diseases and organ transplantation. little is known about the pathogenesis and epidemiology of azathioprine - induced cholestatic hepatitis, a rare but dramatic iatrogenic complication. a 73-year - old man presented to the emergency department at a tertiary center in south india with fever, vomiting, pruritus, and dark urine for seven days. before six weeks graft rejection was subsequently diagnosed and 15 days prior to the presentation, he was started on azathioprine 50 mg and prednisone 40 mg once daily. past medical history revealed chronic bronchitis, essential hypertension, dyslipidemia, and left ventricular diastolic dysfunction for two years. the patient was on aspirin, atorvastatin, losartan, hydrochlorothiazide, and inhaled salmeterol plus fluticasone, ipratropium and salbutamol medications for the past two years with no history of adverse drug events or allergies. examination revealed temperature 100.4f, pulse rate 100/min, blood pressure 106/70 mm hg, and respiratory rate 20/min. cholangitis, sepsis with cholestasis, acute hepatitis (of viral, drug or autoimmune aetiology), and systemic febrile illnesses like malaria, leptospirosis, and scrub typhus were considered as differential diagnosis. investigations revealed conjugated hyperbilirubinemia, elevated transaminases and alkaline phosphatase, prolonged prothrombin time, and bilirubinuria [table 1 ]. blood smears for plasmodium and complete blood counts were unremarkable except for mild eosinophilia (549 cell/l). blood cultures, hepatitis b surface antigen (hbsag) and serology for hepatitis a, c and e, human immunodeficiency virus, scrub typhus, and leptospirosis were negative. the patient was discharged after 72 hours of intravenous antibacterials. selected serial hematologic and biochemical parameters thirty - five days later, he again presented with persisting low - grade fever, severe pruritus, and jaundice with pale stools. clinical examination and worsening of hyperbilirubinemia, normalization of transaminases and prothrombin time, and improvement in alkaline phosphatase were observed [table 1 ]. anti - nuclear antibody (ana) and liver kidney microsomal (lkm-1) antibody for autoimmune hepatitis and anti - mitochondrial antibody (ama) for primary biliary cirrhosis were negative. percutaneous liver biopsy showed perivenular hepatocanalicular cholestasis with mild lobular inflammation [figure 1 ]. perivenular canalicular cholestasis : bile plugs within dilated canaliculi (arrows) 400- h&e symptomatic treatment for pruritus with ursodiol and hydroxyzine were started from day 35. an ophthalmology consultation was reassuring ; topical prednisone for the operated eye was started (the patient had stopped prednisone after the first visit). by day 42, follow - up at 4.5 and 7.5 months showed that the patient was asymptomatic with normal liver function tests (lfts), except for alkaline phosphatase which showed a declining trend. hepatotoxicity due to azathioprine encompasses a spectrum of syndromes including predominant cholestasis, hypersensitivity (characterized by hepatocellular injury) and peliosis hepatis (reflecting endothelial cell injury). hepatic idiosyncratic drug reactions are believed to be due to genetic polymorphisms in drug - metabolizing pathways, or due to immunologic mechanisms. azathioprine - induced cholestatic hepatitis has been reported in various settings including renal and cardiac transplantation, wegener granulomatosis, systemic lupus erythematosus, and inflammatory bowel disease. the latency from the initial exposure to azathioprine to the onset of jaundice ranges from two weeks to three months in previous reports. one author reports shortening of the latent period on rechallenging with azathioprine, characteristic of hypersensitivity reactions. although this was our patient 's first exposure to azathioprine, a short latency of seven days preceded jaundice and fever. wide variation is also noted in the time for clinical and biochemical recovery following drug withdrawal. our patient had peak bilirubin levels seven weeks after starting azathioprine (five weeks after stopping it), and clinical resolution at four months, with gradual subsidence of alkaline phosphatase beyond seven months. previous authors document shorter recovery periods ranging from two weeks to two months with one case that had abnormal alkaline phosphatase and gamma - glutamyl transpeptidase beyond four months after cessation of azathioprine. our patient was not rechallenged because there are no guidelines on rechallenge in azathioprine - related cholestatic hepatitis ; the toxicity was severe, and alternative drugs were available. the exclusion of other causes of hepatitis with cholestasis in our resource - limited setting was governed by clinical relevance. epstein - barr virus has been reported to cause cholestatic hepatitis, but not prolonged cholestasis. the temporal profile of fever concurrent with jaundice and the negative serology argue against viral hepatitis. an elevated serum angiotensin - converting enzyme raised the possibility of sarcoidosis ; this enzyme is also elevated in cholestatic disorders. the histological absence of granulomas and normal serum calcium levels and chest radiographs decreased the likelihood of sarcoidosis. however, the patient had been taking this medication for two years and it was not stopped during the episode. our report highlights a rare hepatic complication of azathioprine that has a wide range of latency, severity and duration, and is probably reversible. recent guidelines on monitoring for azathioprine toxicity require weekly blood counts and lfts until a maintenance dose is achieved, thereafter reducing to a minimum of once every three months. we would also stress the need for patient education and regular clinical assessment. rechallenge following an idiosyncratic hepatic reaction to azathioprine is usually unwarranted in the current era of multiple therapeutic alternatives for immunomodulation. | we report a case of cholestatic hepatitis developed one week after exposure to azathioprine. the subsequent prolonged cholestatic phase was followed by full clinical remission. current knowledge on pathogenesis and epidemiology and the diagnostic challenges presented by this rare complication are discussed, followed by recommendations for monitoring and management. |
nocturia is defined as waking one or more times to void during the period between going to bed with the intention of sleeping and waking with the intention of arising (1, 2). recent multinational population - based, cross - sectional survey (epic) of adults aged 18 yr using 2002 international continence society (ics) definitions showed overall prevalence of any lower urinary tract symptoms (luts) was 62.5% in men and 66.6% in women and nocturia (2 times / night) was the most commonly reported storage symptom for both men (20.9%) and women (24.0%) (3). similarly, the epidemiological data in adult korean aged 40 - 89 yr national community showed that of 2005 subjects interviewed, 33.5% reported voiding once per night and 48.2% twice or more per night and in addition, the impact of nocturia on daily life was reported by 14.6% of subjects (4). nocturia is associated with sleep disruption and consequently impairs the quality of life (qol) and potentially increases mortality (5, 6). although this condition should be evaluated and treated, it has only recently been recognized as a clinical entity in its own right, rather than just one of the features of luts. relatively few patients with nocturia seek medical care because of a lack of awareness or misconceptions about nocturia. furthermore, there is a general lack of knowledge about treatment options for nocturia (4, 7, 8). because the pathophysiology of nocturia is multifactorial and complex, it is important to adopt a systematic approach to identify the possible causal factors of nocturia and to treat them accordingly. patients with nocturia can be categorized as having one of the following three disorders : (1) nocturnal polyuria (np) in which the voided urine volume during the hours of sleep exceeds 33% of the 24-hr output (2), decreased nocturnal bladder capacity (nbc), which results in a nocturnal urinary volume greater than the maximum voided volume (mvv), (3) or mixed nocturia, a combination of the preceding two categories (9, 10). in one retrospective study of 194 patients with nocturia, 13 (7%) had pure np, 111 (57%) had diminished nbc, and 70 (36%) had a combination of the two (9, 11). desmopressin is a synthetic analogue of arginine vasopressin (avp), but desmopressin has a more potent antidiuretic effect than avp. desmopressin has been shown to be an effective treatment for nocturia with np by decreasing night - time urine production (12 - 16). however, a patient with a significantly decreased nocturnal bladder capacity index (nbci) > 2 may have bladder outlet obstruction, detrusor overactivity (do), sensory urgency, and/or primary bladder disorders such as infection, or malignancy (16). specific treatment of the underlying urological condition would therefore be expected to have a mitigating effect on decreased nbc as a component of nocturia. although clinicians usually treat nocturia patients with decreased nbc based on their personal experience and use anticholinergic and hypnotic agents as well as desmopressin, these treatments have not been studied extensively clinically. in the present study, we investigated the safety and efficacy of oral desmopressin for the treatment of mixed nocturia in patients with both np and a decreased nbc. patients aged 18 yr with mixed nocturia (2 voids / night, nocturnal polyuria index [npi ] > 33% and nbci > 1) were enrolled in this open label, prospective, seven - center study. the formula for calculating npi is simply nocturnal urine volume (nuv) divided by 24-hr urine volume, and nbci, more complicated formula that addresses voids at night in patients with decreased nbc, is defined as the difference between the predicted number of nightly voids (pnv) and the actual number of nightly voids (anv). pnv is derived by calculating (nuv / mvv) and subtracting 1 (17). the following exclusion criteria were used : nocturia due to other defined causes of increased urinary frequency, primary polydipsia (> 40 ml / kg/24 h), neurogenic bladder dysfunction, urge incontinence, continued post - voiding residual urine > 150 ml, serum sodium levels 5 hr of undisturbed sleep, and the patient 's overall impression about his / her quality of sleep expressed as feeling fresh in the morning or feeling tired in the morning. clinical and laboratory assessments including body weight, blood and urine analysis, serum sodium monitoring, and adverse event reports were recorded at baseline and after 4 weeks of treatment. the patients with serum sodium levels 1, the decrease in the mean number of nocturnal voids (from 3.0 to 1.7), mean nocturnal diuresis (from 1.5 to 0.9 ml / min), and mean ratio of night/24-hr urine volume (23%) were significantly different between the desmopressin and placebo groups (all p 5 hr of unbroken first sleep period per night was experienced by 27% of desmopressin - treated patients (placebo group : 9%), resulting in a highly significant between - group difference (25). these authors reported that two questions concerning the quality of sleep and tiredness favored desmopressin over the placebo (p=0.02). among the physiological theories associated with nocturnal urine output, elderly people with frequent awakenings have a larger part of their 24-hr urine output at night than those with fewer awakenings (26). the ability to sleep without waking is more indicative of quality sleep than interrupted longer periods of total sleep time during the night. it seems that not only spontaneous sleep, but also less fragmented sleep with undisturbed sleep duration, and deep stages of sleep with a decreased nocturnal voiding frequency improve after desmopressin treatment as a result of the effects of endogenous in vivo adh secretion on nu. however, in another study to investigate the effects of long - term oral desmopressin on baseline adh secretion in elderly patients with nocturia, administration of desmopressin for 1 yr in elderly patients did not affect baseline adh secretion in spite of significantly decreasing nocturnal urine output and the number of nocturia episodes (p<0.01) (27). the practical importance of the increase in endogenous adh with regard to the treatment of np requires further investigation. finally, the safety analysis of desmopressin in the present study confirmed that adverse events associated with treatment are infrequent and usually mild. significant hyponatremia occurred in only one case, and this particular patient was asymptomatic. in addition, other side effects such as body weight gain and electrolyte imbalance including sodium abnormalities and decreased renal function did not occur. in a meta - analysis of the risk of hyponatremia in older adults using desmopressin for treatment of nocturia, hyponatremia was found to be a relatively common adverse event with a frequency of 7.6% (28). however, other studies have suggested that desmopressin has antidiuretic activity for one night or less ; a daytime pause in treatment may therefore be sufficient to maintain water and electrolyte balance (29, 30). in addition, at least a 10-fold higher - thanrecommended dose did not cause changes in the serum sodium concentration. in conclusion, oral desmopressin treatment results in a significant improvement in the number of mixed nocturia episodes by reducing nuv and increasing nbc. this drug improves the quality of sleep by prolonging the period of sleep until the first void, and is shown to have an acceptable safety profile. | to investigate the efficacy and safety of desmopressin in patients with mixed nocturia, patients aged 18 yr with mixed nocturia (2 voids / night and a nocturnal polyuria index [npi ] > 33% and a nocturnal bladder capacity index [nbci ] > 1) were recruited. the optimum dose of oral desmopressin was determined during a 3-week dose - titration period and the determined dose was maintained for 4 weeks. the efficacy was assessed by the frequency - volume charts and the sleep questionnaire. the primary endpoint was the proportion of patients with a 50% or greater reduction in the number of nocturnal voids (nv) compared with baseline. among 103 patients enrolled, 94 (79 men and 15 women) were included in the analysis. the proportion of patients with a 50% or greater reduction in nv was 68 (72%). the mean number of nv decreased significantly (3.20 to 1.34) and the mean nocturnal urine volume, nocturia index, npi, and nbci decreased significantly. the mean duration of sleep until the first nv was prolonged from 118.444.1 to 220.390.7 min (p<0.001). the overall impression of patients about their quality of sleep improved. adverse events occurred in 6 patients, including one asymptomatic hyponatremia. desmopressin is an effective and well - tolerated treatment for mixed nocturia. |
a 38-year - old female from puerto rico with complaints of hypoxia was transferred to our facility. she had been diagnosed 5 years earlier with hermansky - pudlak syndrome (hps) subtype 1, complicated by pulmonary fibrosis. she had experienced a slow deterioration in pulmonary function, manifesting as worsening dyspnea, cough, and increasing oxygen dependence. in addition to pulmonary fibrosis, she had other manifestations of hps, including multiple major bleeding episodes requiring blood transfusions, and oculocutaneous albinism with legal blindness. she required 24 h supplemental home oxygen with a baseline flow requirement of 6 l / min via nasal cannulae and was under evaluation for lung transplantation prior to her terminal hospital admission. pulmonary function tests [table 1 ] showed increasing severe restrictive ventilatory deficit over the 2 years prior to admission, with total lung capacity going from 64% in 2010 to 57% in 2011 and 44% in 2012. pulmonary function test results over a course of 2 years revealing progressive worsening of restrictive ventilatory deficit the patient presented with increasing dyspnea, pleuritic chest pain, and cough. she was transferred to the intensive care unit due to hypoxia, with an oxygen saturation of 44% on 6 l nasal cannula. she was started on empiric antibiotics and supported with noninvasive ventilation, but continued to desaturate and was subsequently intubated. the patient developed multiorgan failure as evidenced by elevated creatinine, transaminitis, and right heart failure. blood and urine cultures on admission were negative, and white blood cell count was normal at 10.9 10 cells / l. chest x - ray upon terminal admission demonstrated diffuse reticular interstitial opacities throughout the lungs [figure 1 ]. baseline ct of the chest obtained 16 months [figure 2a ] and 3 months before admission [figure 2b ] demonstrated progressive worsening of pulmonary fibrosis, as evidenced by increasing traction bronchiectasis, interlobular septal thickening, and consolidative opacities. chest ct upon admission showed multifocal ground glass opacities, subpleural cysts, septal and peribronchovascular thickening, reticulation, and traction bronchiectasis involving greater than two - thirds of the lungs. also, there was new honeycombing in the bilateral upper lobes suggesting end - stage fibrosis [figure 2c ]. 38-year - old female with a known history of hermansky pudlak syndrome complicated by pulmonary fibrosis, admitted with hypoxia. posteroanterior chest radiograph performed on admission demonstrates bilateral, mid to lower lung predominant, diffuse reticular interstitial opacities (arrows). 38-year - old female diagnosed with hermansky pudlak syndrome complicated by pulmonary fibrosis, as evidenced by worsening dyspnea, cough, and increasing oxygen dependence. baseline high - resolution ct (hrct) of the chest : (a) axial image on lung windows at the level of the carina demonstrates scattered, predominantly peripheral, ground glass opacities with associated fine peripheral reticulation in the lungs bilaterally (curved arrows) ; (b) high - resolution ct of the chest from the same level obtained 13 months later demonstrates progression of the peripheral opacities that have become more consolidative (curved arrows) with more conspicuous subpleural opacity in the right upper lobe (straight thin arrow) ; there is new bronchiectasis (arrowhead) and septal thickening along the left main fissure (thick arrow) ; and (c) axial ct of the chest on lung windows at terminal admission, performed as part of a ct pulmonary angiogram protocol (16 months after initial imaging), demonstrates further progression with increased size of peripheral consolidative opacities and increasing bronchiectasis. there is new honeycombing in the upper lobes bilaterally (curved arrow). of note, both lungs were found to be heavy with diffuse nodular parenchymal fibrosis [figure 3a and 3b ]. there was end - stage remodeling, characterized by cystic spaces lined by bronchial epithelium and surrounded by well - developed fibrosis on microscopic examination. the cysts, airspaces, and bronchioles were filled with mucus, suppurative exudate, and abundant vacuolated macrophages [figure 4a and b ]. 38-year - old female with history of hermansky pudlak syndrome and end -stage pulmonary fibrosis, admitted with hypoxia and was terminally extubated after sustaining multiorgan failure. (a and b) photographs of coronal cut sections of a lung demonstrate diffuse nodular parenchymal fibrosis (arrows) with apparent bronchiolocentric distribution. 38-year - old female with history of hermansky pudlak syndrome and end - stage pulmonary fibrosis, admitted with hypoxia. photomicrographs of postmortem microscopic examination of lung tissue stained with hematoxylin eosin stain : (a) 200 magnification demonstrates end - stage remodeling, with honeycomb cystic spaces lined by metaplastic bronchial epithelium and surrounded by well - developed fibrosis (arrows) ; (b) 400 magnification shows alveolar septal thickening associated with prominent diffuse vacuolization of type ii pneumocytes (arrow). chest x - ray upon terminal admission demonstrated diffuse reticular interstitial opacities throughout the lungs [figure 1 ]. baseline ct of the chest obtained 16 months [figure 2a ] and 3 months before admission [figure 2b ] demonstrated progressive worsening of pulmonary fibrosis, as evidenced by increasing traction bronchiectasis, interlobular septal thickening, and consolidative opacities. chest ct upon admission showed multifocal ground glass opacities, subpleural cysts, septal and peribronchovascular thickening, reticulation, and traction bronchiectasis involving greater than two - thirds of the lungs. also, there was new honeycombing in the bilateral upper lobes suggesting end - stage fibrosis [figure 2c ]. 38-year - old female with a known history of hermansky pudlak syndrome complicated by pulmonary fibrosis, admitted with hypoxia. posteroanterior chest radiograph performed on admission demonstrates bilateral, mid to lower lung predominant, diffuse reticular interstitial opacities (arrows). 38-year - old female diagnosed with hermansky pudlak syndrome complicated by pulmonary fibrosis, as evidenced by worsening dyspnea, cough, and increasing oxygen dependence. baseline high - resolution ct (hrct) of the chest : (a) axial image on lung windows at the level of the carina demonstrates scattered, predominantly peripheral, ground glass opacities with associated fine peripheral reticulation in the lungs bilaterally (curved arrows) ; (b) high - resolution ct of the chest from the same level obtained 13 months later demonstrates progression of the peripheral opacities that have become more consolidative (curved arrows) with more conspicuous subpleural opacity in the right upper lobe (straight thin arrow) ; there is new bronchiectasis (arrowhead) and septal thickening along the left main fissure (thick arrow) ; and (c) axial ct of the chest on lung windows at terminal admission, performed as part of a ct pulmonary angiogram protocol (16 months after initial imaging), demonstrates further progression with increased size of peripheral consolidative opacities and increasing bronchiectasis. there is new honeycombing in the upper lobes bilaterally (curved arrow). of note, at postmortem examination, both lungs were found to be heavy with diffuse nodular parenchymal fibrosis [figure 3a and 3b ]. there was end - stage remodeling, characterized by cystic spaces lined by bronchial epithelium and surrounded by well - developed fibrosis on microscopic examination. the cysts, airspaces, and bronchioles were filled with mucus, suppurative exudate, and abundant vacuolated macrophages [figure 4a and b ]. 38-year - old female with history of hermansky pudlak syndrome and end -stage pulmonary fibrosis, admitted with hypoxia and was terminally extubated after sustaining multiorgan failure. (a and b) photographs of coronal cut sections of a lung demonstrate diffuse nodular parenchymal fibrosis (arrows) with apparent bronchiolocentric distribution. 38-year - old female with history of hermansky pudlak syndrome and end - stage pulmonary fibrosis, admitted with hypoxia. eosin stain : (a) 200 magnification demonstrates end - stage remodeling, with honeycomb cystic spaces lined by metaplastic bronchial epithelium and surrounded by well - developed fibrosis (arrows) ; (b) 400 magnification shows alveolar septal thickening associated with prominent diffuse vacuolization of type ii pneumocytes (arrow). hps is a heterogeneous group of autosomal recessive disorders characterized by oculocutaneous hypopigmentation, platelet dysfunction (easy bruising and prolonged bleeding time despite normal platelet count), and lysosomal accumulation of ceroid - lipofuscin in the lungs, which primarily accounts for the associated morbidity. in north america, most patients with hps are from puerto rico, where the prevalence is estimated to be 1 in 1800 or occurring in 5 of every 6 albinos. groups of affected individuals have also been identified in japan and a small village in switzerland. patients usually present in childhood, often with recurrent epistaxis or prolonged bleeding after dental procedures. nine subtypes of hps are described, most of which are associated with a mutation in the hps gene on the long arm of chromosome 10. this genotype leads to a high risk of pulmonary disease, hemorrhage, and, in approximately 15% of patients, granulomatous colitis. only type 4 approaches type 1 in severity, with the remaining subtypes behaving more mildly clinically and with little risk of restrictive lung disease. while most patients with hps1 die of pulmonary complications, approximately 13% die of complications from bowel inflammation, with this granulomatous colitis mimicking crohn 's disease and often occurring in adolescence or early adulthood. the pathophysiology is believed to be impaired intracellular trafficking of melanosomes, platelet dense bodies, and lysosomes. impaired formation of platelet - dense bodies leads to the bleeding dyscrasia, whereas impaired intracellular trafficking of melanin in the melanosomes of the skin and retina is postulated to be the cause of oculocutaneous albinism. systemic complications are associated with accumulation of ceroid - lipofuscin, an amorphous lipid protein complex in multiple organs and the reticuloendothelial system, leading to pulmonary fibrosis, granulomatous colitis, cardiomyopathy, and renal failure. ceroid - lipofuscin accumulation in the pulmonary alveolar macrophages is believed to be the primary mechanism for the development of pulmonary fibrosis. recurrent hemorrhage with resulting hemosiderosis and inflammatory response is also suggested as an alternative mechanism. pulmonary fibrosis in hps is twice as common in women, occurring between the third and fifth decades. it initially presents as dyspnea on exertion or restrictive lung disease on pulmonary function tests, and is the most common cause of death in affected patients. the radiographic appearance of hps is nonspecific and chest radiographs may be normal at presentation. various radiographic abnormalities have been described, including reticular or reticulonodular opacities, pleural thickening, interstitial infiltrates, and perihilar fibrosis. chest radiograph in our patient at the time of admission demonstrated symmetric reticular opacities throughout both lungs [figure 1 ]. as chest radiographs sometimes underestimate the extent of disease, high - resolution ct is the imaging modality of choice for characterizing the parenchymal abnormalities and evaluating the extent of pulmonary involvement. early stages of the disease are characterized by subtle reticulations, interlobular septal thickening, and peripheral ground glass opacities. in more advanced stages, there are more severe reticulations, peribronchovascular thickening, subpleural cysts, and bronchiectasis involving central airways. abnormalities on high - resolution ct tend to be evenly distributed throughout the lungs, with a slight predilection for the middle and lower lungs. chest ct from our patient demonstrated diffuse septal thickening and ground glass opacities throughout both lungs, while honeycombing was predominantly seen in the bilateral upper lobes [figure 2a c ]. hps is more likely than the usual interstitial pneumonias or collagen diseases to also involve the upper lobes, particularly later in the course of the disease. pulmonary hemorrhage, which typically presents with diffuse ground glass opacity on ct, should be considered in these patients given the underlying platelet dysfunction. other entities that might have overlapping appearance on ct images include idiopathic pulmonary fibrosis (ipf), which features irregular reticulation, fibrosis and honeycombing in the posterior subpleural lung bases, and nonspecific interstitial pneumonitis (nsip), which has bilateral and symmetric ground glass opacity and less honeycombing. however, ipf most commonly occurs in older patients and nsip is not a progressive process. hps has been classified by the american thoracic society and the european respiratory society as a mimic of ipf and one of the few diseases that has histological features similar, but not identical to usual interstitial pneumonia. a prior imaging study examining patients with hps used a four - point score to grade the severity of ct appearance of disease. grade 1 showed minimal changes (thickened interlobular septa, reticular disease, subpleural cysts, and areas of ground glass). grade 2 had moderate disease (traction bronchiectasis, peribronchovascular thickening, and tracheal retraction in one - third or two - thirds of the lungs). grade 3 had the findings of grades 1 and 2, but involved more than two - thirds of the lungs. the study found that patients under 20 years of age typically had no ct findings (grade 0) and those between 20 and 29 years had minimal ct changes (average grade 0.25 0.16). among patients older than 30 years, those with hps1 mutations had significantly more severe disease on ct compared with those patients without this mutation (p = 0.001). worsening ct appearance correlated highly with poorer pulmonary function ; as ct severity score increased, the forced vital capacity (fvc) fell dramatically (p < 0.001). also, higher ct severity score correlated with time to death, with over half the patients with grade 3 disease dying within 4 months. our patient had a chest ct 16 months prior to admission, which showed grade 2 findings [figure 2a ]. subsequent chest ct 3 months before admission showed significant worsening of fibrosis, now involving greater than two - thirds of the lungs and meeting the criteria for grade 3 disease [figure 2b ]. final chest ct upon admission revealed extensive traction bronchiectasis, peribronchovascular thickening, and honeycombing, consistent with end - stage fibrosis [figure 2c ]. there is no effective treatment for pulmonary fibrosis due to hps, besides lung transplantation. high - dose corticosteroids are administered to patients with advanced disease, though their efficacy has not been proven. cigarette smoke and other pulmonary irritants must be avoided, and the antibiotic pirfenidone may slow the progression of pulmonary fibrosis in patients with significant residual lung. our patient had an objective and symptomatic decline over the last 12 months of her life, which rendered her oxygen dependent with a baseline requirement of 6 l / min and was under evaluation for lung transplantation prior to her terminal admission. in summary, hps is a rare congenital disorder, most often seen in puerto ricans, that manifests as easy bruising, epistaxis, prolonged bleeding, granulomatous colitis, and a high risk of pulmonary fibrosis. patients with mutations in hps1 are much more likely to develop more severe disease, typically after 30 years of age. ct correlates well with pulmonary function, and more severe ct changes significantly correlate with time to death. | hermansky pudlak syndrome (hps) is a rare autosomal recessive disorder characterized by oculocutaneous hypopigmentation, platelet dysfunction, and in many cases, life - threatening pulmonary fibrosis. we report the clinical course, imaging, and postmortem findings of a 38-year - old female with hps - related progressive pulmonary fibrosis, highlighting the role of imaging in assessment of disease severity and prognosis. |
the incidence rates of oral cancer are 3.7% for men and 2.6% for women in the sudan. several lifestyle risk factors for the development of oral cancer are familiar, including tobacco products, alcohol, infections, dietary factors, chemical irritants and frank carcinogens. prevalence of oral cancer is 3.2% in sudan and the disease is mainly attributed to n - nitrosamine rich oral snuff consumption. there are mainly 4 smokeless tobacco products : loose leaf or chewing tobacco, snuff, plug tobacco and twist or roll tobacco. chewing tobacco and snuff are by far the most widely distributed types of smokeless tobacco. however, snuff is now habitually used orally by insertion it between lower gum and cheek or lip (dipping). the oral use of snuff in north america and western europe is causally associated with an increased risk for cancer of the oral cavity and pharynx. snuff dipping has also been incriminated as being associated with cancer of the nasal cavity, oesophagus, pancreas, kidney and urinary bladder [2 - 6 ], and other pre - neoplastic changes such as leukoplakia. however, all these risk factors are beyond the scope of the present review. the main goal of this paper was to review the studied risk factors that linked to aetiology of oral cancer in sudan. in this review 33 studies published up to august 2012 in the aetiology of oral cancer from sudan were included. they were identified through searches of the medline database, using the keywords : " sudan ", " toombak ", " infection ", " hpv ", " oral hygiene ", " alcohol ", " hot meals ", " cancer ", " oral squamous cell carcinoma " and " risk factors ". papers were also searched among those quoted as references in the retrieved studies, as well as, in a few previous reviews. only papers in english were considered. in the sudan, oral snuff, known locally as toombak, is home - made from finely ground leaves of nicotiana rustica, a tobacco species with a particularly high content of nicotine and minor alkaloids. this tobacco is mixed with natron or atron (sodium bicarbonate) (about 4:1), then water is added to the mixture, and after a period of about 2 hours or longer the mixture, called " saffa ". atron, opposed to lime in other parts of the world, is probably added to toombak for its alkaline effects. it has been shown that at high ph (11.0 - 11.8) nicotine is completely protonated and its rate of absorption is increased. n - nitrosamines : the study by idris. have analyzed the tobacco specific nittrose amine (tsna) levels in toombak and found unusually high levels of these tsnas compared to the reported levels in any snuff. these high levels of tsnas found in toombak were partially attributed to the use of tobacco species, nicotiana rustica, and fermentation of toombak at elevated temperature, prolonged storage, and contamination during processing [13 - 15 ]. therefore, assuming chronic toombak use, the minimum daily dose of nnk to which these users were exposed was 0.12 - 0.44 mg. epidemiological evidence suggests that toombak is a risk factor for cancer of the oral cavity and possibly of the oesophagus in the sudan [17 - 19 ]. data from 1,916 cases of oral neoplasms occurring in the sudan in a 16-year period, from january 1970 to december 1985, were retrieved and analyzed. the study revealed a relatively high frequency of oral neoplasms in comparison with neighbouring countries. squamous - cell carcinoma was the most common oral malignancy (66.5%), followed by tumours of the salivary gland (14.7%), neoplasms of non - odontogenic and non - epithelial origin (9.6%) and odontogenic neoplasms (8.6%). female toombak use is considered social stigma in sudan, consequently the 95% of toombak users were males, which supports its etiological effects. in sweden, snus (locally known as snus), was introduced since the year 1637. the study by idris. compared between snus (sweden) and toombak. snus and toombak dippers develop a clinically and histologically characteristic lesion at the site of dipping. snus was associated with a lower risk of cancer of the oral cavity (relative risk : rr 5 - 6-fold) compared to high risk of toombak (rr 7.3 - 73.0-fold). clinical (n = 281) and histopathological (n = 141) characteristics of toombak - associated oral mucosal lesions detected in an epidemiological study in northern sudan in 1992/93 found parakeratosis, pale surface staining of the epithelium and basal cell hyperplasia were commonly observed, but epithelial dysplasia was infrequent (10/141). ultra structural features oral toombak dipper s lesions with distinctive sub epithelial hyaline deposits their bulk is made up of collagen, as typical cross - striated fibrils. the pathogenesis of this deposit could therefore be interpreted as over - production and/or reduced turnover of collagen by resident fibroblasts, which is further altered by the ingredients of toombak. in a study investigated the effects of toombak on primary normal human oral keratinocytes, fibroblasts, and a dysplastic oral keratinocytic cell line, compare to swedish snuff, a potential for toombak, higher than for swedish snuff, to damage human oral epithelium. furthermore, in oscc, apoptosis was associated with bax expression and was unaffected by p53 gene status or toombak use in oscc from the sudan. the study by ibrahim. analyzed 14 oral squamous - cell carcinomas (osccs) and 8 pre - malignant oral lesions from different sudanese patients for prevalence of mutations in exons 5 to 9 of the p53 gene in relation to toombak - dipping status. osccs (14 from sudan, 28 from scandinavia), and 3 pre - malignant oral lesions from sudanese non - dippers were used as controls. a statistically significant (p < 0.05) increased incidence in mutations of the p53 gene was found in osccs from toombak dippers (93% ; 13/14), as compared with those from non - dippers in sudan (57% ; 8/14) and in scandinavia (61% ; 17/28) respectively. several studies from sudan have proved that toombak use is a major risk factor that responsible of high frequencies of potential malignant oral lesions and oral cancers and in particular osccs in the sudan. most of tumours were observed at the site of dip application (lower lip). oral cancer seems to be gender - specific, as the majority of cases were males [26 - 31 ]. however, all of the preceded discussed literatures support the criminal role of toombak in the aetiology of oral cancer in the sudan. probably toombak has a major role but it is not alone responsible of oral cancer in the sudan, particularly in the recent years with dramatic increase in overall cancer risk. most oral cancer cases and deaths are due to both individual predisposition, linked to specific genetic characteristics, and exposure to carcinogens, caused by lifestyle behaviours, particularly tobacco. nicotine is only a minor component of tobacco leaves and constitutes about 5% of the total weight of dry plant leaves. when tobacco smoke is inhaled, 25% of the nicotine reaches the brain in about seven seconds. nicotine functions by binding to nicotinic acetylcholine receptors, causing increased heart rate, vasoconstriction, and alertness. nicotine induces dependence among genetically, mentally and socially predisposed individuals with addictive personalities. tobacco carcinogenicity is more than evident and about one fourth of oral cancer cases are attributable to cigarette smoking. specifically, tobacco products are causally linked to a variety of cancers, including those of the lung, oral cavity, nasal cavity, larynx, oropharynx, hypopharynx, oesophagus, stomach, liver, pancreas, bladder, ureter, kidney, cervix, and myeloid leukaemia. more than 60 carcinogens are present in cigarette smoke and at least 16 in unburned tobacco have been identified. the most important are tobacco - specific nitrosamines, such as 4-(methylnitrosamino)-1-(3-pyridyl)-1-butanone (nnk) and n - nitrosonornicotine (nnn), polycyclic aromatic hydrocarbons in particular, nnk, nnn and pah have been causally linked to oral cancer. few women admit to tobacco use due to cultural norms in the society, and among males the estimated prevalence is around 20%. in most samples from indoor air polluted with environmental tobacco smoke (ets), the highest concentration of an individual tsna is that of nnk. when nonsmokers had remained for up to 2 hours in a test laboratory with high ets pollution, they excreted measurable amounts of nnk metabolites in the urine, indicative of the uptake of tsna. a house to house cross - sectional survey of a random population sample of 4,535 households was performed, as the first survey on tobacco in the sudan. of the 23,367 household members identified, 21,648 (92.6%) eligible individuals were questioned about tobacco use. results showed that, among children and adolescents (4 - 17 years) prevalence of tobacco use was quite low (2%, range 1 - 2%), but there was an abrupt increase up to 25% in late adolescence. among the adult population aged 18 years and older the prevalences of toombak use (34%) and cigarette smoking (12%) among males were significantly higher than among females (2.5 and 0.9%, respectively). as part of the development of a screening procedure for oral cancer and precancer, exfoliative cytology (efc) was applied to a retrospective cohort to assess the presence and severity of oral epithelial atypia (et) in 300 subjects (100 toombak dippers ; 100 cigarette smokers ; 100 non - tobacco users) without prior knowledge of the subjects tobacco exposure. et was ascertained in 29 subjects and could not be ascertained in the remaining 271. among the 29 subjects with et, there were 11 (38%) toombak dippers, 14 (48%) cigarette smokers and 4 (14%) non - tobacco users. among the 271 subjects without et, there were 89 (33%) toombak dippers, 86 (32%) cigarette smokers and 96 (35%) non - tobacco users. for the et among toombak dippers and cigarette smokers, adjusted or and the 95% ci were found to be 3 (0.91 - 9.7) and 4 (1.2 - 12.3), respectively. both in tobacco smoke and smokeless tobacco, carcinogenic n - nitroso compounds (noc) are implicated as dna - damaging agents in cancers of the aerodigestive tract and the pancreas. poor oral hygiene was found to contribute to the formation of nitrosamines in the oral cavity. the evidence so far accumulated demonstrates that tobacco habits increase endogenous noc formation, thus adding to the burden of exposure by preformed carcinogenic noc in tobacco products. however, in view of these studies, smoked tobacco has a role in aetiology of oral cancer, but how far this is contributing in oral cancer in sudan and how far it is significant is unknown, since there is lack of data in this context. alcoholic beverages are a heterogeneous group of beverages, with variable number, type and concentration of components. carbon dioxide, minerals (mostly potassium, phosphates and sodium), amino - acids, organic and inorganic acids, polyphenols and carbohydrates are prevailing in beers. alcohols, carbohydrates (mainly sugar and pectin), organic acids, minerals (mostly potassium, iron, phosphates and calcium), polyphenols, vitamins and carbon dioxide are the major wine components, while spirit and liqueur composition is very assorted, with common components being alcohols, acids (mainly fatty and acetic acid), esters, aldehydes, terpenes, ethereal oils and volatile bases. indeed, among individuals consuming 4 - 5 drinks daily, the risk for cancer of the oral cavity is 2 - 3 folds higher than among non - drinkers [44 - 47 ]. overall, 7 - 19% oral cancer cases are attributable to heavy alcohol drinking.. the major alcohol metabolising enzymes are alcohol dehydrogenase, that oxidises ethanol to acetaldehyde, and aldehyde dehydrogenase, that detoxifies acetaldehyde to acetate. acetaldehyde is responsible for the oral carcinogenic effect of ethanol, owing to its multiple mutagenic effects on dna. in addition, ethanol is not the only carcinogen present in alcoholic drinks, other minor components, such as nitrosamines, acrylamide, oxidized polyphenols are classified as probable carcinogenic to humans, with animal experiments showing mutagenic activity on oral epithelial cells. since alcohol consumption is considered illegal in sudan, no available data about the relationship between alcoholic beverage consumption and oral cancerous development. the study evaluated cytological atypical changes in apparently healthy oral mucosa exposed to smoking, alcohol, hot meals, and peppers. the features of cytological atypia were verified among 10 individuals, including 5 smokers, 2 alcohol users, 2 hot meals and peppers consumers, and one non - exposed. for atypia among tobacco smokers, the adjusted odds ratio (or) and the 95% ci were found to be 2 (0.246 - 16.24). increased keratinisation was detected among 27 (45%) of the smokers (p < 0.0001), 17 (32.7%) of the pepper and hot meals consumers (p < 0.005), 4 (11.8%) of the alcohol consumers, and among 2 (3.7%) of the non - exposed group. statistical analyses revealed a greater mean number of agnors per nucleus in smokers (3.68) followed by (2.82) alcohol consumers, compared to the habitual peppers and hot meal consumers (2.28) and the non - exposed group (2.00). however, no available data about alcoholic consumers in the sudan, approximately 95% of the habitual use to drink in hide, since it is considered as illegal and social stigma. consequently, nobody can estimated or even imagine its burden as a risk contributes to the aetiology of oral cancer in the sudan. human papilloma viruses (hpv) infection the hpv are a large family of non - enveloped dna viruses, mainly associated with cervical cancers. recent epidemiologic evidence has suggested that hpv may be an independent risk factor for oropharyngeal cancers. hpv has a wide disease spectrum affecting the cutaneous and mucosal areas of the body, ranging from benign common warts to invasive carcinoma. hpv infections have been reported in a number of body sites, including the anogenital tract, urethra, skin, larynx, tracheobronchial mucosa, nasal cavity, paranasal sinus, and oral cavity. hpv is one of the most prevalent infections in the world with several new cases diagnosed every year. there are more than 120 genetically different, yet closely related hpvs that are referred to as genotypes. by definition, each type is defined by having less than 90% dna base pair homology with any other identified hpv type. the hpv genome encodes dna sequences for six early (e1, e2, e4, e5, e6, and e7) proteins associated with viral gene regulation and cell transformation, two late (l1 and l2) proteins which form the shell of the virus, and one region of regulatory dna sequences. the different hpv types are characterized by genotypic variations in the dna base - sequences of e6 and e7. it is these genotypic differences that permit stratification of the virus oncogenic phenotype into high, intermediate-, and low - risk types. in the case of high - risk hpv infection and under favourable conditions, the viral genome is integrated into the host genome, which is the necessary event for the keratinocytes immortality. during this process of integration, the circular form of viral genome breaks at the level of the e1 and e2 regions, never at the level of the e6 or e7 region. different studies have shown that the integrated part of the genome corresponds to e1, e6, and e7, while the regions from e2 to e5 are lost and are not transcribed in the tumours. the loss of e2 during this process of integration produces the loss of e6 and e7 control. therefore, the sequences e6 and e7 are directly involved in the cellular cycle by inhibiting the normal functions of p53 and prb, respectively. the protein p53 is known as the " genome s guard " and in the case of dna damage, the p53 can provoke the arrest of cellular division and assure the time necessary for dna repair. if damage can not be repaired, p53 is able to induce the programmed cellular death and prevent the propagation of dna damage in subsequent generations of cells. in the case of other types of tumours, p53 is usually mutated and acts as a real oncogene. in the case of hpv infection, e6 suppresses the properties of p53 gene product, achieving the functional equivalent of the two hits required to knock out both alleles of a tumour suppressor gene. the e7 protein interacts with retinoblastoma protein (prb), which is the crucial factor for the cellular cycle control. this interaction causes the release of the transcription factor e2f, which is now free to act and can stimulate the cellular division. e7 is also able to bind and inactivate the protein kinase inhibitors p21 and p27 and can interact with different proteins whose significance has still not been determined. hpv has gained much interest recently, because it is accepted as important correlates of cervical cancer. oral hpv infections have not been studied to the degree as those of the genital tract, although the evidence of association between certain tumours and hpv infection today is indisputable. oncogenic hpvs are associated with oral malignancies, but its prevalence varies widely in different studies. although study results are mixed, it seems possible that smoking and alcohol use may interact with hpv infection to increase a person s risk of oral cancer. so, oral hpv infections need to be studied and investigated deeply so that it can guide us for future cancer prevention programs, including oral hpv vaccination for oral hpv infections. nevertheless, few studies have investigated the role of viruses in general and hpv in particularly in the aetiology of oral cancer in the sudan. the first study investigated potentially malignant oral mucosal lesions from sudanese patients (9 hyperplasias/40 dysplasias). hpv was found in only 2 sudanese cases, both of which harboured both type 6 and type 11 : both these cases demonstrated mild epithelial dysplasia. another study from sudan investigated a total of 40 cases (patients with osccs) and 15 controls (patients with benign lesions) were included in the study. the cases included ; carcinomas of the oral cavity proper, carcinomas of the lip vermilion. hpv was detected among 6 (15%) of the cases, four were hpv type 16 and two were hpv type 18. a recent study from sudan reported the presence of hpv in head and neck cancers (hncs) in general and in oral in particular. hpv16 was the most prevalent type, with single infections present in 3/6 (50%) cases, whereas hpv18 and hpv33 were detected in 2/6 (33%) and 1/6 (17%), respectively. hpv infections were detected in 3 (50%) cases of oral cavity and 3 (50%) cases of pharynx. furthermore, very limited studies have investigated the role of other viruses such as, epstein - barr virus (ebv) and herpes simplex virus (hsv) in the aetiology of oral cancer in sudan beside hpv. using pcr / dna sequencing, a study has investigated the prevalence of hpv, hsv and ebv dna in brush biopsies obtained from 150 sudanese toombak dippers and 25 non - toombak dippers in formalin - fixed paraffin - embedded tissue samples obtained from 31 patients with oral dysplasias (25 toombak users and 6 non - users), and from 217 patients with oral cancers (145 toombak users and 72 non - users). in the brush tissue samples from toombak users, hpv was detected in 60 (40%), hsv in 44 (29%) and ebv in 97 (65%) of the samples. the corresponding figures for the 25 samples from non - users were 17 (68%) positive for hpv, 6 (24%) positive for hsv and 21 (84%) for ebv. the formalin - fixed samples with oral dysplasias were all negative for hpv. in the 145 oral cancer samples from toombak users, hpv was detected in 39 (27%), hsv in 15 (10%) and ebv in 53 (37%) of the samples. the corresponding figures for the samples from non - users were 15 (21%) positive for hpv, 5 (7%) for hsv and 16 (22%) for ebv. these findings illustrate that prevalence of hsv, hpv and ebv infections are common and may influence oral health and cancer development. these observations warrant further studies involving toombak - associated oral lesions, to uncover the possible mechanisms of these viral infections in the development of oral cancer, and the influence of toombak on these viruses. in another study investigated the prevalence of hpv, hsv and ebv in 155 osccs from eight different countries from different ethnic groups, continents and with different socioeconomic backgrounds, the highest prevalence of hpv was seen in sudan (65%). genetic factors have enhancing roles in many cancers including oral cancer. limited studies have dealt with the investigation of association between genetic factors and level of risk for development of oral cancer in sudan. in a study aiming to explore possible range of gene expression profiles in head and neck squamous cell carcinomas (hnscc) and pair wised normal controls from sudanese (n = 72) and norwegian (n = 45) patients using a 15k cdna microarray and to correlate the findings with clinicopathologic variables. analysis of the two population groups revealed a common set of 73 genes within three main biological pathways. this indicates that the development of hnsccs is mediated by similar biological pathways regardless of differences related to race, ethnicity, lifestyle, and/or exposure to environmental carcinogens. of particular interest, however, was the valuable association of gene expression signature found with toombak use and anatomic site of the tumours. the prevalence of mutations in exons 2 and 3 of the s100a4 gene was analyzed in the 14 osccs from toombak - dippers and in 25 cases of osccs from the control non - snuff - dippers. of the 14 osccs investigated from toombak - dippers, mutations in the p21waf1 exon 2 were found in 43% (6 out of 14), compared to 14% (2 out of 14), 22% (6 out of 27) and 14% (5 out of 35) found in those from non - snuff - dippers from the sudan, scandinavia and the usa / uk, respectively. the study by lazarus. suggested that toombak components such as tsnas may induce p53 mutations in head and neck sccs and are likely contributors to the tobacco - induced carcinogenic load in humans. a recent study investigated the prevalence of p53 codon 72 polymorphism in brush biopsies obtained from a sudanese population. a total of 174 individuals were included in the study ; chronic toombak users (n = 152) and non - users (n = 22). dna was extracted from all the samples and genotyped for the codon 72 polymorphism by polymerase chain reaction / restriction fragment length polymorphism. the arg / pro genotype was found in 53% of the 174 study participants, compared to 21% found with arg / arg and 26% found with pro / pro. stratifying by toombak use, 28 (18%), 45 (29%) and 79 (52%) of the 152 samples from toombak users had arg / arg, pro / pro and arg / pro respectively, compared to 9 (41%), 0 (0%) and 13 (59%) found in the 22 samples from non users. the differences between the samples from toombak users and non users in arg / arg and pro / pro codon 72 polymorphism and hpv infection were statistically significant (p < 0.05). the study indicated that a high prevalence of the genotype arg / pro at the p53 codon 72 may contribute to susceptibility to oscc, especially in combination with the use of carcinogenic tobacco - specific nitrosamine (tsna)-rich toombak. in another study, 9 genes related to apoptosis, cell cycle regulation and intermediate filament proteins were selected and their differential expression status was examined by real - time quantitative rt - pcr in 26 samples of sudanese osccs and their matched normal controls. the mrna levels of bcl2, keratin 1, keratin 13 and p53 were significantly lower and the level of survivin was significantly higher in the oscc samples of the toombak users compared to their paired control samples. a significant down - regulation in keratin 1 and keratin 13 expression levels was found in the oscc samples of the non - toombak users compared to their normal control samples. the differential expression of genes related to apoptosis, cell cycle regulation and types i and ii keratin could be useful diagnostic markers and provide valuable information for the understanding of oral malignancy in relation to toombak use. besides tobacco and alcohol, other risk factors have been studied in relation to oral cancer in sudan. among these factors, diet and nutrition so far there is a lack of studies focusing on the prevalence of a wide spectrum of oral mucosal lesions (oml) in patients with dermatologic diseases. this is noteworthy as skin lesions are strongly associated with oral lesions and could easily be neglected by dentists. a study from sudan found that oml were frequently diagnosed in skin diseased patients and varied systematically with age, gender, systemic condition and use of toombak. epidemiological studies conducted in various populations reported an inverse association between intake of fruit and vegetables and the risk of cancer of the oral cavity. thus, fruit and vegetables appear to be the most consistent favourable dietary aspect on oral and pharyngeal neoplasm. in a network of studies from italy, about 20 - 25% of cancers of the oral cavity and pharynx were attributed to low fruit and vegetable consumption, and the population attributable risk, rose to 85 - 95% when tobacco and alcohol consumption were also considered. the evidence on the role of milk and dairy products, as well as coffee and tea, on the risk of cancer of the oral cavity and pharynx is scanty, and does not indicate any consistent association., none of these factors has been investigated in the sudan. as sudan is very large country with different climates toombak plays a major role in the aetiology of oral cancer in sudan, since it is a potent carcinogenic, as well as it acts as a synergistic factor enhances the carcinogenesis of other etiological factors, such as genetic factors and viruses. the role of human papilloma virus in aetiology of oral cancer is reasonable, but the question how far is it without toombak ? the real contribution of other risk factors such as viruses other than human papilloma virus, smoked tobacco and alcohol still need more investigation other factors that have increased risk or reverse risk need more investigation. | abstractobjectivesto review the studied risk factors that linked to aetiology of oral cancer in the sudan. there have been numerous reports in the increase in the incidence of oral cancer from various parts of the world. a recent trend for a rising incidence of oral cancer, with the absence of the well established risk factors, has raised concern. although, there are inconsistent data on incidence and demographical factors, studies suggest that the physiologic response to risk factors by men and women vary in different populations.material and methodsthis review principally examines 33 publications devoted to aetiology of oral cancer in the sudan, in addition to some risk factors that are commonly practiced in the sudan.resultsseveral studies examining risk factors for oral cancer include tobacco use (smoked and smokeless), alcohol consumption, occupational risk, familial risk, immune deficits, virus infection and genetic factors.conclusionstoombak use and infection with high risk human papilloma virus (hpv) were extensively investigated and linked to the aetiology of oral cancer in sudan. |
understanding how the vertebrate nervous system develops from the embryonic ectoderm has been one of the most intensely investigated developmental processes. the process of neural induction was discovered by hans spemann and hilde mangold ; when dorsal mesoderm was transplanted adjacent to ventral ectoderm that normally would give rise to skin instead it formed a neural tube (spemann and mangold, 1923, reprinted 2001). this result was interpreted to mean that dorsal mesoderm releases a signaling factor that converts the adjacent ectoderm into neural tissue. while the identity of this signal was sought for decades, only in recent years have we come to appreciate the signaling and transcription factors that mediate neural induction and control neural differentiation. recently it has been possible to place these factors in a gene regulatory network that controls the progression of cells from newly induced neural precursors, to proliferative, multipotent neural plate stem cells, to specified neural progenitors. these early steps of neural development are critical for producing a pool of multipotent cells that later give rise to the multitude of neurons and glia in the central nervous system that have diverse and distinct functions, and are selectively affected in a number of human neurodegenerative diseases. research to produce a population of cells in vitro that recapitulate the differentiated progeny seen in the intact nervous system has defined a neural stem cell (nsc), which is a self - renewing cell whose progeny can produce neurons, oligodendrocytes and astrocytes (fujita, 2003). in the embryonic neural tube these cells reside in the ventricular zone (or subventricular zone in the developing cerebral cortex), and in both the embryo and in nsc cultures, these cells are activated to produce new progeny by the integration of a number of signaling pathways, such as sonic hedgehog, bmp, wnt/catenin, notch, and fgf (fig. 1 ; brault., 2001 ;, 2008 ; mizutani., 2007 ; ohtsuka., 2001 ; palma and ruiz i altaba, 2004 ; zechner., 2003) the self - renewal ability of these cells is regulated by numerous transcription factors, including he s, sox, bmi1, tlx, and gli proteins (fig. 1 ; ahmed., 2009 ; bylund., 2003 ;, 2004 ; ishibashi., 1995 ; miyagi., 2008 ; the recognition of this cell has inspired numerous researchers to develop new culturing approaches to obtain specific kinds of neurons or glia for therapeutic purposes, such as striatal neurons for huntington s disease and dopamine neurons for parkinson s disease. while there has been much success (e.g., reviewed in chan., 2014 ; ross and akimov, 2014), the potential presence of pluripotent stem cells in these cultures remains a concern because they might cause tumors if transplanted into patients. therefore, in this review we will focus on what we know about the functions of the signaling and transcriptional factors that control the earliest steps in neural development in the vertebrate embryo. these are the steps that control commitment to a neural fate, proliferation of immature cells, and initiation of differentiation. if we can determine whether the molecular interactions that occur in the embryo to regulate how cells initially become committed to a neural fate are similarly employed when embryonic stem cells (escs) and induced pluripotent stem cells (ipscs) are differentiated into nscs, we will be able to eliminate tumorigenic cells. therefore, we will relate the findings from embryos to studies of cultured mammalian stem cells in expectation that the knowledge gained from the embryo will significantly enhance in vitro stem cell approaches and thereby yield transplantable neural precursors for regenerative applications. the vertebrate neural ectoderm forms on the dorsal side of the embryo in response to signals from the adjacent dorsal mesoderm, which is called the organizer in frog and fish and the node in chick and mouse (reviewed in de robertis and kuroda, 2004 ; itoh and sokol, 2007 ; levine and brivanlou, 2007 ; moody., 2013 ; rogers., small diffusible proteins that are secreted from the organizer bind to either bmp or wnt ligands and prevent them from activating their receptors in the adjacent ectoderm. embryonic ectodermal cells become epidermis (skin) in the presence of bmps and wnts and become neural ectodermal (ne) precursors in their absence (fig. 1). the newly induced ne precursors express a large number of neural transcription factors (ntfs), including members of the fox, sox, zic and irx families, that are co - expressed in broad overlapping domains. some of these factors regulate the competence of the ne precursors to respond to neural inducing signals and cause the newly induced neural cells to become refractory to bmp and wnt signals (moody., 2013 ; rogers. fgf signaling is another important pathway that facilitates neural induction (fig. 1 ; streit., 1998 ; 2000 ; one of the downstream effectors of the fgf neural promoting activity is a protein called churchill (churc), which up - regulates early neural genes and down - regulates early mesodermal genes (sheng., 2003). its activity is mediated, at least in part, by a smad - interacting protein called sip1 (verschueren., 1999). sip1 is expressed in the dorsal ectoderm at the time that neural induction is occurring, converts nave ectoderm to neural ectoderm, represses mesoderm genes and inhibits bmp signaling (eisaki. the fgf - mediated switch from mesodermal to neural fates also is regulated by a pluripotency transcription factor, pou91/oct4, upstream of churc and sip1(snir., 2006). thus, these factors appear to be activated at the time cells are switching off a pluripotency program. adding neural inducing factors to esc or ipsc cultures also directs these cells to become ne precursors (also called primitive neural stem cells) (devine., 2011 ; tropepe., some of the cells in esc cultures secrete bmps and wnts, and maintaining ne precursors depends on inhibiting these signals (tropepe., 2001), just as in the embryo. maintaining the expression of sox1 in human esc and ipsc cultures also requires continuous down - regulation of bmp signaling (chaddah., 2012 ; neely., 2012). however, unlike in the embryo, fgf signaling inhibits human escs from forming neural tissues, at least under some culture conditions (greber., 2011). it is not clear whether this means that fgf signals have different effects in embryos versus escs, or whether fgf signaling acts to promote mesoderm formation under these culture conditions. it will be very informative to more precisely determine the roles of fgf signaling in neural induction of cultured stem cells. to our knowledge, no studies have been published regarding churc function in stem cell cultures. but, sip1 plays a key role in the decision between neural ectoderm and mesendoderm in human escs and in mouse epiblast stem cells (chng., 2010). thus, the evidence so far suggests that the induction of the ne precursor state in esc and ipsc cultures relies upon some of the same signaling factors as in the embryo. further study of the timing and effectiveness of these factors in both embryos and stem cell cultures may reveal how to drive cells towards a stabilized neural fate and prevent the formation of unwanted lineages. once the neural ectoderm is induced and the ntfs are expressed, the tissue continues to be exposed to both bmp and wnt signals from the surrounding mesoderm and ectoderm. it has been proposed that some of the earliest expressed ntfs somehow stabilize the neural fate program, and thereby prevent cells from reverting to a non - neural fate (sasai, 1998). since several of the ntf genes can be directly repressed by bmp - regulated transcription factors (rogers., 2008 2006), stabilizing a cell s commitment to a neural fate is important as the signaling environment changes in the embryo. one such factor is zic1, which causes the embryonic ectoderm to be more sensitive to neural inductive signals (kuo., 1998). to date, the mechanism by which this occurs has not been defined, but perhaps zic1 attenuates some aspect of bmp or wnt signaling. gmnn and irx1/xiro1 reduce bmp4 mrna levels when ectopically expressed in epidermal domains (glavic., 2001 ; gomez - skarmeta., 2001 ; kroll., sox3 down - regulates the expression of the bmp target, vent2, which is required for epidermis formation (rogers., 2008 ; 2009b). foxd4 down - regulates the expression of several genes in the bmp pathway, including their epidermal gene targets, and also reduces the nuclear localization of the phosphorylated smads that are the effectors of bmp signaling (yan., 2009 ; 2010). sox11 interacts with the map kinase nlk to antagonize wnt signaling by phosphorylating the tcf/catenin complex (hyodo - miura., 2002). ectopic expression of foxd4 mrna in the epidermal lineage, which is a field of high bmp / wnt expression, induces several other ntfs in presumptive epidermal cells, indirectly indicating that the bmp and/or wnt pathways have been repressed (yan., 2009). in fact, we recently found that mouse foxd4 has the same effects in xenopus embryos, indicating conservation of this protein s function (unpublished). these studies in embryos demonstrate that the combined anti - bmp and anti - wnt activities of several of the earliest expressed ntfs maintain a permissive neural ectoderm environment by dampening the effects of inhibitory bmp and wnt signals that persist in the embryo after neural induction. maintenance of a bmp / wnt - free environment effectively stabilizes the neural fate of the ne precursors, which allows them to transition into neural plate stem cells and prevents them from converting back to a non - neural state. to our knowledge, the roles of these early ntfs in stabilizing neural fates in mammalian esc cultures have not been explored. we hypothesize that ntfs have a role because maintaining ne precursors depends on inhibiting bmp and wnt signals (tropepe., 2001), and maintaining the expression of sox1, another ntf, in human esc and ipsc cultures requires continuous down - regulation of bmp signaling (chaddah., 2012 ; neely., 2012). thus, the maintenance of the ne precursor state in esc and ipsc cultures may rely upon some of the same signaling and transcription factors as in the embryo. once the neural ectoderm has been induced, the cells become highly proliferative and take on a columnar shape, thus forming the neural plate, a morphologically distinct domain that will give rise to the central nervous system. numerous experiments in embryos show that when the level of each ntf is experimentally increased in ne precursors the neural plate is expanded. this phenotype may occur because the ntfs have broadened the domain in which bmp signaling is repressed, as described above. however, neural plate expansion also appears to result from the ability of some ntfs to promote the proliferation of neural plate stem cells, and/or delay their differentiation into neural progenitors. for example, in xenopus embryos, gmnn maintains ne precursors in a proliferative state by modulating interactions between the swi / snf complex and the bhlh transcription factors that promote neural differentiation (seo and kroll, 2006 ; seo., 2005). foxd4 also increases the number of proliferating cells, expands markers of immature neural ectoderm, and down - regulates bhlh neural differentiation genes (moody., 2013 ; sullivan., 2001 interestingly, gmnn and foxd4 are down - regulated as neural plate stem cells exit the cell cycle and differentiate into neural progenitors (kroll., 1998 ; sullivan., 2001), indicating that their activities must be suppressed for neural differentiation to proceed. zic2 also represses bhlh neural differentiation genes and counteracts the formation of extra neurons when bhlh genes are ectopically expressed in the epidermis (brewster., 1998). these studies indicate that gmnn, foxd4 and zic2 cause neural plate expansion by promoting ne precursor and neural stem cell proliferation and delaying the bhlh - mediated establishment of differentiating neural progenitors. other ntfs appear to promote the transition of neural plate stem cells to neural progenitor cells, a process that requires cells to exit the cell cycle and initiate bhlh neural differentiation gene expression (imayoshi and kageyama, 2014). in several animals, soxb1 family members (sox1, sox2, sox3) maintain neural stem populations and must be down - regulated for neural progenitor differentiation to proceed (bylund. when expressed at high levels, each maintains neural stem cells in a proliferative state upstream of neuronal differentiation genes (bani - yaghoub., 2006 ; ellis., 2004 ; graham., 2003 ; li., 1998;wang., 2006 ; wegner and stolt, 2005 ; zappone. likewise, sox11, a member of the soxc subfamily, is up - regulated as neural stem cells transition to neural progenitor cells, and later maintains neuronal progenitors (bergsland. thus, together these sox genes may function downstream of foxd4, gmnn and zic2 to promote the initial step from neural plate stem cell to neural progenitor cell. several studies show that other zic genes and the irx genes function in neural progenitors downstream of the soxb1 genes to promote the onset of bhlh neural differentiation gene expression. zic1 is required for the expression of soxd (a member of the soxg group), which causes ectopic neural masses that express bhlh neural differentiation genes (mizuseki., 1998b), and zic1 and zic3 promote the expansion of neural progenitors in the spinal cord and forebrain (aruga., 2002 ; nakata., 1997 ; 1998). in drosophila, iroquois genes are required for the activation of the proneural bhlh genes (gomez - skarmeta., 1996), and in xenopus the homologous irx / xiro genes are expressed just prior to the earliest expressed bhlh neural differentiation genes(bellefroid., 1998). while irx genes promote the onset of neural differentiation (bellefroid., 1998 ; 1998), they also suppress terminal differentiation into neurons (de la calle - mustienes., 2002). these studies demonstrate that the ntfs have important roles in maintaining a ne precursor state, regulating the size of the neural plate stem cell population and controlling the onset of differentiation of the neural progenitor cells. to date, most emphasis has been put on forcing stem cells to a neural fate and then differentiating them into specific neuronal and glial types (chan., 2014 ; ross and akimov, 2014 ; wichterle., 2002). however, a common problem with these protocols is the difficulty in maintaining neural precursors in an immature, undifferentiated state. for many therapeutic applications this would be advantageous because it would expand neural precursor cells without the spontaneous and premature differentiation that commonly occurs in these cultures. in the embryo, next, as the neural plates forms, these cells transition into neural plate stem cells which subsequently differentiate into regionally - specified neural progenitor cells. whereas in the embryo, proliferative, undifferentiated neural precursors can be maintained, it is rare to see this kind of cell in esc cultures. however, similar primitive neural precursors do form in mouse esc cultures when neural inductive signaling is maintained either by adding anti - bmp factors or growing cells at a low enough density to dilute endogenously produced bmps (tropepe., 2001). the molecular mechanisms by which this primitive state is acquired and maintained, and by which these cells transition to the better characterized neural stem cell state will be important to elucidate in light of the potential to eliminate tumor - producing pluripotent cells. the studies summarized above took a gene - by - gene approach to understand the functions of each ntf during the progressive transition from neural inductive signaling to neural progenitor differentiation. based on their sequential and overlapping activities it seems likely that the ntfs coordinately function in a gene regulatory network (fig. 2). an initial set of ntfs maintains cells in a stable, immature ne precursor state that proliferates to form an appropriately sized neural plate. these are then down - regulated and a different set of ntfs is activated to promote the formation of the neural stem cell constituents of the neural plate. finally, neural stem cell ntfs are down - regulated and a third set of ntfs is activated that initiate neural differentiation. our understanding of the functional and transcriptional relationships between these proteins is woefully incomplete, yet this information is fundamental for understanding how the balance between neural precursor, neural stem and neural progenitors is molecularly regulated. developmental events often are controlled by regulatory networks of transcription factors that control temporal and region - specific gene expression (levine and davidson, 2005). therefore, we sought to place these ntfs in a network to determine their distinct roles in the progression from ne precursors to differentiating neural progenitors by analyzing the epistatic position of foxd4 amongst these other ntfs. using protein knock - down approaches we found that foxd4 is required for the expression of each of the other ntfs(yan., 2009), thus placing it upstream in the network (fig. 2). using gain - of - function approaches, we found that elevated foxd4 levels in ne precursors up - regulated gmnn and zic2, delayed the expression of sox2, sox3 and sox11, which resulted in an expanded neural plate ; and down - regulated the zic and irx genes that promote neural progenitor differentiation. we also found that foxd4 expression is down - regulated at early neural plate stages, suggesting that its repression is required for ne precursors to progress to neural plate stem cells and to neural progenitors. additional experiments demonstrated that gmnn, zic2 and sox11 together can mediate the foxd4 effects on the remaining ntfs(yan., 2009). from these results we can construct a gene regulatory network in which foxd4 is a critical upstream component. it directly activates a transcriptional triad consisting of gmnn, zic2 and sox11, which in turn regulates the more down - stream components that promote the transition to neural stem and neural progenitor cells (fig., structure - function analyses of the foxd4 protein show that it has separate domains in the n - and c - terminal regions that account for its ability to both activate or repress targets (klein. these findings illustrate how a single transcription factor can regulate the transition of immature, ne precursors to neurally - committed stem cells, and then to neural progenitors that are beginning to differentiate. the experiments described above demonstrate that foxd4 is a critical element in the gene regulatory network that regulates the balance between ne precursors, neural plate stem cells and regionally specified neural progenitor cells. elucidating how these different ntf genes interact to regulate this balance between expanding precursor populations and initiating differentiation should ultimately prove useful for preventing the formation of mesodermal and endodermal cells in neuronal culture protocols, expanding the number of neural precursors available and preventing their premature differentiation. one would predict from the experiments performed in embryos that during the differentiation of escs into nscs foxd4, gmnn, zic2 and sox11 would act early to promote immature, proliferative ne precursors and impede neural differentiation (fig. our preliminary work analyzing the role of foxd4 in embryoid bodies derived from mouse escs and differentiated along a neural lineage indicates that this protein is key to the transition between pluripotency and neurally committed cells, just as in the embryo (moody., 2013, and unpublished observations). in the embryo, gmnn promotes an uncommitted state by promoting polycomb mediated repressive histone marks at differentiation - promoting genes (lim., 2011). it also prevents premature neurogenesis by antagonizing the interaction between brg and bhlh neural differentiation factors (seo., 2005). just like in the embryo, in mouse escs gmnn is required for the acquisition of a neural fate, and can promote the formation of neural precursor cells by maintaining the chromatin in a hyper - acetylated state and in an open conformation that allows other ntfs access to their binding sites on the dna (yellajoshyula., 2011). subsequent work showed that gmnn promotes both activating and repressive histone modifications at neural differentiation genes, suggesting that gmnn makes the chromatin accessible to both factors that promote ne precursors as well as to factors that will subsequently activate neural differentiation genes (yellajoshyula., 2012). these results explain why gmnn - deficient neural progenitors in the mouse cortex are defective in cell proliferation and differentiation (spella., 2011), whereas gmnn - deficient nscs in neurospheres can still divide and differentiate into neurons and glia (schultz., 2011). whereas the role of zic2 has not been examined in esc culture, soxb1 family proteins (sox1, sox2, sox3) are expressed throughout mouse and chick embryo ne precursors and neural plate stem cells, and when expressed at high levels they prevent cells from differentiating into neurons (uwanogho., 1995 ; wegner, 1999). in addition, sox2 has an earlier function in escs : maintenance of pluripotency(avilion., 2003). sox11 is expressed through - out ne precursors and neural plate stem cells, overlapping completely with sox2 and sox3 expression in xenopus (hyodo - miura., 2002), whereas in chick and mouse embryos, it is expressed later in neural progenitors and differentiating neurons, and does not overlap with sox2 or sox3 expression in the neural tube (uwanogho., 1995 ; wegner, 1999) ; in mouse it also is expressed in several other tissues to support progenitor cell survival (bhattaram., 2010). a recent study showed that sox2, sox3 and sox11 bind to the promoters / enhancers of the same neural genes expressed in mouse nscs, neural progenitors and differentiating neurons in a defined temporal sequence (bergsland., 2011). in escs, sox2 binds to nsc genes to mark sites for later sox3 binding. subsequently in nscs, sox3 binds to neuronal differentiation genes to mark sites for later sox11 binding. in each case, sox protein binding results in biva - lent chromatin marks, indicating that part of their effect is making the appropriate neural genes accessible to other transcriptional factors that are expressed later. there is a large literature that describes the ntfs that are involved in neural progenitor cell (npc) formation that has been extensively reviewed elsewhere (florio and huttner, 2014 ; imayoshi and kageyama, 2014). as mentioned above, the bhlh factors, particularly neurog and neurod are critical for npcs to exit the cell cycle and activate neuron or glial specific genes. in addition, pattern formation genes are important in providing npcs with their spatial address within the neural tube. depending upon whether the npc is located in the anterior or posterior, dorsal or ventral parts of the neural tube, the expression of patterning genes will impact whether it differentiates into the forebrain versus spinal cord, and sensory versus motor neuron. once the neural ectoderm is induced and the ntfs are expressed, the tissue continues to be exposed to both bmp and wnt signals from the surrounding mesoderm and ectoderm. it has been proposed that some of the earliest expressed ntfs somehow stabilize the neural fate program, and thereby prevent cells from reverting to a non - neural fate (sasai, 1998). since several of the ntf genes can be directly repressed by bmp - regulated transcription factors (rogers., 2008 ; taylor., 2006), stabilizing a cell s commitment to a neural fate is important as the signaling environment changes in the embryo. one such factor is zic1, which causes the embryonic ectoderm to be more sensitive to neural inductive signals (kuo., 1998). to date, the mechanism by which this occurs has not been defined, but perhaps zic1 attenuates some aspect of bmp or wnt signaling. gmnn and irx1/xiro1 reduce bmp4 mrna levels when ectopically expressed in epidermal domains (glavic. sox3 down - regulates the expression of the bmp target, vent2, which is required for epidermis formation (rogers., 2008 ; 2009b). foxd4 down - regulates the expression of several genes in the bmp pathway, including their epidermal gene targets, and also reduces the nuclear localization of the phosphorylated smads that are the effectors of bmp signaling (yan. sox11 interacts with the map kinase nlk to antagonize wnt signaling by phosphorylating the tcf/catenin complex (hyodo - miura., 2002). ectopic expression of foxd4 mrna in the epidermal lineage, which is a field of high bmp / wnt expression, induces several other ntfs in presumptive epidermal cells, indirectly indicating that the bmp and/or wnt pathways have been repressed (yan., 2009). in fact, we recently found that mouse foxd4 has the same effects in xenopus embryos, indicating conservation of this protein s function (unpublished). these studies in embryos demonstrate that the combined anti - bmp and anti - wnt activities of several of the earliest expressed ntfs maintain a permissive neural ectoderm environment by dampening the effects of inhibitory bmp and wnt signals that persist in the embryo after neural induction. maintenance of a bmp / wnt - free environment effectively stabilizes the neural fate of the ne precursors, which allows them to transition into neural plate stem cells and prevents them from converting back to a non - neural state. to our knowledge, the roles of these early ntfs in stabilizing neural fates in mammalian esc cultures have not been explored. we hypothesize that ntfs have a role because maintaining ne precursors depends on inhibiting bmp and wnt signals (tropepe., 2001), and maintaining the expression of sox1, another ntf, in human esc and ipsc cultures requires continuous down - regulation of bmp signaling (chaddah., 2012 ; neely. thus, the maintenance of the ne precursor state in esc and ipsc cultures may rely upon some of the same signaling and transcription factors as in the embryo. once the neural ectoderm has been induced, the cells become highly proliferative and take on a columnar shape, thus forming the neural plate, a morphologically distinct domain that will give rise to the central nervous system. numerous experiments in embryos show that when the level of each ntf is experimentally increased in ne precursors the neural plate is expanded. this phenotype may occur because the ntfs have broadened the domain in which bmp signaling is repressed, as described above. however, neural plate expansion also appears to result from the ability of some ntfs to promote the proliferation of neural plate stem cells, and/or delay their differentiation into neural progenitors. for example, in xenopus embryos, gmnn maintains ne precursors in a proliferative state by modulating interactions between the swi / snf complex and the bhlh transcription factors that promote neural differentiation (seo and kroll, 2006 ; seo., 2005). foxd4 also increases the number of proliferating cells, expands markers of immature neural ectoderm, and down - regulates bhlh neural differentiation genes (moody., 2013 ; sullivan., 2001 interestingly, gmnn and foxd4 are down - regulated as neural plate stem cells exit the cell cycle and differentiate into neural progenitors (kroll., 1998 ; sullivan., 2001), indicating that their activities must be suppressed for neural differentiation to proceed. zic2 also represses bhlh neural differentiation genes and counteracts the formation of extra neurons when bhlh genes are ectopically expressed in the epidermis (brewster., these studies indicate that gmnn, foxd4 and zic2 cause neural plate expansion by promoting ne precursor and neural stem cell proliferation and delaying the bhlh - mediated establishment of differentiating neural progenitors. other ntfs appear to promote the transition of neural plate stem cells to neural progenitor cells, a process that requires cells to exit the cell cycle and initiate bhlh neural differentiation gene expression (imayoshi and kageyama, 2014). in several animals, soxb1 family members (sox1, sox2, sox3) maintain neural stem populations and must be down - regulated for neural progenitor differentiation to proceed (bylund., 2003 ; kishi., 2000;mizuseki., 1998a ; penzel., each maintains neural stem cells in a proliferative state upstream of neuronal differentiation genes (bani - yaghoub., 2006 ; ellis., 2004 ; graham., 2003 ; li. likewise, sox11, a member of the soxc subfamily, is up - regulated as neural stem cells transition to neural progenitor cells, and later maintains neuronal progenitors (bergsland., 2006 ; uwanogho., thus, together these sox genes may function downstream of foxd4, gmnn and zic2 to promote the initial step from neural plate stem cell to neural progenitor cell. several studies show that other zic genes and the irx genes function in neural progenitors downstream of the soxb1 genes to promote the onset of bhlh neural differentiation gene expression. zic1 is required for the expression of soxd (a member of the soxg group), which causes ectopic neural masses that express bhlh neural differentiation genes (mizuseki., 1998b), and zic1 and zic3 promote the expansion of neural progenitors in the spinal cord and forebrain (aruga., 2002 ; nakata., 1997 ; 1998). in drosophila, iroquois genes are required for the activation of the proneural bhlh genes (gomez - skarmeta., 1996), and in xenopus the homologous irx / xiro genes are expressed just prior to the earliest expressed bhlh neural differentiation genes(bellefroid., 1998). while irx genes promote the onset of neural differentiation (bellefroid., 1998 ; 1998), they also suppress terminal differentiation into neurons (de la calle - mustienes., 2002). these studies demonstrate that the ntfs have important roles in maintaining a ne precursor state, regulating the size of the neural plate stem cell population and controlling the onset of differentiation of the neural progenitor cells. however, their functions in cultured stem cells has not been extensively explored. to date, most emphasis has been put on forcing stem cells to a neural fate and then differentiating them into specific neuronal and glial types (chan., 2014 ; ross and akimov, 2014 ; wichterle., 2002). however, a common problem with these protocols is the difficulty in maintaining neural precursors in an immature, undifferentiated state. for many therapeutic applications this would be advantageous because it would expand neural precursor cells without the spontaneous and premature differentiation that commonly occurs in these cultures. in the embryo, next, as the neural plates forms, these cells transition into neural plate stem cells which subsequently differentiate into regionally - specified neural progenitor cells. whereas in the embryo, proliferative, undifferentiated neural precursors can be maintained, it is rare to see this kind of cell in esc cultures. however, similar primitive neural precursors do form in mouse esc cultures when neural inductive signaling is maintained either by adding anti - bmp factors or growing cells at a low enough density to dilute endogenously produced bmps (tropepe., 2001). the molecular mechanisms by which this primitive state is acquired and maintained, and by which these cells transition to the better characterized neural stem cell state will be important to elucidate in light of the potential to eliminate tumor - producing pluripotent cells. the studies summarized above took a gene - by - gene approach to understand the functions of each ntf during the progressive transition from neural inductive signaling to neural progenitor differentiation. based on their sequential and overlapping activities it seems likely that the ntfs coordinately function in a gene regulatory network (fig. 2). an initial set of ntfs maintains cells in a stable, immature ne precursor state that proliferates to form an appropriately sized neural plate. these are then down - regulated and a different set of ntfs is activated to promote the formation of the neural stem cell constituents of the neural plate. finally, neural stem cell ntfs are down - regulated and a third set of ntfs is activated that initiate neural differentiation. our understanding of the functional and transcriptional relationships between these proteins is woefully incomplete, yet this information is fundamental for understanding how the balance between neural precursor, neural stem and neural progenitors is molecularly regulated. developmental events often are controlled by regulatory networks of transcription factors that control temporal and region - specific gene expression (levine and davidson, 2005). therefore, we sought to place these ntfs in a network to determine their distinct roles in the progression from ne precursors to differentiating neural progenitors by analyzing the epistatic position of foxd4 amongst these other ntfs. using protein knock - down approaches we found that foxd4 is required for the expression of each of the other ntfs(yan., 2009), thus placing it upstream in the network (fig. 2). using gain - of - function approaches, we found that elevated foxd4 levels in ne precursors up - regulated gmnn and zic2, delayed the expression of sox2, sox3 and sox11, which resulted in an expanded neural plate ; and down - regulated the zic and irx genes that promote neural progenitor differentiation. we also found that foxd4 expression is down - regulated at early neural plate stages, suggesting that its repression is required for ne precursors to progress to neural plate stem cells and to neural progenitors. additional experiments demonstrated that gmnn, zic2 and sox11 together can mediate the foxd4 effects on the remaining ntfs(yan., 2009). from these results we can construct a gene regulatory network in which foxd4 is a critical upstream component. it directly activates a transcriptional triad consisting of gmnn, zic2 and sox11, which in turn regulates the more down - stream components that promote the transition to neural stem and neural progenitor cells (fig., structure - function analyses of the foxd4 protein show that it has separate domains in the n - and c - terminal regions that account for its ability to both activate or repress targets (klein. these findings illustrate how a single transcription factor can regulate the transition of immature, ne precursors to neurally - committed stem cells, and then to neural progenitors that are beginning to differentiate. the experiments described above demonstrate that foxd4 is a critical element in the gene regulatory network that regulates the balance between ne precursors, neural plate stem cells and regionally specified neural progenitor cells. elucidating how these different ntf genes interact to regulate this balance between expanding precursor populations and initiating differentiation should ultimately prove useful for preventing the formation of mesodermal and endodermal cells in neuronal culture protocols, expanding the number of neural precursors available and preventing their premature differentiation. one would predict from the experiments performed in embryos that during the differentiation of escs into nscs foxd4, gmnn, zic2 and sox11 would act early to promote immature, proliferative ne precursors and impede neural differentiation (fig. our preliminary work analyzing the role of foxd4 in embryoid bodies derived from mouse escs and differentiated along a neural lineage indicates that this protein is key to the transition between pluripotency and neurally committed cells, just as in the embryo (moody., gmnn promotes an uncommitted state by promoting polycomb mediated repressive histone marks at differentiation - promoting genes (lim., 2011). it also prevents premature neurogenesis by antagonizing the interaction between brg and bhlh neural differentiation factors (seo., 2005). just like in the embryo, in mouse escs gmnn is required for the acquisition of a neural fate, and can promote the formation of neural precursor cells by maintaining the chromatin in a hyper - acetylated state and in an open conformation that allows other ntfs access to their binding sites on the dna (yellajoshyula., subsequent work showed that gmnn promotes both activating and repressive histone modifications at neural differentiation genes, suggesting that gmnn makes the chromatin accessible to both factors that promote ne precursors as well as to factors that will subsequently activate neural differentiation genes (yellajoshyula., 2012). these results explain why gmnn - deficient neural progenitors in the mouse cortex are defective in cell proliferation and differentiation (spella., 2011), whereas gmnn - deficient nscs in neurospheres can still divide and differentiate into neurons and glia (schultz., 2011). whereas the role of zic2 has not been examined in esc culture, soxb1 family proteins (sox1, sox2, sox3) are expressed throughout mouse and chick embryo ne precursors and neural plate stem cells, and when expressed at high levels they prevent cells from differentiating into neurons (uwanogho., 1995 ; wegner, 1999). in addition, sox2 has an earlier function in escs : maintenance of pluripotency(avilion., 2003). sox11 is expressed through - out ne precursors and neural plate stem cells, overlapping completely with sox2 and sox3 expression in xenopus (hyodo - miura., 2002), whereas in chick and mouse embryos, it is expressed later in neural progenitors and differentiating neurons, and does not overlap with sox2 or sox3 expression in the neural tube (uwanogho., 1995 ; wegner, 1999) ; in mouse it also is expressed in several other tissues to support progenitor cell survival (bhattaram., 2010). a recent study showed that sox2, sox3 and sox11 bind to the promoters / enhancers of the same neural genes expressed in mouse nscs, neural progenitors and differentiating neurons in a defined temporal sequence (bergsland., 2011). in escs, sox2 binds to nsc genes to mark sites for later sox3 binding. subsequently in nscs, sox3 binds to neuronal differentiation genes to mark sites for later sox11 binding. in each case, sox protein binding results in biva - lent chromatin marks, indicating that part of their effect is making the appropriate neural genes accessible to other transcriptional factors that are expressed later. there is a large literature that describes the ntfs that are involved in neural progenitor cell (npc) formation that has been extensively reviewed elsewhere (florio and huttner, 2014 ; imayoshi and kageyama, 2014). as mentioned above, the bhlh factors, particularly neurog and neurod are critical for npcs to exit the cell cycle and activate neuron or glial specific genes. in addition, pattern formation genes are important in providing npcs with their spatial address within the neural tube. depending upon whether the npc is located in the anterior or posterior, dorsal or ventral parts of the neural tube, the expression of patterning genes will impact whether it differentiates into the forebrain versus spinal cord, and sensory versus motor neuron. a large literature describes the signaling and transcriptional factors that are responsible for differentiating neural progenitor cells into the wide range of neurons and glia that are responsible for the complex function of the vertebrate central nervous system. these factors are important for understanding how the brain functions, and for designing stem cell protocols to replace specific cells affected by neurodegenerative disease and injury. in contrast, much less in know about the molecular regulation that guides a newly induced neural ectodermal cell to commit to its new fate, expand the neural plate stem cell population and transition to a neural progenitor cells. as work in the embryo reveals more details about the regulation of these earliest steps in neural development, we need to test these details in cultured stem cell paradigms to facilitate the manipulation of nscs for therapeutic uses. it is now important to test these possibilities experimentally, and elucidate the essential gene regulatory network in mammalian esc - and ipsc - derived neural cell cultures. using the information we have obtained from the embryo | the early steps of neural development in the vertebrate embryo are regulated by sets of transcription factors that control the induction of proliferative, pluripotent neural precursors, the expansion of neural plate stem cells, and their transition to differentiating neural progenitors. these early events are critical for producing a pool of multipotent cells capable of giving rise to the multitude of neurons and glia that form the central nervous system. in this review we summarize findings from gain- and loss - of - function studies in embryos that detail the gene regulatory network responsible for these early events. we discuss whether this information is likely to be similar in mammalian embryonic and induced pluripotent stem cells that are cultured according to protocols designed to produce neurons. the similarities and differences between the embryo and stem cells may provide important guidance to stem cell protocols designed to create immature neural cells for therapeutic uses. |
heart failure is considered the last stage of heart disease and a significant cause of worldwide mortality and morbidity.1,2,3 the end - stage of heart failure, which is marked by a lack of response to medical treatment, disabling symptoms, and repeated hospital stays, is associated with high morbidity and mortality.4 heart transplantation is an acceptable gold standard treatment for select patients in the terminal stages.5 the exacerbated neurohormonal activity plays an important role in disease progression and prognosis in heart failure.6,7,8,9 this neurohormonal axis has become one of the biggest targets in heart failure interventions.10,11,12,13,14,15 it is well known that beta - blockers provide survival improvement and, because of this, have become one of the main drugs for treating heart failure.16 despite the significant improvement in quality of life and survival provided by heart transplantation, the neurohormonal profile is not restored to normal values.6 the mechanisms involved and the neurohormonal profile after heart transplantation have been poorly described, especially during the 6-minute walking tests that could represent the effort relative to daily activities.17 the aim of this study was to evaluate the neurohormonal activity in heart transplant recipients and compare it with that in heart failure and healthy subjects during rest and just after the 6-minute walking test. a total of 20 sedentary heart transplant recipients (18 men, 4911 years), 11 sedentary heart failure patients (8 men, 4310 years), and 7 sedentary healthy subjects (5 men 398 years) were included in this study. heart transplant recipients and heart failure patients with atrial fibrillation, a pacemaker, and noncardiovascular functional limitations like osteoarthritis were excluded from this study. heart failure patients whose drug therapy was not optimized were also excluded from this study. optimization was considered 50 mg / day or more of carvedilol for at least 3 months.18 healthy subjects did not have any risk factors for cardiovascular disease or noncardiovascular functional limitations. this protocol was approved by the ethics committee of the heart institute, incor hcfmusp. all patients were asked to refrain from both strenuous physical activity and the consumption of any stimulants (eg, coffee, tobacco, alcohol) 24 hours prior to the 6cwt. the patients last meal was ingested at least 2 hours before the start of the test. before starting the 6cwt and blood sample collection the 6cwt using the borg scale was performed on a treadmill with zero inclination and patient - controlled speed in a temperature - controlled room (2123c) in the afternoon (between 13:00 hours and 15:00 hours). all patients were advised to keep walking during the test at a pace between relatively easy and slightly tiring (between 11 and 13 on the borg scale). the distance walked was recorded by the treadmill microprocessor (series 2000, marquette electronics, milwaukee, wi, usa). encouragement was standardized with phrases like if you can walk faster, increase the speed, you are doing very well, and if it is tiring, you can reduce the speed. blood pressure was measured at rest and at the sixth minute by the auscultation method. the electrocardiography (max 1, marquette electronics), ventilatory and gas exchange variables were continuously evaluated breath - by - breath by a computerized system (vmax 229 model, sensormedics, yorba linda, ca, usa) during the entire test but only collected at rest (stand position) and just after the 6-minute walking test. the 6-minute walking test has been shown to reflect submaximal effort in heart failure patients19 and to reproduce the daily activities.20 blood samples were collected immediately before the 6-minute walking test with patients in the upright position and just after the exercise by the antecubital vein. the norepinephrine assay was performed as previously reported.21 all heart failure patients were receiving carvedilol associated with an ace inhibitor or losartan. patients took carvedilol, angiotensin - converting enzyme inhibitors, losartan, and isosorbide 5-mononitrate twice per day, one half of the daily dose in the morning (9:00 a.m.) and the other half at night (9:00 p.m.). diuretics, digoxin, and spironolactone were taken in the morning (9:00 a.m.). all heart transplant recipients were receiving immunosuppressive therapy two times per day, one half of the daily dose in the morning and the other half at night. the descriptive analysis is presented as the mean and standard deviation. to compare the norepinephrine levels between the 3 groups data were analyzed using the statistical package for social sciences for windows, 11.5 (spss inc, chicago, il). a total of 20 sedentary heart transplant recipients (18 men, 4911 years), 11 sedentary heart failure patients (8 men, 4310 years), and 7 sedentary healthy subjects (5 men 398 years) were included in this study. heart transplant recipients and heart failure patients with atrial fibrillation, a pacemaker, and noncardiovascular functional limitations like osteoarthritis were excluded from this study. heart failure patients whose drug therapy was not optimized were also excluded from this study. optimization was considered 50 mg / day or more of carvedilol for at least 3 months.18 healthy subjects did not have any risk factors for cardiovascular disease or noncardiovascular functional limitations. this protocol was approved by the ethics committee of the heart institute, incor hcfmusp. all patients were asked to refrain from both strenuous physical activity and the consumption of any stimulants (eg, coffee, tobacco, alcohol) 24 hours prior to the 6cwt. the patients last meal was ingested at least 2 hours before the start of the test. before starting the 6cwt and blood sample collection the 6cwt using the borg scale was performed on a treadmill with zero inclination and patient - controlled speed in a temperature - controlled room (2123c) in the afternoon (between 13:00 hours and 15:00 hours). all patients were advised to keep walking during the test at a pace between relatively easy and slightly tiring (between 11 and 13 on the borg scale). the distance walked was recorded by the treadmill microprocessor (series 2000, marquette electronics, milwaukee, wi, usa). encouragement was standardized with phrases like if you can walk faster, increase the speed, you are doing very well, and if it is tiring, you can reduce the speed. blood pressure was measured at rest and at the sixth minute by the auscultation method. the electrocardiography (max 1, marquette electronics), ventilatory and gas exchange variables were continuously evaluated breath - by - breath by a computerized system (vmax 229 model, sensormedics, yorba linda, ca, usa) during the entire test but only collected at rest (stand position) and just after the 6-minute walking test. the 6-minute walking test has been shown to reflect submaximal effort in heart failure patients19 and to reproduce the daily activities.20 blood samples were collected immediately before the 6-minute walking test with patients in the upright position and just after the exercise by the antecubital vein. patients took carvedilol, angiotensin - converting enzyme inhibitors, losartan, and isosorbide 5-mononitrate twice per day, one half of the daily dose in the morning (9:00 a.m.) and the other half at night (9:00 p.m.). diuretics, digoxin, and spironolactone were taken in the morning (9:00 a.m.). all heart transplant recipients were receiving immunosuppressive therapy two times per day, one half of the daily dose in the morning and the other half at night. the descriptive analysis is presented as the mean and standard deviation. to compare the norepinephrine levels between the 3 groups data were analyzed using the statistical package for social sciences for windows, 11.5 (spss inc, chicago, il). during rest, norepinephrine plasma level was higher in heart transplant recipients and healthy subjects. just after the 6-minute walking test, heart transplant recipients norepinephrine plasma level was not different than that of heart failure patients and both these groups had a higher level of norepinephrine than did healthy subjects. the main finding of this study is that norepinephrine remained increased just after the 6-minute walking test in heart transplant recipients. this is the first report of the norepinephrine profile during a 6-minute walking test in heart transplant recipients. that it s why we studied the neurohormonal profile in a 6-minute walking test. this way the intensity of the 6-minute walking test, between 11 and 13 on the borg scale, was similar for all groups, which are expected to spend about the same metabolic or vo2, regardless of physical status. moreover, they were all clinically stable outpatients with optimized medication.22 prez - villa studied the neurohormonal profile of 37 heart failure patients on the waiting list for heart transplantation before and 1, 4, 9, and 12 months after heart transplantation. the authors concluded that the neurohormonal activation did not normalize after heart transplantation. in this study, plasma norepinephrine level was only recorded before and 1 month after heart transplantation, showing a tendency to decrease. in our study, the heart failure group had a higher level of resting norepinephrine compared with heart transplant recipients and healthy subjects. the norepinephrine level of the heart transplant recipients did not differ from that of healthy subjects, probably by the partial reinnervation that occurred during the 8.5 years. the progressive heart reinnervation through the years could have progressively decreased the necessity for norepinephrine plasma levels in heart transplant recipients. the study by ferretti evaluated the neurohormonal profile of 17 heart transplant recipients and 9 healthy subjects at rest and at maximal exercise. the resting norepinephrine plasma level was higher in heart transplant recipients than in healthy subjects. in our study the time of heart transplantation, ie, different stages of reinnervation, between our heart transplant recipients and the ones in the study by ferretti was different (8.5 versus 3.4 years). the norepinephrine plasma level was also higher in heart transplant recipients compared with that in healthy subjects. this is in accordance with our results, despite the fact that our heart transplant recipients had performed a 6-minute walking test. this study was limited by the study design (cross - sectional), small study population, and the use of a single method of neurohormonal evaluation. this study was limited by the study design (cross - sectional), small study population, and the use of a single method of neurohormonal evaluation. | objective : we sought to evaluate the neurohormonal activity in heart transplant recipients and compare it with that in heart failure patients and healthy subjects during rest and just after a 6-minute walking test.introduction:despite the improvements in quality of life and survival provided by heart transplantation, the neurohormonal profile is poorly described.methods:twenty heart transplantation (18 men, 4911 years and 8.53.3 years after transplantation), 11 heart failure (8 men, 4310 years), and 7 healthy subjects (5 men 398 years) were included in this study. blood samples were collected immediately before and during the last minute of the exercise.results:during rest, patients norepinephrine plasma level (659225 pg / ml) was higher in heart transplant recipients (463167 pg / ml) and heathy subjects (512132), p<0.05. heart transplant recipient s norepinephrine plasma level was not different than that of healthy subjects. just after the 6-minute walking test, the heart transplant recipient s norepinephrine plasma level (1248692 pg / ml) was not different from that of heart failure patients (1174653 pg / ml). both these groups had a higher level than healthy subjects had (54595 pg / ml), p<0.05.conclusion : neurohormonal activity remains increased after the 6-minute walking test after heart transplantation. |
we do not understand yet why some patients develop multiple myeloma and others a single plasmacytoma 1, 2, but that might be related to differences in the chemokine receptor expression profiles of the malignant plasma cells or cellular adhesion molecules 3. solitary extramedullary plasmacytomas (sep) are extremely rare tumors that arise outside of the bone marrow in the absence of any sign of multiple myeloma 4, 5. they are solitary lesions, and are often seen in the head and neck regions 6, 7, mainly in the upper aerodigestive and to a lesser extent in the gastrointestinal tract (gi) tract, bladder, central nervous system (cns), thyroid, breast, testes, parotid gland, lymph nodes, as well as in skin. solitary extramedullary plasmacytomas tends to occur during the fifth and seventh decades of life, rarely in younger population 8. extramedullary plasmacytomas (emps) can arise in patients with multiple myeloma at any time during the course of the disease and in onethird of the cases, resulting in a worse clinical outcome that should not be confused with sep 9, 10. this particular case involves a 40yearold hispanic male with past medical history of mental retardation, anxiety, and dyslipidemia who lives in a group home. patient presented initially to his primary care physician (pcp) for an annual physical. during routine examination, a neck mass was identified on the base of the lateral right side of the neck anterior of the sternocleidomastoid that measured 3 cm by 3 cm nontender with slight mobility. he denied any constitutional symptoms such as fever, chills, sweating, weight loss or change in diet or bowel habit, easy bruising, or hoarseness. patient was referred to ent, and neck soft tissue ct scan with and without contrast was performed which showed enhancing 3.4 2.8 5.2 cm mass right retromandibular region just anterior to the sternocleidomastoid muscle with mass effect pushing the carotid vessels posteriorly with adjacent bony destruction ; metastasis was not excluded by this test. also, the scan was showing anterior adjacent mass approximately 0.8 1.1 cm that may present metastasis or mildly enlarged lymph node. upon seeing ent doctor, fine needle aspiration (fna) was done on the site and referred to a medical oncologist for this suspicious mass indicating the malignant pathology. fna result was not diagnostic as it showed cuboidal to columnar histologically benign appearing cells along with many small to medium size lymphocytes with differential diagnosis of : (i) salivary gland neoplasm, (ii) branchial cleft cyst, or (iii) possibility lymphoid proliferative disorder. based on the fna result, lymphoma could not be ruled out and surgical biopsy was recommended at that time. initial blood work up was done showing monocyte was elevated of 11.3%. blood work up : cr0.9wbc5.9hb15.5hct45.3mcv90.5plt283granulocytes62.8%lymphocytes28.2%monocytes8.0%glucose87na142k4.3ast19alt27alkphos80 initial differential diagnoses by ent physician involved castleman disease versus plasma cell neoplasia. open biopsy was done with frozen section and it was found to have an extensive plasma cells with no carcinoma but some cells showing binucleated forms and atypical nuclei with final diagnosis of primary lymph node plasmacytoma (figs 1 and 2). the following observations were made : cd138 staining : positive flow cytometry : nondiagnostic.beta2 microglobulin : normal range with no elevation immunofixation.quantitative immunoglobulins igg / a / m panel : normal.serum k / l light chains ratio : normal.pet ct scan showing mildly enlarged right level 2b lymph nodes measuring 14 mm with suv of 4.1 and few adjacent smaller level 2b lymph nodes which are a normal size with lowgrade metabolic activity.bm bx was done and showed no myeloma, norm cellular marrow with maturing trilineage.hematopoiesis no definitive morphologic or immunephenotypic evidence of clonal expansion of plasma cells or involvement by a mature bcell nonhodgkin lymphoma mildly increased iron storeskappa and lambda : stain a few polytypical plasma cells in normal number and distribution (figs 3, 4, 5, 6). pet ct scan showing mildly enlarged right level 2b lymph nodes measuring 14 mm with suv of 4.1 and few adjacent smaller level 2b lymph nodes which are a normal size with lowgrade metabolic activity. bm bx was done and showed no myeloma, norm cellular marrow with maturing trilineage. hematopoiesis no definitive morphologic or immunephenotypic evidence of clonal expansion of plasma cells or involvement by a mature bcell nonhodgkin lymphoma mildly increased iron stores kappa and lambda : stain a few polytypical plasma cells in normal number and distribution (figs 3, 4, 5, 6). initial recommendation by the oncologist was radiation therapy for definitive treatment for localized plasmacytoma of the neck which should likely provide adequate disease control onsite. patient received 4500 cgy in 25 fractions of radiation treatments on the right neck with 3060 cgy in 17 fractions. the primary tumor site received additional 1440 cgy in eight fractions of the total of 4500 cgy in 25 fractions. after radiation, patient was observed and monitored periodically and clinically did not show any evidence of residual disease other than superficial skin changes from the radiation which resolved later. the patient will be at risk for developing plasmacytoma at other locations, hence he will be under close follow up. in this report, we present a case of sep with no involvement of the bony structure or bone marrow. the case is quite rare as it represents 210% of all the multiple myeloma cases 11, 12. solitary extra osseous plasmacytoma has a unique characteristic of being undetectable disease elsewhere on pet / cat scan. a plasmacytoma is a unique solitary mass of neoplastic monoclonal plasma cells in either bone or soft tissue (extramedullary). the most common type is multiple myeloma, which is usually a disseminated disease and characterized by abnormal m protein. the other two types, solitary plasmacytoma of the bone and emp of the soft tissue, are considerably less common. emps present in 80%) originate in the head and neck region 16. anatomically, we can divide the sep into two groups : (i) plasmacytoma of the skeletal system (sbp) and (ii) emp 21, 24. the diagnosis of emp of the soft tissue has been based on the following criteria : (i) pathological tissue evidence of monoclonal plasma cells involving a single extramedullary site ; (ii) no bone marrow involvement ; (iii) no anemia, hypercalcemia or renal impairment caused by plasma cell dyscrasias ; (iv) negative skeletal survey results ; and (v) low serum or urinary levels of monoclonal immunoglobulin 2. the etiology of this disease remains unknown, but factors, such as chronic irritation from inhaled irritants or viral pathogenesis, have been previously indicated 17, 18. radiotherapy remains the mainstay for the management of emp. as it considered radiosensitive, with a local control rate of 90100% with a rate of conversion into mm moderate dose rt of at least 40 gy using limited radiation fields is recommended 14, 15, 17, 19, 20. because of the high rate of recurrence and progression to multiple myeloma, followup radiological and electrophoresis assessment is required following treatment 22. a literature search revealed no publications supporting the use of surgery alone to treat emp 24. | key clinical messageplasma cell neoplasms are characterized by a neoplastic plasma cell lineage which produces a monoclonal immunoglobulin. these neoplasms can present as a single lesion (solitary plasmacytoma) or as multiple lesions (multiple myeloma). solitary plasmacytomas most frequently occur in bone (plasmacytomas of bone), but can also be found outside bone in soft tissues (extramedullary plasmacytomas). |
upper gastrointestinal bleeding ia a major emergency and represents a major public health problem, its prevalence being 100170:100000 people. bleeding from esophageal varices is only 511% of all gastrointestinal bleeding and in the u.s. approximately 30% of patients with compensated cirrhosis have esophageal varices when first diagnosed. in cirrhotic patients, life prognosis in a patient with upper gi bleeding from esophageal varices depends on the severity of the bleeding, the hepatic functional reserve (stage of cirrhosis), the extent and location of the varices (esophageal or gastric), their age, the existence of associated diseases and the treatment prescribed. the aim of this study was to monitor the risk factors that trigger variceal bleeding in cirrhotic patients, to assess the severity of the bleeding and the efficacy of the endoscopic hemostasis techniques, to assess the relapses of bleeding episodes and the mortality rate. the study was a prospective one, and it was conducted in the period 11.200412.2006 in the o. fodor regional institute of gastroenterology and hepatology, cluj - napoca ; 273 patients with variceal bleeding underwent upper gastrointestinal endoscopy for diagnosis and treatment in emergency conditions, at the office of digestive endoscopy of the o. fodor regional institute of gastroenterology and hepatology, cluj - napoca ; the study included only patients who had upper gastrointestinal bleeding because of variceal rupture, which was assessed by emergency endoscopic examination, performed within 6 hours of the patient s admission ; after identifying the variceal source of the upper gi bleeding, endoscopic therapy was performed (elastic ligatures or endoscopic sclerotherapy with hypertonic glucose solution, depending on the possibilities under emergency conditions) ; during periodical check - ups, the risk factors for variceal bleeding were monitored in cirrhotic patients, along with the efficacy of endoscopic treatment techniques, the relapses of bleeding episodes, the number and cause of deaths of cirrhotic patients who experienced upper gastrointestinal bleeding because of rupture of varices ; the patients included in the study met the clinical, biochemical, endoscopic and ultrasound criteria for liver cirrhosis ; the etiology and the staging of the disease were documented according to the patient s observation charts ; the results of the study were processed using excel ; quantitative data were expressed as median se ; the study was approved by the local ethics board of the prof. the average age of the patients included in the study was 57.86 years, the extremes being represented by 26 and 82 years (figure 1). regarding the gender distribution of patients, there were 167 male patients (61.17%) and 106 female patients (38.83%). out of the 273 cases with variceal bleeding, 255 (93.4%) were cases of bleeding from esophageal varices and 18 (6.6%) were cases of gastric varices. thus, it occurs more frequently in patients with varices grade ii and iii. in our series there were 2 patients (0.73%) with bleeding from esophageal varices grade i, 131 patients (47.98%) with hemorrhage from esophageal varices grade ii and 122 patients (44.7%) with bleeding from varices grade iii. gastric varices hemorrhage was found in 18 patients (6.59%) (figure 2).. however, the risk of variceal rupture increases with the increased severity of the liver disease. thus, variceal bleeding occurred in 63 (23%) patients in child class a, 109 (40%) patients in child class b and 101 (37%) patients in child class c. mortality caused by variceal hemorrhage was 2.93% in the study group. death occurred in 8 patients as follows : 1 was in child class a, 2 were in child class b and 5 in child c class. hence, the mortality of patients with variceal hemorrhage is closely related to the child class. variceal bleeding was more frequent in patients with alcoholic liver cirrhosis - 140 cases (51.28%) ; the next highest frequency was in patients with c virus cirrhosis - 69 cases (25.27%), b virus liver cirrhosis - 21 cases (7.69%), followed by other etiologies - 43 cases (15.76%). some of the 273 patients with variceal bleeding also had significant comorbidities : liver carcinoma (45 cases), gastric cancer (2 cases), lung cancer (1 case), liver metastases (1 case), pregnancy 28 weeks (1 case), peritoneal carcinomatosis (1 case). initial variceal bleeding was arrested in 269 of the 273 patients admitted under emergency conditions, using endoscopic techniques for hemostasis therapy. the failure of the endoscopic techniques imposed the use of the sengstaken - blakemore probe in 4 patients. a number of 148 patients were treated by sclerotherapy and 125 patients by performing elastic ligatures. out of the 148 patients treated by sclerotherapy during the first bleeding episode, in 28 cases sclerotherapy was performed when the first bleeding relapse occurred, while in 16 cases elastic ligatures were performed. out of the 125 patients treated by elastic ligature during the first bleeding episode, in 9 cases sclerotherapy was performed when the first bleeding relapse occurred, while in 12 cases elastic ligatures were performed. three porto - caval shunt interventions were performed, in 3 male patients of 41, 46 and 71 years of age. each of these three patients presented three episodes of variceal bleeding. in the case of the 71-year old patient bleeding recurrence occurred in 65 (23.8%) patients of the study group, 43 men and 22 women (figure 3). the average age of those who experienced bleeding relapses was close to the average age of the group, which was 58.69 years. thirty - seven patients underwent endoscopic treatment by sclerosis of varices with 2 ml of hypertonic glucose in 812 points intra- and para - varices, while 28 patients underwent elastic ligatures (46 elastic rings). there were 17 cases of bleeding recurrence within 7 days of the first bleeding episode, 19 cases in the first month, 11 cases after 6 months and 18 cases after 1 year. the failure of emergency endoscopic treatment is defined as continued or recurrent bleeding within less than five days, in spite of the completion of two sessions of sclerotherapy. there are no predictors of the failure to indicate and require treatment change. in the study group there were 14 cases of bleeding relapses within the first 4 days of the endoscopic hemostasis. bleeding relapses were favored by the presence of hepatocarcinoma (26 patients out of the 65 patients with recurrent bleeding) and by the child class c (23 patients). in the present study, out of the 65 patients with recurrent bleeding, 13 were not treated with propranolol while 9 were treated with small doses of propranolol. alcoholic cirrhosis was the dominant etiology in patients with recurrent bleeding (36 cases - 55.38%), while the child class c was found in 22 patients (33.84%). the results of the current study show that bleeding from esophageal varices is more frequent than from gastric varices (93.4% versus 6.6%). mention must be made that the rupture of the varices depends on the size of the variceal veins, and thus rupture is more frequent in patients with esophageal varices grade ii and iii. the data obtained in the present study are in keeping or similar to those shown in the speciality literature. variceal bleeding was more frequent in male patients, the data obtained being in keeping or similar to those shown in the specialty literature. the variceal hemorrhage prevailed in each clinical class and the variceal rupture risk increased with the increase in the severity of the liver disease. thus, the majority of patients with variceal bleeding were child class b or c (77%), our results overlapping those mentioned in literature. variceal bleeding was more frequent in patients with alcoholic liver cirrhosis (51.28%), the results being similar to those reported in literature. in the current study, 19% of the patients who had variceal bleeding also had significant comorbidities, the most frequent being hepatocarcinoma which has been reported as a significant predictive factor for death in decompensated cirrhosis and early rebleeding in several studies. bleeding relapses were frequent (23.8%) and raised special problems related to the type of treatment (sclerotherapy or ligation), to the therapeutic booster in order to eradicate esophageal varices, to the adjuvant medication therapy for portal hypertension (beta - blockers) or to invasive procedures for the treatment of portal hyper - tension (tips) or liver cirrhosis (liver transplantation). bleeding recurred more frequently after sclerotherapy than after the application of elastic ligatures (29.72% versus 16.8%), the results of the current study being similar to those published in literature. approximately 34% of our patients experiencing bleeding relapses did not receive beta - blocker therapy or were given low - dose propranolol. for secondary prevention, a meta - analysis that compared beta - blockers plus isosorbide mononitrate versus endoscopic band ligation showed no significant differences between treatments in preventing rebleeding or in preventing deaths. however, in trials using a mean beta - blocker dose of less than 80 mg / day, endoscopic band ligation significantly reduced rebleeding compared with beta - blockers plus isosorbide mononitrate. in addition, earlier meta - analyses usually included trials in abstract form, of which complete data were not available. therefore, a further update of studies comparing endoscopic band ligation and pharmacological therapy should be performed. the study also showed that the severity and etiology of the liver disease (cirrhosis) may play a role in the risk for bleeding relapses. in the study group there were three porto - caval shunt interventions, each patient presenting three bleeding relapses during the period studied. mortality due to variceal hemorrhage was 2.93% in the study group, which was lower than that in the data published in the speciality literature. many studies have reported decreasing incidence and mortality rates of variceal bleeding by progress of treatment modalities. the significant decrease in mortality in other studies is probably due to the improvement in treatment modalities such as endoscopic treatment, transjugular intra - hepatic portosystemic shunt, and pharmacologic treatment including the use of prophylactic antibiotics and vasoactive drugs. the mortality of patients with variceal hemorrhage is closely related to the child class (62.5% - child c). however, the risk of variceal rupture increases with the increased severity of the liver disease. hence, the mortality of patients with variceal hemorrhage is closely related to the child class. variceal bleeding occurred more frequently in patients with alcoholic cirrhosis stage child c. in our study mortality was relatively low. hence, prevention methods against variceal rupture and the application of elastic ligatures are required as a means of treatment. | objectivesthe aim of this study was to monitor the risk factors that trigger variceal bleeding in cirrhotic patients, assess the severity of the bleeding and the efficacy of the endoscopic hemostasis techniques, as well as the recurrence of bleeding episodes and the mortality rate.materials and methodsthe current study was a prospective one, and it was conducted in the period november 2004 - december 2006 in the o. fodor regional institute of gastroenterology and hepatology, cluj-napoca.it included 273 patients with upper gastrointestinal bleeding because of variceal rupture, assessed by emergency endoscopy. the patients included in the study met the clinical, biochemical, endoscopic and ultrasound criteria for liver cirrhosis. its etiology and staging were documented from the patients observation charts.resultsout of the 273 cases of variceal bleeding there were 255 (93.4%) cases of bleeding from esophageal varices and 18 (6.6%) from gastric varices. variceal bleeding episodes were more frequent in patients with alcoholic liver cirrhosis (51.28%). most patients with variceal bleeding were in child class b or c (77%). mortality because of variceal hemorrhage was 2.93% in the study group. a number of 148 patients were treated by sclerotherapy and 125 patients with elastic ligatures. bleeding relapses occurred in 65 (23.8%) patients within the study group, 43 men and 22 women. variceal bleeding relapses were more frequent after sclerotherapy than after elastic ligatures.conclusionsvariceal bleeding occurred more frequently in patients with alcoholic cirrhosis stage child c. in the current study mortality was relatively low. the treatment of bleeding recurrence is more difficult, hence variceal rupture prevention and application of elastic ligatures represent a therapeutic necessity. |
when ischemic stroke occurs, motor function is one of the most important brain functions to be protected together with language, sensory, visual, and hearing function. since the most motor cortex and motor neuron pathway are located in the middle cerebral artery territory, infarction of this territory causes contralateral hemiplegia. therefore, to treat acute cerebral infarction, recanalization of occluded blood vessels should be performed, and neuroprotective therapies that prevent reperfusion injury should be performed. nevertheless, if it could not be treated within limited times, neurons could not be restored permanently. in such manners, the regeneration capacity of neurons is very low, and thus methods that could substitute neurons are required. therefore, studies that transplant diverse stem cells to the cerebral ischemic area and induce them to differentiate to neurons or to substitute neuronal function are ongoing. several studies reported that in adult rats, bmscs transplanted after cerebral infarction accelerated neuroplasticity and facilitated neuronal regeneration as well as functional recovery [27 ]. in other words, it has been reported that intravenous injection of bmscs reduced the cerebral infarct volume and improved motor functions, and cerebral infarct size could be reduced noticeably by the injection within 3 hours after mca occlusion (mcao). in addition, one - hour mcao rat models, similarly, when bmscs were injected to the ipsilateral carotid artery immediately after reperfusion, the cerebral infarct volume was reduced and motor function was improved. all investigators reported that as the cerebral infarct volume became smaller, the neurobehavior was improved more [911 ]. nevertheless, evaluation of the recovery of motor function was conducted with subjective neurobehavioral tests, determined and scored by examiners [911 ]. objective evaluation studies on the recovery of motor function in cerebral ischemic animals after transplantation of stem cells have not been conducted. therefore, for the objective evaluation of the recovered neuronal function after stem cell transplant, we applied meps representing the level of electrophysiological response. meps have been used to measure the motor nerve function in animals with lower limb paralysis at bmscs transplantation after spinal cord injury [1214 ]. however, it has been rarely used to measure the motor neuronal function in the ischemic rat brain [1517 ]. recently, it has been reported that in normal rats, by measuring the sensory - evoked potential, the corticomotor - evoked potential (cmep), and the brainstem - derived mep (bmep) serially, cmep was originated from the motor cortex. in addition, through monopolar as well as bipolar stimulation, meps originated from the brainstem could be measured during suprathreshold stimulation, and after focal stimulation of the motor cortex, the mep in the brain stem was measured, and thus studies on electrophysiological changes after reperfusion in transient ischemic animal models have been conducted [15, 20 ]. therefore, if meps with a different origin could be measured continuously through focal monopolar as well as bipolar stimulation of the motor cortex, the integrity of the motor pathway may be examined. in our study, in acute cerebral ischemic rats, electrophysiological effects on motor - neuron pathway of transplanted bmscs were assessed by measuring mep. all experimental protocols used in this study were designed according to animal guidelines established by the institutional animal care and use committee of the catholic university medical school. ten adult male sprague - dawley rats weighing 270 to 320 g were employed in the study. meps were measured in all animals of each group before surgery for establishment of comparative baseline waves. after transient mca occlusion surgery, experimental animals were assigned randomly to one of the following two groups : (1) the saline injection group as the control group (n = 5, control group) and (2) the bmsc injection group as the experimental group (n = 7, bmsc group). rats weighing 150200 g were sacrificed by 15% urethane ; bmscs were collected from femurs and tibias by flushing the shaft with 15 ml buffer (phosphate - buffered saline supplemented with 2% fetal bovine serum) using a sterile hypodermic syringe. cells were diluted with buffer up to 50 ml and centrifuged through a density gradient (ficoll - paque - plus ; 1.077 g / ml ; pharmacia) for 30 minutes at 1000 g. cells were plated at 7 10 cells / cm in a coating - culture dish and maintained in dulbecco 's modified eagle medium/20% fetal bovine serum. for cell - surface ag phenotyping, cells were detached and stained with fluorescein isothiocyanate or fluorescein phycoerythrin - coupled ab. labeled cells were analyzed by facscalibur flow cytometer (becton dickinson, san jose, ca, usa) using cellquest software (becton dickinson). for detection of surface ag, aliquots of bmscs were washed 3 times with pbs, ph 7.4, following treatment with 0.25% trypsin. for direct assays, cells were immunolabeled with antirat ab cd45 (fitc) and cd90 (pe). rats were anaesthetized using nitrous oxide / oxygen gas, 70 and 30%, respectively, administered through an inhalation mask. during all procedures, body temperatures were maintained under controlled conditions (37 0.4c) via the use of a rectal thermometer and a heating pad (harvard apparatus inc., holliston, ma, usa). each animal was placed in a stereotactic frame (narishige scientific instrument lab, tokyo, japan). an incision of about 1.5 cm was made in the scalp under asceptic conditions, and two holes were carefully drilled into the skull of each animal ; a 1.0 mm diameter drill bit (saeshin precision co., ltd., daegu, korea) was used in order to avoid direct injury by perforation of the dura mater. cerebral blood flow (cbf) in each occlusion model was measured using a transonic laser doppler (blf21 series, transonic systems inc. after measurement of cerebral blood flow, transient mca occlusion was induced using the intraluminal filament technique described by longa.. briefly, this involves making an approximately 2 cm ventral incision, exposing the right common carotid artery (cca), external carotid artery (eca), and internal carotid artery (ica). a 30 mm length 30 nylon monofilament (ethicon, johnson & johnson medical ltd., livingston, uk) with a heat - blunted round tip was inserted through a small opening in the eca stump and gently advanced into the ica until it blocked the bifurcating origin of the mca. the wound was closed temporarily to enable bmsc injection following withdrawal of the inserted nylon through the eca after an occlusion period of 90 minutes. each animal was then fixed again in a stereotactic frame, and the scalp wound was reopened. 35% of the levels recorded prior to mca occlusion (26.5 5.73%). one screw (coronal) was positioned at the following coordinates : 2.5 mm right from midline and 1 mm anterior to the bregma over the forelimb area of the right motor cortex [22, 23 ]. the other (posterior) screw was positioned 12 mm posterior to the lambda. commercial stainless steel screws (10 mm in length with a 1.2 mm outer diameter) were implanted as the anode for elicitation of mep at the burr hole site at a depth of 1.5 mm (figure 1). conshohocken, pa, usa) with a diameter of 0.4 mm (27 g) and a needle length of 12 mm. however, the uninsulated part of the needle tip proved too long for insertion into the forelimb and hind limb muscles ; therefore, we insulated the tip using a rubber tube to expose just 3 mm of the tip. one pair of 1 mm uninsulated electrodes was used to record the motor evoked potential at the brain stem (bsmep), and the interpolar distance was 2 mm. intra - arterial injection of bmscsafter a 90-minute period of mca occlusion, the wound was reopened, and the cca and ica were loosely ligated with 40 silk to prevent bleeding from the eca opening. the 30 monofilament was then gently withdrawn from the eca opening through the ica lumen, and 100 l of saline containing 1 10 cells was introduced using a hamilton syringe (harvard apparatus, holliston, ma, usa) to which a polyethylene (pe) 10 tube (harvard apparatus, holliston, ma, usa) was connected. the pe 10 tube was then advanced into a small opening of the eca to the lumen of the ica to a depth of approximately 15 mm, and blood was retrieved through the pe 10 tube to confirm a connection with the mca. stem cells were slowly injected into the lumen, and the blunt needle was withdrawn from the eca. the eca was ligated with 40 silk, and pulsation of the ica was confirmed. after injection of stem cells or saline, reperfusion was identified by measuring the cortical blood flow with a transonic laser doppler in both groups (62 10.4%). after a 90-minute period of mca occlusion, the wound was reopened, and the cca and ica were loosely ligated with 40 silk to prevent bleeding from the eca opening. the 30 monofilament was then gently withdrawn from the eca opening through the ica lumen, and 100 l of saline containing 1 10 cells was introduced using a hamilton syringe (harvard apparatus, holliston, ma, usa) to which a polyethylene (pe) 10 tube (harvard apparatus, holliston, ma, usa) was connected. the pe 10 tube was then advanced into a small opening of the eca to the lumen of the ica to a depth of approximately 15 mm, and blood was retrieved through the pe 10 tube to confirm a connection with the mca. stem cells were slowly injected into the lumen, and the blunt needle was withdrawn from the eca. the eca was ligated with 40 silk, and pulsation of the ica was confirmed. after injection of stem cells or saline, reperfusion was identified by measuring the cortical blood flow with a transonic laser doppler in both groups (62 10.4%). meps were recorded using the cadwell cascade (cadwell laboratories, inc., kennewick, wa, usa). after a fast of 12 hours, a dosage of ketamine 60 mg / kg and xylazine 5 mg / kg was administered intraperitoneally to rats for induction of anesthesia. the level of anesthesia was assessed by scoring of the withdrawal reflex described in a previous report. this index assigns a score of 16 based on factors such as level of muscle contraction and limb withdrawal as well as flexion. a withdrawal reflex score below 3 was required before fixing any animal on the stereotaxic frame. monopolar electrical stimulationmbmeps were evoked by a single, short train of supramaximal intensity impulses (stimulation type, tcs-1 ; pulse width, 50 s ; 620 ma ; 0.5 hz ; interstimulus interval 2 milli seconds, maximal intensity 20 mv) applied via the coronal screw (anode) referenced to the electrode around the nasion (cathode). latency was defined as the interval from administration of the electrical stimulation to the starting point of initial deflection (figure 3(a)). mbmeps were evoked by a single, short train of supramaximal intensity impulses (stimulation type, tcs-1 ; pulse width, 50 s ; 620 ma ; 0.5 hz ; interstimulus interval 2 milli seconds, maximal intensity 20 mv) applied via the coronal screw (anode) referenced to the electrode around the nasion (cathode). latency was defined as the interval from administration of the electrical stimulation to the starting point of initial deflection (figure 3(a)). an additional midline skin incision was made over the neck, and the posterior atlantooccipital membrane was exposed in order to enable placement of mep recording electrodes. signals were filtered at a bandpass frequency of between 10 and 3,000 hz and stored for later computer analysis. electrical stimuli were discharged at 30-second intervals in order to avoid signal deterioration [25, 26 ]. amplitude (mv) was defined as the height from negative peak to positive peak. latency was defined as the interval from administration of electrical stimulation to the starting point of the initial deflection. meps were evoked by a single short train of low intensity impulse (stimulation type : electrical ; pulse width : 100 s ; stimulation intensity : 0.0510 ma ; repetition rate : 15.1 hz) applied via the coronal screw (anode) referenced to the posterior screw (cathode) in the manner used by schlag.. bcmep was elicited by focal bipolar stimulation of the cortex with two cranial screws and recorded from the contralateral forelimb. stimulation intensity was increased by 0.5 ma and was stopped when both bbmeps were recorded or movement of both forelimbs developed (figure 4). an additional midline skin incision was made over the neck, and the posterior atlantooccipital membrane was exposed in order to enable placement of mep recording electrodes. signals were filtered at a bandpass frequency of between 10 and 3,000 hz and stored for later computer analysis. electrical stimuli were discharged at 30-second intervals in order to avoid signal deterioration [25, 26 ]. amplitude (mv) was defined as the height from negative peak to positive peak. latency was defined as the interval from administration of electrical stimulation to the starting point of the initial deflection. meps were evoked by a single short train of low intensity impulse (stimulation type : electrical ; pulse width : 100 s ; stimulation intensity : 0.0510 ma ; repetition rate : 15.1 hz) applied via the coronal screw (anode) referenced to the posterior screw (cathode) in the manner used by schlag.. bcmep was elicited by focal bipolar stimulation of the cortex with two cranial screws and recorded from the contralateral forelimb. stimulation intensity was increased by 0.5 ma and was stopped when both bbmeps were recorded or movement of both forelimbs developed (figure 4). daily adhesive - removal tests and treadmill tests were performed in 10 adult rats for 5 days prior to cranial screw implantation. after mcao, behavior tests were conducted at 1- and 7-day follow - up. adhesive - removal testtwo small adhesive paper dots (12 mm diameter) as bilateral tactile stimuli were firmly attached to each wrist of the forelimbs of the rat so that they covered the hairless part of the forepaw.the time required to remove both paper dots from each limb was recorded in five trials per day for 3 days. all animals can remove the dot within 10 seconds (sec.) at the end of training. three trials were conducted after ischemic stroke, with a cutoff time of 180 seconds. two small adhesive paper dots (12 mm diameter) as bilateral tactile stimuli were firmly attached to each wrist of the forelimbs of the rat so that they covered the hairless part of the forepaw. the time required to remove both paper dots from each limb was recorded in five trials per day for 3 days. all animals can remove the dot within 10 seconds (sec.) at the end of training. three trials were conducted after ischemic stroke, with a cutoff time of 180 seconds. treadmill (panlab, barcelona, spain) acceleration was 5 to 80 cm / sec within 4 minutes, and the end of the test was 85 cm / sec. treadmill (panlab, barcelona, spain) acceleration was 5 to 80 cm / sec within 4 minutes, and the end of the test was 85 cm / sec. maximum velocity of tolerance for each animal was checked 3 times. at 7 days after mcao, all rats (n = 5 and 4 in each group) were anesthetized with 15% urethane and sacrificed by decapitation. the brain was immediately removed and sectioned into 2 equally spaced, 2 mm coronal blocks using a rodent brain matrix. these sections were stained with 0.1 m pbs containing 2% solution of 2 - 3 - 5-triphenylterazolium (ttc ; sigma, st. louis, mo, usa) for 15 minutes at 37c. scanned brain images were quantified using meta - morph imaging software (molecular devices inc, downingtown, pa, usa). to reduce errors due to cerebral edema, the infarct area in each slice was corrected by subtracting the ischemic lesion site hemisphere area from the contralateral hemisphere area. seven days after mcao, three animals into which pkh26-labeled stem cells had been injected were sacrificed for histological examination. another was anesthetized with 15% urethane and then perfused transcardially with 0.01 m pbs (ph 7.4), and with 4% paraformaldehyde (pfa) in 0.01 m pbs. postfixed tissue was cryoprotected in 0.1 m phosphate buffer (ph 7.4) containing 10, 20, and 30% sucrose solutions, respectively, at 4c. brain tissues were embedded in tissue - tech oct compound (ps002) and stored at 70c. for immunohistochemical staining of bmsc - injected brain lesions, after cleaning the brain with a pbs solution, nuclei were stained with 4, 6 diamidino-2-phenylindole (dapi ; sigma). statistical analysis was performed using commercially available software (pasw statistics 16.0 ; spss inc., behavior tests, cerebral infarct volume, and mep parameters in each group were subjected to one - way anova with posthoc analysis, independent t - test, or mann - whitney u test. size of infarcts was evaluated by measurement of infarction volumes at 7 days after mcao. rat brains were stained with ttc in order to obtain the infarction volume, which was calculated by measurement of the area of the infarcted region. infarction volume of the bmsc group decreased significantly compared with that of the control group (control : 393.18 155.4 ; bmsc : 123.1 55.0, prior to mcao, neurological scores among the 10 animals were similar. at postoperative day 1, the performance of the bmsc group in the adhesive removal test was superior to that of the control group (control group : 174.3 10.2, bmsc group : 74.5 12.6, p <.05). at day 7, the difference between the two groups was significant (control group : 149.85 50.3, bmsc group : 26.7 4.7, p <.05). however, in relation to the treadmill test, there were no significant differences between the two groups at postoperative days 1 and 7 (figures 2(b) and 2(c)). in the case of bipolar electrical stimulation, meps following anesthetic injection were recorded at intervals ranging from 24 to 42 minutes (36.4 5.2). meps produced by monopolar electrical stimulation were then recorded (45.7 8.5 minutes). latency of left forelimb mbmep was 4.0 0.4 milli seconds, and the amplitude of mbmep was 3,770 1,518 v. no significant difference was observed between right and left sides in the preoperative mep parameters of all animals of both groups (table 1, figure 3). at postoperative day 7, mean mbmep values between the control group and the bmsc group showed significant differences in left forelimb amplitude (p =.013) and right forelimb latency (p =.021) (table 2). figure 4(c) shows a characteristic mbmep signal with a conspicuous reduction of amplitude of left forelimb mep recorded in the control group at day 7 after mcao. in contrast, the bmsc group showed relatively larger amplitude of left forelimb mep (table 2, figure 3(d)). in comparison with preoperative baseline meps, the control group at postoperative day 7 showed a significant reduction in the amplitude of mbmep and prolonged latency in the left limbs, although not in amplitude in the left hind limbs. the bmsc group, however, displayed significant prolongation of latency at both the left forelimbs and the hind limbs (p <.05) (table 3). focal bipolar stimulation of the cortex via the cranial screw resulted in movements of the contralateral forelimb. on gradual increase of stimulation intensity, the current interval required to elicit bcmep ranged from 0.1 to 12.0 ma (7.1 2.9). however, a pure bcmep occurred at a current interval ranging from 0.5 to 5.0 ma (2.9 1.4) (figure 4(a)). movement in both forelimbs was accompanied by the appearance of short - latency peak waves. d and i waves were produced at a 1.0 ma threshold (figure 4(a)(c)) in response to gradual stimulation. larger d and i waves were produced at greater stimulation intensities (figure 4(a)). at a different current intensity, d wave could be measured in each group (preoperative baseline, 3.6 1.9 ma ; control group, 5.8 1.2 ma ; bmsc group, 5.0 0.3 ma). preoperative baseline amplitudes of d and i waves were 59.9 18.9 and 36.1 16.9 v, respectively (table 4). with regard to latency, mean values of d and i waves in the normal group were 2.4 0.7 milli seconds and 5.6 1.8 milli seconds, respectively (table 4). at day 7 after stroke, the control group showed no bcmep, except for a vestige signal in one animal (figure 4(b)). bsmep of this group contained larger amplitudes of d waves than in the normal group (116.5 25.4 v) ; however, no i wave was detected at sub- and supramaximal thresholds (figure 4(b)). control group at postoperative day 7 had a trend with larger amplitudes of d wave (195.6 70.6 v, p =.06) than those of the bmsc group ; however, no differences in latency were observed between the two groups (table 4). in two of five rats, i wave and bcmep turned up and were measured (figure 4 (c)(g) and (i)). in our experiment, we aimed to determine the distribution of bmscs in the motor cortex following intra - arterial injection of pkh26-labeled bmscs. at 7 day after injection of bmsc, sectioned rat brain around 1 mm anterior to the bregma abundant pkh26-labeled cells were detected using fluorescence microscopy at the left forelimb primary motor cortex in the infarction hemisphere (figure 5(c)) and external capsule in the contralateral hemisphere (figure 5(d)). this rat was proved to be one of two rats whose i wave and bcmep were restored. in additional two rats, the presence of pkh26-labeled cells in the right mortor cortex was identified. in this study, the electrophysiological effect of bmsc transplanted during reperfusion to acute phase cerebral ischemia rats through the carotid artery on the motor cortex and the motor nerve pathway was examined by measuring the mep one week after transplantation of stem cells. at 7 days after surgery, the cerebral infarct volume of the bmsc group was significantly decreased in comparison with the control group (figure 2(a)). together with such histological effects, based on the serial record of mbmeps, bcmeps, bbmeps, and bsmeps in response to monopolar as well as bipolar stimulation, it could be proven that the bmsc group was associated with the partial recovery of the long - lasting synaptic transmission defect of the motor cortex in comparison with the control group. konrad and tacker have reported that the transcranial mep generated in response to monopolar suprathreshold stimulation was measured at the 2nd lumbar vertebra and the latency was lower than 3.5 milli seconds. have reported that the latency of mep with a distinct negative peak measured in hind limbs was 57.5 milli seconds. our preoperative baseline latency of mbmeps measured at forelimbs and hind limbs (3.9 0.4 ms, 7.3 0.5 ms) concurred to other studies, and the amplitude was larger (table 1). in addition, after focal bipolar stimulation, the latency of bbmeps measured in the left forelimbs was 4.2 0.7 milli seconds, which was shorter than the latency of mbmep measured in the hind limb (7.3 0.5 ms), and slightly longer than the latency of mbmep measured in the forelimbs (3.9 0.4 ms) (table 4). this phenomenon is thought to be due to the fact that the stimulation intensity of our study was stronger than that of other studies, and monopolar stimulation was stronger than bipolar stimulation, and thus the latency became shorter, and such phenomena have been reported in other studies. therefore, mbmep and bbmep could be considered to be the evoked potentials originated from the brainstem. on the other hand, schlag. transected the corticospinal tract of spinal cord and measured the cmep in the hind limbs and proved that the origin of this mep generated in response to focal bipolar stimulation is the motor cortical area, based on the observation that cmep disappeared and only bmep was recorded. in our study, the latency of bcmep of left forelimbs in response to focal bipolar stimulation was 15.1 2.3 milli seconds, which was shown to be shorter than the latency of cmep (1722 ms) measured in hind limbs by schlag., and because of the delay comparable to the difference of the latency of the mbmeps of hind limbs and forelimbs (3.4 ms), bcmeps were proven to be originated from the motor cortex. furthermore, when the intensity of bipolar stimulation was increased gradually, in response to low currents, bcmeps were generated first, and simultaneously, only the movement of left forelimbs was observed. when the stimulation intensity was increased more, bcmep following bbmep was observed, and the simultaneous movement of both forelimbs was observed, which also supports that bcmep was originated from the motor cortex. the bsmep measured in the atlantooccipital membrane after bipolar stimulation consisted of the d wave and the i wave, similar to the studies reported by amassian. and the preoperative baseline latency of the d wave was 2.4 0.7 milli seconds, which was slightly longer than results of other studies (1.81 0.11 ms) [15, 20 ]. in addition, the threshold of the generation of bsmep was 1.0 ma, which was comparable to results of previous studies [15, 20, 29 ]. moreover, the difference of the beginning of d wave and the beginning of i wave was 3.2 milli seconds, and this value comparable to the result reported by amassian. (3.5 ms) was shown. in our study, it was epidural stimulation by screws, and in other studies, it was intracortical stimulation, and thus the latency was different. combined together, bsmep is considered to be originated from the motor cortex. on the day 7 after reperfusion, the mbmep latency of the two groups became significantly longer than the preoperative baseline values. the mbmep amplitude in left forelimbs of the bmsc group was longer significantly in comparison with that of the control group (table 3). many investigators believe that meps generated in response to suprathreshold monopolar or bipolar electrical stimulation are developed in the brainstem and associated with the activation of the extrapyramidal tract [18, 19, 27, 3032 ]. since it is difficult to induce cerebral infarction in the brainstem by mcao because of the distribution of blood vessels, the amplitude of mep corresponding to the extrapyramidal tract associated with the left forelimb of the bmsc group was decreased relatively less, which may imply that the nerve conduction into the extrapyramidal tract at the upper levels in the bmsc group was recovered more than that in the control group. such phenomenon could be explained with the enhanced corticofugal plasticity after the development of a unilateral lesion in the extrapyramidal tract in the brain [31, 33 ]. andrews. reported that treatment with human adult bone marrow - derived somatic cell after ischemic stroke in adult rats results in recovery of forelimb function, which is positively correlated with increased axonal outgrowth of the intact, uninjured corticorubraltract. in immunofluorescent staining performed on the day 7, stem cells were detected even in the contralateral external capsule, and thus the effect on the corticofugal plasticity of the ipsilateral side as well as the contralateral side could be considered. on the other hand, the mbmep latency of the right forelimb in the bmsc group was significantly longer than that in the control group, which was thought to be due to that the stimulation intensity in the bmsc group was relatively stronger than that in the control group, and even if the stimulation intensity was considered, the amplitude difference of the left forelimbs of the two groups was significant. such relative recovery could be explained better in bipolar stimulation, and in regard to bsmep measured in the atlantooccipital membrane, the amplitude of d waves of the bmsc group and the control group was significantly increased than the preoperative baseline values, and the tendency that the amplitude of the bmsc group was lower than the control group was shown ; nonetheless, it was not significant (p =.06). in addition, concerning the bmsc group, the i wave and the bcmep measured in the left forelimbs were observed in 40%. it could be considered that such recovery of bsmep and bcmep in the bmsc group may imply the relative recovery of the subcortical and cortical area. claimed that the d wave is the spike trigger zone (initial segment) present in the axon, it is generated by the first or deeper nodes in the white matter, or the excitation of the branch formation of the axon collaterals in the gray matter, and the i wave is generated in response to the excitement of the specific thalamocortical projection, corticocortical projection, or intrinsic, tangentially oriented fibers. in addition, bolay. have reported that during reperfusion after temporary mcao, the d wave was recovered but the i wave was not recovered, the amplitude of d wave became larger after reperfusion, and the mep in the contralateral paralyzed muscles was not observed [15, 20 ]. therefore, such recovery of the i wave and bcmep implies that bmsc transplant acts to protect the neurons in the cortical area, and as shown in figure 5, bmsc transplant to the motor cortical area corresponding to forelimbs facilitated the recovery of nerve conduction. in addition, the amplitude of d wave of the two groups was increased in comparison with the preoperative baseline values, which implies that the axonal excitability caused by cerebral infarction is increased, which could be explained as the decrease of the inhibitory synapses in the cortical area. therefore, the tendency that the bmsc group showed smaller amplitudes than the control group could be considered to indirectly demonstrate the possibility of the relative recovery of the cortical area. in our study, in adhesive removal tests, differences between the bmsc group and the control group were shown ; nonetheless, differences in the treadmill test were not detected, which suggests the association with our mep results. in other words, the results of mbmep, bsmep, and cmep indirectly suggest that the adhesive removal tests assess primarily the sensation and skilled motor function of forelimbs, and thus significant differences were detected, and the treadmill test is associated with the motor ability of all four limbs. such differences are considered due to the collateral circulation through the anterior cerebral artery after mcao. in addition, the i wave and bcmep were observed after bmsc transplantation, and thus the possibility of the migration of bmscs to the motor cortical area of forelimbs through the anterior cerebral artery could not be ruled out. the latency of d wave of both groups became shorter, but they were not significantly different, which implies that the distance from the area of the development of d wave to the atlantooccipital membrane was comparable, which shows that the area of the initiation of d wave is similar. such fact indirectly suggests that up to the day 7 after bmsc transplantation, it could not mediate direct effects to show electrophysiological improvement in subcortical area. our limitation was that we could not demonstrate the neuroprotective mechanism of bmsc transplatation. in conclusion, by measuring mep, we were able to imply that bmsc transplanted to the transient ischemic rat brain could reduce the cerebral infarct volume as well as mediate electrophysiological effects on the motor cortical area and the motor pathway. in addition, it is thought that to elucidate the mechanism of transplanted bmscs mediating effects on the motor neuron, more studies are required. in the future, in stem cell therapy on the ischemic rat brain, mep may become an objective marker of the functional recovery. | this study investigated the effect of bone marrow mesenchymal stem cells (bmscs) on the motor pathway in the transient ischemic rat brain that were transplanted through the carotid artery, measuring motor - evoked potential (mep) in the four limbs muscle and the atlantooccipital membrane, which was elicited after monopolar and bipolar transcortical stimulation. after monopolar stimulation, the latency of mep was significantly prolonged, and the amplitude was less reduced in the bmsc group in comparison with the control group (p <.05). meps induced by bipolar stimulation in the left forelimb could be measured in 40% of the bmsc group and the i wave that was not detected in the control group was also detected in 40% of the bmsc group. our preliminary results imply that bmscs transplanted to the ischemic rat brain mediate effects on the functional recovery of the cerebral motor cortex and the motor pathway. |
stroke refers to cerebrovascular bleeding and a neural disease caused by nontraumatic reasons and is accompanied by brain damage or functional loss of lower body muscles on the paretic side1, 2. despite the development of modern medicine, economic growth, and widening interests in health, its occurrence is increasing quickly due to unbalanced lifestyles, environmental pollution, and excess stress in fast - paced societies1, 2. it has been reported that loss of motor ability and balance due to stroke can limit some daily living activities such as walking and may lead to secondary injury like falls. generally, gait in stroke patients is characterized by reduced gait parameters3,4,5, and decreased gait function following stroke in particular is known as the fundamental reason why patients fail to safely return to society6,7,8. yang.9 reported that because hemiplegia patients with lesions support more than 60% of the load on the non - paretic limb during standing, it could result in an asymmetric gait pattern. they also reported that a stroke patient group showed lower gait velocity, stride time, stride length, and cadence compared with a group of healthy elderly. grasies10 and vattanasilp.11 showed that patients with stroke tend to reduce the range of motion (rom) of paretic lower limb joints during walking, while fonseca.12 reported that increased stiffness of the hip joint can reduce energy efficiency, which results in limited walking and running movement. in addition, turnbull.13, who analyzed the gait cycles of eight elderly hemiplegia patients for ten years, reported that the patients showed increased double support time and decreased single support time during walking, indicating that patients tended to walk as slowly and stably as possible to secure stability. as reported from previous studies, the predominant characteristic of hemiplegia patients with stroke is the difficulty with walking due to weakened lower limb muscles and imbalance. therefore, many researchers have conducted studies focusing on the recovery of gait ability as the ultimate purpose of rehabilitation of patients with stroke14,15,16. pilates exercise is also called contrology and it is based on the idea of muscle control. it focuses to make a neutral spine of body to prevent excess flexion and extension of the spine when walking upright17, 18. since pilates exercise has been shown to have positive effects on development of muscular strength and endurance and improvement of flexibility, it has been used not only for exercise programs for healthy adults but also for rehabilitation for the elderly19, 21. moreover, unlike other exercise programs, pilates exercise can be performed with various tools to adjust the fitness level for the elderly, who have weaker physical strength and it can be performed at home without having to visit a rehabilitation center22, 23. as pilates is reported to develop the deep core muscles, which helps to improve the stability of spine, reduce back pain, and control the pelvis and hip joints24,25,26,27,28,29,30, it is thought to have a positive influence on gait ability, and many researchers have actually reported the effectiveness of pilates exercise on gait ability for the elderly17, 18, 21. despite the various positive effects of pilates, however, it is still not widely applied to stroke patients. therefore, the purpose of this study was to investigate the effects of an 8-week program of pilates exercise on gait in chronic hemiplegia patients and to determine whether or not it can be used for rehabilitation in poststroke patients. twenty poststroke participants (age, 66.1 4.4 yrs ; height, 162.3 8.3 cm ; weight, 67.4 12.3 kg) were recruited for this study. the exclusion criteria for the poststroke participants included moderate / severe chronic white matter disease on mri or orthopedic and other gait - influencing diseases such as arthrosis or history of lower - extremity joint replacement. participants who were involved in other studies or rehabilitation programs were also excluded from this study. the objective and requirements of this study were explained to all participants, and written informed consent was obtained from each participant prior to participation. the university s institutional review board approved the study protocol. after completing participants selection, the participants were randomized into two matched number groups : the pilates exercise group (eg) and control group (cg). during the intervention period, the eg performed pilates exercise, while the cg did not perform any kind of exercises or receive any treatment. the pilates exercise program used in this study was conducted by two certified pilates instructors and one physical therapist three times a week, 60 minutes per session, for 8 weeks. the exercise was composed of warm - up exercise, the main exercise, and cool - down exercise. for improving core muscles stability, breathing exercises were conducted in a sitting posture before and after the main exercise. the details of the program are provided in table 1table 1.pilates exercise programprogramwarm - up exercise1. breathing : 8 sets10 min2. spine stretch side : 8 setsexercise2. draw a sword : 8 sets40 min3. swan : 8 sets. in order to investigate the effects of the 8 weeks of pilates exercise, a 3-d motion analysis with 8 infrared cameras (oqus, qualisys, sweden) was performed twice (a week before and after the pilates exercise period). the subjects were required to wear black spandex shorts, a dark t - shirt, and a pair of walking / running shoes during data collection. to identify lower limb movements, 12 reflective markers were attached to the lower body. after sufficient warm - up, each subject was asked to walk on a treadmill (instrumented treadmill, bertec, usa), and his / her walking trials were captured with sampling rate of 100 hz. the cameras were positioned around the treadmill for sufficient tracking redundancy and were calibrated using the nlt (non - linear transformation) method. the treadmill speed was set at each subjects preferred speed which was measured before the experiment. each subject was recorded while walking for 30 seconds, and five strides from the middle of the recording were used for analysis. to evaluate the asymmetric pattern of gait parameters, the asymmetry index proposed by robinson.31 was used. xparetic and xnon - paretic are the values of a gait parameters recorded for the paretic and non - paretic limbs, respectively. to verify the effect of the 8-week pilates exercise on the gait of poststroke patients, the two - way anova with repeated measure was used, and statistical significance was set at =0.05. the gait parameters and asymmetry indexes before and after the 8 weeks of exercise are presented in tables 2 and 3table 2.kinematic gait parameters between exercise periodsvariablescontroltrainingpre - trainingpost - trainingstride length (cm)33.25 12.9043.91 19.35stride time (s)1.52 0.361.40 0.32stride velocity (cm / s)21.54 3.4131.48 12.81step length (cm)paretic17.22 4.2424.63 8.25non - paretic16.03 8.6919.28 11.28hip rom (deg)paretic22.55 5.9122.77 6.84non - paretic26.01 10.4330.61 5.47knee rom (deg)paretic25.30 12.5520.38 3.08non - paretic48.39 8.8952.03 8.72ankle rom (deg)paretic12.14 2.8311.67 6.76non - paretic14.69 2.3919.96 6.07values are group mean standard deviation. significant difference between pre- and post- pilates training. significant difference between groups (p0.05). significant difference between groups (p0.05, table 2). the lack of statistically significant improvements in asymmetry indexes was probably the result of a limited number of subjects or a short exercise period. in this study, we chose eight weeks as the exercise period because many previous intervention studies for elderly have shown the effectiveness of 8-week interventions. as pilates exercise strengthens the deep muscles, however, it is thought to require a longer exercise period compared with resistance training, cardiovascular exercises, or underwater exercises, which strengthen the superficial muscles. this issue should be examined in a follow - up study. in conclusion, this study proved the effect of pilates exercise, which has not been used as a means of intervention to rehabilitate motor functions of poststroke patients. the 8-week program of pilates exercise had a positive influence on improving the gait ability of poststroke patients, and the intervention could be applied to poststroke patients with various levels of physical disability by adjusting the intensity of exercise. | [purpose ] the purpose of this study was to investigate the effects of an 8-week program of pilates exercise on gait in chronic hemiplegia patients and to determine whether or not it can be used for rehabilitation in postsrtoke patients. [subjects and methods ] twenty individuals with unilateral chronic hemiparetic stroke (age, 66.1 4.4 yrs ; height, 162.3 8.3 cm ; weight, 67.4 12.3 kg) participated in this study and were randomly allocated equally to either a pilates exercise group or a control group. to identify the effects of pilates exercise, a 3-d motion analysis with 8 infrared cameras was performed. [results ] for the gait parameters, improvements were found in the pilates exercise group for all variables, and statistical significance was observed for stride length, gait velocity, knee range of motion and hip range of motion. for the asymmetry indexes, insignificant improvements were found for all variables in the pilates exercise group. [conclusion ] in conclusion, an 8-week program of pilates exercise had a positive influence on improving the gait ability of poststroke patients, and the intervention could be applied to poststroke patients with various levels of physical disability by adjusting the intensity of training. |
many women use natural health products for relief of menopausal symptoms and perceive these products as safe.phytotherapeutic products for menopausal complaints can contain plants with phytoestrogenic properties.the use of phytoestrogens can be related to endometrial hyperplasia and subsequent vaginal hemorrhage. several treatment options exist for the treatment of menopausal symptoms in women. the use of hormone - replacement therapy (hrt) has decreased substantially after publication of the women s health initiative (whi) trial in 2002 [13 ]. many post - menopausal women often perceive phytoestrogens in food supplements as a safer alternative than hrt, because these products are phytoestrogens represent a diverse group of non - steroidal natural products that have some estrogenic effects and are often marketed as food supplements. phytoestrogens are secondary metabolites that can mimic or modulate the actions of endogenous estrogens, usually by binding to estrogen receptors (ers). the three major classes of phytoestrogens found in typical human diets are isoflavones (such as genistein and daidzein), which are concentrated in soybeans and soy products, but are also found in other legumes ; lignans, which are distributed in seeds, whole grains, berries, fruit, vegetables, and nuts ; and coumestans, which are found in broccoli and sprouts. the flavonoid substance 8-prenylnaringenin (hopein) in hop also has phytoestrogenic properties. of all phytoestrogens, 8-prenylnaringenin has the strongest er activity [5, 6 ]. in the period from november 2011 to april 2015, the netherlands pharmacovigilance centre lareb received seven reports of vaginal hemorrhage caused by endometrial hyperplasia related to the use of the same hop - containing phytotherapeutic product (menocool). the first three cases were reported by the same gynecologist and published in a dutch medical journal in 2012. since then, lareb has received four more cases concerning the same association : two from gynecologists from different hospitals in the netherlands and two directly from a patient. patient a, a 54-year - old woman (weight 79 kg, height 177 cm, body mass index [bmi ] 25.2 kg / m) had been using the phytotherapeutic product menocool (two times daily ; morning one tablet, evening 0.5 tablet per oral administration) for the indication hot flushes and vaginal dryness due to the menopause. she had bought the product online through the manufacturer s website (http://www.menocool.nl)., she had not menstruated for 2 years but had experienced hot flushes for approximately 4 years. the patient had never had an abnormal cervix smear, except a pap iii result 20 years previously. she had no medical history of vaginal bleeding, endometrial hyperplasia, endometrial polyps, or uterine leiomyomas. her concomitant medication consisted of fexofenadine 120 mg once daily and mometasone 50 g / dosage nasal inhalation spray, both since 2005. the patient started using menocool in april 2014. after using menocool for 2 months, the patient started to experience abdominal cramps and vaginal hemorrhage. these complaints were diagnosed by a gynecologist as related to a high proliferation of the endometrium. after the curettage, the patient went for a follow - up appointment to her gynecologist and a significant increase in the endometrium was found again. norethisterone has been effective in the treatment of dysfunctional uterine bleeding in patients with a proliferative or hyperplastic endometrium and, as expected, the patient had a heavy withdrawal bleeding following withdrawal of norethisterone that lasted for several days. the following control appointment showed proliferation of the endometrium again but still within the normal range ; this time no action was deemed necessary. according to the patient, the gynecologist in the hospital had no explanation for the rapid and extreme growth of the endometrium. the patient ceased the use of the phytotherapeutic product in september 2014 and had fully recovered by the time follow - up information was received by the pharmacovigilance center in february 2015. after a consultation in a women s health clinic, the patient was told by another gynecologist that she should not use drugs against hot flushes containing phytoestrogens to prevent a repeat of her complaints. her endometrium was diagnosed at this clinic as thick but within the normal range, and there has been no hyperplasia of the endometrium or vaginal bleeding since. patient b, a 54-year - old woman (weight 70 kg, height 176 cm, bmi 22.6 kg / m), had been using menocool (1.5 tablets daily) for the indication hot flushes due to the menopause. she also bought the product online, through the same website as patient a. the patient had not previously used the suspected drug. her concomitant medication consisted of another supplement from the same manufacturer : menocool extra oral lyophilisate (2 tablets daily) containing a non - estrogenic flavone extract 22 mg, vitamin d3 5 g, biotine 25 g. in addition, she had been using alendronic acid 70 mg once weekly since 2012. the menocool extra oral lyophilisate was added shortly after the start of menocool, following the advice of the manufacturer, because the patient experienced abdominal pain / cramps. a cervix smear, internal examination, and ultrasound were performed. due to the thickness of the endometrium the use of menocool was ceased on 2 april 2015 ; on 13 april 2015 follow - up information received from the patient indicated that she had almost entirely recovered from the abdominal pain / cramps and vaginal hemorrhage. according to the product s manufacturer (standby vital bv, based in the netherlands), the phytotherapeutic product menocool consists of a combination of three kinds of hops (humulus lupulus) selected for their flavonoid content, a natural isoflavones complex, buckwheat (fagopyrum esculentum), black oat (avena sativa), malt (semen hordei vulgaris germinatum), barley (hordeum vulgare), rye (secale cereale), wheat (triticum aestivum), corn (zea mays), and dietary fiber. exact ingredients of the product, as specified by the manufacturer, are given in table 1.table 1ingredients of the product menocool as described by the manufacturer (tablet weight 1010 mg)ingredientspercentagehop (humulus lupulus)41.40dietary fiber9.90buckwheat (fagopyrum esculentum)8.48black oats (avena sativa)8.48malt (semen hordei vulgaris germinatum)8.48rye (secale cereale)4.24barley (hordeum vulgare)4.24wheat (triticum aestivum)4.24corn (zea mays)4.24silicon (as an excipient for hair and nails)4.10natural isoflavones complex1.80vegetable magnesium stearate0.40 ingredients of the product menocool as described by the manufacturer (tablet weight 1010 mg) vaginal bleeding is not uncommon in post - menopausal women and occurs in approximately 411 % of women who have reached menopause [9, 10 ]. the incidence of bleeding appears to correlate with time since menopause, with the likelihood of bleeding decreasing over time. recurrence of bleeding is very common in the time immediately after menopause, being the 12 months of amenorrhea after what is currently defined as the final menstrual period, declining to low frequencies more than 3 years after the final menstrual period. the differential diagnosis of bleeding in post - menopausal women can include endometrial cancer, polyps, leiomyomata uteri (fibroids), and infection, among others. endometrial hyperplasia may manifest clinically as uterine bleeding ; however, since post - menopausal women should be estrogen deficient, endometrial hyperplasia at this time is abnormal and should be further investigated. post - menopausal hormone therapy can be a cause of endometrial hyperplasia and subsequent post - menopausal bleeding. soy and other phytoestrogens in large doses may be associated with stimulation of the endometrial lining, as has also been described by the netherlands pharmacovigilance centre lareb in a previous case series. a randomized trial of 376 post - menopausal women who received soy versus placebo over a 5-year period showed that the occurrence of endometrial hyperplasia was significantly higher in the group receiving soy (3.37 vs. 0 %). hops are the female flowers (seed cones, strobiles) of the hop species humulus lupulus l. the principal estrogen from hops is the 1:1 racemate, ()-8-prenylnaringenin 8pn, also called hopein. this is one of the most potent in vitro estrogenic substance known from the plant kingdom [5, 6, 13 ], with an estrogenic activity in vitro greater than that of other phytoestrogens such as coumestrol, genistein, and daidzein. nowadays, hops are used primarily for their bitter and aromatic properties in the manufacture of beer and are cultivated by the brewing industry. most beer does not contain 8-prenylnaringenin in detectable quantities ; the highest concentration found being 19.8 g l. although the percentage of hop in the product used in this case report is known (41.40 %), the 8-prenylnaringenin amount is not. in addition to hop, the product also contains other plants that have phytoestrogenic activities. the plant lignans secoisolariciresinol (sec) and matairesinol (mat), which can be found in cereals, are converted to the metabolites enterodiol (ed) and enterolactone (el), known as the mammalian lignans, in the gastrointestinal tract. these compounds are bioactive as phytoestrogens because of their structural and functional similarity to 17-estradiol. in vitro, mammalian lignans may have both estrogenic and anti - estrogenic properties. for cereals like oats (a. sativa), barley (h. vulgare), and rye (s. cereale), the presence of lignans with phytoestrogenic properties has been described [1618 ]. isoflavones are also found in cereals and are an additional ingredient in this product. the exact nature of the natural isoflavones complex described on the packaging of the product is unknown. dietary flavonoids are also known for their antiplatelet activity, although the specific mechanisms by which this inhibition occurs has not been fully established. inhibition of the platelet function through binding to the thromboxane a2 receptor has been described. it is possible that all the ingredients with phytoestrogenic properties in the product could have had an additional effect to each other in causing the endometrial proliferation and subsequent vaginal hemorrhage. estrogenic activity of the product was found in an er assay with 17--estradiol as a comparator by the company phytogenix, which is a spin off contract research organization originating from the university of utrecht (department of medicinal chemistry, faculty of pharmacy) in the netherlands. the following is stated on the products website (http://www.menocool.nl) : only in the event that your uterus is removed, the starting dose is suitable for use indefinitely. in europe, this phytotherapeutic product is not officially registered through the dutch medicines evaluation board, but is sold as a food supplement, which falls under the jurisdiction of the dutch netherlands food and consumer product safety authority (nvwa). the commodities act requires that these products are safe, but no mandatory safety testing takes place before they come on the market. pharmacovigilance systems for registered drugs are well - defined. for food supplements, although they can contain pharmacologically active ingredients, such systems (nutrivigilance) are less developed. for pharmacovigilance, engaging consumer reports can be especially helpful in the recognition of blind spots such as adverse reactions caused by (adulterated) herbal product and counterfeit medicines [22, 23 ]. in addition, awareness among healthcare professionals that patients can use herbal drugs or other products, bought without a prescription, is important. with products bought over the internet however, we deem it unlikely that this company has adulterated the herbal product with western medicines. the product itself is compounded by dusart pharma (katwijk, the netherlands). for the department dry formulations (tablets and capsules) dusart pharma adheres to the pharma gmp regimen of nutritional supplements in accordance with a quality guideline issued by the european federation of associations of health product manufactures (ehpm). dusart pharma complies with the hazard analysis and critical control points (haccp) requirements [24, 25 ]. also, the owner of the company selling the product, among other herbal products such as a natural breast size enhancer, has been featured in both national and local newspapers in the netherlands [2628 ], and a skeptical article about their products was published in a dutch medical journal. a complaint about menocool this was upheld and the manufacturer had to remove some of the health claims from the product s website. a limitation of this case description is that some of the characteristics of the herbal product, as recommended in the consort (consolidated standards of reporting trials) statement, are unknown to the authors. since lareb received spontaneous reports on the menocool tablets and did not investigate the products in a trial setting, the authors have no information on what parts of the plants were used in this product. from a dutch newspaper article, we know some limited details on the production process by dusart pharma for products they made for standby vital bv. the described production process, at the time of publication of the article, was rather simple : plant materials were ground and the created thick paste was used to press tablets. the manufacturer of the product declares that it consists of 100 % plant materials. based on a visual inspection of the tablets, we presume that the tablets indeed consist of pressed plant materials. however, the netherlands pharmacovigilance centre lareb has not performed any qualitative testing on the product. based on the described cases and five earlier case reports to the dutch pharmacovigilance center, we suggest a causal relation between vaginal bleeding due to endometrial proliferation after menopause associated with the use of this hop - containing product. a naranjo assessment score of 5 was obtained for both case a and b, indicating a probable relationship between the vaginal hemorrhage and endometrial proliferation and their use of the suspect drug. phytoestrogenic products for relief of menopausal symptoms are available as over - the - counter preparations, and consumers often mistakenly believe that they do not cause adverse drug reactions. in the differential diagnosis, it is important to be aware that the use of a dietary supplement or an herbal drug with phytoestrogenic properties may be a possible cause of post - menopausal bleeding. | two 54-year - old women developed abdominal cramps and vaginal hemorrhage as a result of endometrial hyperplasia during treatment with a hop - containing phytotherapeutic product (menocool) for post - menopausal complaints. the women used the hop - containing phytotherapeutic product (418 mg of hop per tablet) twice daily (1 and 0.5 tablets by both patient a and b). patient a developed abdominal cramps and vaginal hemorrhage after 2 months of use. after gynecological examination, she was diagnosed with endometrial hyperplasia. the patient was treated with a curettage. the hop - containing phytotherapeutic product was discontinued, and the patient recovered. patient b developed abdominal pain / cramps and vaginal hemorrhage after 5 months of use. a cervix smear, internal examination, and ultrasound were performed. due to the thickness of the endometrium, a pipelle endometrial biopsy was performed. results showed no indication for cervix cancer. the use of menocool was ceased ; follow - up information received from the patient shortly thereafter indicated that she had almost entirely recovered from the abdominal pain / cramps and vaginal hemorrhage. hop (humulus lupulus) has phytoestrogenic properties that may be the cause of endometrial hyperplasia and subsequent vaginal hemorrhage. a naranjo assessment score of 5 was obtained for both cases, indicating a probable relationship between the patient s endometrial proliferation and subsequent vaginal hemorrhage and their use of the suspect drug. |
inflammatory bowel disease (ibd), cd, and uc are chronic relapsing inflammatory disorders of the intestine with unknown causes. ibd develops upon the interaction between genetic predisposition leading to immunological abnormalities, environment, and microbiome. none of these factors alone are sufficient to induce disease development.1 the occurrence of ibd is increasing in north america, europe, and worldwide.23 the incidence and prevalence of ibd in asia remain low compared with those in western countries, but a rising trend has been recognized in recent years, providing strong evidence in support of the effect of the environment.4567 the characteristics of western and asian patients with ibd differ in epidemiology, phenotype, clinical course, and genetic susceptibility.48910 korea has a relatively small land area and an ethnically homogeneous population. since the 1960s, when the first ibd cases in korea were reported, the incidence and prevalence of ibd in korea has been lower than that in western countries but have demonstrated rapidly increasing trends. several distinct characteristics of patients with ibd in korea have been reported, such as thiopurine - induced leukopenia and the risk of opportunistic tuberculosis (tb) infection in patients administered tumor necrosis factor- (tnf-) inhibitor.111213 these distinct characteristics can provide important clues about etiology and pathophysiology, and comprehensive population - based studies are required to clarify these differences.6 a cohort study is generally useful for estimating the incidence, prevalence, disease course, prognosis, and other clinically distinct features. cohort studies conducted in western europe and america have described the ibd clinical course and identified its pathophysiology. some unique features of patients with ibd in korea have been well demonstrated by cohort studies in korea, especially by members of the korean association for the study of intestinal diseases (kasid). however, most of the currently available data came from single - center and a few multi - center studies those were primarily retrospective in design.6 potential problems with the retrospective cohort approach include selection and misclassification bias. moreover, there have been no well - designed prospective cohort studies in korea. from this background, here we prepare the key findings of cohort studies of patients with ibd in korea to date and explore future potential ibd cohort studies in korea. the first systematic population - based cohort study was performed in the songpa - kangdong district of seoul in 1986.14 according to a recent update on the descriptive epidemiology of the songpa - kangdong district, the mean annual incidences of cd and uc increased significantly from 0.05 and 0.34 per 100,000 inhabitants, respectively, in 1986 - 1990 to 1.34 and 3.08 per 100,000 inhabitants, respectively, in 2001 - 2005. during this 20-year period, the ratio of the incidences of uc to cd was decreasing from 6.8 in 1986 - 1990 to 2.3 in 2001 - 2005, indicating that cd demonstrated a trend of a more accelerated incidence compared than uc in korea.15 the clinical features of uc at diagnosis are reportedly similar in koreans and westerners. for example, there are no differences in disease extent and severity at diagnosis or in the male - to - female ratio of patients with uc between korea and western countries. the cumulative risk of proximal extension in patients with proctitis or left - sided colitis was 33.0% after 5 years and 44.5% after 10 years. the cumulative probability of colectomy was 2.0% after 1 year, 2.8% after 3 years, and 3.3% after 5 - 15 years. the cumulative survival rates after 1, 5, and 10 years were 100%, 99.4%, and 97.4%, respectively. korean patients with uc look like to have a milder course that results in a reduced risk of colectomy compared to the western population.16 approximately 1.1% of patients with uc are diagnosed with any uc - related symptoms. most of these asymptomatic uc patients become symptomatic during follow - up but appeared to have better prognosis than symptomatic uc patients.17 a male predominance in the prevalence of cd was observed in the korean population, a finding similar to those reported in recent studies from china and japan,18 while the incidence of cd in females is predominant or equal to that in males in western countries.419 patients with cd may have a similar clinical course to westerners, especially regarding the intestinal resection rate. the surgery rate has decreased as time passes, which was related to the early administration of azathioprine/6-mercaptopurine (aza/6-mp).20 however, korean patients with cd are more likely than western patients to have jejunal involvement. jejunal involvement is a factor of poor prognosis in both korean and western patients with cd.21 moreover, perianal fistulas seemed more common in korean patients than in westerners.22 the early administration of biological agents showed improved outcomes of ibd ; consequently, increasing numbers of patients with ibd are expected to receive tnf- inhibitor therapy.2324 one of the most feared side effects is an increased risk of opportunistic tb infection, especially in an ibd population from a tb endemic area.2526 the kasid conductive retrospective cohort study aimed to define the risk of tb in a large cohort of 873 ibd patients using tnf- inhibitors compared with the matched general population and identify the detailed clinical characteristics of and risk factors for newly developed tb in patients with ibd. a total of 25 newly developed tb cases-21 (84%) pulmonary and 4 (16%) extra - pulmonary infections - were identified in the cohort. the adjusted standardized incidence ratio (sir) of tb was 41.7 (95% ci, 25.3 - 58.0), but the outcomes of anti - tb treatment were primarily favorable.27 the risks and characteristics of colorectal cancer (crc) in ibd were also assessed.28 the population - based nationwide study was conducted to investigate the incidence of crc in patients with uc. the kasid reviewed 7,061 cases of uc that occurred between 1970 and 2005 and found a total of 26 cases of crc. they reported that the cumulative risk of uc - associated crc was 0.7% for patients that had uc for 10 years, 7.9% for 20 years, and 33.2% for 30 years. the mean duration of uc before the development of crc was 11.5 years.29 another study from a well - defined hospital - based cohort was recently published. a total of 5,212 patients with ibd (2,414 with cd and 2,798 with uc) were followed up for 39,951 person - years, thus a mean follow - up duration of 7.7 years. the sir of crc was 6.0 (95% ci, 3.10 - 10.48) for cd and 1.68 (95% ci, 1.00 - 2.66) for uc. the sir of crc in korean patients with uc may be similar to that in western patients with uc. in contrast to western patients with cd, the most common site of crc in korean patients with cd is the low rectum, probably in accordance with the higher prevalence of perianal fistulas in korean patients with cd.30 thiopurine - associated leukopenia could be a life - threatening side effect of aza/6-mp in ibd treatment, and korean patients seem to experience myelotoxicity more frequently (31 - 56%) than patients of european descent.11 although thiopurine methyltransferase (tpmt) genotypes and activity could not clarify the thiopurine - induced leukopenia completely, it was proposed that tpmt genotypes and activity measurements before aza/6-mp administration might help to predict the development of leukopenia.31 recently, using the sample from cohort of 978 cd patients treated with thiopurine, a nonsynonymous single nucleotide protein (snp) in nudt15 (encoding p.arg139cys) was strongly associated with thiopurine - induced early leukopenia in both korean and united states patients with ibd.32 these findings explain, in part, the higher prevalence of thiopurine - associated leukopenia in asians despite the lower prevalence of tpmt mutations. these results highlight genetic or biomarker studies require blood or tissue specimens from a large number of patients with accurate, matched clinical data. in addition to leukopenia, growing evidence supports the existence of a correlation between thiopurine treatment and the increased risk of developing lymphoma. non - hodgkin lymphoma was diagnosed in 5 patients (71.4%) and hodgkin disease in 2 patients (28.6%). the sir of lymphoma was 2.03 (95% ci, 0.81 - 4.18) in the entire ibd patients and the sir of lymphoma in patients with cd was 9.31 (95% ci, 1.13 - 33.62). the sir of lymphoma in patients who were exposed to thiopurines was 5.93 (95% ci, 1.61 - 15.18). the risk of lymphoma in patients with cd seems to be increased and thiopurine may be related with the risk of lymphoma in korean patients with ibd.33 since genome - wide association studies (gwas) have been introduced in an effort to identify the etiological factors of ibd,34 many genetic polymorphisms related to an increased risk of ibd have been identified based on an established ibd cohort.35 tnfsf15 is a proven susceptibility gene for cd. the associations among five tnfsf15 snps and various clinical parameters were investigated using a total of 906 patients with cd and tnfsf15 genotype data with clinical information. in korean patients with cd, non - risk allele homozygotes of the tnfsf15 snps rs6478108 and rs4574921 are independent genetic predictive factors for the development of strictures / non - perianal penetrating complications and perianal fistula.36 gwas was performed in the korean population comprising a total of 2,311 patients with cd, 2,442 controls derived from the ibd clinic of asan medical center, and 792 from the korea research network for crohn 's disease. in this study, three new susceptibility loci demonstrated genome - wide significance : rs6856616 at 4p14 (or, 1.43 ; combined p=3.60 10), rs11195128 at 10q25 (or, 1.42 ; combined p=1.55 10), and rs11235667 at 11q13 (or, 1.46 ; combined p=7.15 10).37 attempts to comprehensively understand the epidemiologic, clinical, and genetic characteristics of korean patients with cd first started in 2008. won ho kim, president of the kasid at that time, earned a national grant entitled the " research network for crohn 's disease. " during the study period, active translational research was performed using clinical data from about 2,000 blood samples of 1,316 korean patients with cd. the study team also established the korean diagnostic and therapeutic guidelines for cd with the help of the ibd study group of kasid that published the diagnostic guidelines for intestinal tb, a disease often misdiagnosed as cd.6 after the research network for crohn 's disease study, a subsequent study named " establishment of crohn 's disease in korea and characterization of clinical features with long - term follow - up " was conducted from january 2012 to december 2013 led by dong soo han. retrospective clinical data before 2009 and prospective data for patients enrolled during and after 2009 were collected using a novel web - based well - recorded, high - quality electronic case report form system. data for 1,388 retrospective patients and 890 prospective patients including 635 blood specimens were collected and analyzed. in 2012, korea centers for disease control and prevention solicited grant applications for a 6-month demonstration project aimed at establishing a cd network. the kasid obtained this grant, and in 2012, a 3-year national grant from the korea centers for disease control and prevention. the new study funded by these grants was named " establishment and management of crohn 's disease research network ". recently, the latter two studies discussed above were integrated into a single study referred to as the " crohn 's disease clinical network and cohort " (connect) study led by joo sung kim (fig. a total of 34 institutions are now participating in this study, and more than 2,000 blood specimens have been collected to date. more than 20 studies using a retrospective cohort of 1,382 patients are underway and some results were presented at the kasid, asian organization for crohn 's and colitis, and european crohn 's and colitis organization meetings. the baseline characteristics of patients with a follow - up period 6 months are shown in table 1. a total of 2.1% of patients had a family history of ibd. according to the montreal classification, ileocolonic location (l3) was the most common, followed by the ileal (l1) and colonic (l3) locations. in 1,016 (79.5%) patients, the disease behavior at diagnosis was inflammatory. of the patients, 560 (43.8%) and 144 (11.3%) had perianal disease and stenosis at diagnosis, respectively. the first result led by dong il park was recently published about long - term clinical outcomes of korean patients with cd in urban versus rural environments. although numerous studies have reported a positive correlation between urban environment and cd development, no differences were found in terms of disease presentation and natural history between korean urban and rural populations except for a higher rate of surgery in the urban population of patients recently diagnosed with cd.38 you sun kim. conducted the study to determine the incidence of free perforation, the clinical characteristics of patients with cd and free perforation, and the risk factors associated with free perforation in patients with cd in korea. eighty - eight (6.4%) of the 1,382 patients had free perforation, and multivariate analysis revealed that free perforation was significantly associated with age 30 years at diagnosis (or, 2.082 ; p=0.002) and bowel stricture (or, 1.982 ; p=0.004).39 these retrospective data - based studies will be the background for future prospectively enrolled cohort studies to investigate the clinical course and pathophysiology of korean patients with cd. as of march 2015, 1,302 patients diagnosed with cd after 2009 with 850 serum samples are registered and prospectively followed - up in the national connect study.6 we regularly audit the data and send a newsletter to the investigators to remind them of the importance of accurate and complete data acquisition. investigators from the connect study anticipate that the typical clinical characteristics and genetic causes of cd in korean patients will be elucidated. despite the established nationwide cd cohort study in korea, there has been little nationwide systematic data on epidemiology, clinical characteristics, and genetics of korean patients with uc. there is an urgent need for the nationwide collection of well - defined clinical characteristics of korean patients with uc and their blood samples. hyo - jong kim, as coordinating investigator, and the other associated kasid members initiated the first nationwide " cohort study of newly diagnosed moderate to severe ulcerative colitis in korea " (mosaik). this is a multicenter, nationwide, prospective, 5-year follow - up longitudinal, disease - oriented, and hospital - based cohort study. patients newly diagnosed with moderate to severe uc in tertiary referral centers are to be included in this study. the aim of the mosaik study is to establish a uc research cohort for the investigation of epidemiology, clinical characteristics, and genetics of korean patients with uc by collecting well - defined clinical and follow - up characteristics of korean patients with uc and their blood samples with a goal of recruiting 400 patients. as of april 2015, > 100 patients diagnosed with uc from 27 hospitals are registered. the mosaik cohort will be closed when the last patient completes the last scheduled 5-year follow - up visit. the clinical characteristics and genetic causes of korean patients with uc could be elucidated through the mosaik cohort study results. it is widely accepted that a cohort study with a prospective design would be a very appropriate for epidemiologic study. no well - designed nationwide cohort study of ibd has been published in korea to date. however, recent long - term follow - up results from the ibd cohort study might help clinicians and researchers to make more clear decisions. these findings, together with the distinct demographic and phenotypic characteristics of korean patients with ibd, may provide clues to the etiology of these diseases as well as useful information for health care policy. even though the initiation of ibd cohort studies in korea was quite behind compared to western studies, there are some clinical and genotypic characteristics unique to korean patients with ibd like thiopurine - induced side effects and opportunistic tb infections. these unique features could provide important implications about the etiology and pathophysiology of ibd through a comprehensive population - based study. moreover, studies regarding biosamples might strengthen the impact of the cohort study. in an ibd cohort study, the typical clinical characteristics and genetic causes of korean patients with ibd could be elucidated, which will be a basis for further research into the clinical and genetic characteristics of ibd in korea. further diagnosis and treatment guidelines and new therapeutic agents for korean patients with ibd will be developed based on these cohort findings. | inflammatory bowel disease (ibd) is defined as a chronic and relapsing inflammatory disorder of the intestine. intestinal inflammation in ibd has been proposed to be attributable to the interplay between microbial, genetic, environmental, and immunological factors. the incidence and prevalence rates of ibd are rapidly increasing apparently in other parts of the world, with dramatic increases especially in east asia. generally, cohort studies are useful for estimating the incidence, prevalence, natural course, prognosis, and risk factors of diseases. in particular, cohort studies performed in western countries have well described the prevalence, risk factors, and natural course of ibd and investigated its genetic pathophysiology. however, the outcomes of ibd cohort studies performed in korea are not as persuasive as those of western studies because of the relatively low prevalence of ibd and short follow - up periods of the cohorts in korea. despite this critical limitation, members of the korean association for the study of intestinal diseases have demonstrated outstanding results. some unique features of ibd patients in korea are well demonstrated, such as thiopurine - induced leukopenia or risks of opportunistic tuberculosis infection in patients receiving tumor necrosis factor- inhibitors. in this review, the present authors summarized the key points of the results of the cohort studies performed in korea and explored future perspectives. |
breast cancer (bc) continues to remain the commonest cause of cancer death in women worldwide. bc is increasing in regions that until recently had low rates of the disease. in asia, bc incidence peaks among women in their forties, whereas in the united states and europe, it peaks among women in their 60 's. similarly, in iran, bc is most common cancer among females, which comprising 24.4% of all neoplasms. the mortality rate of bc was 5.8/100,000 women in tehran in 1998, 2.5/100,000 for a female population, and 7762 years life lost in the 18 provinces of iran in 2001. the latest data on age - specific incidence rate of bc showed that its crude incidence rate and asr are 17.4 and 23.1/100,000, respectively. data from the cancer institute show that bc highest rate is occurring in those aged between 35 and 44 years in iran. it is suggested that bc incidence among iranian females is rising and affects at least one decade earlier than their western counterparts, with a mean age ranging from 47.1 to 48.8 years. various risk factors for bc have identified, but a positive family history remains among the most important ones established for bc, with first - degree relatives of patients having an approximately two - fold elevated risk. it is estimated that approximately 20 - 25% of this risk is explained by known bc susceptibility genes, mostly those conferring high risks, such as brca1 and brca2. germline mutations in the brca1 (bc 1, early onset) and brca2 (bc 2, early onset) genes are the most important cause of hereditary breast and ovarian cancer. there is not a direct procedure to estimate the prevalence of brca1/2 mutations in the general population. in the absence of such measurement procedure, it can be assumed that nearly 50% of the mutations would be in brca1 and 50% in brca2. since the isolation of these two genes, more than 2000 different mutations have been identified. the most common types of mutation are attributed to small insertion / deletion frameshift, non - synonymous truncation, and disruption of splice site leading to entire nonfunctional brca proteins. early works of the bc linkage consortium have shown that respectively 52% and 32% of families with at least four cases of bc diagnosed tt1915 m in exon 11, ivs16 - 14 t > c ivs16 - 6 t > g in exon 17, 8345a > g n2706s in exon 18 and 9266c > tt3013i in exon 23 mutations in brca2. (2002) studied a family with four cases of ovarian cancer and one case of bc in close relatives for common mutations in the brca1 and brca2 genes using sscp of exons 2 and 20 of the brca1 gene and ptt on exon 11 of the brca1 gene and exons 10 and 11 of the brca2 gene. ptt demonstrated an abnormal result in exon 11 of the brca1 gene in the three sisters. upon sequencing a novel c. 2031 t > it was a nonsense mutation which leads to a truncated protein by replace glutamic acid with a stop codon. for the first time, pietschmannl. have performed a study that screened complete coding sequences and 3 and 5 utr regions of brca1 and brca2 genes in iranian bc patients. in addition, they used semi - quantitative fluorescent multiplex pcr to detect large rearrangements of the genes. the patient population was selection account of individuals from 10 unrelated high - risk families, which had at least three breast, breast and ovarian cancer cases as first - degree relatives, multiple cases as first, second and other relatives, bilateral bc, and early onset of breast and/or ovarian cancer (a) has detected as the second common brca1 gene defect (6.7%). the other mutations have identified as single nucleotide replacement including : c. 792a > c, c. 825 g > c, c. 822 t > a, c. 1068a > g, c. 969a > t and c. 966 t > c. keshavarzi. studied 27 patients with either early onset bc (at age 35 year) or personal and/or family history of breast or ovarian cancer. all of the brca1 and brca2 genes except exons 1 and 4 in brca1 and exon 1 in brca2 were analyzed. they identified a total of 13 missense mutations, 9 in brca1 and 4 in brca2. two of these were novel (c. 3538a > g in brca1 and c. 4350 g > a in brca2). in addition, c. 3232 g > a and c. 3538a > g brca1 mutations were found in large series of breast and ovarian cancer and matched controls. in another study, keshavarzi. investigated entire coding sequences and each intron / exon boundaries of brca1/2 genes in 85 patients from high risk iranian families. thirty - six patients had a family history of bc, while 49 patients had early onset bc (a) and brca2 (c. 2600 - 70 t > g) were identified in one patient with early bc and one control. in addition, a missense mutation within exon 11 of the brca1 (c. 3419ga) was identified in 20% of patients and have studied 39 patients with bc and 29 high risk healthy women, related to the patients to find to find 185delag and 5382insc founder mutations. in addition, a 251bp fragment of brca1 's exon 11 was analysed to determination new mutations. two out of 39 bc patients (5.1%) and 1 out of 29 relatives (3.4%) were carriers of 185delag mutations, and 1 patient (2.6%) was a carrier of a 5382insc mutation. in addition, 2 patients (5.1%) and 3 (10.3%) of relatives were carriers of unclassified mutations in the brca1 gene 251bp fragment [tables 1 and 2 ]. descriptive characteristics of 13 studies on the brca1 mutations in iranian breast cancer patients descriptive characteristics of 6 studies on the brca2 mutations in iranian breast cancer patients the present review provides the identified brca mutations rates in iranian bc patients to date. the 13 studies included in this study, screened a total of 1183 and 454 female and male bc patients, respectively [tables 1 and 2 ]. the prevalence of brca mutations carriers in the general population is estimated at between 0.12% and 0.1%. it is estimated that in all women, the prevalence of brca1 mutation is 1 in 800 - 1 in 1400 and the prevalence of brca2 mutation is slightly lower at 1 in 450 - 1 in 80). genetic linkage analyses have shown that the prevalence of brca gene mutation in familial bc and/or ovary cancer rate ranging from 45% to 90%. while, many of screening studies have shown brca1 mutation rate is about 6 - 45% and in familial bc varies from 1% to 35% worldwide. among early onset familial cases, 10 - 40% in contrast, among sporadic early - onset bc patients, the frequency of brca1/2 mutation ranges from 1% to 10%. of the total of 1183 patients who were included in the studies 377 cases therefore, based on the previous studies, the overall brca1 mutation rate in iranian bc patients with various levels of family history range (with or without bc diagnosed in relatives) was estimated 31.8% (377/1183). while, this gene mutation rate for male patients is less than 0.01%. this is comparable to that of algerian and tunisian families brca1 mutation frequency, which were reported 36.4% and 37.5%, respectively. the prevalence of brca1 mutations reported in a study conducted on french hereditary bc and/or oc families (10.3%) is approximately 3 times less than that estimated for our community. this high prevalence may be due to limited particular region of the genes, and the techniques were used. identification of brca mutations in a substantial proportion of iranian patients indicates that these genes play a role in the incidence of bc in the iranian population. in addition, according to these studies, there is heterogeneity in brca mutations in iranian bc patients like other population. results shown that a total of 104 brca1/2 distinct mutations were identified in the interim 13 articles analysis among 377 females bc patients with or without family history, which 71 (68.8%) mutations were in brca1, and 33 (31.2%) were in brca2. thirty - two mutations were putatively novel mutations that previously not reported. of these, 28 of 71 in brca1 (39.4%) and 4 of 33 in brca2 (12.2%) were putatively novel mutations which previously not reported. as shown in tables 3 and 4, the most common recurrent mutations in iranian bc patients, which were repeatedly reported twice or more in different articles included : c. 4837a > g, c. 3419 g > a, c. 3119 g > a, c. 2612c > t, c. 3113a > g, c. 2311 t > c, c. 4301 t > c and c. 4308 t > c in brca1, c. 4771c > t and c. 6494 g > c in brca2. however, brca1 c. 3419 g > a mutation was identified as a novel mutation, but its prevalence was higher than expected (28/377). incidence of more frequent brc1 mutations in the iranian bc patients incidence of recurrent brc1 mutations in the iranian breast cancer patients the frequency range of seven common mutations is shown in table 4. the frequency range for c. 4837a > g was between 13.6 and 30.0, for c. 3119 g > a was between 18.5 and 25.0, for c. 2311 t > c was between 5.9 and 30.0, for c. 5213 g > a was between 4.7 and 18.5, for c. 2612c > t was between 13.6 and 31.8, for c. 3113a > g was between 22.2 and 34.1, and for c. 4308 t > c was between 11.2 and 30.0. to the best of our knowledge, it has not introduced a founder mutation in iranian breast and ovarian patients yet. therefore, these mutations could represent candidate founder mutations and support testing these mutations at least for those with a family history of bc. the effect of most of the brca mutations on protein is unknown and making it difficult to predict the consequences on risks of breast and ovarian cancers. thus, many individuals undergoing genetic testing for brca mutations receive test results reporting a variant of uncertain clinical significance, leading to issues in risk assessment, counseling, and preventive care. based on the virtual analyses of functional compatibility for amino acid changes using sift and gvgd programs, 4 of the 8 most frequent variants in brca1 (i.e. p.ser1613gly, p.ser1040asn, p.pro871leu, and p.glu1038gly) were predicted to have an impact on protein structure (align - gvgd, http : www.//agvgd.iarc.fr/) and sift (http://www.sift.bii.a-star.edu.sg/). in well - defined populations based on ethnicity, founder mutations in the brca1 and brca2 genes have been found to account for a higher proportion of breast and ovarian cancer than in the general population. three specific mutations (185delag, 5382insc in brca1, and 6174delt in brca2 occur in 36% of breast / ovarian cancer families. interestingly, it is reported that 185delag mutation occurs at a very high frequency of 18.0% in families of ashkenazi jews with breast / ovarian cancer. based on the previous studies results, ashkenazi jewish founder mutations 185delag and 5382insc (brca1) rates among iranian bc patients were estimated approximately 0.75% (7/924) and 0.13% (1/774), respectively. however, some studies have supported that ashkenazi jewish founder mutations is not restricted to this particular ethnic subgroup and occurs in non - ashkenazi jewish ethnic groups at rates similar to the ashkenazi population, with a similar genetic background for all jewish mutation carriers. these mutations are not frequent mutation in iranian bc patients. according to the pietschmann. (2012) studies, which have sequenced most coding regions of brca1 and brca2, exons 2 and 11 in brca1 harbor 13.2% (7/53) and 28.3% (15/53) of the brca mutations, respectively. for the brca2, the overall frequency of 4075delgt was 0.02 (95% ci : 0.000.03, p = 0.51 for heterogeneity test). until date, no other such analysis study on bc in iran has been published. the major advantage of this review is help to determining the spectrum of brca mutations in iranian bc patients and must be strengthened with further studies with a larger cohort in order to determine the rate of the brca mutations in the iranian general population and to determine the existence of founder mutations. although, these studies were limited by some reasons, they are useable to provide appropriate cancer prevention, screening, and counseling strategies based on the mutation data. therefore, a new bc risk assessment based brca mutations data were created for the iranian population. all authors equally contributed in the conception of the work, revising the draft, approval of the final version of the manuscript, and agreed for all aspects of the work. hn contributed in the conception of the work, revising the draft, approval of the final version of the manuscript, and agreed for all aspects of the work. sms contributed in the conception of the work, revising the draft, approval of the final version of the manuscript, and agreed for all aspects of the work. smk contributed in the conception of the work, revising the draft, approval of the final version of the manuscript, and agreed for all aspects of the work. | background : brca1/2 genes mutation prevalence varies among ethnic groups and may be influenced by founder mutations. understanding brca1/2 genes mutations is important for reducing breast cancer (bc) incidence, accurate risk assessment and counseling. this systematic review of the literature was conducted to addressing brca1/2 mutations in iranian bc patients.materials and methods : a search for relevant articles was run on before january 2014 using medline, pubmed, science iranian database, google, and web sites related to the study topic. the key words included : bc and iran with genes, brca genes, brca1 and brca2 ; cancer genes, and iran.results:thirteen articles retrieved from this search strategy were eligible for this review. the overall brca1 mutation rate for iranian female bc patients was detected 31.8% (377/1183). although this gene mutation rate for male patients is g, c. 3419 g > a, c. 3119 g > a, c. 2612c > t, c. 3113a > g, c. 2311 t > c, c. 4301 t > c and c. 4308 t > c in brca1, and one brca2 mutation (c. 6494 g > c) were found in multiple case subjects and represent candidate founder mutations.conclusion:according to these studies, there is heterogeneity in brca mutations in iranian bc patients. |
the arterial stiffness is an index of vascular health and has been shown to confer additional independent predictive value for adverse cardiovascular outcomes in patients with uncomplicated essential hypertension as well as in the general population [2, 3 ]. the measurement of carotid to femoral pulse wave velocity (pwv) is considered the gold standard method for arterial stiffness assessment in daily practice because of its easy use, low cost, and high reproducibility. reference values of pwv come mostly from multicenter registries obtained in asia, usa [6, 7 ], australia, and europe [9, 10 ]. however, several studies in latin and hispanic populations have shown significant differences in size, structure, and arterial stiffness of large and small arteries [1115 ]. in addition, there are very few population - based studies that evaluate arterial stiffness from populations of south america [16, 17 ]. at present, there are no reference values of pwv based on healthy normotensive argentinean population studies involving adolescents, young adults, and older people that take into account factors influencing pwv values such aging. the aims of this work were (a) to establish normality values of carotid - femoral pwv in a large cohort of healthy and normotensive population with no cardiovascular risk factor according to age and (b) to determine the relationship of carotid - femoral pwv with aging and identify the rate of change of this parameter in a healthy population of a community - based study. this project was part of an epidemiological study aimed at determining the prevalence of cardiovascular risk factors in a well - characterized population. the present analysis is a descriptive, observational, and cross - sectional population - based study carried out in healthy people from the population of tandil city. the first was focused on evaluating cardiovascular risk factors in the elementary, middle, and high school populations (children and adolescents) [18, 19 ]. the second phase consisted of blood pressure measurements and cardiovascular risk assessment in the adult population. tandil is the main city of the homonymous department, located in argentina in the southwest of buenos aires province. this city is located 180 meters above sea level and 360 km from buenos aires city (371908s 90805w). the tandilean population is originated from native inhabitants and a large immigration influx from italy, spain, france (basques), and denmark. the population is distributed along an urbanized area of 22.07 km (13.7 mi) surrounded by a suburban area of 30.3 km (18.8 mi). the latest population data from the national institute of statistics and census reported 123.871 inhabitants in 2010. the protocol was approved by the institutional ethics and research committee, and it was conducted according to the declaration of helsinki and the good clinical practice guidelines. written consent describing the health examination and type of data collected was obtained from all participants. between march 2010 and december 2012, a total of 780 consecutive healthy subjects were enrolled. during their routine checkup, we measured the carotid - femoral pwv. our normal population was defined as asymptomatic nonsmoking subjects, having optimal or normal blood pressure values, without diabetes or dyslipidemia and no history of hypertension in first - degree relatives. asymptomatic subjects from 10 to 98 years old without history of cardiovascular, pulmonary, and renal disease, serum total cholesterol (tc) levels 1.5 mg / dl, smokers, lipid profile with one or more of the following conditions : tg 150 mg / dl, tc 200 mg / dl, subjects with bmi 30 kg / m. subjects with high blood pressure at the time of the examination, history or symptoms of cardiovascular disease (including arrhythmias), serum creatinine levels > 1.5 mg / dl, lipid profile with one or more of the following conditions : tg 150 mg / dl, tc 200 mg / dl, subjects with bmi 30 three blood pressure measurements were made, with the patient at rest seated for at least 10 minutes. an individual was defined as being normotensive when presenting with systolic blood pressure (sbp) 25 and < 29.9 and, finally, obesity was diagnosed when bmi was higher than 30 venous blood samples were obtained in the forearm by standard techniques and processed for determination of serum triglycerides, total cholesterol, glucose, and creatinine. measured and calculated values were expressed as mean value sd. statistical analyses were done using statistical package for the social sciences 19.0 (chicago, il, usa). demographic characteristics of the seven hundred eighty healthy subjects included in this research (age 39.8 18.5 years, range 1098 years) are summarized in table 1. the mean pwv found was 6.84 m / s 1.65 (range : 3.1213.4). table 2 shows the mean pwv, the range, and the 95% confidence intervals in normal subjects divided into 7 age groups. note that the standard deviation of the first four age groups (10 to 49 years) is smaller than that of the last three age groups (50 to 98 years) indicating an increase in the scatter of the pwv with the aging process. similarly in subjects older than 70 years minimum and maximum values of the 95% confidence interval show a greater dispersion than that observed in younger healthy participants. figure 1 shows that the pwv increases linearly with aging with a high degree of correlation (r = 0.61 ; p < 0.05) with less dispersion in younger subjects. in our population, pwv progressively increases averaging 68% with each decade of life, and this tendency is more pronounced after 50 years in which the average pwv increased by 18%. no differences in pwv values were found between men and women (6.81 m / s versus 6.89 m / s, resp.). pwv values were higher in subjects over 50 years (8.35 versus 5.92 m / s). this difference remained statistically significant in both men (8.52 1.39 versus 5.86 1.17 m / s) and women (8.20 1.13 versus 6.03 1 m / s). figure 3 shows that the differences are mainly determined by the aging and are significant between young and elderly subjects regardless of gender (p < 0.05). the measurement of pwv is a well - known method for the quantification noninvasive arterial stiffness and is currently considered the gold standard of arterial stiffness due to its simplicity, accuracy, reproducibility, and predictive value [24, 25 ]. most studies that establish reference values of pwv include data obtained from retrospective analysis of patients evaluated in different specialized centers. the mentioned reports includes several selection bias that difficult comparative studies with other patient populations. despite the recognized value of pwv for predicting cardiovascular risk, in south america moreover, only the republic of uruguay has reference values of pwv based on urban population. the present research clearly shows the normal values of pwv with the corresponding confidence interval of 95% for each age group providing relevant clinical information in terms of aortic stiffness. since pwv is an age dependent parameter, the clinician needs to know the mean value and calculated dispersion for each decade of human life in order to orientate both diagnosis and preventive strategies. first, this work represents the first argentine record based on urban and rural population that determines normal values of pwv in a large number of normotensive and healthy subjects free of family history of hypertension. on the other hand, the results of this based population study could only be extrapolated to those communities with similar demographic characteristics. it should be noted that several demographic and sociocultural aspects of the population of the city of tandil have similarities with the general population of argentina and south america (table 3). indeed, the percentage of women and the elderly as well as literacy rates, infant mortality, unemployment, and incomes (gross domestic product) is comparable to those of the general population of argentina and several countries of latin america. moreover, the prevalence of hypertension in tandil city and in rural areas is comparable to that reported in population - based epidemiologic studies of our country in the last two decades [27, 28 ] and similar to that reported in other urban and rural populations in latin america and the caribbean region. one of them is similar number of subjects but covering a wider range of ages (10 to 98 years). reported the reference values of pwv in 429 subjects selected from a hospital population from urban uruguay. they used a methodology similar to ours but considered six age groups and they analyzed subjects older than 60 years as a single group. in addition, the number of individuals in each group is less than our work. on the other hand, the european registry these normal values emerged from a retrospective analysis of pwv obtained with different methodologies in 13 european centers of high complexity. this record included several methods of measurement of pwv and required validation between different methodologies with international comparative study subjects to verify measurement accuracy. in the research here reported, all data collection was obtained by only one research group, always using the same technology. the third important aspect is the different rate of increase in pwv observed across different age groups. in our population, there is a deterioration of arterial elasticity associated directly with age across all the ages studied ; however, in agreement with previous reports, pwv presents a different behavior before and after the age of fifty (figure 3). fourth, with respect to gender difference in arterial stiffness, the authors are aware of the controversial reports about this interesting issue [9, 30 ]. in our study, there were no statistically significant differences in pwv values linked to gender. this finding is in concordance with reported large population - based studies in which the gender difference in pwv was absent or without clinical significance (< 0.1 m / s) [8, 13, 31 ]. in the framework of the anglo cardiff collaborative study and multiethnic study of atherosclerosis, it was demonstrated that gender might not directly influence arterial stiffening in healthy normotensive individuals. fifth, in our study, we included an important aspect to define normal values of pwv as it is the absence of first - degree relatives with a history of hypertension, coronary heart disease, or sudden death before age 65. several studies have shown a significant hereditary burden on several indexes of arterial function independently of blood pressure values and there is an association between pwv and genetic polymorphisms [6, 33, 34 ]. both associations justify analyzing a redefinition of the inclusion criteria to define a population as the reference standard. with respect to the methodological approach used in this work, the authors take into account previous reports in which comparative studies with invasive methods show that the use of direct measurement of carotid - femoral distance results in an overestimation of up to 25.4% of the distance traveled by the pulse wave and consequently results in overestimation of pwv values in 2 - 3 m / s [4, 23 ]. in consequence, we used the corrected carotid - femoral distance (x 0.8) which is the distance that has proven to be the best correlate with the real distance traveled by the pulse wave. finally, the relevance of routine pwv measurements under standardized conditions was emphasized in the last expert consensus on aortic stiffness measurement. consequently, the availability of pwv values that characterized each decade of the human life in healthy person acquires great relevance in order to establish the degree of functional impairment in different pathological states. this research is the first population - based study of an urban and rural population in latin america that provides normal values of the pulse wave velocity in healthy, normotensive subjects without family history of hypertension. in the analyzed sample, our data provide relevant clinical information to daily clinical practice setting with pwv cut - off values for each age group with the ci 95%. moreover, a significant increase in the pwv growth rate after the fifth age decade was confirmed. this supports the idea for an increase in the cardiovascular risk accompanying the ageing process. | in medical practice the reference values of arterial stiffness came from multicenter registries obtained in asia, usa, australia and europe. pulse wave velocity (pwv) is the gold standard method for arterial stiffness quantification ; however, in south america, there are few population - based studies. in this research pwv was measured in healthy asymptomatic and normotensive subjects without history of hypertension in first - degree relatives. normal pwv and the 95% confidence intervals values were obtained in 780 subjects (39.8 18.5 years) divided into 7 age groups (1098 years). the mean pwv found was 6.84 m / s 1.65. pwv increases linearly with aging with a high degree of correlation (r2 = 0.61 ; p < 0.05) with low dispersion in younger subjects. pwv progressively increases 68% with each decade of life ; this tendency is more pronounced after 50 years. a significant increase of pwv over 50 years was demonstrated. this is the first population - based study from urban and rural people of argentina that provides normal values of the pwv in healthy, normotensive subjects without family history of hypertension. moreover, the age dependence of pwv values was confirmed. |
the human thumb is opposable and prehensile, making it the most unique digit of the hand, responsible for hand functions, grasp, manual dexterity, and fine motor skills. the thenar eminence constitutes the intrinsic muscles of the hand that are responsible for complex movements of the thumb. congenital anomaly of the thumb and/or of the thenar muscles can be quite a disabling condition. in 1979, arminio was the first to report congenital absence of the flexor pollicis longus (fpl) tendon. since then, many cases have been published with congenital absence of the fpl tendon with or without associated anomalies of the thumb and thenar muscles. most of the reports have documented unilateral absence of fpl tendon ; bilateral absence is extremely rare. only three cases of bilateral congenital absence of fpl tendon have been reported so far ; no thenar atrophy was seen in these cases. we present a case of a 9-year - old female child with bilateral congenital absence of fpl tendon and associated thumb hypoplasia and thenar eminence atrophy. a 9 year old female child presented to our hospital with difficulty in performing tasks like writing or holding an object. there was absence of flexion movement at interphalangeal (ip) joints of bilateral thumbs. local examination revealed minor hypoplasia of the thumbs and absence of dorsal wrinkles [figure 1a ] and flexion creases [figure 1b ] at the ip joints of both the thumbs. minor hypoplasia of both the thumbs and absence of dorsal wrinkles are seen (arrows). (b) palmar surface of the hands show absent flexion creases (arrows) of bilateral thumbs. (c) functional limitation in holding a pen ; note the absence of active flexion of the interphalangeal joints of both the thumbs radiograph of hands showed mild skeletal and soft tissue hypoplasia of bilateral thumbs [figure 2 ]. radiograph of both hands shows mild skeletal and soft tissue hypoplasia of bilateral thumbs magnetic resonance imaging (mri) of hands showed absence of bilateral fpl muscles. in addition, there was absence of flexor pollicis brevis (fpb) and abductor pollicis brevis (abpb) muscles and hypoplasia / atrophy of bilateral opponens pollicis (op) muscles. bilateral adductor pollicis brevis (adpb) muscles appeared normal in morphology [figure 3 ]. t1w coronal image of both the hands shows absence of bilateral flexor pollicis longus (fpl) tendons. normally, fpl tendon is seen between the lateral head of flexor pollicis brevis (fpb) and the oblique head of adductor pollicis brevis (adpb) muscles, and is inserted into the base of the distal phalanx of thumb (see inset images). bilateral flexor pollicis brevis (fpb) and abductor pollicis brevis (abpb) muscles were also absent. bilateral adpb muscles appeared normal in morphology after pre - anesthetic checkup, the patient was planned for fpl reconstruction in bilateral thumbs. two - staged surgery was planned ; in the first stage, flexor pulley was reconstructed, and 2 months later, fpl reconstruction was done using flexor digitorum superficialis (fds) tendon to the ring finger. postoperative physiotherapy was done to increase the range of motion (rom) of the ip joints of both thumbs. at this moment, the patient has achieved ~20 of flexion at the ip joints bilaterally. congenital inability to flex the ip joint of the thumb may be due to several causes, including congenital absence of fpl, anomalous insertion of fpl, congenital tenovaginitis of the flexor tendon sheath, partial anterior interosseous nerve paralysis, traumatic rupture of the fpl, and anomalous connection between the tendons. among these, congenital absence of the fpl is extremely rare. absence of fpl may occur with or without associated anomalies of the thumb and thenar muscles. thumb hypoplasia with congenital absence of the fpl tendon, but without hypoplasia of the thenar muscles is the rarest variation. congenital absence of thenar muscles (e.g. fpb and abpb) without absence of fpl has also been reported in literature. most cases are reported in the pediatric age group and only a few are described in adults. clinically, the affected thumb shows absent or less evident dorsal wrinkles and flexion creases. skeletal and soft tissue hypoplasia of thumb may be present. according to blauth., thumb hypoplasia may be classified as isolated minor hypoplasia (type 1), associated with thenar hypoplasia and metacarpophalangeal (mcp) joint instability (type 2), musculotendinous / osseous deficiency with absent active motion at mcp or ip joint (type 3), floating thumb (type 4), and complete absence of the thumb (type 5). imaging may show absence (agenesis), hypoplasia, or atrophy of the muscles and tendons. ultrasonography (usg) is a low - cost, safe, non - invasive, and rapid method of evaluating musculoskeletal system. it is indicated in patients with cardiac pacemakers and metal implants where mri is contraindicated. computed tomography (ct) and mri may be used to confirm and support the usg findings. surgical reconstruction followed by rigorous rehabilitation the preferred surgical technique is one- or two - staged tendon transfer using the fds tendon of the ring finger. the range of flexion achieved at ip joint after the surgery varies between 20 and 35. following surgery, planned physiotherapy is a must to obtain satisfactory results. this case is unique as it demonstrates bilateral congenital absence of fpl tendon in association with thumb hypoplasia and thenar atrophy in a child. thenar atrophy was due to selective absence of bilateral fpb and abpb, and hypoplasia / atrophy of op. | congenital absence of flexor pollicis longus with or without associated anomalies of thenar muscles and thumb is of rare occurrence. inability to flex the interphalangeal joint of the thumb and absent dorsal wrinkles and flexion creases of the thumb are important clues to the diagnosis. routine radiography and cross - sectional imaging help to confirm and document the condition. this article presents an extremely rare case of bilateral congenital absence of flexor pollicis longus tendon with thumb hypoplasia and thenar atrophy. |
nearly 200 cases of calcifying epithelial odontogenic tumor have been reported in literature since pindborg described it as a separate pathologic entity in 1955. it has been identified under different denominations, such as, ameloblastoma of unusual type with calcification, calcifying ameloblastoma, malignant odontoma, cystic complex odontoma, and has also been considered as a variant of simple ameloblastoma. the eponym pindborg tumor was first introduced to the literature in 1967, to describe this interesting and unique odontogenic tumor. the calcifying epithelial odontogenic tumor is a benign odontogenic tumor of epithelial origin that accounts for approximately 1% of all odontogenic tumors. it is more common in the posterior part of the mandible of adults in the fourth to fifth decades. it is characterized by squamous epithelial cells, calcifying masses, and homogeneous acellular material admixed with the tumor epithelium and stroma that have been identified as amyloid. the origin of this neoplasm is not clearly known, although it is generally accepted to be derived from oral epithelium, reduced enamel epithelium, stratum intermedium or dental lamina remnants.[13 ] the differential diagnosis for ceot should include adenomatoid odontogenic tumor (aot), calcifying odontogenic cyst (coc), ameloblastic fibro odontoma (afo), odontoma, and in our case amyloidoma. this article describes a case of ceot involving the left maxilla associated with multiple myeloma in a middle - aged patient. the correlations between clinical, radiological, and histological patterns are discussed and a brief review of the literature is presented. a 55-year - old man presented at the ent department of sri ramchandra university with a one - and - a - half year history of painless left nasal swelling. it was hard in consistency, gradually increasing in size to attain the current size of 4 cm. he was diagnosed to have multiple myeloma stage iiia in 2004, for which he underwent chemotherapy for a year. he had a fall in 2008, and underwent left bipolar hemiarthroplasty and left lateral rhinotomy for the nasal mass. computed tomography (ct) imaging of the maxillofacial region revealed a rounded heterogeneous mass over the left side of the nasal dorsum and frontal process of the left maxilla. bone marrow aspirate and biopsy were consistent with a diagnosis of multiple myeloma (36% plasma cells). intra - operatively, a mass was found to be over the left side of the dorsum of the nose and the frontal process of the maxilla was eroded. ct imaging of the maxillofacial region shows a heterogeneous mass over the left side of the nasal dorsum and frontal process of the left maxilla histopathological examination revealed a well - circumscribed neoplasm, predominantly made up of acellular eosinophilic material, arranged as nodules, with extensive calcification and ossification [figure 2, figure 3 ]. increased number of osteoclast type of giant cells was noted admixed with the acellular material [figure 4 ]. occasionally individual and cords of squamoid cells were seen with nuclear pleomorphism [figure 5, figure 6 ]. congo red stained the acellular material, confirming the amyloid nature [figure 7 ]. the acellular material was negative for cytokeratin stain by immunohistochemistry. with these features, a differential diagnosis of amyloidoma and ceot the presence of extensive calcification, ossification, osteoclast type of giant cells, and occasional atypical squamoid cells, strongly favored a diagnosis of ceot. photomicrograph showing acellular eosinophilic material arranged as nodules with ossifi cation (h and e, 20) photomicrograph showing acellular eosinophilic material arranged as nodules with extensive calcifi cation (h and e, 20) photomicrograph showing osteoclast type of giant cells admixed with acellular material (h and e, 20) photomicrograph showing cords of squamoid cells with nuclear pleomorphism (h and e, 20) photomicrograph showing cords of squamoid cells with nuclear pleomorphism and acellular material (h and e, 40) photomicrograph showing acellular material arranged as nodules (congo red, 20) previously, the uncertainty regarding the histological characteristics of ceot was reflected in the variety of terms for the disease, including unusual ameloblastoma, cystic odontoma, and adenoid adamantinoma. pindborg tumor is a rare, benign, but locally aggressive odontogenic tumor, which accounts for less than 1% of all odontogenic tumors. most investigators believe that the tumor cells originate from the stratum intermedium of the normal dental germ. this idea is based on the morphological similarity of the tumor cells to the normal cells of the stratum intermedium, and high activity of alkaline phosphatase and adenosine triphosphate. ceot occurs most commonly between 20 and 60 years of age with the mean age around 40 years. most investigators agree that the central type is usually located in the premolar and molar regions with a mandibular to maxillary ratio of 2:1 or 3:1. the literature reports that this intraosseous tumor usually manifests as a painless swelling that causes slow bone expansion. when located in the maxilla, patients may sometimes complain of nasal stuffiness, epistaxis, and headache. few (5%) extra osseous cases have been described, all of which have been involved peripherally in the anterior maxillary or mandibular gingiva.[79 ] fifty - two percent of the reported cases have been associated with an unerupted or embedded tooth. in the index case the tumor most commonly appears as either a diffuse or a well - circumscribed unilocular radiolucent area. in some cases, the lesion becomes multilocular with a honeycomb pattern. in others, multiple radio - opacities there is no doubt that the epithelial cells of ceot are of squamous epithelial origin, as is manifested by their morphological and immunohistochemical characteristics. an amyloid deposit is well documented in association with squamous cell carcinoma at different locations namely ; skin, vagina, uterine cervix, and nasopharynx. some reports have indicated that the squamous cell carcinoma associated with amyloid is immunoreactive for cytokeratin, and amyloid is regarded as the degradation product of keratin. it is also known that in primary cutaneous amyloidosis, the amyloid is derived from filamentous degeneration of keratin filaments. therefore, it is reasonable to regard all these findings, including ceot amyloid, as similar expressions of the same process, that is, filamentous degeneration of keratin and conversion into amyloid. with this view and since pindborg tumor is an odontogenic tumor, one draws back to previous suggestions that the ceot amyloid is somehow connected to enamel formation, which is related to keratin. another idea that emerges from the analysis of the cytokeratin staining pattern of amyloid is the concept of aging. this thought is based on several observations, among which is the resemblance between the cytokeratin staining pattern of the amyloid and the epithelial gland - like structures of the tumor. it seems that the tumor passes through various stages of development, commencing with epithelial degeneration and conversion of keratin filaments into amyloid. at first, the amyloid is periodic acid - schiff - negative and stains positively for cytokeratin. however as it ages and the amyloid deposits coalesce into globules, the amyloid loses its immunoreactivity for cytokeratin and becomes pas - positive. at this stage, the amyloid also mineralizes and gives rise to calcifications that exhibit the liesegang ring phenomenon. the idea of aging has been proposed in the past, but it gains a new dimension now that it relies on more facts. however the relative significance and contribution of keratin and basement membrane protein to the development of ceot - associated amyloid are still not clear. it is for future studies to reveal this and other secrets concerning this rare and unique tumor. the closest differential diagnosis of ceot in the present case is amyloidoma of the bone, which is a rare condition characterized by the massive destructive deposition of al amyloid in bones. although extensive amyloid was present, plasma cells were not seen in the maxillary lesion. this along with the other features, mentioned wide supra, favored a diagnosis of ceot. the literature search depicts 200 cases of ceot in the jaw and only two cases in case of amyloidoma. differences between ceot and amyloidoma the deposits in amyloidoma proved to be composed of al amyloid showing kmno4-resistant congophilia. immunohistochemistry showed immunoglobulin igg (lambda), igg (kappa), and igm (lambda) monoclonality of the plasma cells, and lymphoid infiltrate. the treatment for ceot has ranged from simple enucleation or curettage to radical and extensive resection, such as, hemimandibulectomy or hemimaxillectomy.in the present case lateral rhinotomy was done. malignant behavior is extremely rare ; only two cases have been reported in the literature so far. although it has not been established in the literature, five years should be the absolute minimum follow - up necessary to assess the behavior of this type of odontogenic tumor. overall, a range of clinical, imaging, and histopathological findings should be used to help in the diagnosis of ceot and to evaluate its extension to different sites. | calcifying epithelial odontogenic tumor (ceot), also known as pindborg tumor, is a rare benign odontogenic tumor of locally aggressive behavior. it is more common in the posterior part of the mandible of adults, typically in the fourth to fifth decades. its origin as well as its true malignant potential is not clearly known. it usually starts as a painless swelling and is often concurrent with an impacted tooth. a case of ceot in a 55-year - old man with multiple myeloma is presented. clinical, radiological, and pathologic findings are discussed. |
24-hour abpm plays an important role in determining cardiovascular prognosis and has been shown to be a better predictor of cardiovascular morbidity and mortality as compared to office blood pressure measurements [13 ]. the objective was to study the relationship between 24 h abpm and cardiovascular outcomes in patients from chesterfield royal hospital. over 12 months from the 1st of august 2002, 1187 individuals had 24-hour abpm performed. these individuals represented a typical spectrum of patients attending for 24 h abpm with blood pressure at different stages and with varying durations of hypertension. cardiovascular outcomes were studied in a subset (297) of the original cohort, made up by every 4th consecutive subject. individuals must have one of the recognized indications for 24-hour abpm, as outlined in table 1. every 4th consecutive patient was entered into the study, giving a total of 297, patients. table 2 outlines demographics and blood pressure criteria between the original cohort and the study cohort. clinical case notes to study prognostic information were not available from 52 subjects and they were excluded leaving 245 patients. the following abpm - related prognostic features were studied (table 3) high day time systolic and diastolic bp (135, 85 mmhg), high night time systolic and diastolic bp (120 mmhg, 75 mmhg), absence of nocturnal dip (10% fall in night time sbp), high early morning sbp (140 mmhg), and morning surge (20/15 mmhg rise in the first two morning readings from 7 am as compared to average night time bp). the cardiovascular outcomes studied (table 4) included fatal and nonfatal mi, new diagnosis of angina, acute coronary syndrome, sudden cardiac death, cardiac arrhythmias, acute lvf, cerebrovascular events, peripheral vascular disease, abdominal aortic aneurysm, and ckd stage 3 or above. over a followup period of 2015 116 days (17202305 days) 82 cardiovascular events occurred in 61 subjects. cardiac arrhythmias were the most common cv outcome (34 events) followed by cerebrovascular events (15). statistically significant associations found were between cerebrovascular events and absent nocturnal dip 10% (p =.05) and high day time dbp (p =.029), peripheral vascular disease and morning surge 20/15 mmhg (p =.014), cardiac arrhythmias and high day time and night time dbp (p =.009 and, 033, resp.). in this study, cardiac arrhythmias were the most commonly observed event accounting for 13.9% of the total events. atrial fibrillation was the most common cardiac arrhythmia seen in 14/38 (52.9%) patients with cardiac arrhythmias, followed by symptomatic ventricular ectopics in 13 subjects (38.2%) and supraventricular tachycardia and sinoatrial pause in 1 patient each. it has been shown to be associated with systolic hypertension and high pulse pressure. atrial fibrillation may complicate even mildly raised blood pressure, and it would be reasonable to assume that there is no threshold below which the risk of atrial fibrillation is not increased. to the best of our knowledge, our study is the first one to show an increased risk of atrial fibrillation with high day time and night time diastolic blood pressure. having said that, one of the recent japanese studies has shown that control of both systolic and diastolic blood pressure is important in reducing risk of new onset atrial fibrillation. over the years a variety of other risk factors for atrial fibrillation have been identified such as large left - atrial size, obesity, thyrotoxicosis, and high alcohol. the exact mechanism for atrial fibrillation in hypertensive subjects is not understood but is believed to be related to left ventricular hypertrophy and an increase in left - atrial size, left - atrial fibrosis secondary to high systolic blood pressure and changes in autonomic tone with higher in - treatment heart rate on serial ecgs. atrial fibrillation is an important cardiovascular risk factor for thromboembolic cardiovascular disease and adds to the existing risk from hypertension itself. treatment of hypertension exclusively with ace inhibitors, angiotensin - ii - receptor blockers, and beta blockers was shown to be associated with a lower risk of developing atrial fibrillation than current exclusive therapy with calcium - channel blockers. in summary, our study shows that diastolic hypertension plays an important role in leading to cardiac arrhythmias, in particular atrial fibrillation, and should be treated as vigorously as systolic hypertension. the study 's main limitation is that it did not take into account presence or absence of other cardiovascular risk factors, such as diabetes mellitus, smoking, hyperlipidaemia, or family history, and has relied entirely on blood pressure criterias. the authors would like to acknowledge that this may have had bearing on some of the findings. | introduction. 24-hour ambulatory blood pressure monitoring (abpm) plays an important role in assessing cardiovascular prognosis, through presence or absence of abpm - related prognostic features. objectives. to study relationship between 24-hour abpm and cardiovascular outcomes in patients from chesterfield royal hospital. material and methods. over 12 months from the 1st of august 2002, 1187 individuals had 24-hour abpm performed. cardiovascular outcomes were studied in a subset (297) of the original cohort, made up by every 4th consecutive subject. the following abpm - related prognostic features were studied high day time systolic and diastolic bp (135, 85 mmhg), high night time systolic and diastolic bp (120 mmhg, 75 mmhg), absence of nocturnal dip (10% fall in night time sbp), high early morning sbp (140 mmhg), and morning surge (20/15 mmhg). the cardiovascular outcomes studied in the fourth table included fatal and nonfatal mi, new diagnosis of angina, acute coronary syndrome, sudden cardiac death, cardiac arrhythmias, acute lvf, cerbrovascular events, peripheral vascular disease, abdominal aortic aneurysm, and ckd stage 3 or above. results. over a followup period of 2015 116 days (17202305 days) 82 cardiovascular events occurred in 61 subjects. cardiac arrhythmias were the most common cv outcome (34 events) followed by cerebrovascular events (15). statistically significant associations found were between cerebrovascular events and absent nocturnal dip 10% (p =.05) and high day time dbp (p =.029), peripheral vascular disease and morning surge 20/15 mmhg (p =.014), cardiac arrhythmias and high day time and night time dbp (p =.009 and.033, resp.). conclusion. significant associations were found between cerebrovascular events and absent nocturnal dip 10% and high day time dbp, peripheral vascular disease and morning surge 20/15 mmhg, cardiac arrhythmias and high day time and night time dbp. |
acute renal failure (arf) is associated with adverse outcomes, especially in children admitted to the pediatric intensive care unit (picu). due to usage of multiple definitions of arf in the literature, causing large variations in the reported incidence and outcome, the term arf was replaced by acute kidney injury (aki) recently, to provide the uniformity of definition and standardize the care of patients [figure 1 ]. many studies on the incidence of aki in critically ill - children have been conducted in developed countries and are often retrospective in nature. only a few retrospective studies have been conducted to determine the incidence and profile of aki, in critically ill - children from the developing world in recent years. this study was performed considering the paucity of data available on the incidence and determinants of aki in indian children and taking into account the retrospective nature of previous studies. this prospective observational study was conducted over a period of 10 months from june 2010 to march 2011. informed consent was obtained from the parents prior to inclusion of subjects into the study. to determine the incidence of aki as defined by the acute kidney injury network (akin) classification in critically ill pediatric patients admitted to the picu ; aged 1 month to 13 years. to determine the predictors of fatality in aki in critically ill - childrento study the etiology and short term outcome of aki in critically ill - childrento compare the demographic and clinical parameters among survivors and non - survivors in aki. to determine the predictors of fatality in aki in critically ill - children to study the etiology and short term outcome of aki in critically ill - children to compare the demographic and clinical parameters among survivors and non - survivors in aki. patients with known chronic kidney disease stage 5 (estimated glomerular filtration rate 5 mg / dl. patients with known chronic kidney disease stage 5 (estimated glomerular filtration rate 5 mg / dl. the incidence of aki was estimated to be around 30% in picu patients on the basis of current literature. assuming a variation of 7% in picu, i.e., absolute precision d = 0.07, and the chance of this to be at least 95%, the sample size was calculated to be 165 subjects. results were analyzed using the spss version 16 (ibm corporation, new york, u.s.a). values for continuous data were expressed as mean sd (if normally distributed) and median (range) (if non - normally distributed). the incidence of aki was defined as its occurrence as a proportion of total admissions. continuous variables with normal distribution were compared using student t - test while those not normally distributed were analyzed using mann whitney u test. multivariate binary logistic regression models were used for multivariate analysis of statistically significant variables in univariate analysis (p 60 mmhg), uncontrollable or poorly controlled seizures, hypotension requiring inotropic support, requirement of renal replacement therapy (rrt) and fulminant hepatic failure. the diagnosis of aki was based on akin definition and classification [figure 1 ]. either serum creatinine or urine output was used to diagnose and stage aki, using a criterion that led to a higher stage classification. a total of 2 ml of intravenous blood was withdrawn and centrifuged at 3000 rpm for 10 min. estimation of serum creatinine was repeated every 24 6 h for 3 consecutive days and daily thereafter until discharge from hospital. an absolute increase in serum creatinine of 0.3 mg / dl or an increase in serum creatinine of more than or equal to 1.5-fold from the initial serum creatinine was considered as aki. similarly, a decrease in serum creatinine of more than or equal to 0.3 mg / dl or a decrease in serum creatinine of 1.5-fold from the initial serum creatinine was also considered as aki. if there was a progressive rise in serum creatinine values, re - classification and progression to maximum aki stage during the hospital stay was recorded. the provisional diagnosis at admission and final diagnosis (at discharge / death) were recorded. the diagnosis of sepsis was made according to the international pediatric sepsis consensus conference definition. demographic parameters and short term outcomes (complete renal recovery, partial renal recovery and death) were recorded. shock was defined as the presence of at least two of the following : tachycardia (heart rate > 2 sd for age), feeble pulses, cool peripheries and hypotension (blood pressure 3 s. hypertension was defined as > 95 percentile blood pressure for age, height and gender. complete renal recovery was defined as normal serum creatinine for age (0.2 - 0.4 mg / dl for infants, 0.3 - 0.7 mg / dl for 1 - 12 years, 0.5 - 1 mg / dl for > 12 years) and normal blood pressure at discharge. partial renal recovery was defined as elevated serum creatinine for age or persistent hypertension at discharge. to determine the incidence of aki as defined by the acute kidney injury network (akin) classification in critically ill pediatric patients admitted to the picu ; aged 1 month to 13 years. to determine the predictors of fatality in aki in critically ill - childrento study the etiology and short term outcome of aki in critically ill - childrento compare the demographic and clinical parameters among survivors and non - survivors in aki. to determine the predictors of fatality in aki in critically ill - children to study the etiology and short term outcome of aki in critically ill - children to compare the demographic and clinical parameters among survivors and non - survivors in aki to determine the incidence of aki as defined by the acute kidney injury network (akin) classification in critically ill pediatric patients admitted to the picu ; aged 1 month to 13 years. to determine the predictors of fatality in aki in critically ill - childrento study the etiology and short term outcome of aki in critically ill - childrento compare the demographic and clinical parameters among survivors and non - survivors in aki. to determine the predictors of fatality in aki in critically ill - children to study the etiology and short term outcome of aki in critically ill - children to compare the demographic and clinical parameters among survivors and non - survivors in aki patients with known chronic kidney disease stage 5 (estimated glomerular filtration rate 5 mg / dl. patients with known chronic kidney disease stage 5 (estimated glomerular filtration rate 5 mg / dl. the incidence of aki was estimated to be around 30% in picu patients on the basis of current literature. assuming a variation of 7% in picu, i.e., absolute precision d = 0.07, and the chance of this to be at least 95%, the sample size was calculated to be 165 subjects. results were analyzed using the spss version 16 (ibm corporation, new york, u.s.a). values for continuous data were expressed as mean sd (if normally distributed) and median (range) (if non - normally distributed). the incidence of aki was defined as its occurrence as a proportion of total admissions. continuous variables with normal distribution were compared using student t - test while those not normally distributed were analyzed using mann whitney u test. multivariate binary logistic regression models were used for multivariate analysis of statistically significant variables in univariate analysis (p 60 mmhg), uncontrollable or poorly controlled seizures, hypotension requiring inotropic support, requirement of renal replacement therapy (rrt) and fulminant hepatic failure. the diagnosis of aki was based on akin definition and classification [figure 1 ]. either serum creatinine or urine output was used to diagnose and stage aki, using a criterion that led to a higher stage classification. a total of 2 ml of intravenous blood was withdrawn and centrifuged at 3000 rpm for 10 min. estimation of serum creatinine was repeated every 24 6 h for 3 consecutive days and daily thereafter until discharge from hospital. an absolute increase in serum creatinine of 0.3 mg / dl or an increase in serum creatinine of more than or equal to 1.5-fold from the initial serum creatinine was considered as aki. similarly, a decrease in serum creatinine of more than or equal to 0.3 mg / dl or a decrease in serum creatinine of 1.5-fold from the initial serum creatinine was also considered as aki. if there was a progressive rise in serum creatinine values, re - classification and progression to maximum aki stage during the hospital stay was recorded. the provisional diagnosis at admission and final diagnosis (at discharge / death) were recorded. the diagnosis of sepsis was made according to the international pediatric sepsis consensus conference definition. demographic parameters and short term outcomes (complete renal recovery, partial renal recovery and death) were recorded. shock was defined as the presence of at least two of the following : tachycardia (heart rate > 2 sd for age), feeble pulses, cool peripheries and hypotension (blood pressure 3 s. hypertension was defined as > 95 percentile blood pressure for age, height and gender. complete renal recovery was defined as normal serum creatinine for age (0.2 - 0.4 mg / dl for infants, 0.3 - 0.7 mg / dl for 1 - 12 years, 0.5 - 1 mg / dl for > 12 years) and normal blood pressure at discharge. partial renal recovery was defined as elevated serum creatinine for age or persistent hypertension at discharge. 54 children had aki, giving incidence of 25.1%. the median age of patients with aki was 21 months (range 1 - 144 months) and 53.7% of patients were boys. the mean level of maximum creatinine value during the hospital stay was 1.9 (sd 1.7) mg / dl. aki stage 1, stage 2 and stage 3 were detected in 19 (35.2%), 14 (25.9%) and 21 (38.9%) of patients respectively. demographic parameters of critically ill - children with aki the etiology of aki observed is summarized in table 2. tropical febrile illnesses (dengue, scrub typhus, tuberculosis (tb) and malaria) constituted 9.3% of aki patients. sepsis (without localizing signs) was diagnosed in 9 children, 5 were culture positive. organisms isolated were pseudomonas aeruginosa (2 patients), escherichia coli (1 patient), klebsiella pneumoniae (1 patient) and streptococcus pneumoniae (1 patient). other common etiologies were acute post - streptococcal glomerulonephritis (psgn), snake envenomation, hemolytic uremic syndrome (hus) and congestive cardiac failure. etiological profile of aki in critically ill children (n=54) mortality rate in children with aki was 46.3%. in aki stage 1, 7 (36.8%) patients died while in stage 2 and stage 3, 8 (57.1%) and 10 (47.6%) patients died respectively (differences not significant). mortality was nil in psgn, diarrhea, snake envenomation and tb, but highest in pneumonia (66.7%). mortality in babies aged less than 10 months was 65% while it was 35.3% above 10 months (p = 0.049). the age, etiological profile and maximum serum creatinine values were not different among the survivors and non - survivors [table 3 ]. the length of hospital stay was 10.1 5.8 days among survivors. among the non - survivors, the corresponding value was 6.3 4.3 days. comparison of survivors and deaths in critically ill - children with aki (n=54) a total of 23 (79.3% of survivors) children with aki had complete renal recovery while 6 (20.7% of survivors) had partial renal recovery at discharge. in aki stage 1, out of the survivors, 11 (91.7%) had complete renal recovery while 1 (8.3%) had partial renal recovery at discharge. in aki stage 2, 4 (66.7%) had complete renal recovery while 2 (33.3%) had partial renal recovery at discharge. in aki stage 3, 8 (72.7%) had complete renal recovery, while 3 (27.3%) had partial renal recovery at discharge (differences not significant). the mortality among children requiring rrt was similar to children not requiring rrt (46.7% vs. 46.2%). requirement of rrt was not related to age or the etiology of aki. the complications and co - morbidities observed among the study subjects included severe metabolic acidosis in 32 (59.3%), hyponatremia in 14 (25.9%), hypernatremia in 8 (14.8%), hyperkalemia in 13 (24.1%), hypertension in 8 (14.8%), encephalopathy in 19 (35.2%), thrombocytopenia in 21 (38.9%), mechanical ventilation in 43 (79.6%) and shock in 47 (87%) children. there was no correlation between stages of aki and etiological profile, mortality or length of hospital stay. the predictors of mortality on univariate analysis were : age less than 10 months, shock and requirement of mechanical ventilation [table 4 ]. in the multivariate model, requirement of mechanical ventilation was found to be an independent predictor of fatality (r = 24.3% ; odds ratio 9.7 ; 95% confidence interval [ci ] : 1.1 - 85.5 ; p value 0.041). predictors of fatality in aki on univariate analysis detection of incidence, etiological profile and outcome of aki is important for the institution of appropriate management as well as comparison of epidemiological studies for improved clinical decision making. the present prospective observational study from a tertiary center in southern india found the incidence of aki to be 25.1% in critically ill - children admitted to the picu. some recent pediatric studies on aki using risk, injury, failure, loss, end - stage criteria or its modifications have reported the incidence of aki to be widely varying from 10% to 82%, highlighting the heterogeneity of patient populations, diverse regional differences, sample sizes and study designs. one of these studies was a retrospective analysis of prospectively collected clinical data in 3396 critically ill - children ; 15.7% had some degree of aki at admission and 10% had aki develop during hospital course. another study from texas in 150 mechanically ventilated children found the incidence of aki to be 82%. of these children, 11 required dialysis. in another retrospective study from the netherlands, among 103 children requiring mechanical ventilation, 58% developed aki ; 6 patients received rrt. in yet another study from california, in 123 children with burn injury of 10% or more of body surface area, incidence of aki was 45.5%. a wide spectrum of etiologies for aki has been found in studies across the world. while sepsis, glomerulonephritis, hus and acute tubular necrosis predominate in developing countries, these have been replaced by hemato oncologic complications and pulmonary failure as causes of aki in the west. one study from turkey on 100 children with aki described the most common causes as bone marrow transplantation, renal disease, dehydration, nephrotoxic medication and cardiac surgery. in another study from the same country, on 472 children with aki (including 32.6% neonates), hypoxic ischemic injury and sepsis were leading causes of aki. at kolkata, india, glomerulonephritis and snake bite were the two most important causes of aki in 37 children, making up 70% of all cases. we found that the common etiologies were infections, psgn, snake envenomation, hemolytic uremic syndrome (hus) and congestive cardiac failure. pneumonia constituted one - fourth of all infections associated with aki and was associated with high mortality. mortality was not seen in psgn, diarrhea, snake envenomation and tb, but was quite common in pneumonia (66.7%). increased risk of developing aki has been mentioned with pneumonia, but seems to have been under - reported in children. in a prospective study from scotland, out of 1241 tropical febrile illnesses have been significantly associated with aki, especially in adults. in our study too, tropical febrile illnesses (dengue, scrub typhus, tb and malaria) constituted 9.3% of children with aki. mechanisms for aki in tropical illnesses include direct invasion by the micro - organism leading to acute interstitial nephritis (in dengue and tb), hemodynamic perturbation and renal ischemia (in malaria). severe diarrhea and psgn form a large proportion of children with aki in india. in our study, acute glomerulonephritis (predominantly psgn) accounted for 7.6% of patients. probably, awareness regarding the usage of oral rehydration solution has led to fewer cases of severe dehydration and thereby aki, being referred to our institute. three patients developed aki owing to snake envenomation in our study, which is an important problem in some regions of india. the mortality in aki in children also has been reported to vary widely from 16% to 43.8%. in our study, it was 46.3%, which is comparable to a recent study from kuwait reporting 43.8% mortality. a retrospective study of 311 children with arf over a 22 year period from thailand reported mortality as 41.5%. much of the data on the incidence and mortality of aki are limited to the developed world. apart from mortality, 20.7% of children with aki who survived, had partial renal recovery at discharge (majority being aki stage 3), pointing toward significant morbidity resulting from aki. in the present study, requirement of mechanical ventilation however, the wide ci of the odds of death indicates that the estimate lacks precision since the study was not powered to look at the predictors of fatality. though age less than 10 months and shock predicted fatality on univariate analysis, they were eliminated on multivariate logistic regression analysis. mortality in aki is primarily related to etiology ; psgn and gastroenteritis having a much better outcome than sepsis, malignancy or major surgery. multiple factors have been described as predictors of outcome in aki, again reflecting heterogeneity of patient populations. sepsis and hus with prolonged anuria have been associated with poor outcome. in rapidly progressive glomerulonephritis delay in seeking health - care, infections and cardiovascular / respiratory complications result in poor outcome. in critically ill - patients with aki undergoing hemodialysis, cardiovascular co - morbidities, metabolic acidosis and acute respiratory distress syndrome led to poor outcome. age below 2 years, shock, fluid overload, need for mechanical ventilation, multi - organ failure and late referral predicted poor outcomes in a study from kuwait. children with aki may have long - term residual renal injury e.g., microalbuminuria, hypertension or elevated creatinine levels. lack of information on the long - term outcome does not permit evaluation of the impact of mild aki on renal function. secondly, the study was conducted at a tertiary hospital ; the clinical profile of patients would be affected by a referral bias. we did not compare children with aki and those without aki as this was not the objective of our study. thirdly, data regarding non aki patients was not collected and hence increased odds of mortality with aki could not be analyzed. finally, the study was not powered to examine predictors of mortality in aki as this was not the primary outcome variable ; and larger studies would be required. in summary, we emphasize that the incidence of aki is high in critically sick hospitalized children. a clearer understanding of the long - term outcomes of this condition would allow optimization of follow - up strategies. the present prospective observational study from a tertiary center in pondicherry, southern india found the incidence of aki to be 25.1% in critically ill - children admitted to the picu. the mortality was 46.3% in children with aki admitted to the picu and 20.7% of children with aki who survived (majority being aki stage 3), had partial renal recovery at discharge. the etiological profile of aki was dominated by infections, including pneumonia, sepsis, meningoencephalitis and tropical febrile illnesses. on multivariate logistic regression analysis, requirement of mechanical ventilation was found to be an independent predictor of fatality in children with aki. | background : although the term acute renal failure was replaced by acute kidney injury (aki) recently, there is a paucity of data on the incidence and profile of aki in critically ill children from the developing world.objectives:the objective of this study is to determine the incidence, etiology, short term outcome and predictors of fatality in critically ill children admitted to the pediatric intensive care unit (picu) with aki, aged 1 month to 13 years.materials and methods : in this prospective observational study, from june 2010 to march 2011, 215 children admitted to the picu were screened for aki, defined according to the aki network criteria. the patients with aki were followed - up until discharge / death. their clinical and biochemical data were recorded.results:the incidence of aki among 215 patients screened was 54 (25.1%). the common etiologies were infections, [34 (62.9%) ], acute glomerulonephritis (7.6%), snake envenomation (5.7%), hemolytic uremic syndrome (3.8%) and congestive cardiac failures (3.8%). among infections, pneumonia and septicemia constituted 26.5% each, meningoencephalitis accounted for 23.5%, and dengue, scrub typhus, tuberculosis and malaria constituted 9.3% of children with aki. 27.8% of patients required dialysis. overall mortality was 46.3%. on logistic regression analysis, requirement of mechanical ventilation was an independent predictor of fatality in aki.conclusions:besides the high incidence of aki in critically ill - children admitted to the picu (25.1%), the condition was associated with adverse outcomes, including high mortality (46.3%) and need for dialysis (27.8%). infections dominated the etiological profile. requirement of mechanical ventilation predicted an adverse outcome in our patient population. |
chronic heart failure (chf) is the common end - result of many cardiovascular diseases. despite the improvements in the methods of prevention and the new strategies in the treatment of patients, the incidence of chf is increasing in many countries.1, 2 in recent years, the focus of research for the treatment of chf patients has been on drug therapies and devices, while other factors may affect the rise in the number of chf patients. for example, oxidative stress may contribute to the pathogenesis of chf;3, 4 and some clinical and experimental studies have shown that in chf patients, free radical formation is increased and antioxidant defenses are reduced.3, 5, 6 also, dietary trials that assessed the impact of high intakes of natural antioxidants showed that the incidence of chf was reduced.7, 8 selenium (se), an essential trace mineral, is mainly obtained from seafood, meat, and cereals. se deficiency has been identified as a major contributing factor in the pathogenesis of certain chf syndromes. for instance, in areas with low soil se contents such as eastern china and western africa, se deficiency is associated with cardiomyopathy.9, 10 moreover, there are cases of cardiomyopathy associated with low se levels in malnourished hiv - infected patients in western countries11 and in patients on chronic parenteral nutrition.12 it has also been reported that serum levels of se were lower in idiopathic dilated and also ischemic cardiomyopathy patients compared with levels in healthy controls.13 on the other hand, in another report, the mean serum se levels in 30 patients with idiopathic cardiomyopathy did not show any difference from those of healthy individuals.14 it has been suggested that se may be involved in the deconditioning of skeletal and cardiac muscles and in chf symptoms such as fatigue and low exercise tolerance, rather than in ventricular dysfunction.15, 16 regarding the controversy over the prevalence of se deficiency among chf patients, the present study aimed at assessing serum se levels in a relatively large population of advanced chf patients and comparing them with those of healthy controls. this study was performed at mazandaran heart center, sari, iran, during an 11-month period beginning in march 2010. all the patients had chf symptoms and a left ventricular ejection fraction (lvef) of < 0.40. in the chf group, 39 patients had chronic atrial fibrillation (af) and 38 patients were in sinus rhythm. the patients were on medications, including angiotensin converting enzyme (ace) inhibitors, diuretics, digitalis, beta blockers, and angiotensin receptor blockers (arbs). the calculation of the lvef was according to simpson s rule.17 the healthy controls were recruited from the family members of the patients at the hospital or those undergoing cardiovascular screening at the clinic. for the control group, in addition to echocardiography, we took a complete medical and drug history and performed a complete physical examination. none of the control group members had chronic diseases such as diabetes mellitus, hypertension, and hyperlipidemia. nearly all the patients and controls were permanent residents in the northern part of iran at the time of study. the volunteers did not have a history of consuming ace inhibitors, arbs, digitalis, and diuretics and did not receive any se - containing supplements within the prior three weeks. a 10 ml blood sample was taken from the patients and the control group, which was then heated in special tubes of water bath (37 c) for 1 h. the samples were centrifuged (1500 rpm) and frozen at 20 c after serum isolation. varian aa240fs atomic absorption spectrometer with the graphite tube atomizer (mulgrave victoria, australia) was used to determine se concentrations. five hundred micro liters of the sample were diluted with 2 ml of the trinon - ascorbic acid reagent. a 50 ppb standard was prepared by diluting the 1000 ppm standard, with distilled water. the limit of measurement was 5 g / l.18 statistical analysis was performed using the spss 16 software. an independent samples t - test was used for the comparison of the quantitative variables between the patient and control groups. a p value < 0.05 was considered significant, and the power of study was 10% (z1 = 1.28). the demographic, echocardiographic data, and se levels of heart failure and control groups are shown in table 1. the mean values of left atrial areas (laa) and left ventricular systolic and diastolic diameters were higher in patients than in the controls, which was statistically significant. also, the average heart rate and systolic and diastolic blood pressures of the patients were higher than the values in the control group and it was statistically significant. as is presented in figure 1, the median se concentrations in the chf patients were not significantly different from those of the control group. also, we did not find any statistically significant difference between the se concentrations in the chf patients with atrial fibrillation and those who had sinus rhythm (p value = 0.39). we also made separate comparisons between the se concentrations in patients with and without af and those of the control group, with the se concentrations showing no statistically significant difference between the groups (figure 2). there was no correlation between serum se concentrations and ef in both the normal group and chf patients (p value = 0.96 and 0.99 ; r = 0.006 and 0.002 for patients and healthy volunteers, respectively). this is the first study in the northern part of iran to compare se concentrations in a relatively large number of chf patients with those of a control group. it had previously been shown that in both chf and healthy groups there was a similar relationship between dietary and blood se levels.14 the northern part of iran has a mediterranean climate and diet. the mediterranean diet is a se - rich diet including seafood and vegetables, and it is considered a cardioprotective diet.8 the relatively wider dispersion of se concentrations in chf patients may partially be related to the consumption of a wider range of se - containing food. it is now widely accepted that diet - derived antioxidants may play a role in the development, progress, and prevention of chf. for instance, some clinical studies have suggested that chf may be associated with increased free radical formation and reduced antioxidant defenses.15, 19, 20 still, it is important to know whether any specific micronutrient deficiency might have a causal relationship with chf and/ or whether it can aggravate chf symptoms. for example, hypozincemia in chf patients may be an effect of diuretic drugs, but there are no definite data on the clinical effect of zinc supplementation in these patients. nevertheless, there are some chf syndromes in which se deficiency has been identified as a contributing factor in the etiology. for example, in china an endemic cardiomyopathy called keshan disease has been illustrated to be the result of se deficiency.10, 21 but, in keshan disease se levels correlate with the clinical severity of chf, rather than the degree of left ventricular dysfunction. interestingly though, se supplementation resulted in a reduced mortality rate.12, 21, 22 in endemic areas, however, after raising se levels in residents, clinically latent cases were still found. thus, although se deficiency can also influence the clinical severity of keshan disease, it is not the specific etiologic factor for the occurrence of it. also, in western countries, cases of congestive cardiomyopathy associated with low serum vitamins and trace elements have been reported in malnourished hiv - infected patients and in subjects on chronic parenteral nutrition.15 in a relatively small number of chf patients, de lorgeril. found lower dietary intake and blood levels of se compared with those of healthy controls.15 also, blood levels of se were lower in chf patients. there was a positive correlation between se intake and blood levels, and the low dietary intake accounted for the low blood se levels. in our study, we did not find any statistically significant difference regarding the blood se levels between the chf and healthy controls. also, the blood se levels in both chf and healthy controls in our study were higher than those in the study by lorgeril. thus, we think that the mediterranean diet in the northern part of iran, which includes seafood and vegetables, has resulted in normal, rather than deficient, blood se levels in this area. in addition, in our study, se concentrations did not show any association with the severity of ventricular dysfunction assessed by echocardiography. on the other hand, blood se was strongly related to maximum oxygen consumption and exercise tolerance in chf patients,15 and also in keshan disease even a mild deficiency in se could influence the severity of the disease. be that as it may, the mechanism by which se deficiency results in some chf symptoms is not well defined and it has been suggested that se may be involved in skeletal and cardiac muscle deconditioning rather than in left ventricular dysfunction.16, 21 se is an essential trace element. its primary role is that of an antioxidant in the enzyme glutathione peroxidase (gp), the main intracellular antioxidant. selenoprotein p is another selenoprotein and its primary role is as an extracellular antioxidant.23 also the se - dependent thioredoxin reductase system may be involved in ascorbate regeneration.24 consequently, in addition to their role in vascular endothelial function,25 selenoproteins act as antioxidants. considering the fact that chf is associated with peripheral vasoconstriction and impaired skeletal muscle metabolism,26 se - dependent systems are very important defense mechanisms in humans. however, there are some limitations to the present study. for example, the relationship between dietary and blood se levels in chf and healthy groups has previously been shown. in our study, we did not have a definite dietary comparison between patients and controls ; chf patients may have been advised to take more vegetables and seafood after developing the disease. we do realize that this amount of the power is really low ; it is worthy of note, however, that choosing the number of cases was based on the previous similar studies (study of sahin.,13 heart failure : n = 54, healthy volunteers : n = 30 ; study of de lorgeril.,15 heart failure : n = 21, healthy volunteers : n = 18). the number of cases included in our study was 77 patients and 73 healthy volunteers, which was higher than those in the aforementioned studies. the low power of our study can be explained by the great sd of the se levels, especially in heart failure patients. in conclusion, although we did not find a causal relationship between se and chf in the northern part of iran, we recommend that monitoring dietary determinants can be helpful in the treatment and management of chf patients in all areas. | backgroundselenium (se) is an essential trace element mainly obtained from seafood, meat, and cereals. se deficiency has been identified as a major contributing factor in the pathogenesis of certain congestive heart failure (chf) syndromes. since there is controversy over the prevalence of se deficiency among patient with chf, the aim of this study was to assess the serum se concentrations in patients with chf and compared them with the se status of healthy controls.methods:the study included 77 patients (age, 68.4 10.4 years old ; 40.3% female) and 73 healthy volunteers (64.9 4.7 years old ; 35.6% female). a complete medical / drug history and physical examination were performed for all patients and healthy volunteers. all patients had symptoms and signs of chf and had a left ventricular ejection fraction (ef) of < 40% obtained by echocardiography. the se concentration was assessed by atomic absorption spectrometer with the graphite tube atomizer. the limit of measurement was 5 g / l.results : the se concentrations in chf patients did not show a significant difference from those of healthy controls (185.9 781.2 g / l vs. 123.3 115.5 g / l, respectively ; p value = 0.499). there was no correlation between serum se concentrations and ef in both the normal group and the patients with heart failure (p value = 0.96 and 0.99 ; r = 0.006 and 0.002 for patients and healthy volunteers, respectively).conclusion : in this study, serum se levels in chf patients were similar to those of controls and the se concentrations did not correlate with the degree of left ventricular dysfunction. |
it is a significant challenge even to the most experienced anaesthesiologist to intubate patients in whom the movement of the cervical spine is not desirable or restricted. in cases of cervical spine immobility or instability, the use of direct laryngoscopy is reserved : it requires flexion of the cervical spine and atlanto - occipital extension for alignment of the oral, pharyngeal and laryngeal axis to create a direct line of vision from the mouth to the vocal cords. tracheal intubation in patients with suspected neck injuries should achieve two contradicting goals : sufficient laryngeal exposure and the least cervical spine movement. as the former involves movement of the cervical vertebrae, intubation has to be performed using cervical spine immobilisation to prevent exacerbation of spinal cord injuries. protective measures to avoid deleterious compression forces on the spinal column include application of rigid collar, a forehead tape and manual - in - line stabilisation (mils). application of cervical collars may reduce cervical spine movements, but it hinders tracheal intubation with the standard laryngoscope. the cervical collar also significantly reduces the mouth opening, rendering laryngoscopy difficult. besides, the neck collar lifts up the chin and tips the larynx anteriorly. removing the anterior portion of the collar can facilitate tracheal intubation. however, this jeopardises the safety of the cervical spine. fibreoptic intubation is the most reliable method in patients with cervical trauma, but it may be difficult in patients with restricted neck movement. the other drawbacks of fiberoptic intubation are lack of availability of equipment, requires lack of expertise in its use and difficulty in using it if the patient is not co - operative or if there is blood or secretions in the airway. it possesses considerable advantages in the setting of cervical spine immobilisation when direct laryngoscopy is difficult or not recommended. it provides a full view of the glottis without requiring to align the airway axis. in addition, the airtraq laryngoscope also appears to cause less cervical spine movements during tracheal intubation when compared with the macintosh or mc coy laryngoscopes. the airtraq facilitates tracheal intubation with the neck in neutral position, which is similar to the neck position maintained by a rigid cervical collar. however, a rigid cervical collar in combination with forehead strapping and mils virtually obliteratess even the small neck movements which normally facilitate airway insertion. the present study evaluates the efficacy of airtraq in patients undergoing cervical spine immobilisation with rigid cervical collar and mils, simulating the situation of cervical trauma, and compares it with the mc coy laryngoscope. the study was designed as an open - labelled, randomised, cross - over trial. following institutional ethics committee 's (iec) approval, informed consent was obtained from 60 asa i and ii patients, aged between 18 and 50 years, belonging to either gender and undergoing elective surgery requiring general anaesthesia with oral endotracheal intubation. patients with anticipated difficult airway (mallampatti grade iv, mentohyoid distance (mhd) 42 cm), obese (body mass index (bmi) > 30) patients, patients with risk of pulmonary aspiration of gastric contents, pregnant patients, and patients with cervical spine pathology, airway distortion or trauma were excluded from the study. an appropriate - sized rigid cervical collar (ambulance collar, mgrm medicare limited, hyderabad, india) was placed as per manufacturer 's instructions. standard monitoring included ecg, non - invasive arterial pressure, spo2, and measurement of end - tidal carbon dioxide. before induction of anaesthesia, all patients were given fentanyl 11.5 g / kg and glycopyrrolate 5 g / kg i.v. after induction of anaesthesia, all patients were manually ventilated with sevoflurane 2.02.5% in oxygen, and vecuronium 0.1 mg / kg was administered. each patient was intubated twice, once using airtraq and the other time with mc coy laryngoscope. patients were randomised using online randomisation (http : //www.randomization.com) to undergo airtraq intubation first or intubation with mc coy first to reduce the influence of one intubation on the other. anaesthesiologists with adequate experience (> 40 intubations) in both techniques performed all intubations. the trachea was intubated with a 7.5-mm tracheal tube in females and an 8.0-mm tracheal tube in males. in case of mc coy, if cormack lehane grade 1 or 2, intubation proceeded without using hinge of the laryngoscope. if cormack lehane grade > 2, then hinge of mc coy was used to improve laryngeal visualisation. if glottic view did not improve even with hinge, an intubating bougie was used. the ease of introduction of mc coy blade or airtraq in the presence of collar was graded for both techniques on a likert scale from 2 to + 2. a modification intubation difficulty score (ids) described by adnet and colleagues to suit mc coy and airtraq aided intubation as given in appendix 1 was noted. the intubation time (time from removal of face mask for intubation to successful intubation and connection of circuit to the endotracheal tube) was noted for both techniques. the intubating anaesthesiologist graded the ease of intubation for both techniques on a visual analogue scale from 1 to 10, 10 being most difficult or failed intubation and 1 being very easy intubation. if introduction of the intubating device was not possible or there were more than three attempts for intubation or intubation time was more than 120 sec, it was considered to be a failure. intubation was then performed with the second technique. in case of failure with the second technique also failure to intubate (> 3 attempts or > 120 sec), episodes of desaturation (spo2 5. fewer manoeuvres were required with airtraq to improve the glottic exposure, compared to laryngoscopy [table 3 ]. there was no difference in the incidence of complications in the two groups [table 2 ]. removal of cervical collar was not required in any of the patients with both the techniques. demographic data and airway characteristics of patients in the study comparison of intubation between laryngoscopy and airtraq comparison of intubation difficulty score between laryngoscopy and airtraq spinal cord injury has been reported in association with the airway management of patients with cervical spine instability in whom cervical spine immobilisation was not performed. mils prevents head extension and neck flexion, which are necessary for optimal alignment of the three airway axes and exposure of the vocal cords using direct laryngoscopic techniques. the use of a rigid collar, tape and sandbags may result in an increased incidence of grade 3 and 4 laryngoscopic views (up to 64%) with conventional laryngoscopy owing to the combination of decreased inter - incisor distance and cervical spine immobility. consequently, manoeuvres to stabilise the neck in patients at risk of cervical spinal injury may result in failure to secure the airway, which may result in substantial morbidity and even mortality in this patient group. these issues have prompted, in part, the development of a number of alternative approaches to securing the airway in patients at risk of cervical spine injury. we evaluated the relative efficacies of this intubation technique when used by experienced anaesthesiologist in the clinical setting of cervical spine immobilisation using rigid cervical collar and mils and compared it with the commonly used mc coy laryngoscope. airtraq has an exaggerated curvature of the blade and an internal arrangement of optical components, which provide a high - quality view of the glottis without the need for alignment of the oral, pharyngeal and tracheal axes, and therefore requiring application of less force during laryngoscopy, less external neck pressure and less manoeuvres to facilitate intubation, as seen in this study. there are published reports that airtraq intubating device is superior to laryngoscopy in patients with normal airways and difficult airway scenarios simulated in manikins. airtraq has also been shown to produce less haemodynamic stimulation, a potentially important advantage in certain clinical situations. in a study of morbidly obese patients where tracheal intubation was compared using the airtraq and macintosh laryngoscopes, the mean time taken for tracheal intubation was found to be shorter in the airtraq group. there was no statistically significant difference in the duration of intubation compared to that with mc coy blade. however, it should be noted that the authors were more familiar with the latter technique. the presence of cervical collar could have also resulted in slightly longer intubation times with airtraq as compared to earlier studies. studies have demonstrated that the airtraq reduces the difficulty of tracheal intubation in patients undergoing cervical spine immobilisation with mils when compared with the macintosh laryngoscope. koh. reported higher success rate of intubation with airtraq in patients with cervical immobilisation with collar. arslan. evaluated the effectiveness of the airtraq and ctrach in lean patients with simulated cervical spine injury after application of a rigid cervical collar. our study confirms and extends these findings to application of cervical immobilisation as recommended in advanced trauma life support (atls) guidelines using rigid cervical collar along with mils and head taped to the table. this study demonstrated that the airtraq reduced the ids, improved the cormack and lehane grade and reduced the number of optimisation manoeuvres compared with the laryngoscopy. the reduced interdental distance due to rigid cervical collar did not hamper the intubation quality with airtraq. this was the principal reason for the increased duration of tracheal intubation in some of these patients. there were two failures to intubate in the airtraq group, which were not related to poor view of vocal cords but to an inability to advance the tracheal tube within 120 sec despite using the manoeuvres described. these patients were intubated using mc coy laryngoscope and bougie. however, the ids was high. another problem with airtraq was fogging on the distal lens which reduced the image quality. difficulty in intubation despite good glottic visualisation is a problem reported with most video laryngoscopes. most video laryngoscopes can achieve a better view of the glottis and have a similar success rate. overall, the time to tracheal intubation was not different between the video laryngoscopes and direct laryngoscopy. different video laryngoscopes such as glidescope ranger, storz c - mac, ambu pentax aws, airtraq and mcgrath series 5 were compared with macintosh blade for ease and time of intubation in a cervical spine immobilised manikin. the time to first effective ventilation was fastest when using macintosh laryngoscope (21.07.6 sec). it was 33.223.9 sec with airtraq, 32.414.9 sec with pentax airway scope, and 34.123.9 sec, 101.7108.3 sec and 46.359.1 sec with storz c - mac, mcgrath series 5 and glidescope ranger, respectively. the investment cost is lower with airtraq when compared to other video laryngoscopes. we acknowledge that it is impossible to blind the anaesthesiologist to the device being used, thus there is a potential bias. furthermore, some subjectivity is involved with certain measurements used in this study, such as laryngoscopic grading. though cormack and lehane classification has an advantage of being used widely in clinical practice, the appropriateness of using this classification with indirect laryngoscopes is open to question. have considered only the cormack and lehane grading of glottis visualisation in their study using airtraq. in addition to cormack and lehane glottic view classification, we have also used ids which is more objective and found to have a good agreement between subjective indices of difficulty of intubation. secondly the results seen may differ in the hands of less experienced users. finally, the relative efficacy of these devices in comparison to other promising devices has not been determined. airtraq improves the ease of intubation when compared to laryngoscopy with mc coy blade in patients immobilised with cervical collar and mils simulating cervical spine injury with the same rapidity as mc coy laryngoscope and can aid intubation without the need to remove the cervical collar. | background : it is difficult to visualise the larynx using conventional laryngoscopy in the presence of cervical spine immobilisation. airtraq provides for easy and successful intubation in the neutral neck position.objective:to evaluate the effectiveness of airtraq in comparison with the mc coy laryngoscope, when performing tracheal intubation in patients with neck immobilisation using hard cervical collar and manual in - line axial cervical spine stabilisation.methods:a randomised, cross - over, open - labelled study was undertaken in 60 asa i and ii patients aged between 20 and 50 years, belonging to either gender, scheduled to undergo elective surgical procedures. following induction and adequate muscle relaxation, they were intubated using either of the techniques first, followed by the other. intubation time and intubation difficulty score (ids) were noted using mc coy laryngoscope and airtraq. the anaesthesiologist was asked to grade the ease of intubation on a visual analogue scale (vas) of 110. chi - square test was used for comparison of categorical data between the groups and paired sample t - test for comparison of continuous data. ids score and vas were compared using wilcoxon signed ranked test.results:the mean intubation time was 33.27 sec (13.25) for laryngoscopy and 28.95 sec (18.53) for airtraq (p=0.32). the median ids values were 4 (interquartile range (iqr) 16) and 0 (iqr 01) for laryngoscopy and airtraq, respectively (p=0.007). the median cormack lehane glottic view grade was 3 (iqr 24) and 1 (iqr 11) for laryngoscopy and airtraq, respectively (p=0.003). the ease of intubation on vas was graded as 4 (iqr 35) for laryngoscopy and 2 (iqr 22) for airtraq (p=0.033). there were two failures to intubate with the airtraq.conclusion:airtraq improves the ease of intubation significantly when compared to mc coy blade in patients immobilised with cervical collar and manual in - line stabilisation simulating cervical spine injury. |
we obtained data on persons with hiv / aids and tb from the national databank at the centers for disease control (cdc taiwan) of the department of health, taiwan. coinfection with hiv and tb was defined as hiv infection in persons in whom tb was later diagnosed. a total of 660 persons with both hiv and tb were reported during 19932006. we used microsoft excel xp spreadsheet (microsoft, redmond, wa, usa) and sas version 9.1 (sas institute inc., the goodness - of - fit test with type i error = 0.05 was used to examine differences in demographic, clinical, and behavioral characteristics of persons with hiv and tb coinfection during 19932006. multivariates for analysis were sex and age, results of sputum smear and sputum culture, pulmonary radiographic diagnosis, tb types (extrapulmonary and nonextrapulmonary), mode of hiv transmission, sexual behavior, compliance with haart, and use of the surveillance system (table 1). we used the kaplan - meier method (4) from sas to evaluate and compare the effect on survival rates of different factors in persons coinfected with hiv and tb 1 year after reported tb diagnosis. goodness - of - fit test with type i error = 0.05 used to examine differences in demographic, clinical, and behavioral characteristics. kaplan - meier analysis yielded the following results : 63% of persons coinfected with hiv and tb survived during 19931996 ; 78% survived during 19982000 ; and 93% survived during 20022006 (p<0.0001) (figure). we then applied cox proportional hazards modeling (5) to each variable to assess the effect on survival rates after implementation of haart and the surveillance system. age < 45 years, negative sputum smear, availability of free haart, and implementation of the national surveillance system substantially increased survival rates of persons coinfected with hiv and tb (table 2). kaplan - meier analysis of survival of hiv - infected patients with tuberculosis in taiwan during 3 different periods : before free highly active antiretroviral therapy (haart) was available (19931996, black line) ; b) after free haart was available but before the national web - based reporting and management surveillance system was implemented (19982000, red line) ; and c) after free haart and the surveillance system were available (20022006, blue line). many factors can increase survival rates of hiv - infected persons, such as haart (69), prevention of opportunistic infections, patient attitude, healthcare worker knowledge, and promotion of health education. our data indicate that national web - based surveillance reporting and management, coupled with the availability of free haart, increase survival rates of persons coinfected with hiv and tb (p<0.0001). taiwan s national web - based surveillance system enables healthcare workers to follow, record, and understand the conditions of patients without geographic limitations. physicians, public health nurses, health administrators, and other healthcare professionals in local through federal government agencies can use the system to follow up and manage the condition of persons coinfected with hiv and tb. for example, public health nurses from national healthcare centers visit such patients regularly, record treatments, and assess their conditions and compliance with therapy ; staff from central health department monitor and supervise the condition of each patient through the system. in this way, the system may increase patients compliance and thus their survival rates (1014). | in 1997, taiwan made highly active antiretroviral therapy (haart) available without cost to hiv - infected persons ; in 2001, a national web - based surveillance system was implemented. healthcare workers use the system to monitor patients ' conditions and can intervene when necessary. free haart, coupled with the surveillance system, appears to have increased survival rates of hiv - infected persons with tuberculosis in taiwan. |
small organic molecules with specific interactions with dna have become antitumor, antiviral, and antibiotic drugs [1, 2 ]. duplex dna - binding drugs interact in two main ways, through groove binding and through intercalation. medicinal chemistry has made a considerable effort in searching for and testing of a large number of drugs with increased selectivity to a range of dna sequences or structures. more recently, some of this interest has moved to the search of new ligands for g - quadruplexes. this structure motif is formed by the planar association of four guanines in a cyclic hoogsteen hydrogen bonding tetrad. guanine - rich sequences form g - quadruplex structures and have been found in telomeres and in transcriptional regulatory regions of critical oncogenes such as c - myc and c - kit [5, 6 ]. ligands that selectively bind and stabilize these structures have become anticancer drugs of interest. the g - quadruplex stabilization occurs in most cases by - stacking and electrostatic interaction. g - quadruplex ligands are normally planar aromatic molecules that are prone to stacking with g - tetrads. some of them are also positively charged or have hydrophilic groups to favor electrostatic interaction. although there is a long way to go in the development of potent drugs that target g - quadruplexes, some promising lead compounds have been achieved. several ligand structures have been studied, such as anthraquinones, cationic porphyrins, perylene derivatives, and a large number of compounds. among the acridine compounds, 3,6,9-trisubstituted acridines have inhibitory activity in the nanomolar range and they have entered preclinical studies [8, 10, 11 ]. in previous studies we described the preparation of sequence specific oligomers of dna - intercalating drugs using protocols based on solid - phase synthesis in an attempt to facilitate the preparation of compounds with improved dna - binding selectivity [12, 13 ]. it has been proposed that bis- and tris - intercalating drugs show promising activity and selectivity [14, 15 ]. here we described solid - phase synthesis protocols for the preparation of several acridine oligomers linked through 2-aminoethylglycine units as well as their dna - binding properties. although the acridine derivatives described in this study are not cytotoxic, they show a clear affinity for several dna g - quadruplex structures, especially those sequences found in the promoter regions of c - myc and bcl-2 [17, 18 ] oncogenes. the phosphoramidites and ancillary reagents used during oligonucleotide synthesis were obtained from applied biosystems (usa) and link technologies ltd. the slide - a - lyzer mini dialysis units 3500 mwco were purchased from pierce. 2-(acridine-9-carboxamide)acetic acid was prepared by reaction of acridine-9-carboxylic acid with glycine methyl ester and subsequent saponification of the methyl ester as described. boc-(2-aminoethyl)glycine(fmoc) (boc - aeg(fmoc)-oh) was obtained from iris biotech and fmoc - glycine (fmoc - gly - oh) was obtained from bachem. oligonucleotide sequences (table 1) were prepared on an automatic applied biosystems 3400 dna synthesizer on 1 mol (cpg resin) scale using commercially available 2-cyanoethyl phosphoromidites. after the assembly of the sequences, oligonucleotide - supports were deprotected using 32% aqueous ammonia at 55c for 16 h. ammonia solutions were concentrated to dryness and the residue was desalted by a nap-10 (sephadex g-25) column. acridine dimers and trimers (14, figure 1) were prepared with the 2-aminoethylglycine scaffold, which allows the growth of a polyamide skeleton on solid - phase and the following incorporation of acridine unit. the assembly of 2-aminoethylglycine derivatives was carried out on methylbenzhydrylamine (mbha) polystyrene-1%-divinylbenzene solid support applying an fmoc / boc hybrid strategy using boc - aeg(fmoc)-oh, fmoc - gly - oh, and acridine-9-carboxylic acid as building blocks (figure 2). fmoc - sarcosine - oh (5 eq) was coupled to the resin using standard coupling conditions (5 eq. diea, 1 h), then the fmoc group was removed (20% of piperidine in dmf, 30 min), and boc-6-aminohexyl hemisuccinate (boc nh(ch2)6ococh2ch2cooh) was coupled (5 eq r - cooh, 5 eq. pybop and 10 eq. the residual unreacted amino groups were acetylated with 5 eq. of acetic anhydride and 5 eq. next, the boc group was removed (40% trifluoroacetic acid in dichloromethane) and the 2-aminoethylglycine skeleton was synthesized by repetitive couplings of boc - aeg(fmoc)-oh until reaching the desired dimer or trimer compound. the last boc group of the sequence was removed and the resulting amino group was acetylated (5 eq. once the aminoethylglycine backbone was built, the fmoc - protecting groups of the side chains were removed (20% of piperidine in dmf, 30 min), and fmoc - gly - oh followed by 9-acridine carboxyl acid was coupled to the support. the progress of the coupling reactions was followed by ninhydrine test and by uv monitoring of the 9-methylene-9h - fluorene released during deprotection, which allowed optimization of the coupling conditions. the acridine dimer 1 and trimer 3 were obtained by treatment of the appropriate solid supports with hf anhydrous at 0c. finally the acridine dimer 2 and trimer 4 were obtained by treatment of dimer 1 and trimer 3 respectively with 32% aqueous ammonia (1 h, 55c). good yields and purities were obtained for the products (around 85% for 1 and 2, 75% for 3 and 4). hplc and maldi - tof spectra are shown in supplementary material available onlie at doi : 10.4061/2010/489060. the compounds were analyzed by maldi - tof, 1 [m+na ] = 1080.3 (expected 1054.2), 2 [m ] = 885.2 (expected 884.0), 3 [m ] = 1418.9 (expected 1416.5), and 4 [m ] = 1247.4 (expected 1246.4). maldi - tof spectra were obtained using a perseptive voyager detmrp mass spectrometer, equipped with nitrogen laser at 337 nm using a 3 ns pulse. the matrix used contained 2,5-dihydroxybenzoic acid (dhb, 10 mg / ml in water). analytical hplc was performed using xbridge ost c18 (waters), 2.5 m, 4.6 50 mm column using a 10-minute linear gradient from 9% to 45% b, flow rate 1 ml / min ; solution a was 5% acn in 0,1 m aqueous teaa, and b 70% acn in 0.1 m aqueous teaa. fluorescence spectra were recorded using a jasco fp-6200 spectrofluorometer equipped with a peltier temperature controller, em = 435 nm (exc = 252 nm). uv spectra were recorded using a jasco spectrophotometer v-650, max = 252 nm, 360 nm. the in vitro cytotoxicity of the compounds (figure 1) was evaluated by colorimetric assays with tetrazole salts (mtt) on jurkat clone e6 - 1 (human leukemia), glc-4 clone (human lung carcinoma) cell lines, and one mouse fibroblast cell line (niht-3t3). glc4 and jurkat cell lines were cultured in rpmi and nih3t3 in dmem and supplemented with 10% fetal calf serum, 10000 u / ml penicillin, 10 g / ml streptomycin and 200 mm l - glutamine. cells were grown in a humidified atmosphere of air containing 5% co2 at 37. cells were plated in triplicate wells (1.510 cells well) in 100 l of growth medium in 96-well plates and proliferate for 24 h and then treated with increasing concentrations of acridine oligomers. after 72 h of incubation, 10 m of mtt (5 mg / ml in phosphate buffer saline 10%) was added for an additional 4 h. the absorbance at 570 nm was measured on a multiwell plate reader after addition of 100 l of isopropanol : 1n hcl (24 : 1). cell viability was expressed as a percentage of control and ic50 was determined as the concentration of drug that produced a 50% reduction of absorbance at 570 nm. 100 l of a 50 m oligonucleotide (table 1) in potassium phosphate buffer (185 mm nacl, 185 mm kcl, 2 mm nah2po4, 1 mm na2edta, 6 mm na2hpo4 at ph 7) was introduced into a separated dialysis unit. all 12 dialysis units were then placed in the beaker containing the 1 m solution of the appropriate acridine derivative. sds is usually added to denature the dna sample and release the acridine oligomer, but in our case the presence of k ions induced the formation of a white precipitate, which interfered with the measurement of the fluorescence spectra of the samples. in order to measure the compound retained in the dialysis unit, samples were treated with snake - venom phosphodiesterase to degrade the dna and release the acridine oligomer. 350 l of potassium phosphate buffer (without edta), buffer at ph 8.5, 50 l of 100 mm mgcl2, and 1 m of snake venom phosphodiesterase solution were added for an additional overnight incubation at 37c. finally, the fluorescence of each sample was measured (ex and em were set to 252 and 435 nm, resp.). the study of the interaction equilibrium of acridine derivatives and oligonucleotides consists of recording the fluorescence spectra of a 0.2 m solution of the acridine derivative after the addition of increasing amounts of oligonucleotide (from 0 to 10 m) in potassium phosphate buffer (185 mm nacl, 185 mm kcl, sodium phosphate, 1 mm edta, ph 7). these experiments were carried out by adding small volumes of an oligonucleotide stock solution to the 0.2 m solution of the acridine derivative. after 24 h the emission spectra of the resulting solutions were recorded from 300 to 500 nm at 252 nm excitation wavelength at 25c. the macroscopic binding constant (k) corresponding to the reaction (1)dna+ligand interaction complex was calculated from the multivariate analysis of fluorescence data recorded in the range 300390 nm using the hard - modeling program equispec. this program performs a nonlinear least squares optimization of k and of the pure fluorescence spectra corresponding to each of the species considered (dna, ligand, and interaction complex). a 1 : 1 stoichiometry dna : ligand for the interaction complex was assumed. the logarithms of the binding constants calculated are given as their weighted means with twice their standard errors (units of the least significant digit). an increasing amount of 1 (from 0 to 8 m) in potassium phosphate buffer (185 mm nacl, 185 mm kcl, sodium phosphate, 1 mm edta, ph 7) was added to a 1 m solution of the oligonucleotide. the cd spectra were recorded after 24 h on a jasco j-810 spectropolarimeter attached to a julabo f/25hd circulating water bath in 1 cm path - length quartz cylindrical cells, using a 50 nm / min scan rate, a spectral band width of 1 nm, and a time constant of 4 s. all the spectra were corrected with the buffer blank, normalized to facilitate comparisons and noise - reduced using matlab software. the synthesis of acridine dimer and trimers has been described previously using a boc-(2-aminoethyl)glycine derivative carrying the 2-(acridine-9-carboxamide)acetyl residue. an alternative method developed is to first assemble the boc-(2-aminoethyl)glycine backbone on solid - phase, and then the intercalating agent is added. this strategy is more convenient for rapid synthesis, as it is unnecessary to construct each intercalating monomer. thus, acridine oligomers were assembled using the methylbenzhydrylamine (mbha) resin by applying an fmoc / boc hybrid strategy and using boc-(2-aminoethyl)-(fmoc)glycine [boc - aeg(fmoc)-oh ] as building block (figure 2). the boc group was used to protect the aminoethyl group of each unit, which thus facilitated elongation of the backbone. fmoc was the semipermanent protecting group for the amino group of glycine through which the intercalating compound was introduced. it has been described that an intramolecular side reaction can lead to premature loss of the oligomers during the base treatment used to remove the fmoc group. thus, n - methylglycine (sarcosine) was incorporated between the amino - support and the succinyl linker. 6-aminohexanol was used to connect the succinyl linker and the oligomer backbone, as described for the synthesis of peptide nucleic acid (pna) oligomers. the (2-aminoethyl)glycine backbone was assembled by consecutive additions of boc - aeg(fmoc)-oh to obtain the dimer and trimer sequence. acetylation of the n - terminal position was carried out using acetic anhydride and a base. next, the removal of the fmoc group allowed the addition of a fmoc - glycine unit as a spacer, which was followed by the addition of acridine-9-carboxylic acid. the desired oligomers were not released from the support even after prolonged time and high temperatures. we therefore treated the supports with anhydrous hf to yield the acridine oligomers 1 and 3, which contain the sarcosyl succinyl linker. at this point hplc spectra showed a major peak that had the expected molecular weight for the oligomers 1 and 3 carrying the sarcosyl succinyl linker (see supplementary data). ammonia treatment of acridines 1 and 3 in solution now yielded the desired acridine dimer 2 and trimer 4 in excellent yields and purity (see supplementary data). the pka of the acridine ring of acridine-9-carboxylic acid and 2-(acridine-9-carboxamide)acetic acid was measured by uv titration. we therefore estimated that the acridine rings of compounds 14 are mainly unprotonated at ph 7.0. the in vitro cytotoxicity of the compounds was evaluated by colorimetric assays with tetrazole salts (mtt). this assay is based on the capacity of living cells to incorporate and reduce mtt. this reaction can be followed by the change of absorbance of the reduced and oxidized forms. this reaction is done by the action of the mitochondrial enzyme succinatehydrogenase, which is active only in living cells. the intensity of color is directly correlated with the number of living cells in the sample. no cytotoxicity activity was observed in compounds 14 at concentrations up to 50 m. in order to evaluate the selectivity of the compounds for dna structures, a competitive dialysis experiment was performed using 11 oligonucleotides (table 1) representing several nucleic acid structures. the more acridine accumulated in the dialysis unit indicates a higher binding affinity to the oligonucleotide present in the dialysis unit. as model for single stranded structures we used t20 and the c - rich complementary strand of bcl-2 (24bclc) this last oligonucleotide folds in an i - form quadruplex structure at acidic ph but has no structure in the conditions used in the dialysis (ph 7). as duplexes we used the self - complementary sequences dickerson - drew dodecamer (dickerson) and a 26 mer (ds26). a parallel triplex (tc triplex) and an antiparallel triplex (ga triplex) were also prepared by mixing a hairpin watson - crick sequence and the corresponding triplex - forming sequence. finally, the following g - quadruplex sequences were prepared : the tetramolecular parallel g - quadruplex tg4 t, the antiparallel thrombin - binding aptamer (tba), the human telomere sequence (ht24), and the promoter sequences of c - myc (cmyc) and bcl-2 (24bcl) [17, 18 ]. the buffer solution was similar to that described in but k was included (185 mm) to ensure the formation of the most stable g - quadruplex structures. at the end of the dialysis experiment, the amount of acridine derivative bound to the dna was analyzed by fluorescence measurement of the acridine compound. after dialysis, we observed that the spectra of the acridine derivatives of some samples (especially those from g - quadruplexes) differed greatly from the fluorescence spectra of the initial compounds. this difference is attributed to the interaction of the acridine derivatives with the g - quadruplex. in order to release the acridine oligomer, the presence of k ions resulted in the formation of a white precipitate with sds which did not allow the measurements of the fluorescence spectra. in order to solve this problem, the oligonucleotide was digested with snake venom phosphodiesterase at the end of the dialysis experiment. thus, the fluorescence spectra of the acridine derivatives were recorded with high accuracy without the interference of dna and without the use of sds. we measured the binding preferences of compounds 14 and the acridine monomers acridine-9-carboxylic acid and 2-(acridine-9-carboxamide)acetic acid (figure 1). unexpectedly, the addition of the glycine residue, used as spacer, induced a change in the affinity. 2-(acridine-9-carboxamide)acetic acid showed the highest affinity for the g - quadruplex sequences cmyc and 24bcl and less affinity for duplex ds26. dimer 1 (dimer with the sarcosylsuccinyl linker) has a similar profile as 2-(acridine-9-carboxamide)acetic acid. in this case, the g - quadruplex 24bcl is preferred to the cmyc sequence. surprisingly, dimer 2 (without the sarcosylsuccinyl linker) lost most of the selectivity although some residual higher affinity for g - quadruplex 24bcl was observed. in contrast, trimer 3 (trimer with the sarcosylsuccinyl linker) presented lower binding affinity than the other compounds. however, trimer 4 (without the sarcosylsuccinyl linker) recovered most of the affinity for 24bcl and cmyc showing a similar profile to that observed for dimer 1. dialysis experiments suggest that some of the acridine derivatives prepared have special affinity for 24bcl and cmyc g - quadruplexes. in order to confirm this observation, hence, increasing amounts of oligonucleotides 24bcl, ht24, cmyc, and dickerson were added to a solution with a fixed concentration of the acridine derivatives, and the fluorescence spectra were recorded at excitation wavelengths 252 and 360 nm. at both wavelengths, changes in the fluorescence spectra upon the addition of oligonucleotides were observed. figure 4 shows the changes in the fluorescence spectra of acridine dimer 1 at excitation wavelengths 252 nm when oligonucleotides 24bcl, ht24, cmyc, and dickerson were added. a dramatic increase in fluorescence intensity was observed around 360 nm upon addition of g - quadruplex dna sequences (24bcl, ht24, and cmyc). similar results were found with compounds 2, 3, and 4 as well as 2-(acridine-9-carboxamide)acetic acid (see supplementary data). interestingly, when the dickerson dodecamer was added, no changes in the fluorescence spectra were detected. the progressive modification of the fluorescence spectrum of these compounds reflects their interaction with the g - quadruplex. fluorescence data obtained at an excitation wavelength of 252 nm were analyzed with the hard - modelling equispec program in order to calculate the corresponding binding constants (table 2). the values of the logarithm of the binding constant (log k) obtained lie in the range 46, suggesting a weak interaction with dna. of all the compounds, dimer 1 showed the highest binding constants, thereby suggesting a stronger interaction with dna than the other compounds studied. however these values were slightly lower than the binding constants calculated for other similar ligands, such as the acridine monomers braco-19 (log k = 7.4) and bsu6048 (log k = 6.5) or a hemicyanine - peptide ligand (log k = 7.1), when interacting with human telomere quadruplex. changes in the fluorescence spectra at an excitation wavelength of 360 nm were also recorded. fluorescent emission at this wavelength 360 nm was much lower than that recorded at 252 nm ; so the fluorescent signal was low. upon addition of the oligonucleotide to a solution of compounds 14, the formation of a new maximum at 442 nm was observed (see supplementary data). although the fluorescent intensity was low, we could estimate the binding constant of the stronger interactions of compound 1 with 24bcl (7.2 0.4) and cmyc (5.5 0.2) sequences (see supplementary data). these values are in agreement with those recorded at an excitation wavelength of 252 nm (table 1). finally, cd spectra of the dna : ligand mixtures showed no significant differences in relation to those of dna (see supplementary data). this observation suggests that the dna g - quadruplex structure is not altered significantly upon binding of the acridine derivatives. in summary, here we have described a new optimized protocol for the synthesis of acridine oligomers with a (2-aminoethyl)glycine backbone. in this method, the boc-(2-aminoethyl)glycine backbone is first assembled on solid - phase, and then the intercalating agent is assembled on the backbone. this strategy is faster and more efficient than the one described previously and yields the desired oligomers with good yields. the succinyl linker attached to sarcosine was unexpectedly too stable and oligomers could not be directly released from the support by a single ammonia treatment. instead a two - step protocol was used obtaining the desired compounds and an intermediate oligomer carrying a long succinyl sarcosine chain at the c - terminal position. competitive dialysis experiments have shown differences on the affinity of acridine oligomers to g - quadruplexes. higher affinities are found in g - quadruplex sequences present on the promoter regions of c - myc and bcl-2 oncogenes. this affinity is modulated by the number of acridines and the presence of the succinyl sarcosine chain at the c - terminal position, dimer 1 and trimer 4 being the more relevant compounds for g - quadruplex binding. the monomer 2-(acridine-9-carboxamide)acetic acid also shows binding properties of interest and it is the simplest compound to be prepared. unfortunately, the compounds synthesized in this study did not have antiproliferative activity in spite of their affinity to quadruplex. this observation contrasts with other reported quadruplex - binding acridine derivatives, such as braco-19 [11, 30 ] which shows anticancer activity. the lower affinity to telomere g - quadruplex sequence and the larger size of the acridine derivatives described in the present study may hinder cellular uptake and may explain the absence of antiproliferative activity. depending on the substituents, the acridine nitrogen can be charged when bound to dna, and with the ring stacked on a g - tetrad, the charge will occupy a position similar to that of the potassium cation that stabilizes the g - quadruplex. the introduction of protonable side chains on the acridine ligand enhances binding by electrostatic interactions. in our case the acridines had no protonable substituents and the acridine nitrogen was not charged when bound to dna. for this reason, the affinity of oligomeric acridines to g - quadruplexes is due to the multimeric nature of the compounds as well as the addition of a glycine to acridine-9-carboxylic acid. an interesting possibility for future development is the introduction of protonable sites at the oligomeric acridines, which may increase solubility in water, affinity to target, and cellular uptake. the method described here will contribute to accelerating the preparation of potential active oligomeric compounds. | the synthesis of oligomers containing two or three acridine units linked through 2-aminoethylglycine using solid - phase methodology is described. subsequent studies on cell viability showed that these compounds are not cytotoxic. binding to several dna structures was studied by competitive dialysis, which showed a clear affinity for dna sequences that form g - quadruplexes and parallel triplexes. the fluorescence spectra of acridine oligomers were affected strongly upon binding to dna. these spectral changes were used to calculate the binding constants (k). log k were found to be in the order of 46. |
pseudohypoparathyroidism (php) was first reported in 1942 by albright who presented 3 cases of hypocalcemia with no response to parathyroid extract and albright hereditary osteodystrophy (aho) phenotype of short stature with round facies, obesity, brachydactyly, mental retardation, and subcutaneous calcification. subsequently it was proved that php is caused by resistance to action of parathyroid hormone (pth) in selective organs with monoallelic expression of gs (proximal renal tubules, thyroid, gonads, pituitary), later resistance to pth was reported in patients with php cases without aho phenotype and normal gs activity and it was classified as php type-1b (php1b) caused due to imprinting defects in gnas1 gene located on chromosome 20q13.3. classically, php1b is characterized by blunted nephrogenous cyclic - amp (camp) and phosphaturic response to exogenous pth. here we report a case of php1b confirmed by blunted urinary camp response to exogenous pth but had normal phosphaturic response on treatment with calcium and vitamin d. we discuss possible mechanisms to explain normal phosphaturic response. a 34-year - old male presented with recurrent carpopedal spasms of 4 years duration with perioral numbness, cramps, muscle twitching and generalized weakness for which he was evaluated 2 years back and found to have hypocalcemia, hyperphosphatemia, hypercalciuria, high pth without aho phenotype. on examination he was 184 cm tall with weight of 78 kg (bmi 23 kg / m). latent signs of tetany in the form of positive chvostek 's sign and trousseau 's sign were present. we decided to re - evaluate him in view of late age of onset of symptoms, absence of aho phenotype, and to confirm the diagnosis. investigations revealed normal renal parameters (serum creatinine, 88.4 mol / l) and normal serum magnesium (0.86 mmol / l), 25(oh)d levels (137.2 nmol / l) and raised pth (63 pmol / l). his bone mineral density (t score,0.5 at l1-l4 and 0.2 at femur neck) and thyroid function were normal (t3 0.87 ng / ml, t4 7.59 g / dl, tsh, 2.20 miu / l). in view of hypocalcemia with raised pth three possibilities were entertained idiopathic hypoparathyroidism with heterophile antibody against pth, php1b, or php2. for the first possibility his serum was precipitated with polyethylene glycol (peg) for detection of heterophile antibodies. his peg precipitated serum pth levels were also high (61.5 pmol / l). modified ellsworth howard test was planned to differentiate between php1b and php2. since this test was performed first time in this institute, it was planned in three subjects : one present case, one healthy age- and sex - matched control and another known case of postoperative hypoparathyroidism. the protocol starts with oral hydration with 200 ml water every 30 min starting at 0600 hours in the morning. starting from 0830 hours, timed urine collection is done every 30 min and blood samples are collected at midpoint of each till 1100 hours. from 1000 to 1010 hours, teriparatide is infused intravenously in the dose of 5 units / kg (200 units maximum ; conversion factor 20 g = 67 units). each sample the patient had blunted urinary camp response to exogenous pth (recombinant 1 - 34) as compared to healthy control and patients with postoperative hypoparathyroidism [figure 1 ]. however, our patient had normal tubular reabsorption of phosphate and percentage fall in the ratio of maximum rate of renal tubular reabsorption of phosphate to glomerular filtration rate (tmpo4/gfr) [figure 2 ] when compared to two other subjects, indicating normal phosphaturic response. in view of hypocalcemic tetany with raised pth levels and blunted nephrogenous response to exogenous pth, the patient was diagnosed as a case of php1b. basic mineral parameters of case, control, and known hypoparathyroid patients urinary cyclic - amp response to exogenous parathyroid hormone fall in tubular reabsorption of phosphate and tmpo4/gfr in response to exogenous parathyroid hormone php is a genetic disorder characterized by hypocalcemia, hyperphosphatemia, increased concentration of pth with insensitivity to action of pth at different sites depending on maternal or paternal imprinting and reduced gs activity. gnas-1 gene is located on ch 20q13.3 and encodes gs. gs has biallelic expression in most tissues but paternal gene is silenced in renal proximal convoluted tubule, thyroid, gonads, and pituitary. php1a has mutated maternal gs. php1b is characterized by epigenetic defects in imprinting of gnas locus of maternal allele due to microdeletions at gnas-1 locus with normal gs activity everywhere but renal proximal tubule. gs deficiency in renal proximal tubules is due to the lack of an active maternal allele, but has no effect on gs expression in the majority of other tissues, thus lack aho phenotype, may present as pseudohypohyperparathyroidism and may have mild resistance to tsh and calcitonin. biochemically, php1b is classically characterized by blunted urinary camp and phosphaturic response to pth. in our patient, hypocalcemic tetany, raised pth, and blunted urinary camp response to exogenous pth confirmed the diagnosis of php1b. php usually presents at an early age but late onset php1 at age of 54 years and php1b has been reported at 46 years of age. this is the second case in the literature where presentation of the disease delayed beyond third decade. also about 50% cases of php1b are sporadic so family history of similar illness may not be there. we propose following possible explanations for normal phosphaturic response in our case who, was on treatment. first, it is reported that during in vitro tests, adding serum of pseudohypoparathyroid patient to exogenous pth leads to blunted phosphaturic response due to the presence of some unknown inhibitory factor in serum of these patients, which reverts on treatment of these patients with vitamin d. it is speculated that this unknown factor is amino - truncated fragments of pth, which gets accumulated as part of raised pth. vitamin d treatment leads to normalization of serum calcium level, which by feedback inhibition of parathyroid glands reduces truncated fragments of pth. hence, interference to exogenous pth is reduced leading to normal phosphaturic response [figure 3 ]. second, pth resistance and hyperphosphatemia decrease activity of enzyme 1--hydroxylase, which lead to decreased generation of 1,25(oh)2d. activated vitamin d acts on osteoblast and increases the release of fgf-23, which by inducing phosphaturia restores normal phosphate homeostasis. in our patient, treatment with activated vitamin d might have increased fgf-23 levels leading to restoration of phosphaturic response to pth [figure 4 ]. finally, it has been reported in rat hepatocytes that extracellular hypocalcemia leads to intracellular hypocalcemia, which may lead to decreased activity of calcium - dependent enzymes and restoration of extracellular normocalcemia leads to intracellular normocalcemia and subsequent normal activity of intracellular calcium - dependent enzymes. we postulate that these intracellular calcium - dependent enzymes may be playing important role in phosphaturic response to pth, and thus treatment of patient with vitamin d will lead to restoration of normal phosphaturic response to pth [figure 4 ]. furthermore, correction of phosphaturic effect with vitamin d treatment is supported by the reports of reversal of all abnormalities mimicking php2 in cases of vitamin d deficiency. this suggests that deficiency of 1,25(oh)d might be the cause of hyperphosphatemia in patients with php. in our case, although serum 25(oh)d was in the range above vitamin d insufficiency, we hypothesize that insufficient conversion to activated vitamin d (was not measured) led to its deficiency and supplementation with 1- vitamin d corrected phosphaturic response to exogenous pth. postulate-1 : effect of vitamin d treatment on parathyroid hormone fragments possible mechanism of normal phosphaturic response in pseudohypoparathyroidism-1b | pseudohypoparathyroidism type-1b is a hereditary disorder of clinical hypoparathyroidism without aho phenotype, characterized by blunted nephrogenous cyclic - amp (camp) response to exogenous parathyroid hormone (pth). here we report a young adult presenting with hypocalcemic tetany with raised pth levels. his urinary camp response to exogenous pth (recombinant 1 - 34) was blunted ; however, phosphaturic response was normal. |
a retrocaval ureter or pre - ureteric venacava is a rare anomaly which occurs in 1 per 1000 live births. it is seen three times more often in males and is usually on the right side. a left retrocaval ureter as such is very rare and is usually associated with situs inversus or ivc duplication. these patients usually present in the third or fourth decades and symptoms depend on the degree of obstruction. an 84-year - old male patient presented to us with dull, aching left flank pain of 6 months duration. ct scan of the abdomen revealed transposed ivc to the left side and classic retrocaval ureter with proximal hydro - ureteronephrosis [figures 13 ]. intravenous urogram showing a sea - horse sign coronal ct image showing left retrocaval ureter with pre - caval (arrowhead) and post - caval (arrow) segments axial ct images with arrows showing (a) pre - caval ureter (b) post - caval ureter and (c) left renal vein the embryogenesis of the ivc is a complex process of development involving the posterior cardinal, the subcardinal, and the supracardinal venous systems. aberrant complexation of these veins can result into four anomalies : duplicated ivc, transposition of ivc to left, retroaortic left renal vein, and circum - aortic left renal vein. left retrocaval ureter is rare, and only eight cases have been reported in the literature so far. left retrocaval ureter without situs and without ivc duplication is rarer with only four cases being reported in the literature. pierro. reported a 0.2 to 0.5% prevalence of transposition of ivc to the left and a 0.1% prevalence of left retrocaval ureter. | retrocaval ureter (pre - ureteral vena cava) is an uncommon congenital anomaly that causes ureteral obstruction by external compression. although right retrocaval ureter is a common entity, left retrocaval ureter is extremely rare. a left retrocaval ureter is usually associated with situs inversus or duplicated inferior venacava (ivc). an isolated left retrocaval ureter with single left - sided ivc is even rarer and only four cases have been reported in the literature. we present images of a case with isolated left retrocaval ureter with a single left - sided ivc without situs inversus. |
acute flaccid paralysis (afp) is a clinical entity characterized by areflexia and/or hyporeflexia and weakness which reaches a maximum within days or weeks. polioviruses, enterovirus 71, flavivirus, herpes virus, and rabies virus are well - known as viral etiologic agents. in this article, who presented with lower respiratory tract infection and afp, and was detected to have co - infection of human coronavirus (hcov) 229e and oc43 is reported. afp associated with hcov infections has not been reported previously in the literature, and has been reported for the first time in this article. a 3-year - old girl hospitalized due to the development of shortness of breath and inability to walk 1-day after the beginning of fever, rhinorrhea, cough, and weakness. on the same day, respiratory distress increased, she was intubated, and mechanical ventilation started. on physical examination, she was conscious but swallowing, chewing and speech functions damaged, muscle strength was 0/5 and deep tendon reflexes (dtrs) were absent, the plantar response was flexor. complete blood count, electrolytes and blood biochemistry and urinalysis were normal, c - reactive protein was negative and erythrocyte sedimentation rate was 20 mm / h. no abnormalities in the brain and spinal cord magnetic resonance imaging were detected - so intracranial and spinal pathologies were excluded. after 48 h of hospitalization, a total of 2 g / kg intravenous immunoglobulin (ivig) in 3 days was given. muscle strength improved to 23/5, dtr became norm active, bulbar paralysis and respiratory distress had regressed. at the end of the 2 week, she could mobilize and walk with a guide. lumbar puncture and enmg repeated in the 3 week did not reveal any pathological findings and gullian barre syndrome (gbs) was completely excluded. real - time polymerase chain reaction (pcr) analysis of nasal swab samples was reported to be positive for hcov 229e and oc43 and hcov co - infection was diagnosed. flaccid paralysis occurs by the damage of lower motor neurons in the anterior horn of the spinal cord or peripheral nerves either by direct invasion or parainfectious and/or postinfectious immune - mediated mechanisms. besides gbs, differential diagnosis of afp includes : compressive and inflammatory diseases of spinal cord, the anterior horn motor neurons involvement by infectious (poliomyelitis vaccine - associated poliomyelitis, other enteroviral myelitis, a japanese encephalitis, and west nile virus as agents) and vascular pathology, myasthenia gravis, drug and toxins affecting neuromuscular junction (aminoglycosides, organophosphates, snake venom and botulism, etc.), muscle disease such as viral myositis or inflammatory myopathy, hypokalemic periodic paralysis and electrolyte disorders (hypermagnesemia and hypokalemia). preliminary diagnosis was gbs for this case ; however, csf and enmg findings in the 1 week of paralysis did not support this diagnosis. during the period of mechanical ventilation muscle strength increased and in this short time dtr normalized, as well as after 3 weeks paralysis started no pathologic findings observed by repeating lumbar puncture and enmg so gbs was excluded. history, clinical presentation, physical examination, and laboratory tests were totally normal that ruled out toxicological etiology, polio, and botulismus. members of coronaviridae can cause respiratory, intestinal, hepatic and neurological diseases of various severities in humans and animals. hcovs can cause neurological symptoms such as seizures and meningoencephalitis in children but afp has not been reported until now. hcovs may cause more serious respiratory disease in case of immune suppression and with various respiratory agents rather than single infection. in addition hcov, 229 and oc43 co - infection causing more severe respiratory disease has been demonstrated by a study. we have reported case of co - infection with hcov 229e and oc43 detected by real - time pcr analysis of nasal swab samples, and additional different respiratory pathogen has not been demonstrated, it suggests that co - infection caused lower respiratory tract infection and afp. we believe that hcov 229e and oc43 co - infection may cause respiratory failure and afp. whether ivig have a role in the treatment needs to be supported by more data from the literature. | acute flaccid paralysis (afp) is a life - threatening clinical entity characterized by weakness in the whole body muscles often accompanied by respiratory and bulbar paralysis. the most common cause is gullian barre syndrome, but infections, spinal cord diseases, neuromuscular diseases such as myasthenia gravis, drugs and toxins, periodic hypokalemic paralysis, electrolyte disturbances, and botulism should be considered as in the differential diagnosis. human coronaviruses (hcovs) cause common cold, upper and lower respiratory tract disease, but in the literature presentation with the lower respiratory tract infection and afp has not been reported previously. in this study, pediatric case admitted with lower respiratory tract infection and afp, who detected for hcov 229e and oc43 co - infection by the real - time polymerase chain reaction, has been reported for the first time. |
vitamin d status is highly variable among european countries, largely due to variations in exposure to sunshine, dietary intake of vitamin d, and the use of supplements [1, 2 ]. vitamin d can be synthesized endogenously, and factors affecting its cutaneous synthesis include age, season, latitude, time of day, skin pigmentation, amount of skin exposed, and the use of sunscreen. vitamin d status is normally defined according to the serum concentration of 25-hydroxyvitamin d [25(oh)d ]. usually, vitamin d deficiency is defined as having a serum 25(oh)d concentration lower than 50 nmol / l (20 ng / ml). nevertheless, an evaluation of satisfactory levels of vitamin d in healthy children has not yet been reported by adequate studies [35 ]. based on the current literature, the prevalence of 25(oh)d deficiency varies from 2 to 30% in adults [2, 6 ]. however, in a small - scale italian study, more than 80% of children had insufficient or deficient levels of 25(oh)d. two larger studies providing transversal data have confirmed that 25(oh)d deficiencies are very common among italian children [810 ]. an adequate 25(oh)d status is very important because 25(oh)d deficiency is a risk factor for several chronic diseases in addition to having classic deleterious effects on bone. vitamin d deficiency can cause secondary hyperparathyroidism, which may lead to high bone turnover and bone loss and ultimately increase the risk of fractures [25 ]. emerging evidence suggests that vitamin d also plays an important role in immune system regulation. vitamin d receptors are found on several immune cells, and vitamin d metabolites appear to modulate t - cell proliferation and dendritic cell function [11, 12 ]. vitamin d deficiency may also be a risk factor for the development of autoimmune and other chronic diseases [3, 11, 13, 14 ], loss of muscle mass, and muscle weakness. finally, a number of studies have also suggested that vitamin d may confer protection against diabetes mellitus type 1, hypertension, multiple sclerosis, and cancer. thus, vitamin d insufficiency may have important health consequences not only because of the vitamin 's role in the maintenance of normal bone mass turnover but also because of its role as an immunoregulatory agent. the serum 25(oh)d level is the most commonly measured indicator of vitamin d status because it reflects dietary intake from vitamins d2 and d3 together with cutaneous synthesis of vitamin d3. although data are not available regarding the required vitamin d intake in children, a clinical report from the american academy of pediatrics (aap) recommended a daily intake of 400 i.u./day of vitamin d for all infants, children, and adolescents, in contrast with a previous recommendation of 200 i.u. (5 g)/day. to date, few studies have assessed vitamin d status among italian children [710 ], and, to our knowledge, there have been no longitudinal studies. the available data indicate that, due to lifestyle changes, the main source of vitamin d, via synthesis in the skin from cholesterol after exposure to uv - b light, has been significantly reduced due to the large amount of time spent indoors [18, 19 ]. thus, the purpose of this study was to assess serum 25(oh)d levels in a large cohort of children and adolescents living in the italian province of florence in the tuscany region of italy (latitude 44n) and to identify risk factors for vitamin d deficiency in different age groups. this study also aimed to evaluate whether a normal 25(oh)d value can be restored in 25(oh)d - deficient patients with a daily supplement of 400 i.u. we consecutively evaluated 679 caucasian children and adolescents (326 males and 353 females, aged 2.117.9) from mugello, an area of tuscany, and florence (central italy), italy. all of the subjects were selected by random cluster sampling of the subjects who were seen at the pediatric unit of mugello 's hospital of borgo san lorenzo or the endocrine paediatric unit of the anna meyer children 's university hospital between september 2010 and december 2013 for routine control visits. the hospital ethics committees of anna meyer children 's university hospital and mugello 's hospital approved the study, which was conducted in accordance with the declaration of helsinki guidelines. written informed consent was obtained from the parents / guardians of all of the subjects. the present study aimed first to assess cross - sectionally the serum 25(oh)d levels in a very large cohort of italian children and adolescents and to identify risk factors for vitamin d deficiency in these subjects. for the cross - sectional evaluation, the inclusion criteria were as follows : patients older than 2 and younger than 18. the exclusion criteria were as follows : a recent history of travelling to warmer, sunnier areas prior the study, use of calcium or vitamin d supplements or any drugs affecting calcium or vitamin d metabolism within the past six months, or a positive history of primary hyperparathyroidism or other skeletal diseases, malabsorptive disorders, or neurological or renal diseases. based on the protocol, this study also included a 12-month (t0 t1) controlled longitudinal study comparing the abilities of vitamin d supplementation and the improvement of factors influencing 25(oh)d status to ameliorate 25(oh)d deficiencies. the patients found to have a 25(oh)d deficiency during the cross - sectional evaluation were recruited for the longitudinal study (figure 1). of the 679 caucasian subjects initially included in the cross - sectional evaluation, 398 children and adolescents (186 males and 212 females, aged 3.1 to 18.9) having 25(oh)d levels below 20 ng / ml were enrolled in the interventional study (figure 1). the mean time elapsed between the first (t0) and the second (t1) determinations was 12.1 months (range : 11.812.3 months). randomization was performed using a computer - generated random number table with 1.3 : 1 randomization within strata defined by gender and age. the patients in the first group, arm a, were treated with cholecalciferol 400 i.u. (10 g)/daily. of these subjects (225 subjects, 105 males and 120 females), 12 declined the consent to participate in the study, and 8 dropped out due to noncompliance, lack of follow - up, and so forth. in all, 205 of these individuals (101 males and 104 females) completed the interventional study and constituted the subjects of arm a. the patients in the second group, arm b, did not receive cholecalciferol supplements, but they were instructed to improve variables influencing 25(oh)d levels, including diet, milk intake, hours spent outdoors, sun exposure, and use of sunscreen. of these subjects (173 subjects : 81 males and 92 females), 8 declined the consent to participate in the study, and 5 dropped out due to noncompliance or lack of follow - up. the remaining 160 individuals (71 males and 89 females) completed the interventional study and were the subjects of arm b. the number of subjects required to compare the abilities of the two treatment arms to attain serum 25(oh)d concentrations of at least 30 ng / ml and to detect a change of at least 1.5 nmol / l in serum 25(oh)d levels between groups was calculated with a significance level of 5% and a power of 90%. the aim was to include at least a 1.3 : 1 ratio due to a higher withdrawal expected in arm a. during the cross - sectional evaluation and the interventional study (t0 and t1), the clinical and demographic data were collected from the subjects, including height, weight, body mass index (bmi), pubertal stage, time dedicated to outdoor physical activity, sunlight exposure, and use of sunscreen. furthermore, nutrient diaries were recorded for each subject based on their medical charts and standardized interviews. for both the cross - sectional and interventional (t0 and t1) studies, the subjects were also divided into two age groups : children (212 years) and adolescents (older than 12 years). during the cross - sectional evaluation, the dietary intakes of calcium and vitamin d were assessed through a standardized interview with the parents, recording birth weight, type of feeding during the first year (breast, formula milk, or mixed), the mother 's use of vitamin supplements during pregnancy, and the child 's use of vitamin d supplements [4, 5 ]. depending on the age of the child, the parent or guardian and the child were also interviewed about the frequency of consumption (daily, weekly, and monthly) of each food item with the use of food models, portion booklets, or serving containers to assist in estimating serving size. nutrient analyses were obtained from the food composition database for epidemiological studies in italy (banca dati di composizione degli alimenti per studi epidemiologici in italia bda). a daily intake of more than 200 ml per day of cow 's milk (median content of vitamin d was 40 i.u./l), we recorded the hours / week of outdoor physical activity (consequently subdividing our study group into low level outdoor physical activity (2 hours / day) and high level (> 2 hours / day)) and categorized the activity using a physical activity questioner, as previously described. outdoor exposure was quantified both from questions regarding each child 's average number of daily outdoor hours during each season and from a prospective daily - time - activity diary completed by caregivers during the study. during these interviews, the amount of sun exposure was also calculated and evaluated in terms of the number of days of significant exposure to sunlight before the cross - sectional evaluation and during the period of the study or for the interventional study, the summer previous to study enrolment. at our latitude (44n), cutaneous synthesis of vitamin d takes place only during the summer months (may september) [4, 5, 21 ]. we defined significant exposure to sunlight as the exposure of arms and legs for 15 minutes between 10 a.m. and 3 p.m. without the application of sunscreen, identifying the following three categories : poor (30 ng / ml and defined as severe deficiency, deficiency, insufficiency, and sufficiency, respectively, according to previously established guidelines for bone health (in the absence of a consensus regarding appropriate levels for endocrine and extraendocrine health) [35 ]. to evaluate seasonal variations however, although solar winter is typically defined as the period from november to february, to create larger, more significant statistical groups, we defined winter as the period from november to may and summer as that from june to october because human insolation to uvb radiation is negligible between november and april. the dosage was based on the recommendation of the american academy of pediatrics for all infants, children, and adolescents [18, 19 ]. for arm a, written instructions were provided at the onset of the study, and compliance was evaluated by a written questionnaire completed by the parents. compliance was further verified by e - mails and/or telephone interviews performed by a study nurse and by the bottle count of cholecalciferol performed at the end of the study period. for arm b, after evaluating the questionnaires about calcium and vitamin d dietary intake, outdoor physical activity levels, sun exposure, and the use of sunscreen, the parents and children were informed of how to safely increase vitamin d levels ; this information provided them with new knowledge and had the potential to change their attitudes and behaviors. to eliminate differences in knowledge, at t0, all of the arm b families participating in the study received written and oral information about the importance of vitamin d in the health of children and adults and the amount of vitamin d required for good health (dietary reference intake or dri). to optimize the consumption of nutrients naturally rich in vitamin d, such as fish, fatty fish, eggs, milk, and dairy products, the parents received information about the major dietary sources of vitamin d. this information was provided using the food composition database for epidemiological studies in italy (banca dati di composizione degli alimenti per studi epidemiologici in italia - bda). the equal importance of a certain amount of sun exposure gained through outdoor activities was also explained. for arm b, written instructions were provided at the onset of the study and compliance was evaluated by a written questionnaire completed by the parents. compliance was further verified by e - mails and/or telephone interviews performed by a study nurse. each subject 's weight was measured with electric scales to the nearest 0.1 kg, and the height was measured with a stadiometer. the body mass index (bmi) was calculated using the following formula : bmi = weight (kg)/height (m). the age - related reference values for height, bone age, and bmi obtained from large sample numbers of italian children and currently used in italy were used in the study. as described previously, height and bmi were normalized for chronological age by conversion to standard deviation scores (sds), which were calculated according to the following formula : patient value mean of age - related reference value / standard deviation of the age - related reference value. patient value mean of age - related reference value / standard deviation of the age - related reference value. the pubertal staging was performed according to the criteria of tanner and whitehouse, using an orchidometer for the boys. the plasma concentrations of calcium, phosphate, and b - alp were determined following routine biochemical laboratory protocols. serum 25(oh)d and pth levels were determined by chemiluminescence enzyme - labeled immunometric assays using an immulite 2000 systems analyzer (siemens, gwynedd, uk). the intra- and interassays cvs were 2 hours / day in outdoor physical activity (23.76 7.58, p 2 hours per week outdoors). however, we demonstrated a significant difference in 25(oh)d levels between the groups that spent 2 hours / day in outdoor physical activity (arm a : 10.20 3.78 ng / ml, arm b : 10.14 3.67 ng / ml) and those in the respective arms that spent > 2 hours / day in outdoor physical activity (arm a : 13.80 5.02 ng / ml, arm b : 13.64 4.67 ng / ml, p 2 hours / day in outdoor physical activity (26.49 6.33 ng / ml, p 2 hours / day outdoors. in contrast, in arm a, there was no significant difference between the 25(oh)d level of the subgroup spending 2 hours / day outdoors (13.66 7.78 ng / ml) and that of the subgroup spending > 2 hours / day outdoors (18.79 8.11 ng / ml). furthermore, a significant improvement in the mean intake of vitamin d was observed in arm b (264 53.7 i.u per day at t1 versus 170.1 47.8 i.u. per day at t0, p < 0.0001), whereas no improvement in vitamin d intake was observed in arm a (191.4 53.7 i.u. per day at t1). in particular, there was an increase in the percentage of subjects in arm b who consumed a normal amount of cow 's milk over the course of the study (47.9% versus 37.2%, p < 0.05). however, at t1, only arm b subjects who consumed a normal amount of cow 's milk per day showed significantly higher 25(oh)d levels than those who consumed little or no cow 's milk per day (26.79 8.65 ng / ml versus 17.17 8.18 ng / ml, p < 0.0001). in arm a, at t1, no difference was found in the 25(oh)d levels of those who consumed a normal amount of cow 's milk and those who consumed little or no cow 's milk (20.78 8.33 ng / ml versus 16.59 8.15 ng / ml). in arm b, we demonstrated an increase in the length of the sun exposure, with an increase in the percentage of subjects with moderate sun exposure (48.8% versus 32.7%, p < 0.005). there was a significant increase in 25(oh)d levels compared with those measured at t0 for individuals with low sun exposure (13.12 8.71 versus 9.90 4.41 ng / ml, p < 0.0001), moderate exposure (16.77 6.43 versus 13.19 4.32 ng / ml, p < 0.0001), and good exposure (24.55 6.11 versus 15.39 4.29 ng / ml, p < 0.005). in arm a, the 25(oh)d levels were significantly reduced with respect to those detected in the arm b subjects for each sun exposure subgroup (11.88 8.12 ng / ml in those with low sun exposure, 14.66 9.01 ng / ml in those with moderate exposure, and 21.65 8.84 in those with good exposure, p < 0.0001 for all comparisons). at t1, there was no significant difference in the regular use of sunscreens in the subjects of arm b. we did confirm that the subjects who regularly used sunscreen had significantly lower 25(oh)d levels than nonregular sunscreen users (17.86 7.84 ng / ml versus 26.45 8.56 ng / ml, p < 0.0001). there was a significant difference between the 25(oh)d levels of arm a regular sunscreen users and nonregular users (17.13 8.01 ng / ml versus 26.98 8.77 ng / ml, p < 0.0001) and between the 25(oh)d levels of arm b regular sunscreen users and nonregular users (18.46 7.55 ng / ml versus 26.03 8.79, p < 0.0001). when all of the subjects were included, we demonstrated at t1 a significant reduction of the percentage of subjects with hyperparathyroidism (83/365 patients (22.7%), 41 males and 42 females). pg / ml (versus 53.27 34.78 pg / ml at t0 ; p < 0.05). arm a subjects showed significantly reduced pth levels compared with arm b subjects (43.01 28.81 pg / ml versus 49.74 33.71 pg / ml, p < 0.05). after the interventional study, the mean b - alp level was 112.5 30.6 u / l in arm a and 121.7 32.5 u / l in arm b (p < 0.05). in our evaluation of potential correlations between 25(oh)d levels and factors potentially affecting 25(oh)d levels at t1 (age, sex, bmi, cow 's milk consumption (ml / day), outdoor physical activity (hours / week), use of sunscreens, sun exposure, pth, calcium, phosphorus, birth weight, type of feeding during the first year of life, mother 's use of vitamin supplementation during pregnancy, and child 's past use of vitamin d supplementation), we demonstrated that 25(oh)d levels correlated with age (r = 0.57, p < 0.005), bmi (r = 0.63, p < 0.005), cow 's milk consumption (r = 0.49, p < 0.005), outdoor physical activity (r = 0.66, p < 0.0001), use of sunscreen (r = 0.49, p < 0.005), sun exposure (r = 0.53, p < 0.005), pth levels (r = 0.61, p < 0.001), and calcium levels (r = 0.44, p < 0.005), and b - alp (r = 0.57, p < 0.001). no correlation was found between 25(oh)d levels and sex, phosphorus levels, birth weight, type of feeding during the first year of life, mother 's use of vitamin supplementation during pregnancy, and child 's past use of vitamin d supplementation at t1. our study provides data on the 25(oh)d status in a large sample of children and adolescents in mugello, an area of tuscany, italy. the extent of the deficiency in most of the children and adolescents was surprising, confirming a recent paper of vierucci. in fact, our data shows a very high prevalence of vitamin d deficiency (near 56%) in different age groups. moreover, if we also include individuals with an insufficiency, the percentage rises to 88.6%, very similar to the findings of vierucci. (79.5%) in a different area of tuscany and bellone. in a series of northern italian children (71.7%). as expected, our data also demonstrated a strong seasonal influence on vitamin d status, as observed in some studies performed in countries at northern latitudes and in different age groups [21, 22 ], whereas other studies reported counterintuitive results at different latitudes, probably due to differences in climate. this seasonal effect is probably related to the reduced exposure of children to sunlight and the reduced vitamin d production in the skin during the winter months. in winter, children spend more time indoors, and when they do go out, the amount of skin exposed to the sun is less than that during warmer months. as a consequence, little vitamin d is produced in the skin. furthermore, at our latitude, sun exposure is only effective in promoting vitamin d activation from may through september. thus, the amount of vitamin d produced and stored october through april is probably not sufficient to guarantee an optimal vitamin d status in unsupplemented healthy children and adolescents, especially if other factors limit summer sun exposure. however, the high percentage of vitamin d - deficient and insufficient children in this paper as well as that reported by vierucci. and bellone. [810 ] compared with that reported in previous papers [2428 ] may be the result of changes in the lifestyles of children and adolescents, with many hours each day spent in indoor activities, such as school, watching television, or playing games. however, the use of sunscreen is also an important factor influencing this very high percentage of children with vitamin d inadequacy. in our study, the children had higher 25(oh)d levels than adolescents, most likely due to more outdoors activity, milk intake, and dietary intake of calcium and vitamin d, confirming the results of several previous studies [22, 2931 ]. thus, the presence of such a high percentage of subjects with vitamin d deficiency or insufficiency may be considered a disease of modern society. in fact, modern lifestyles and food habits have probably accentuated the predisposing environmental conditions. the implications of this major inadequacy in the health of children and adolescents need to be better defined. interestingly, our study demonstrated that neither lifestyle changes nor supplementation with 400 i.u. (10 g)/day of cholecalciferol appears to be individually sufficient to restore adequate levels of vitamin d in all italian children and adolescents. although many studies have investigated vitamin d supplementation in healthy adults [32, 33 ] as well as adults [3436 ], children, and adolescents [13, 37, 38 ] with chronic diseases, there are few studies in the literature which focused on the effect of regular daily vitamin d supplementation in healthy children and adolescents [39, 40 ]. in 323 black and white children who randomly received 0, 400, 1000, 2000, or 4000 iu / d of oral vitamin d3 for 12 weeks during the winter, increases in vitamin d concentrations ranging from 10 nmol / l for the placebo to 76 nmol / l for 4000 i.u. the authors reported that supplementation with 400 i.u./day was sufficient to maintain adequate wintertime 25(oh)d concentrations in healthy black but not white children. however, in another study evaluating 150 healthy children with baseline serum 25(oh)d levels < 30 ng / ml, after 4 months of nutritional intervention, the subjects supplemented with 400 i.u / d of vitamin d all reached normal vitamin d levels. in contrast, of the subjects who received dietary counseling alone, only one achieved a normal vitamin d concentration. these results are in contradiction with our data, in which we identified a considerable group of subjects (almost 15% of those included in the study) with severe vitamin d deficiency who did not respond to lifestyle changes or cholecalciferol supplementation. although we demonstrated an improvement in the other classes of 25(oh)d levels (sufficient, insufficient, and deficient levels), over 35% of the subjects remained 25(oh)d - deficient, and over 73% of the patients had levels below the sufficiency level after vitamin d supplementation or implementation of lifestyle changes to improve vitamin d levels. specifically, after implementing an improved diet, higher intake of milk, and increased hours spent outdoors, although we showed an improvement of nearly 25% in 25(oh)d levels, 21% of the subjects were still severely 25(oh)d - deficient. among the available data of italian children and adolescents [4, 5, 810 ], it is interesting to note that one study reported that 54.8% of 93 subjects had serum 25(oh)d levels < 20 ng / ml, despite the fact that 33% of the subjects were receiving vitamin d supplementation at the time of evaluation. in fact, one study demonstrated that two cups (500 ml) of cow 's milk per day was sufficient to maintain 25(oh)d levels above 75 nmol / l. our results demonstrated that subjects who consumed more than 250 ml of milk / day had improved 25(oh)d levels and also identified an inverse association between the intake of cow 's milk and 25(oh)d deficiency, concordant with other studies, indicating the importance of a correct diet in reducing a 25(oh)d deficit. a study of the dietetic habits of our children and adolescents has demonstrated an increased consumption of alternatives to milk (such as soy milk and cereals) that provide little vitamin d and interfere with calcium uptake. the association of a higher risk of 25(oh)d deficit and limited time spent outdoors reflects another modern change of the lifestyle of children and adolescents and, as reported in other studies, may indicate the need for cholecalciferol supplementation to help maintain adequate levels of vitamin d in italian children and adolescents. the promotion of an active outdoor lifestyle may counteract the vitamin d deficiency epidemic, as suggested by a study of saudi children that demonstrated that children who are physically active have higher levels of vitamin d than those who are less active, despite having the same amount of sun exposure. these findings are also confirmed in the report about 414 girls by dahifar., in which the mean serum 25(oh)d concentration increased to 14.4 ng / ml with sunlight exposure. our data demonstrated that the season of blood withdrawal is a significant predictor of vitamin d status, with winter and spring being the seasons associated with lower median 25(oh)d levels, in the range of deficiency. this finding is in accordance with other reports [9, 33, 37, 47, 48 ] and stresses the importance of the promotion of an active outdoor lifestyle, especially from may to september. these findings are of particular concern because of both the well - established and newly identified target effects of vitamin d (bone accrual and health immunomodulatory function in decreasing the risk of many chronic illnesses, including common cancers, autoimmune diseases, infectious diseases, and cardiovascular disease), stressing the importance of maintaining appropriate 25(oh)d levels [1116 ]. our data seem to confirm and suggest that, individually, lifestyle changes or vitamin d supplementation is not sufficient to ensure sufficient 25(oh)d levels. consequently, a global intervention or an increase of daily supplementation by cholecalciferol should be considered. one limitation of our study was that it is an interventional open study rather than a randomized controlled trial. another limitation was the heterogeneous group of subjects (obese, overweight, and normal weight children and adolescents) ; however, this limitation was also a benefit because it allowed us to exhaustively evaluate all of the factors involved in vitamin d status. the large number of children and adolescents enrolled in our study was a strength. the lack of vitamin d supplementation at baseline in all subjects was an additional strength, allowing us to evaluate a large number of vitamin d status predictors and differentiate between the effects of vitamin d supplementation and other factors influencing vitamin d status. in conclusion, deficient or insufficient vitamin d serum levels were found in most of the italian children and adolescents sampled in this study. the presence of inadequate 25(oh)d levels represents a complex problem that reflects lifestyle changes, increasingly poor dietary habits, sunscreen use, and an increase in the obesity rate in the population. our findings suggest that neither changes in lifestyles nor or supplementation with 400 i.u. (10 g)/day of cholecalciferol appears to be individually sufficient to restore adequate levels of vitamin d. thus, daily supplementation with more than 400 i.u. | objective. this paper aims to assess 25(oh)d levels in italian children and adolescents identifying risk factors for 25(oh)d deficiency and to evaluate whether a normal 25(oh)d value can be restored in 25(oh)d - deficient patients. methods. we evaluated 25(oh)d levels in 679 italian children and adolescents (10, 1120, 2130, and > 30 ng / ml were defined as severe deficiency, deficiency, insufficiency, and sufficiency, resp.). of these, 365 25(oh)d - deficient were followed up for 1 year ; 205 were treated with cholecalciferol (arm a : 400 i.u.) and 160 by improving the environmental variables influencing 25(oh)d levels (arm b). results. at cross - sectional evaluation, 11.3% showed sufficiency, 30.0% insufficiency, and 58.7% 25(oh)d deficiency. mean 25(oh)d was 19.08 8.44 ng / ml. at the enrollment time (t0), no difference was found between arms a and b with respect to distribution and 25(oh)d levels. at end time (t1) 26.0% (29.7% in arm a versus 20.6% in arm b) showed sufficiency, 38.4% (42.0% versus 34.4%) insufficiency, and 35.6% (28.3% versus 45.0%) 25(oh)d deficiency. mean 25(oh)d level was 23.71 6.83 ng / ml. conclusions. neither changes of lifestyle nor 400 i.u. cholecalciferol supplementation alone appears to be sufficient to restore adequate 25(oh)d levels. |
the functional asymmetry of the cerebral hemispheres has been reported in patients with many sorts of brain damage, who failed to orient, report, or respond to stimuli located in one hemispace 1. line bisection has been employed as a sensitive test for unilateral neglect 1,3. in this task, lateral deviation from the true center indicate the relative inattention for the contralateral side of space, and a consistent leftward error has been reported in healthy subjects in the western world, indicating a relatively right cerebral dominance in them (reviewed in ref 4). nonetheless, patients with right hemispheric lesions usually place the subjective midpoint to the right of the true center 4. of interest to psychologists and psychiatrists are the possible functional cerebral asymmetries represented in sorts of psychiatric patients and in different emotional states of healthy individuals, which might be due to the different strategies for processing specific stimuli. in healthy subjects, negative emotions are more likely to be associated with the activation of the right hemisphere 5,6. for instance, some studies have shown that the induced anxiety in normal subjects selectively impaired their spatial, but not verbal performance 7,8, other studies in both infants and adults have found that negative affects, such as anxiousness and depression, are more relatively associated with the right hemisphere activation, particularly the frontal lobe 6,9 - 11. moreover, the greater right hemisphere activation in depression patients has been demonstrated in studies that used the consonant - vowel task12, and the electroencephalogram measurements 13,14. individuals with generalized anxiety disorder (gad) are found to be intolerant of uncertainty and perceive more potentially negative situations than the healthy subjects 15 - 17, and people with depression also employ a maladaptive problem - solving method, which contributes to the maintenance of negative emotions they perceived 18,19. one question therefore arises how patients with gad, or with the treatment - resistant depression (trd), a severe form of depression, would perform in the line bisection task. the possible answers might help us to understand better the hemispheric functions that contribute to these pathologies on the one hand, and probably help to further address the overlaps between anxiety and depression on the other. bearing that gad and trd patients often present negative emotions in mind, we have hypothesized that these subjects would bisect lines further leftward than the healthy volunteers would. in addition, given the high prevalence of anxiety / depression in the general population, it might be interesting to compare the levels of anxiety / depression between our patients and healthy subjects. thus, we used the zuckerman - kuhlman personality questionnaire 20 to measure the subject 's anxiety trait, and the plutchik - van praag depressive inventory 21 to measure their depressive mood. moreover, since the sensation seeking trait is correlated with a hemispheric asymmetry 22,23, the zuckerman sensation seeking scales 24 were also used in the present protocol. considering that most chinese people are trained to use their right hands as artful ones in their early lives 25, studies of such training in athletes showed consistently moderate rightward errors in the line bisection task 26,27, we therefore selected 155 moderate to strong right - handed subjects for our study. fifty - six healthy volunteers were recruited among college students, medical staff members or paid volunteers. after a semistructured interview, it was determined that they were not suffering from any kinds of anxiety or depressive disorder. forty - seven outpatients were diagnosed with gad according to the criteria of the diagnostic and statistical manual of mental disorders - version iv - text revision 28. fifty - two outpatients were diagnosed with trd using following criteria (all the four criteria were met) : (1) symptoms met criteria for major depressive disorder 28 ; (2) remission failed after using at least two antidepressants ; (3) patients scored more than 25 on the plutchik - van praag depression inventory ; (4) patients were without comorbidities of psychotic diseases or drug abuse. in addition, patients were ascertained not to have any organic brain lesions after going through computerized tomographic or magnetic resonance imaging scans. about 50% of patients had received anxiolytics or antidepressants before arriving at our clinic, but no participants had ingested alcohol, drugs or medication at least 72 hours prior to the test. there were no significant group differences when referring to age (one - way anova, main effect, f (2,152) =.94, p >.05), or gender (main effect, f =.06, p >.05). this study protocol was approved by a local ethics committee and all subjects gave their written informed consent. each of the 12 items of the inventory were scored 1, 2 or 3 according to the left - hand, either left or right, or right preference. all subjects scored between 29 and 36, and were thus considered to be moderate or strong right - handers., subjects were asked to complete three questionnaires on - site in a quiet room. a brief overview of each questionnaire is described below : the zuckerman - kuhlman personality questionnaire. the test provides five measurements : (a) impulsive sensation seeking (19 items) ; (b) neuroticism - anxiety (19 items) ; (c) aggression - hostility (17 items) ; (d) activity (17 items) ; and (e) sociability (17 items). the internal reliabilities of these scales range from.72 to.86. in this questionnaire, 10 items of another scale of dissimulation (infrequency or lie) were randomly inserted into the test body. any score above 3 on the infrequency scale suggests either inattention to the content of the items and acquiescence or a very strong social desirability set ; therefore, the infrequency scale was used as a test validity indicator for individuals 20. the test has proven to be reliable in the chinese culture 32;the zuckerman sensation seeking scales (form v, 40 items). the test provides four subscales of 10 items each, (i.e., disinhibition, thrill and adventure seeking, experience seeking and boredom susceptibility). the total score in each subject was also calculated as the sum of the four scale scores. the test has proven to be reliable in the chinese culture 34;the plutchik - van praag depression inventory. this inventory contains 34 items ; each item has three scale points (0, 1, 2), corresponding to depressive tendencies. subjects have possible depression if they score between 20 and 25, or depression if they score higher than 25. the test provides five measurements : (a) impulsive sensation seeking (19 items) ; (b) neuroticism - anxiety (19 items) ; (c) aggression - hostility (17 items) ; (d) activity (17 items) ; and (e) sociability (17 items). the internal reliabilities of these scales range from.72 to.86. in this questionnaire, 10 items of another scale of dissimulation (infrequency or lie) were randomly inserted into the test body. any score above 3 on the infrequency scale suggests either inattention to the content of the items and acquiescence or a very strong social desirability set ; therefore, the infrequency scale was used as a test validity indicator for individuals 20. the test has proven to be reliable in the chinese culture 32 ; the zuckerman sensation seeking scales (form v, 40 items). the test provides four subscales of 10 items each, (i.e., disinhibition, thrill and adventure seeking, experience seeking and boredom susceptibility). the total score in each subject was also calculated as the sum of the four scale scores. the test has proven to be reliable in the chinese culture 34 ; the plutchik - van praag depression inventory. this inventory contains 34 items ; each item has three scale points (0, 1, 2), corresponding to depressive tendencies. subjects have possible depression if they score between 20 and 25, or depression the lines, drawn in black and oriented horizontally, ranged from 102 - 144 mm in length, were arranged randomly on a sheet of a4 size paper (in a portrait orientation) one below the other, and differed in their distances from the sheet margins so that their centers were not in alignment. no restrictions were placed on head or eye movements, and no time limits were imposed. subjects were instructed to use their right hand to make a mark indicating the center of the line. there are many classical methods to analyze line bisection performance, for instance the percentage expression of bias errors 35. here briefly, the distance of the line bisecting task mark was measured from the actual center to the nearest millimeter. the frequency of the directional errors (frequency), irrespective of the magnitude, was calculated as (right - left)/ (right + left) ; negative values indicate errors to the left and positive ones indicate errors to the right. the magnitude of line bisection deviation (magnitude) was calculated as an algebraic sum of the distance of marks from the true center divided by the number (e.g., 8) of trials. negative values indicate errors to the left and positive ones indicate errors to the right. two - way anova followed by a post - hoc, duncan 's multiple new range test was applied to the five trait scores of the zuckerman - kuhlman personality questionnaire or four scale scores of sensation - seeking in the three groups. the mean frequency, magnitude, or depression scores in the three groups were submitted to a one - way anova plus duncan 's test. the relationship between the frequency, magnitude, five personality traits, four sensation seeking scales, and depression scores was assessed by the spearman rank order correlation test. with the present sample size, power to detect an effect (e.g., a scale score) was larger than 80% at p.05), or gender (main effect, f =.06, p >.05). this study protocol was approved by a local ethics committee and all subjects gave their written informed consent. each of the 12 items of the inventory were scored 1, 2 or 3 according to the left - hand, either left or right, or right preference. all subjects scored between 29 and 36, and were thus considered to be moderate or strong right - handers. before the line bisection task, subjects were asked to complete three questionnaires on - site in a quiet room. a brief overview of each questionnaire is described below : the zuckerman - kuhlman personality questionnaire. the test provides five measurements : (a) impulsive sensation seeking (19 items) ; (b) neuroticism - anxiety (19 items) ; (c) aggression - hostility (17 items) ; (d) activity (17 items) ; and (e) sociability (17 items). the internal reliabilities of these scales range from.72 to.86. in this questionnaire, 10 items of another scale of dissimulation (infrequency or lie) were randomly inserted into the test body. any score above 3 on the infrequency scale suggests either inattention to the content of the items and acquiescence or a very strong social desirability set ; therefore, the infrequency scale was used as a test validity indicator for individuals 20. the test has proven to be reliable in the chinese culture 32;the zuckerman sensation seeking scales (form v, 40 items). the test provides four subscales of 10 items each, (i.e., disinhibition, thrill and adventure seeking, experience seeking and boredom susceptibility). the total score in each subject was also calculated as the sum of the four scale scores. the test has proven to be reliable in the chinese culture 34;the plutchik - van praag depression inventory. this inventory contains 34 items ; each item has three scale points (0, 1, 2), corresponding to depressive tendencies. subjects have possible depression if they score between 20 and 25, or depression if they score higher than 25. the test provides five measurements : (a) impulsive sensation seeking (19 items) ; (b) neuroticism - anxiety (19 items) ; (c) aggression - hostility (17 items) ; (d) activity (17 items) ; and (e) sociability (17 items). the internal reliabilities of these scales range from.72 to.86. in this questionnaire, 10 items of another scale of dissimulation (infrequency or lie) were randomly inserted into the test body. any score above 3 on the infrequency scale suggests either inattention to the content of the items and acquiescence or a very strong social desirability set ; therefore, the infrequency scale was used as a test validity indicator for individuals 20. the test has proven to be reliable in the chinese culture 32 ; the zuckerman sensation seeking scales (form v, 40 items). the test provides four subscales of 10 items each, (i.e., disinhibition, thrill and adventure seeking, experience seeking and boredom susceptibility). the total score in each subject was also calculated as the sum of the four scale scores. the test has proven to be reliable in the chinese culture 34 ; the plutchik - van praag depression inventory. this inventory contains 34 items ; each item has three scale points (0, 1, 2), corresponding to depressive tendencies. subjects have possible depression if they score between 20 and 25, or depression if they score higher than 25. the lines, drawn in black and oriented horizontally, ranged from 102 - 144 mm in length, were arranged randomly on a sheet of a4 size paper (in a portrait orientation) one below the other, and differed in their distances from the sheet margins so that their centers were not in alignment. no restrictions were placed on head or eye movements, and no time limits were imposed. subjects were instructed to use their right hand to make a mark indicating the center of the line. there are many classical methods to analyze line bisection performance, for instance the percentage expression of bias errors 35. here briefly, the distance of the line bisecting task mark was measured from the actual center to the nearest millimeter. the frequency of the directional errors (frequency), irrespective of the magnitude, was calculated as (right - left)/ (right + left) ; negative values indicate errors to the left and positive ones indicate errors to the right. the magnitude of line bisection deviation (magnitude) was calculated as an algebraic sum of the distance of marks from the true center divided by the number (e.g., 8) of trials. negative values indicate errors to the left and positive ones indicate errors to the right. two - way anova followed by a post - hoc, duncan 's multiple new range test was applied to the five trait scores of the zuckerman - kuhlman personality questionnaire or four scale scores of sensation - seeking in the three groups. the mean frequency, magnitude, or depression scores in the three groups were submitted to a one - way anova plus duncan 's test. the relationship between the frequency, magnitude, five personality traits, four sensation seeking scales, and depression scores was assessed by the spearman rank order correlation test. with the present sample size, power to detect an effect (e.g., a scale score) was larger than 80% at p.05, mse = 9.746) (table 2). the mean depression scores among the three groups also had statistically significant differences from each other (main effect, f (2, 152) = 60.64, p <.001, mse = 6205.02), with that of the trd patients higher than those of both the healthy subjects and the gad patients (also see table 2). on average, trd patients bisected slightly more frequently to the left of the true center, whereas healthy subjects bisected slightly more frequently to the right. in contrast, gad patients bisected significant more frequently to the left of the true center than the healthy subjects did. when the mean frequency errors in the three groups were analyzed, one - way anova detected a significant difference (f (2, 152) = 3.50, p <.05, mse = 1.40). the post - hoc duncan 's test showed that the gad group (-.32.56 s. d.) was significantly different from the healthy control group (.01.67, p <.05). the mean magnitude errors were also significantly different among the three groups (f (2, 152) = 3.31, p <.05, mse = 5.90), post - hoc duncan 's test detected that the mean magnitude in the gad group (-.54 mm 1.15 s.d.) was significantly different from that in the healthy controls ' (.12 1.42), but not from that in the trd (-.33 1.40) ; there was no statistical difference between the mean magnitude errors of the healthy controls and those of the trd either. frequency was negatively correlated with the disinhibition - seeking score (n = 56, r = -.28, p <.05) in healthy subjects, and with total sensation - seeking (n = 47, r = -.30, p <.05) and experience seeking scores (n = 47, r = -.34, p <.05) in gad patients. in addition, the depression score was negatively correlated with magnitude (n = 52, r = -.30, p <.05) in trd patients. no other correlations, such as between the handedness and frequency/ magnitude, or personality trait scores were found in our study. in the present study, patients scored higher on neuroticism - anxiety (with gad patients scoring highest) and on depression (with trd patients scoring highest) than the healthy subjects did. these results lead to the observation that there is a great overlap between anxiety and depression symptoms in clinics 37,38, and that these two disorders might share similar genetic dimensions and disease continuums 39,40. in addition, our patients scored lower on sociability than the healthy subjects did, which was also in line with the previous report that major depression affected the sociability trait 41, and this low extraversion (introversion) 42 tendency has also been suggested as personality endophenotypes in many anxiety disorders, e.g., social phobia and agoraphobia 43. in compliance with our hypothesis, gad patients erred significantly leftward in line bisection, which suggests a right hemispheric overactivation, left hypoactivation, or both for this disorder. as we have noted in our introduction, negative emotions like anxiousness some neuroimaging and electrophysiological data have also shown that patients with anxiety disorders (e.g., panic disorder) displayed lower activation of the left parietal or superior temporal cortex, but relatively greater activation of the right frontal or hippocampal regions than the healthy subjects did 44 - 46. contrary to our hypothesis, trd patients did not show significant leftward line bisection errors in our study. such a result is in line with previous studies, showing that the unipolar depressive patients displayed a non - significant leftward bias in manual line bisection, while schizophrenia patients bisected significantly leftward 47 - 49. however, results in regard to the hemispheric activation in the depressive disorder remain inconclusive up to date 50 - 55. albeit, the slight rightward error found in our healthy subjects was different from those documented in western countries 4, this result is similar to those in other studies conducted in japan 56 and china 30,31. this discrepancy might result from a cultural background where most chinese people are forced to use their right hands during their early lives 25. the negative correlation between frequency and disinhibition - seeking scores in our healthy subjects, and the negative correlation between frequency and the total sensation - seeking and experience - seeking scores in gad patients, contradict the recent neurophysiologic results in sensation seekers. for instance, a greater left frontal eeg asymmetry at rest is related to a tendency to engage in sensation - seeking and risky behaviors in young adults 23. likewise, the association of left hemisphere predominance and risk - taking in healthy university students has also proven by studying the line bisecting performance and zuckerman 's sensation seeking scales in drake and ulrich 's study 36. a rightward bias has been reported in healthy subjects in some eastern countries like japan and china, contradictory to those found in western societies, such a tendency might contribute to our current findings. moreover, whether the reversed correlation in our gad patients was due to the severity of anxiety itself merits further investigation. in our trd patients, this finding is in accordance with the results in the tension - type headache study 31, which might be because many tension - type headache sufferers also displayed signs of depression 57. on the other hand, we could not completely ruled out the medication effects on our findings, since previous studies have shown the effect of anxiolytics or antidepressants on cognition (e.g., attention, memory or learning) 58,59, behavioral aspects (e.g., executive function or motor reaction) 60, and hemispheric asymmetry 48. however, prior to the study, our patients were all medication - free for at least 72 hours, which helped to remove some effects of the anxiolytics or antidepressants. nevertheless, further studies about the medication effects on brain asymmetry in anxiety and depression disorders would be of interest. some limitations in our study should be underlined. firstly, we did not consider the menstrual cycles of our female subjects, since the line bisection performance might be influenced by the menstrual state of women 35,61 ; however, three groups of our subjects were gender - balanced. secondly, we did not measure the disordered personality traits in our subjects, since the dependent personality disorder patients have shown a pronounced leftward line bisection error 30. thirdly, for more extensive comparisons, we would need more data from left - handed subjects, and data obtained using the neuroimaging or other indirect neuropsychological techniques. fourthly, we used lines between 102 - 144 mm which were shorter than what other investigators used, and the line length was demonstrated to have influenced line bisection performance (e.g., ref 62). fifthly, we did not analyze the medication effect on the line bisection task since the individual medication strategies varied among our patients. finally, we did not employ other attention - control paradigms such as using a cue during the task. in conclusion, the leftward line bisection errors in gad might indicate a stronger right, a weaker left hemispheric activation, or both. the task is a non - invasive examination and easy to manipulate in typical clinics. whether it could be used as a diagnostic auxiliary test for anxiety versus depression remains to be determined. | background and objectives the line bisection error to the left of the true center has been interpreted as a relative right hemisphere activation, which might relate to the subject 's emotional state. considering that patients with generalized anxiety disorder (gad) or treatment - resistant depression (trd) often have negative emotions, we hypothesized that these patients would bisect lines significantly leftward. methods we tried the line bisection task in the right - handed healthy volunteers (n = 56), gad (n = 47) and trd outpatients (n = 52). subjects also completed the zuckerman - kuhlman personality questionnaire, the zuckerman sensation seeking scales, and the plutchik - van praag depression inventory. results gad patients scored highest on the neuroticism - anxiety trait, trd patients scored highest on depression, and both patients scored lower on the sociability trait. patients with gad also bisected lines significantly leftward compared to the healthy subjects. the frequency of the bisection error was negatively correlated with disinhibition - seeking in the healthy subjects, and with total sensation - seeking and experience - seeking in gad patients, while the magnitude of the line bisection error was negatively correlated with depression in trd patients. conclusions the study suggests a stronger right hemispheric activation, a weaker left activation, or both in the gad, instead of trd patients. |
uterine leiomyomata, the most common solid pelvic tumors, occur in approximately 20% of women aged 35 years or more. because of the significant operative risk associated with abdominal myomectomy, the operation has been reserved for women who want to preserve or enhance their fertility potential. however, with the general trend towards conserving the uterus and because many women currently choose delayed childbearing, patient demand for this procedure has increased. furthermore, with the improvement in laparoscopic techniques and their associated low morbidity, resection of myomas is now a valid alternative for women suffering from any serious symptoms related to the presence of myomas in the uterus. the only surgical treatment available for intramural and subserous myomas in the past was laparotomy. laparoscopic myomectomy has since been shown to be an effective means of reducing postoperative morbidity, and it expedites recuperation. however, it quickly became apparent that the laparoscopic operation was associated with the prolonged time of anesthesia, increased blood loss, and possibly a higher risk of postoperative adhesion formation. laparoscopic myomectomy was described 2 decades ago and has since been repeatedly shown to provide the recognized benefits of the laparoscopic approach. however, with increasing experience, it has become apparent that the technique demands a high degree of training and skill from the laparoscopic operator. it has been suggested that at present these factors limit the application of laparoscopic myomectomy. despite significant improvements in endoscopic instrumentation, laparoscopic myomectomy remains a technically demanding and time - consuming procedure. in the easiest cases, pedunculated myomas may be simply transected, and some enucleated myomas will pop out through an incision in the uterine wall. the operation is time - consuming, usually because of the difficulty in morcellating and removing the myoma from the abdominal cavity through the laparoscopic trocar ports or a posterior colpotomy. the belief that the strength of the uterine scar may be compromised is based on 2 major considerations and first, the difficulty in adequately reapproximating the incision, as with meticulous multilayer suturing, may lead to the accumulation of an intramural hematoma. second, use of the co2 laser and electrodesiccation, which could lead to thermal injury to surrounding tissue, may result in poor vascularization and tissue necrosis. at second - look laparoscopy, we have observed indentations at the sites from which leiomyomas were removed that were directly proportioned to the size of the myomas removed and may therefore represent structural defects. this is a significant potential problem when the myomectomy is performed to enhance or preserve fertility. however, data are still limited concerning postsurgical adhesion formation and pregnancy outcome although some preliminary data are encouraging. another concern recently raised is the possibility that, after incomplete resection of the uterine myomas, the recurrence of myomas may be higher with the laparoscopic approach. nezhat developed laparoscopic - assisted myomectomy (lam) and reported on it in 1994. it has been advocated as a technique that may lessen these concerns regarding laparoscopic myomectomy while retaining the benefits of laparoscopic surgery, namely, short hospital stay, lower costs, and rapid recovery. herein, the lam technique and possible associated advantages are described. whether to proceed with lam is usually decided in the operating room after first completing the diagnostic laparoscopy and treating any associated pathology. the criteria for lam are myomas larger than 10 to 12 cm or numerous and deep myomas requiring extensive morcellation and necessitating uterine repair with sutures. the leiomyoma, or in patients with multiple myomas, the most prominent one, is injected at its base with 3 to 7 ml of diluted vasopressin to minimize blood loss. an incision is made over the uterine serosa until the capsule of the leiomyoma is reached. a corkscrew manipulator is inserted into the leiomyoma and used to elevate the uterus toward the midline suprapubic puncture. with the trocar and manipulator attached to the myoma after incision of the fascia transversely at 4 to 5 cm, the rectus muscles are separated at the midline. we do not routinely use preoperative gonadotrophin - releasing hormone (gnrh) analogues. for anemic patients, preoperative treatment with gnrh analogues may enable restoration of a normal hematocrit, decrease the size of the myomas, and reduce the need for trans - fusion. however, the benefits of preoperative treatment with gnrh analogues for laparoscopic myomectomy have recently been challenged in a prospective randomized study. the peritoneum is entered transversely, and the leiomyoma is located and brought to the incision using the corkscrew manipulator ; a uterine manipulator is used to raise the uterus. the corkscrew manipulator is replaced with 2 lahey tenacula. the leiomyoma is shelled sequentially and morcellated, gradually exposing new areas. after complete removal of the leiomyoma, the uterine wall defect is seen through the incision. if uterine size allows, the uterus is brought to the skin through the minilaparotomy incision to complete the repair. when multiple leiomyomas are found, as many as possible are removed through a single uterine incision. when the leiomyomas are in distant locations and identification is impossible, the minilaparotomy incision is closed temporarily with 1 layer of running suture or several allis clamps. the laparoscope is reintroduced, and the leiomyomas are identified and brought to the incision. if posterior leiomyomas are difficult to reach through a minilaparotomy incision, they are removed completely laparoscopically. the uterus is reconstructed in layers using 4 - 0 to 2 - 0 and 0-polydioxanone suture without suturing the serosa, and the uterus is palpated to ensure that no small intramural leiomyomas are present. the uterus is returned to the peritoneal cavity, and the fascia and skin are closed in layers. the fascia is closed with 1 - 0 polyglactin suture, and the skin is closed in a subcuticular manner. the pelvis is evaluated to detect and treat endometriosis and adhesions that may have been obscured previously by myomas. laparoscopic - assisted myomectomy outcomes were compared with the results in patients who had either myomectomy by laparotomy or laparoscopic myomectomy. the myoma weight was significantly greater in the lam group than in the patients undergoing laparoscopic myomectomy. it was found that lam could safely replace myomectomy by laparotomy, because patient selection criteria were comparable, and the myoma weights of these 2 groups were similar. in contrast, blood loss among the patients undergoing laparoscopic myomectomy was significantly lower and may be attributed to the smaller myomas removed. although subserosal myomas less than 5 cm can be managed easily laparoscopically, larger and intramural lesions require prolonged morcellation and laparoscopic suturing of the uterine defect. some surgeons have suggested that the size of the myoma to be removed laparoscopically should be limited to 10 cm or even 6 to 7 cm. consequently, it has also been suggested that no more than 4 myomas, 3 cm or more in size, should be attempted to be removed laparoscopically. lam, with conventional morcellation and suturing through the minilaparotomy incisions, allows fast removal of multiple and large myomas and reduces the duration of the operation and the need for extensive laparoscopic experience. similar mean operating times for laparoscopic myomectomy and lam techniques were observed despite larger myomas and their intramural positions, adjunctive laparoscopic procedures, and the smaller incisions in the lam groups. hospitalization was significantly longer for the patients who underwent myomectomy by laparotomy when compared with that for the groups having lam or laparoscopic myomectomy. currently, the hospitalization time is similar for patients undergoing lam and patients having laparoscopic myomectomy. for both procedures, day surgery is used, and the patient is usually discharged on the first postoperative day. the postoperative recovery time is comparable for patients undergoing lam versus laparoscopic myomectomy, despite the differences in the size of the different incisions. the use of lam offers several obvious and potential long and short - term benefits (table 1). the major advantages of lam are ease of repair of the uterus and rapid morcellation of the fibroids. in addition, lam allows more meticulous suturing of the uterus, thus maintaining better uterine wall integrity. the most feared postoperative complication, uterine rupture during pregnancy, has been reported to date in 5 cases after laparoscopic myomectomy. because the number of procedures performed over the last few years remains unknown, it is difficult to determine whether these 5 cases represent an incidence higher or lower than expected after any myomectomy. uterine rupture after myomectomy is rare, but has been reported to account for approximately 2% of all pregnancy - related uterine ruptures. all large series reported to date did not confirm the hypothesis that laparoscopic myomectomy is associated with an increased risk for uterine dehiscence during pregnancy. however, it must be remembered that inadequate approximation of the uterine wall and poor healing may predispose patients to uterine rupture. second - look laparoscopies performed on postmyomectomy patients with pedunculated and superficial subserosal myomas show complete uterine healing. in contrast, intramural and deep subserosal myomas are associated with evidence of granulation tissue and indentation of the uterus proportional to the size of the leiomyoma removed, unless sutures are used to approximate the edges. the use of sutures is associated with a higher rate of adhesions. in a recent study of second - look after laparoscopic myomectomy the factors that influenced the occurrence of an adhesion on the myomectomy site were posterior location of the myoma and the existence of sutures. meticulous suturing techniques facilitated by lam may therefore reduce the rate of postoperative adhesions. in patients with intramural fibroids and significant uterine wall defects, currently, the meticulous suturing made possible during laparotomy is very challenging to perform at laparoscopy. lam therefore may provide a safer approach allowing more complete multilayer correction of the postmyomectomy uterine defects. a similar approach, combined laparoscopic and vaginal myomectomy, has also been suggested for treating extensive and deeply infiltrating fundal and posterior wall leiomyomata. the posterior colpotomy permits delivery of the myomas and allows uterine reconstruction by conventional suturing performed transvaginally. a long - established dogma dictates that, if the endometrial cavity is entered or a submucous or large intrauterine myoma is removed during abdominal myomectomy, the patient should undergo cesarean delivery for subsequent pregnancies. currently, women with large intramural fibroids, who wish to have children, should be strongly cautioned regarding the relative paucity of data regarding the precise risk during pregnancy after laparoscopic myomectomy. lam, in carefully selected cases, is a safe and efficient alternative to both laparoscopic myomectomy and myomectomy by laparotomy. lam may also be a more appropriate approach for women who desire a future pregnancy because uterine healing and adhesion formation remain a significant concern. in such women, lam may offer better management, because it allows careful suturing of the uterine defect in layers and avoids excessive electrocoagulation. by decreasing the technical demands, and thereby the operative time, lam may be more widely offered to patients. thus, providing them with the well - recognized advantages of minimal access surgery, including a shorter hospital stay, and better patient convenience and recovery. | laparoscopic myomectomy has recently gained wide acceptance. however, this procedure remains technically highly demanding and concerns have been raised regarding the prolonged time of anesthesia, increased blood loss, and possibly a higher risk of postoperative adhesion formation. laparoscopic - assisted myomectomy (lam) is advocated as a technique that may lessen these concerns regarding laparoscopic myomectomy while retaining the benefits of laparoscopic surgery, namely, short hospital stay, lower costs, and rapid recovery. by decreasing the technical demands, and thereby the operative time, lam may be more widely offered to patients.in carefully selected cases, lam is a safe and efficient alternative to both laparoscopic myomectomy and myomectomy by laparotomy. these cases include patients with numerous large or deep intramural myomas. lam allows easier repair of the uterus and rapid morcellation of the myomas. in women who desire a future pregnancy, lam may be a better approach because it allows meticulous suturing of the uterine defect in layers and thereby eliminates excessive electrocoagulation. |
one of the most frequently asked questions to those of us at june is what kinds of manuscripts are appropriate for inclusion in the journal. any manuscript with the aim of enabling others to enhance their teaching of neuroscience to undergraduates is appropriate ; those with empirically - tested protocols of innovative pedagogy are particularly welcomed and prioritized for publication in june. beyond manuscripts devoted to new classroom approaches and laboratory exercises, a variety of manuscripts in other aspects that are relevant to undergraduate neuroscience education are welcomed, such as : editorials. june welcomes book reviews ranging from popular press volumes relevant to neuroscience to textbooks.media reviews. june welcomes reviews of media such as films, television shows, websites, and software that may have particular value to neuroscience education.commentaries. june welcomes reviews of media such as films, television shows, websites, and software that may have particular value to neuroscience education. manuscripts must be properly formatted according to the instructions to authors available at the june website (http://funjournal.org). manuscripts should be carefully proofread, and attention should be given to both the flow and appearance of information and positioning of tables, figures, and captions. to facilitate indexing, authors are welcome to provide a list of suggested reviewers, which may or may not be used in the review of their article. the first step in the process is an initial screen by the editors to ensure that the manuscript is appropriate for the journal. if not, a notification indicating why the manuscript does not fit the criteria for inclusion in june is sent to the author(s). sometimes, an editor may provide suggestions of how the manuscript may be modified to better fit the criteria. once the initial reviews are received, the editors make a decision to accept, request revisions and resubmissions, or reject. often times the difference between acceptance and rejection hinges on a few key issues. in the case of manuscripts detailing innovative approaches to teaching, contentions of effectiveness need to be supported with discussion of appropriate methods of assessment and supporting evidence provided by the results obtained. in some cases, an author may inadvertently fail to obtain permission to use and discuss copyrighted material. in all cases if an author has been asked to revise portions of their manuscript, a cover letter indicating how each of the reviewer s comments and concerns have been addressed should be included in the resubmission. one of the most frequently asked questions to those of us at june is what kinds of manuscripts are appropriate for inclusion in the journal. any manuscript with the aim of enabling others to enhance their teaching of neuroscience to undergraduates is appropriate ; those with empirically - tested protocols of innovative pedagogy are particularly welcomed and prioritized for publication in june. beyond manuscripts devoted to new classroom approaches and laboratory exercises, a variety of manuscripts in other aspects that are relevant to undergraduate neuroscience education are welcomed, such as : editorials. june welcomes book reviews ranging from popular press volumes relevant to neuroscience to textbooks.media reviews. june welcomes reviews of media such as films, television shows, websites, and software that may have particular value to neuroscience education.commentaries. june welcomes reviews of media such as films, television shows, websites, and software that may have particular value to neuroscience education. manuscripts must be properly formatted according to the instructions to authors available at the june website (http://funjournal.org). manuscripts should be carefully proofread, and attention should be given to both the flow and appearance of information and positioning of tables, figures, and captions. to facilitate indexing, authors are welcome to provide a list of suggested reviewers, which may or may not be used in the review of their article. the first step in the process is an initial screen by the editors to ensure that the manuscript is appropriate for the journal. if not, a notification indicating why the manuscript does not fit the criteria for inclusion in june is sent to the author(s). sometimes, an editor may provide suggestions of how the manuscript may be modified to better fit the criteria. all manuscripts that are deemed appropriate for the journal are sent to reviewers. once the initial reviews are received, the editors make a decision to accept, request revisions and resubmissions, or reject. often times the difference between acceptance and rejection hinges on a few key issues. in the case of manuscripts detailing innovative approaches to teaching, contentions of effectiveness need to be supported with discussion of appropriate methods of assessment and supporting evidence provided by the results obtained. in some cases, an author may inadvertently fail to obtain permission to use and discuss copyrighted material. in all cases if an author has been asked to revise portions of their manuscript, a cover letter indicating how each of the reviewer s comments and concerns have been addressed should be included in the resubmission. there is a range of ways readers of june can contribute to the journal beyond the submission of manuscripts. one very important way is to use the journal as a resource in your own teaching and to encourage your colleagues to do this as well. in addition, you can promote june to other neuroscience educators, encouraging them to read the journal and to submit manuscripts discussing their own approaches and innovative techniques for teaching neuroscience. it s important to support june in another way as well by joining and holding membership in fun. while june is an open - access journal and free to all, june does cost money to publish. fun members, through a portion of their very reasonable annual dues, provide critical support for june. consider also being a reviewer for june. one is to become an ad hoc reviewer ; to do so, simply send the editor in - chief a short email message expressing your willingness to review articles, indicating your particular areas of expertise. while review board members are regularly called on to complete reviews, such service is typically limited to two or three reviews per year. review board members are also considered to fill vacancies that happen periodically on the editorial board. to be considered for service on the june review board, please contact the june editor - in - chief regarding a possible appointment. june has come a long way since being founded in 2002. through the efforts of our contributing authors, reviewers, and the entire editorial board, june has contributed to the success of undergraduate programs and neuroscience education around the globe. an important milestone for the second decade of june will be the successful completion of our efforts to become indexed across major databases and services, including psych info, scopus, the national science digital library, the directory of open access journals, medline and pubmed (grisham, 2012). at this writing, only indexing in medline and pubmed remain to be accomplished, and the application process is partially completed. as technology changes and new avenues for electronic communication become available, expect june to change as well, adding new features or altering existing ones to reflect latest developments. with support from fun members, the undergraduate neuroscience educational community will continue to have a readily available venue for learning about the latest innovations in laboratory exercises and improved teaching approaches. be sure to visit june online at (http://funjournal.org), in person at the fun and at the society for neuroscience annual meetings, or both. publish in, and review articles for june and join in the fun of promoting undergraduate neuroscience education and research. | in the fall of 2002, the faculty for undergraduate neuroscience (fun) began publication of its flagship journal, the journal of undergraduate neuroscience education (june). for the past ten years, june has been a major forum for the free exchange of information among undergraduate neuroscience educators. numerous articles on laboratory exercises, media, pedagogy, curriculum, and issues pertinent to neuroscience educators have been published in june during the past decade. given the vast expertise in pedagogy amongst the fun membership and within the undergraduate neuroscience education community at large, we strongly encourage all fun members and june readers to become actively involved in june by contributing manuscripts and/or by offering your services as a reviewer. |
the disease known as pneumocystis carinii pneumonia (pcp) is one of the leading causes of illness and death in persons with impaired immunity. the disease has been described in immunocompromised patients for many years, including outbreaks in malnourished young children in orphanages in iran in the 1950s (16). the aids epidemic, however, marked the beginning of the disease s impact on a substantial number of patients. pcp has long been the most common serious aids - defining opportunistic infection in the united states. the introduction of highly active antiretroviral therapy (haart) for the treatment of hiv infection has been accompanied by substantial reductions in mortality and the incidence of opportunistic infections, including pcp (7). despite these advances, pneumocystis remains a major pathogen in hiv - infected persons who either are not receiving or are not responding to haart and among those who are unaware of their hiv status. pcp is also of clinical importance in people immunocompromised for reasons other than hiv, such as organ transplantation or chemotherapy for malignant diseases (8). in addition, pneumocystis infection has been documented recently in persons who are mildly immunocompromised, including those with chronic lung disease (9). pneumocystis organisms were first reported by chagas in 1909 (10), but he mistook them for a morphologic form of trypanosoma cruzi. within a few years of this first report, further studies established that the microbe in question was not a trypanosome but a new species altogether, named pneumocystis carinii (11). from the time of its discovery, until late in the 1980s, pneumocystis these views were based on several criteria : 1) strong similarities in microbe morphology and host pathology, 2) absence of some phenotypic features typical of fungi, 3) presence of morphologic features typical of protozoa, 4) ineffectiveness of antifungal drugs, and 5) effectiveness of drugs generally used to treat protozoan infections. some investigators pointed out that pneumocystis organisms exhibit morphologic similarities to fungi (2). nevertheless, the protozoan hypothesis remained predominant until 1988, when dna analysis demonstrated that pneumocystis is a fungus, albeit an odd one, lacking in ergosterol and very difficult to grow in culture (12,13). soon after the proper classification of pneumocystis had been determined at the kingdom level, additional dna data showed that pneumocystis organisms in different mammals are quite different. these data led to interim name changes (14), but it was not until 1999 that the first valid new binomial appeared. the organism that causes human pcp is now named pneumocystis jiroveci frenkel 1999 (pronounced yee row vet zee), in honor of the czech parasitologist otto jirovec, who is credited with describing the microbe in humans (15). the primary purpose of this article is to explain what led to the name change and why the new name is necessary, useful, and workable for all concerned. for a more extensive review of the systematics and nomenclature of pneumocystis, see stringer s review of workshops on the subject (16). the dna sequence information that led to the renaming of pneumocytsis organisms also provided the tools needed to better understand the relationships between these microbes and the hosts they inhabit. thus, the secondary purpose of this article is to summarize data on these relationships, focusing on current views on the relationship between p. jiroveci and humans. pneumocystis organisms were first reported by chagas in 1909 (10), but he mistook them for a morphologic form of trypanosoma cruzi. within a few years of this first report, further studies established that the microbe in question was not a trypanosome but a new species altogether, named pneumocystis carinii (11). from the time of its discovery, until late in the 1980s, pneumocystis was widely thought to be a protozoan. these views were based on several criteria : 1) strong similarities in microbe morphology and host pathology, 2) absence of some phenotypic features typical of fungi, 3) presence of morphologic features typical of protozoa, 4) ineffectiveness of antifungal drugs, and 5) effectiveness of drugs generally used to treat protozoan infections. some investigators pointed out that pneumocystis organisms exhibit morphologic similarities to fungi (2). nevertheless, the protozoan hypothesis remained predominant until 1988, when dna analysis demonstrated that pneumocystis is a fungus, albeit an odd one, lacking in ergosterol and very difficult to grow in culture (12,13). soon after the proper classification of pneumocystis had been determined at the kingdom level, additional dna data showed that pneumocystis organisms in different mammals are quite different. these data led to interim name changes (14), but it was not until 1999 that the first valid new binomial appeared. the organism that causes human pcp is now named pneumocystis jiroveci frenkel 1999 (pronounced yee row vet zee), in honor of the czech parasitologist otto jirovec, who is credited with describing the microbe in humans (15). the primary purpose of this article is to explain what led to the name change and why the new name is necessary, useful, and workable for all concerned. for a more extensive review of the systematics and nomenclature of pneumocystis, see stringer s review of workshops on the subject (16). the dna sequence information that led to the renaming of pneumocytsis organisms also provided the tools needed to better understand the relationships between these microbes and the hosts they inhabit. thus, the secondary purpose of this article is to summarize data on these relationships, focusing on current views on the relationship between p. jiroveci and humans. one reason that a definitive nomenclature has been slow to develop is that pneumocystis organisms have been difficult to study. cultivation of pneumocystis organisms in vitro requires a large seed population and supports rather modest increases in organism number for a very limited period of time (17). an exception to the rule was recently reported (18) ; however, this method has not been established in other laboratories. fortunately, advances in dna analysis technology allowed progress in the absence of a robust culture system. phenotypic differences between p. jiroveci and other species of pneumocystis were noted decades ago (19). more recent descriptions echo these reports (20). on the basis of phenotypes, the name was not validly published, however, under the then - prevailing specifications of the international code of zoological nomenclature. the first indication of a molecular difference between p. jiroveci and pneumocystis from laboratory animals came from analyses of protein sizes (21,22). however, the importance of these differences was difficult to judge because the pneumocystis was prepared directly from the lung of the host, leaving open the possibility that differences could have been due to extrinsic factors such as contamination with host proteins, host - mediated modification of pneumocystis proteins, or presence of dead pneumocystis organisms. dna analysis provided the information needed to clarify the issue and to establish that the organisms from humans and other animals are quite different (23). wakefield developed primers that amplify dna from all known species of pneumocystis (24,25). when these primers have been used on human - derived samples of pneumocystis, the only dna found has been that of p. jiroveci. moreover, p. jiroveci dna has not been found in lung samples from any other mammals, including nonhuman primates (26). several genes or gene fragments have been cloned from human - derived pneumocystis (2730). in all cases, the gene sequence is very different from its orthologues in pneumocystis organisms from other host species. the 18s rrna sequences from p. jiroveci (i.e., human - derived) and p. carinii (i.e., rat - derived) differ by 5%. this level of divergence is comparable with that between pneumocystis organisms and taphrina deformans (a plant fungal pathogen), whose 18s rrna sequences differ by approximately 6%. in contrast, species in the genus saccharomyces can differ by as little as 1% at the 18s rrna locus. the genetic divergence between p. jiroveci and other pneumocystis organisms is typical of the genus. when pneumocystis from different host species are compared by dna sequence analysis, they always differ (23,25,3133). in addition, experiments with rats, mice, ferrets, and monkeys have demonstrated host - species specificity (3436). for example, when pneumocystis organisms were taken from a rat and transferred to a mouse, proliferation was not evident, and no disease resulted (34). in contrast, when pneumocystis organisms from a rat were transferred to another rat, they proliferated to a very high number and caused severe disease. transfer experiments that seem to show lack of specificity have been reported, but these reports did not show that the proliferating organisms were the same species of pneumocystis as those introduced, leaving open the possibility that endogenous organisms were responsible for the infection. pneumocystis organisms might be obligate parasites that have evolved to survive in a particular host species. note, in this regard, that p. jiroveci is most similar to organisms isolated from other primates (37). however, the host specificity data also fit with an alternative scenario : there could be many free - living species of pneumocystis, one of which is capable of invading humans, others of which are capable of invading nonhuman primates, and the like. in this scenario, the similarity between p. jiroveci and the pneumocystis organisms found in nonhuman primates would reflect the similarities between humans and other primates. if p. jiroveci is not an obligate parasite, finding it outside the human body should be possible. p. jiroveci dna has been detected in samples of airborne fungal spores (24) and in a sample of pond water (38). however, the number of p. jiroveci in the environment seems to be very low, leaving open the possibility that these free forms of the organism may have been deposited by humans. p. jiroveci could be an obligate parasite, spores of which can survive in the environment long enough to infect a new host, should one be encountered. resolving this question awaits the availability of a system capable of detecting infectious pneumocystis organisms in the air, water, or soil. soon after dna sequence data began to appear, name changes were suggested (14,39). however, naming new species seemed premature to many because of concerns about the possibility of creating false species by misinterpreting the importance of a limited amount of dna sequence data. this system referred to the different kinds of pneumocystis organisms as special forms of p. carinii under this system, p. jiroveci was called p. carinii formae specialis hominis (p. carinii f. sp. more dna sequence data were obtained, and by 2001, it became clear that the organism causing pcp in humans should be recognized as a distinct species. the name p. jiroveci had already been published in a valid manner in 1999 (15) ; however, publication of a name does not necessarily lead to its use. therefore, at the 2001 international workshop on opportunistic protists held in cincinnati, ohio, approximately 50 researchers from around the world, including clinicians, epidemiologists, and laboratory scientists, met to discuss the desirability and appropriateness of retaining the currently used trinomial nomenclature system, as opposed to assigning (or using) new species names. the group unanimously endorsed a proposal to rename the organisms currently known as special forms of p. carinii as species in the genus pneumocystis and drew up guidelines for the creation of the new species names (16). consequently, in keeping with the international code of botanical nomenclature, it is no longer correct, either biologically or taxonomically, to refer to the human pneumocystis organism as p. carinii. p. carinii now refers exclusively to the organism formerly known as p. carinii f. sp. the consensus achieved at the workshop will help to make published reports on pneumocystis more uniform with respect to nomenclature. such uniformity will clarify communication among all who are interested in this genus and the disease caused by its members. given the compelling evidence that the human form of pneumocystis is a separate species, the most important objection to designating it as such has been the problem that this name change could create in the medical literature, where the disease caused by p. jiroveci is widely known as pcp, or pcp. this problem can be avoided by taking the species name out of the disease name. under this system this simple modification in the vernacular accommodates the name change pertaining to the pneumocystis species that infects humans. furthermore, adopting this change makes the acronym appropriate for describing the disease in every host species, none of which, except rats, is infected by p. carinii. dna sequence polymorphisms are often observed in isolates of p. jiroveci, suggesting that numerous strains of this species exist. loci that have been favorite targets for sequence analysis include the mitochondrial large subunit ribosomal rna gene, the mitochondrial small subunit rrna gene, the internal transcribed spacer regions of the nuclear rrna gene (its), the arom gene, and the dihydropteroate synthase (dhps) gene. the first three of these loci are considered to be under little if any selective pressure and presumably serve as indicators of genetic changes that are phenotypically neutral. the changes in the arom gene may also be considered neutral because they effect no change in the amino acid sequence of the enzyme. by contrast, the polymorphisms in the dhps gene may be due to selection (see below). these include the use of type - specific oligonucleotide probes to detect variation at the its regions (40) and detection of single - strand conformation polymorphism (sscp) at multiple loci (41). most studies have targeted one locus for analysis, but several multilocus studies have been reported (4144). the allelic sequence polymorphism common in p. jiroveci is not seen in p. carinii (rat - derived pneumocystis). however, p. carinii populations differ with respect to chromosome size, and several different strains have been identified by analysis of chromosome sizes (45,46). the possibility of chromosome size variation in p. jiroveci has not been adequately addressed because this analysis requires more organisms than are typically available from patients. phenotypic differences between p. jiroveci and other species of pneumocystis were noted decades ago (19). more recent descriptions echo these reports (20). on the basis of phenotypes, the name was not validly published, however, under the then - prevailing specifications of the international code of zoological nomenclature. the first indication of a molecular difference between p. jiroveci and pneumocystis from laboratory animals came from analyses of protein sizes (21,22). however, the importance of these differences was difficult to judge because the pneumocystis was prepared directly from the lung of the host, leaving open the possibility that differences could have been due to extrinsic factors such as contamination with host proteins, host - mediated modification of pneumocystis proteins, or presence of dead pneumocystis organisms. dna analysis provided the information needed to clarify the issue and to establish that the organisms from humans and other animals are quite different (23). wakefield developed primers that amplify dna from all known species of pneumocystis (24,25). when these primers have been used on human - derived samples of pneumocystis, the only dna found has been that of p. jiroveci. moreover, p. jiroveci dna has not been found in lung samples from any other mammals, including nonhuman primates (26). the pcr data are supported by the results of sequencing cloned genes. several genes or gene fragments have been cloned from human - derived pneumocystis (2730). in all cases, the gene sequence is very different from its orthologues in pneumocystis organisms from other host species. the 18s rrna sequences from p. jiroveci (i.e., human - derived) and p. carinii (i.e., rat - derived) differ by 5%. this level of divergence is comparable with that between pneumocystis organisms and taphrina deformans (a plant fungal pathogen), whose 18s rrna sequences differ by approximately 6%. in contrast, species in the genus saccharomyces can differ by as little as 1% at the 18s rrna locus. the genetic divergence between p. jiroveci and other pneumocystis organisms is typical of the genus. when pneumocystis from different host species are compared by dna sequence analysis, they always differ (23,25,3133). in addition, experiments with rats, mice, ferrets, and monkeys have demonstrated host - species specificity (3436). for example, when pneumocystis organisms were taken from a rat and transferred to a mouse, proliferation was not evident, and no disease resulted (34). in contrast, when pneumocystis organisms from a rat were transferred to another rat, they proliferated to a very high number and caused severe disease. transfer experiments that seem to show lack of specificity have been reported, but these reports did not show that the proliferating organisms were the same species of pneumocystis as those introduced, leaving open the possibility that endogenous organisms were responsible for the infection. pneumocystis organisms might be obligate parasites that have evolved to survive in a particular host species. note, in this regard, that p. jiroveci is most similar to organisms isolated from other primates (37). however, the host specificity data also fit with an alternative scenario : there could be many free - living species of pneumocystis, one of which is capable of invading humans, others of which are capable of invading nonhuman primates, and the like. in this scenario, the similarity between p. jiroveci and the pneumocystis organisms found in nonhuman primates would reflect the similarities between humans and other primates. if p. jiroveci is not an obligate parasite, finding it outside the human body should be possible. p. jiroveci dna has been detected in samples of airborne fungal spores (24) and in a sample of pond water (38). however, the number of p. jiroveci in the environment seems to be very low, leaving open the possibility that these free forms of the organism may have been deposited by humans. p. jiroveci could be an obligate parasite, spores of which can survive in the environment long enough to infect a new host, should one be encountered. resolving this question awaits the availability of a system capable of detecting infectious pneumocystis organisms in the air, water, or soil. soon after dna sequence data began to appear, name changes were suggested (14,39). however, naming new species seemed premature to many because of concerns about the possibility of creating false species by misinterpreting the importance of a limited amount of dna sequence data. this system referred to the different kinds of pneumocystis organisms as special forms of p. carinii under this system, p. jiroveci was called p.. hominis). after these provisional nomenclature were instituted, more dna sequence data were obtained, and by 2001, it became clear that the organism causing pcp in humans should be recognized as a distinct species. the name p. jiroveci had already been published in a valid manner in 1999 (15) ; however, publication of a name does not necessarily lead to its use. therefore, at the 2001 international workshop on opportunistic protists held in cincinnati, ohio, approximately 50 researchers from around the world, including clinicians, epidemiologists, and laboratory scientists, met to discuss the desirability and appropriateness of retaining the currently used trinomial nomenclature system, as opposed to assigning (or using) new species names. the group unanimously endorsed a proposal to rename the organisms currently known as special forms of p. carinii as species in the genus pneumocystis and drew up guidelines for the creation of the new species names (16). consequently, in keeping with the international code of botanical nomenclature, it is no longer correct, either biologically or taxonomically, to refer to the human pneumocystis organism as p. carinii. p. carinii now refers exclusively to the organism formerly known as p. carinii f. sp. the consensus achieved at the workshop will help to make published reports on pneumocystis more uniform with respect to nomenclature. such uniformity will clarify communication among all who are interested in this genus and the disease caused by its members. given the compelling evidence that the human form of pneumocystis is a separate species, the most important objection to designating it as such has been the problem that this name change could create in the medical literature, where the disease caused by p. jiroveci is widely known as pcp, or pcp. this problem can be avoided by taking the species name out of the disease name. under this system this simple modification in the vernacular accommodates the name change pertaining to the pneumocystis species that infects humans. furthermore, adopting this change makes the acronym appropriate for describing the disease in every host species, none of which, except rats, is infected by p. carinii. dna sequence polymorphisms are often observed in isolates of p. jiroveci, suggesting that numerous strains of this species exist. loci that have been favorite targets for sequence analysis include the mitochondrial large subunit ribosomal rna gene, the mitochondrial small subunit rrna gene, the internal transcribed spacer regions of the nuclear rrna gene (its), the arom gene, and the dihydropteroate synthase (dhps) gene. the first three of these loci are considered to be under little if any selective pressure and presumably serve as indicators of genetic changes that are phenotypically neutral. the changes in the arom gene may also be considered neutral because they effect no change in the amino acid sequence of the enzyme. by contrast, the polymorphisms in the dhps gene may be due to selection (see below). these include the use of type - specific oligonucleotide probes to detect variation at the its regions (40) and detection of single - strand conformation polymorphism (sscp) at multiple loci (41). most studies have targeted one locus for analysis, but several multilocus studies have been reported (4144). the allelic sequence polymorphism common in p. jiroveci is not seen in p. carinii (rat - derived pneumocystis). however, p. carinii populations differ with respect to chromosome size, and several different strains have been identified by analysis of chromosome sizes (45,46). the possibility of chromosome size variation in p. jiroveci has not been adequately addressed because this analysis requires more organisms than are typically available from patients. genotyping samples of p. jiroveci provides a method for exploring epidemiologic issues. for example, one study examined the possibility that the low incidence of pcp in african hiv - infected persons might be due to the presence or absence of certain strains of p. jiroveci. however, samples of p. jiroveci from zimbabwe, brazil, the united states, and the united kingdom have exhibited no major differences in genotypes (47,48). another example is a study in which genotyping at four different genetic loci was used to compare isolates of p. jiroveci collected before (19681981) and after (1982 to present) the beginning of the aids pandemic (47). pre- and postpandemic samples were the same except for a single base polymorphism (in the mitochondrial large subunit rrna gene) found in the pre - pandemic samples only. these data show that the large increase in incidence of pcp was not accompanied by a shift in the kinds or frequencies of strains of p. jiroveci. strain analysis has also led to observations that are difficult to reconcile with the traditional view of the relationship between p. jiroveci and humans. the traditional theory holds that clinically important infection results from reactivation of a latent infection that was acquired during childhood. while infection of young children appears to be common, latent p. jiroveci has not been directly observed in healthy adults. the latency issue is important for several reasons. under the reactivation of latent infection theory, little rationale exists for instituting measures to minimize the risk of infection during adulthood because this infection has already occurred. on the other hand, person - to - person transmission of the disease would have important public heath implications for medical centers that treat hiv - infected patients or other immunocompromised persons (4244,4952). furthermore, transmission from patients who are undergoing treatment for pcp might enhance the opportunity for drug resistance to arise. by contrast, the generation of drug resistance would be less of a concern if most or all infections were due to transmission from an immunocompetent person, such as a young child 's mother, or another child (i.e, someone who is not being treated for pcp). under these conditions, drug - resistant strains, pcp develops in infants infected with hiv perinatally, suggesting that p. jiroveci was present in these infants environments early in their lives (53). evidence of p. jiroveci has also been found in some victims of sudden infant death syndrome (sids) (54). in normal, healthy children most children develop anti - pneumocystis antibodies early in life, and the prevalence of these antibodies appears to increase with age (48,55). recently, p. jiroveci has been linked to clinical illness in normal, healthy infants (51). p. jiroveci dna was identified in nasopharyngeal aspirates obtained during episodes of mild respiratory infection in 24 (32%) of 74 infants. seroconversion developed by 20 months of age in 67 (85%) of 79 infants who remained in the study and occurred in the absence of any symptoms of disease in 14 (18%). although infection of children seems common, little evidence exists for lifelong latency. using pcr, wakefield found no evidence of p. jiroveci in bronchoalveolar lavage fluid from 10 healthy persons (56). peters replicated this result in postmortem lung tissue from 15 immunocompetent adults (56,57). (the techniques used to detect p. jiroveci have found it in hiv - negative adults but only those with other health problems.) studies on recurrent pcp have shown that different p. jiroveci genotypes are present during different pcp episodes in patients with repeat episodes of pcp, a result suggestive of infection proximal to the time of disease (42 - 44). recent infections of adults are also suggested by the high frequency of mutations that cause changes in the sequence of the dhps gene, the enzyme associated with sulfonamide resistance in other pathogens (5961). these mutations have not been detected in patients in whom pcp occurred at a time before the widespread use of sulfonamides to treat and prevent it (62) but are common in today 's patients, even in those with no known exposure to sulfonamides (61,63). mutant dhps genes have been found in a variety of p. jiroveci genetic backgrounds, suggesting that selection for dhps mutations is an ongoing process (64). an alternative approach to exploring the importance of latency is employing population genetics and epidemiology to test the following hypothesis. if lifelong latency is important, adult patients who reside far from their birthplace should have the strain of p. jiroveci common in their place of birth, not in their place of residence. the strains infecting adult patients were more similar to those common in their place of residence than their place of birth, suggesting that infections had been recently acquired, rather than carried since early childhood. latent p. jiroveci have not been found in healthy adults, but proving that they do not exist is practically impossible. a single organism anywhere in the body could be sufficient to maintain a latent infection. however, latent infections may be transitory, and humans who have eliminated the microbe may be subject to reinfection. the observations described above seem more consistent with this transient colonization scenario than with lifelong latency. the microbe that causes pcp in humans is a distinct phylogenetic fungal species called pneumocystis jiroveci. this species has been difficult to find in the environment, has not been found in nonhuman hosts, and is either absent in healthy humans or present at very low levels. in contrast, p. jiroveci is fairly common in humans who have depressed immune function. the number of p. jiroveci in a person appears to be dependent on the degree of immune dysfunction, suggesting that the species is adapted to exploit this dysfunction, growing to very high numbers in the severely immunodeficient and to lesser extents when immune function is less impaired. some patients are infected by genotypes more common in their place of residence than in their birthplace. variable loci include the gene encoding an enzyme targeted by sulfonamides, suggesting transmission from treated patients to others at risk. while these observations, combined with the scarcity of p. jiroveci in healthy adults, do not exclude latency as a cause of pcp, they suggest that long - term latency is not the only source of this disease. | the disease known as pneumocystis carinii pneumonia (pcp) is a major cause of illness and death in persons with impaired immune systems. while the genus pneumocystis has been known to science for nearly a century, understanding of its members remained rudimentary until dna analysis showed its extensive diversity. pneumocystis organisms from different host species have very different dna sequences, indicating multiple species. in recognition of its genetic and functional distinctness, the organism that causes human pcp is now named pneumocystis jiroveci frenkel 1999. changing the organism s name does not preclude the use of the acronym pcp because it can be read pneumocystis pneumonia. dna varies in samples of p. jiroveci, a feature that allows reexamination of the relationships between host and pathogen. instead of lifelong latency, transient colonization may be the rule. |
pruritus is an unpleasant feeling that can cause the desire of scratching in a person and can be the symptoms of systemic, infectious, and neurological diseases. causes and treatments of pruritus have been described by traditional persian medicine scientists. the aim of this study was to derive general principles of the proposed treatment to reduce or relieve pruritus. this descriptive study, review traditional medicine books including al canon fil tibb, al - hawi, makhzan ul - adviyyah, al - abniyah an - haghyegh el - adviyah, tuhfat ul - momineen and exir - e - azam. the above - mentioned documents were derived and classified by keywords such as pruritus, hakka, jarab and sherry. in traditional persian medicine, there are different causes for pruritus such as accumulation of vapors or acute humors in subcutaneous tissue or weakness of expulsive (dafia) faculty and its treatment is based on removing the causes. proper nutrition, bathing, and removing pathogenic humors are involved in the treatment. according to this study, some plants such as cassia fistula, purslane, violets, fumaria, barley, coriander, rose and terminalia chebula are anti - itching. proper nutrition is the most important point in health and treatment of humors production with appropriate quality and quantity. it could be concluded that traditional persian medicine therapies can be effective in the treatment of pruritus with mild side effects. by further investigation and research, we can reach more effective treatment methods in the field of traditional persian medicine along with other new medical therapies. | background : pruritus is an unpleasant feeling that can cause the desire of scratching in a person and can be the symptoms of systemic, infectious, and neurological diseases. pruritus is the most common clinical manifestation of skin diseases. pruritus prevalence is 8 - 38% in the general population. causes and treatments of pruritus have been described by traditional persian medicine scientists. the aim of this study was to derive general principles of the proposed treatment to reduce or relieve pruritus.methods:this descriptive study, review traditional medicine books including al canon fil tibb, al - hawi, makhzan ul - adviyyah, al - abniyah an - haghyegh el - adviyah, tuhfat ul - momineen and exir - e - azam. the above - mentioned documents were derived and classified by keywords such as pruritus, hakka, jarab and sherry.results:in traditional persian medicine, there are different causes for pruritus such as accumulation of vapors or acute humors in subcutaneous tissue or weakness of expulsive (dafia) faculty and its treatment is based on removing the causes. proper nutrition, bathing, and removing pathogenic humors are involved in the treatment. according to this study, some plants such as cassia fistula, purslane, violets, fumaria, barley, coriander, rose and terminalia chebula are anti-itching.conclusion:proper nutrition is the most important point in health and treatment of humors production with appropriate quality and quantity. pruritus can be treated by lifestyle modification and using medicinal plants. it could be concluded that traditional persian medicine therapies can be effective in the treatment of pruritus with mild side effects. by further investigation and research, we can reach more effective treatment methods in the field of traditional persian medicine along with other new medical therapies. |
in june 2006, a 74-year - old woman residing in a house in rural new orleans was bothered by a considerable number (> 50) of insect bites. the woman observed many bugs in the house and showed them to a fumigator, who identified them as triatomines. an internet search showed the potential for transmission of chagas disease, and the woman sought help from a local health sciences center. serum samples from both residents of the house were tested for antibodies to t. cruzi at the centers for disease control and prevention (cdc) by an indirect fluorescent antibody (ifa) test. samples were also tested at loyola university (new orleans, la, usa) and then at cdc. by using an experimental dipstick assay (trypanosoma detect ; inbios international inc., the woman resident was positive for antibodies to t. cruzi by ifa at dilutions of 1:128 (4 weeks after being bitten) and 1:64 (10 weeks after being bitten) and by dipstick assay. she was positive for trypanosomes by hemoculture testing with 10 ml blood and coculture in macrophages (13) 4 months after being bitten. trypanosomes consistent with t. cruzi were observed in culture beginning on day 46 of culture, and amplification of a t. cruzi specific 24s rrna gene target confirmed that the isolate was t. cruzi. the index resident had a history of 5 trips to areas endemic for chagas disease : zacatecas, mexico (1970) ; cozumel, mexico (1990) ; belize (1991) ; guatemala (19988) ; and costa rica (1998), each of < 2 weeks duration, with stays in improved tourist hotels (less likely to harbor triatomines) except for the belize trip, which included an 1-week stay in a palm thatch - roofed cabin. she had not used intravenous drugs or had a blood transfusion or organ transplant, and she is not the daughter of latin american immigrants. except for fatigue, cardiac evaluation that included an electrocardiogram showed normal results, and she decided not to take medication. her residence of 29 years was located on 7.66 acres of bottomland hardwood forest, with many gaps that provided ready access for insects. a house inspection showed fecal streaks characteristic of triatomines on walls, which were identical to what the patient reported seeing on her nightgown. twenty dead adult triatomines were collected in the house (after fumigation) and in another building on the property that contained a bed. no nymphs or eggs were found, which suggests that the house had not been colonized. one live second - stage nymph was collected in a nearby armadillo burrow 50 m from the house. all triatomines collected were identified as t. sanguisuga according to the key of lent and wygodzinsky (6) (figure). because all triatomines except the nymph were dead, pcr was used to determine t. cruzi infection status (14). the last 2 segments of the abdomen were removed from each insect, placed in 200 l 1 pcr buffer (applied biosystems, foster city, ca, usa), boiled for 15 min, and centrifuged. a total of 5 l of supernatant was amplified in a 50-l reaction (3.5 mmol / l mgcl2 and 2 u taq dna polymerase). the primers used anneal to the t. cruzi minicircle dna and were tc3 : 5-ttgaacgcccctcccaaaac-3 and tc4 : 5-gattggggttggtgtaatata-3. the cycling parameters were an initial denaturation step at 94c for 3 min ; 35 cycles at 94c, 55c, and 72c, each for 1 min ; and a 10-min extension at 72c (programmable thermal controller ; mj research, watertown, ma, usa). twenty percent of the pcr product was subjected to electrophoresis on a 1.8% agarose gel and visualized by uv transillumination after staining with ethidium bromide. a positive control of 5 l of t. cruzi parasites boiled in 1 pcr buffer and a negative control without the dna template were included with every pcr. samples that failed to amplify were spiked with 5 l of t. cruzi parasites boiled in 1 pcr buffer and reamplified to ensure that the lack of product was not caused by inhibition of the pcr. more than half of the triatomines were positive for t. cruzi (56%, 10/18 ; 3 failed to amplify), with more positive females (73%, 8/11) than males (50%, 3/6). plasma from the resident dog and 7 other dogs living 1 mile away all tested negative by ifa at cdc. the assertion that the patient contacted t. cruzi in louisiana is strongly supported by limited travel history to disease - endemic areas and stays mostly in improved housing (risk for chagas disease transmission is associated with longer residence in disease - endemic areas), lack of other risk factors, and large numbers of infected t. sanguisuga in the house. however, the streaks on her nightgown consistent with triatomine feces indicate exposure, and the parasite could have been introduced into any of her numerous bite wounds. however, hurricane katrina had hit the area 9 months earlier and increases in domestic infestation with triatomines have been previously reported after a hurricane (15). anecdotally, the armadillo population increased substantially in the months after hurricane katrina, and one can speculate that these hosts supported a larger bug population, who later sought other bloodmeal sources as the armadillo population returned to prestorm levels. | autochthonous transmission of the chagas disease parasite, trypanosoma cruzi, was detected in a patient in rural new orleans, louisiana. the patient had positive test results from 2 serologic tests and hemoculture. fifty - six percent of 18 triatoma sanguisuga collected from the house of the patient were positive for t. cruzi by pcr. |
a 2.5 kb vec promoter sequence was cloned between insulators from the chicken -globin gene 29 and used to drive cre expression. r26r - lacz/+;vec - cre or r26r - yfp/+;vec - cre conceptuses were generated by crossing vec - cre/+ males with r26r - lacz/+ or r26r - yfp/+ females. runx1;vec - cre and runx1;vec - cre conceptuses were generated by crossing runx1 ; vec - cre/+ males with runx1 females. runx1;r26r - yfp/+;vec - cre or runx1;r26r - yfp/+;vec - cre conceptuses were generated from crossing runx1;r26r - yfp/+ ; vec - cre/+ males with runx1 females. conceptuses were suspended in phosphate - buffered saline or in a 1:2 mixture of benzyl alcohol / benzyl benzoate and visualized with a stereomicroscope. x - gal (sigma) staining was performed as described previously 4. in some cases the conceptuses were also incubated with rat anti - mouse pecam1 and abc reagent at 4c 30, then treated with dab peroxidase substrate. conceptuses were embedded in paraffin, sectioned, and counterstained with nuclear fast red. to analyze intra - aortic clusters, embryos were fixed and stained with anti - c - kit and anti - pecam1-antibodies. a 1:2 mix of benzyl alcohol / benzyl benzoate was used to increase the transparency of tissues. three - dimensional reconstructions were generated from z - stacks (6287 serial sections). methylcellulose colony forming assays were performed as described previously 4 and colonies counted after 7 days. immuno - staining was performed with phycoerytherin, allophycocyanin, or alexa fluor 647 conjugated antibodies stained cells were analyzed on a becton dickinson facscalibur flow cytometer. a 2.5 kb vec promoter sequence was cloned between insulators from the chicken -globin gene 29 and used to drive cre expression. r26r - lacz/+;vec - cre or r26r - yfp/+;vec - cre conceptuses were generated by crossing vec - cre/+ males with r26r - lacz/+ or r26r - yfp/+ females. runx1;vec - cre and runx1;vec - cre conceptuses were generated by crossing runx1 ; vec - cre/+ males with runx1 females. runx1;r26r - yfp/+;vec - cre or runx1;r26r - yfp/+;vec - cre conceptuses were generated from crossing runx1;r26r - yfp/+ ; vec - cre/+ males with runx1 females. conceptuses were suspended in phosphate - buffered saline or in a 1:2 mixture of benzyl alcohol / benzyl benzoate and visualized with a stereomicroscope. x - gal (sigma) staining was performed as described previously 4. in some cases the conceptuses were also incubated with rat anti - mouse pecam1 and abc reagent at 4c 30, then treated with dab peroxidase substrate. conceptuses were embedded in paraffin, sectioned, and counterstained with nuclear fast red. to analyze intra - aortic clusters, embryos were fixed and stained with anti - c - kit and anti - pecam1-antibodies. a 1:2 mix of benzyl alcohol / benzyl benzoate was used to increase the transparency of tissues. three - dimensional reconstructions were generated from z - stacks (6287 serial sections). methylcellulose colony forming assays were performed as described previously 4 and colonies counted after 7 days. immuno - staining was performed with phycoerytherin, allophycocyanin, or alexa fluor 647 conjugated antibodies stained cells were analyzed on a becton dickinson facscalibur flow cytometer. | hscs are the founder cells of the adult hematopoietic system, and thus knowledge of the molecular program directing their generation during development is important for regenerative hematopoietic strategies. runx1 is a pivotal transcription factor required for hsc generation in the vascular regions of the mouse conceptus - the aorta, vitelline and umbilical arteries, yolk sac and placenta 1, 2. it is thought that hscs emerge from vascular endothelial cells through the formation of intra - arterial clusters 3 and that runx1 functions during the transition from hemogenic endothelium to hscs 4, 5. here we show by conditional deletion that runx1 activity in vascular endothelial cadherin (vec) positive endothelial cells is indeed essential for intra - arterial cluster, hematopoietic progenitor, and hsc formation. in contrast, runx1 is not required in cells expressing vav, one of the first pan - hematopoietic genes expressed in hscs. collectively these data show that runx1 function is essential in endothelial cells for hematopoietic progenitor and hsc formation from the vasculature, but its requirement ends once or before vav is expressed. |
the main study sample included 190 women aged 33 to 68 years recruited through the elective surgery schedule of the gynecology unit at the laval university medical research center, qubec, canada. surgeries were scheduled for total (n = 184) or subtotal (n = 6) abdominal hysterectomies sometimes accompanied by a salpingo - oophorectomy of one (n = 27) or two (n = 71) ovaries. reasons for surgery included one or more of the following : myoma (n = 90), menorrhagia / menometrorrhagia (n = 83), endometriosis (n = 30), fibroids (n = 29), benign cyst (n = 28), incapacitating dysmenorrhea (n = 18), pelvic pain (n = 11), endometrial hyperplasia (n = 8), polyp (n = 7), pelvic adhesions (n = 5), adenomyosis (n = 3), severe premenstrual syndrome (n = 2), and/or ovarian thecoma (n = 1). menopausal status determination was based on follicle - stimulating hormone levels, menstrual history questionnaires, and medical files. thirty - three women were identified as postmenopausal, whereas the remaining women were identified as pre- or peri - menopausal (n = 157). severely obese women (n = 17) undergoing biliopancreatic diversion were recruited at the laval university cardiology and pulmonology institute. they were aged 47 years on average, and their bmi ranged from 39.9 to 70.5 kg / m. severe obesity was defined using the bmi cutoff of the world health organization (who) (40 kg / m). written informed consent the project was approved by the ethics committees of laval university cardiology and pulmonology institute and laval university medical research center. within a few days before or after the surgery, measurements of total adiposity and body fat distribution were performed in most women (n = 150). classification of normal weight, overweight, and obese women was performed using bmi cutoff points of who. total body fat mass and lean body mass were determined using dual - energy x - ray absorptiometry. abdominal subcutaneous and visceral adipose tissue cross - sectional area measures were obtained at the l4-l5 vertebrae level using a light speed 1.1 ct scanner (general electric medical systems, milwaukee, wi) (20,21). plasma vldl were isolated by ultracentrifugation, and the hdl fraction was obtained after precipitation of ldl in the infranatant with heparin and mncl2. the hdl2 and hdl3 subfractions were obtained after further precipitation of hdl2 with dextran sulfate. cholesterol and triglyceride levels were measured in plasma and lipoprotein fractions by enzymatic methods with a technicon ra-500 analyzer (bayer, etobicoke, canada) as previously described (22). apolipoprotein (apo) b concentrations were measured in plasma, vldl, or ldl fractions by the rocket immunoelectrophoretic method of laurell (20). glucose was measured using the glucose oxidase method and insulin was quantified by radioimmunoassay (linco research, st. the homeostasis model assessment of insulin resistance (homa - ir) index was calculated using the following formula : fasting insulin (in u / ml) fasting glucose (in mmol / l) 22.5 (23). none of the women recruited from the gynecology unit had a previous diagnosis of diabetes or took antidiabetic drugs. however, monitoring fasting plasma glycemia on the morning of surgery revealed that 30 women had impaired fasting glycemia (> 6.1 mmol / l). within 15 min after the beginning of the surgical procedure, paired samples of subcutaneous and omental adipose tissue were respectively collected at the site of surgical incision and the greater omentum. a portion of the tissue sample was digested 45 min at 37c in krebs - ringer - henseleit buffer supplemented with 350 units / ml of type 1 collagenase according to a modified version of the rodbell method (21,24). mature adipocyte suspensions were then washed three times using krebs - ringer - henseleit buffer. mean adipocyte diameter for each adipose tissue sample was calculated from 250 individual measurements using scion image software (frederick, md). sd and skewness of adipocyte size distributions were not independently associated to metabolic alterations and were not included in subsequent analyses. the number of subcutaneous and visceral adipocytes at the l4-l5 vertebrae level was estimated by dividing abdominal subcutaneous and visceral adipose tissue area, respectively, by subcutaneous and omental mean cross - sectional adipocyte surface. nonlinear curve fit of the association between subcutaneous or omental adipocyte size and bmi was performed using a one - phase exponential equation in prism graphpad software. an independent linear regression analysis was performed to predict adipocyte size in each compartment separately using measures of body composition (bmi, fat mass, and lean body mass) and fat distribution (total, subcutaneous, and visceral adipose tissue area at l4-l5 vertebrae) as well as age, menopausal status, and hormone replacement therapy. in models with models were first selected based on the mallows ' cp statistic and further improved using the adjusted r and the predicted residual sum of squares (press). for each model, the study sample was stratified in two subgroups according to whether women had a positive (hypertrophy) or negative (hyperplasia) residual. in each fat depot model, adiposity and metabolic measures between subgroups variables that were not normally distributed based on a significant shapiro - wilk test (p 1.69 mmol / l) using measures of adipocyte size and number in each adipose tissue depot. the model was adjusted for body composition and fat distribution variables as well as age and menopausal status. odds ratios (ors) for each independent variable were computed for a 10% increase. the main study sample included 190 women aged 33 to 68 years recruited through the elective surgery schedule of the gynecology unit at the laval university medical research center, qubec, canada. surgeries were scheduled for total (n = 184) or subtotal (n = 6) abdominal hysterectomies sometimes accompanied by a salpingo - oophorectomy of one (n = 27) or two (n = 71) ovaries. reasons for surgery included one or more of the following : myoma (n = 90), menorrhagia / menometrorrhagia (n = 83), endometriosis (n = 30), fibroids (n = 29), benign cyst (n = 28), incapacitating dysmenorrhea (n = 18), pelvic pain (n = 11), endometrial hyperplasia (n = 8), polyp (n = 7), pelvic adhesions (n = 5), adenomyosis (n = 3), severe premenstrual syndrome (n = 2), and/or ovarian thecoma (n = 1). menopausal status determination was based on follicle - stimulating hormone levels, menstrual history questionnaires, and medical files. thirty - three women were identified as postmenopausal, whereas the remaining women were identified as pre- or peri - menopausal (n = 157). severely obese women (n = 17) undergoing biliopancreatic diversion were recruited at the laval university cardiology and pulmonology institute. they were aged 47 years on average, and their bmi ranged from 39.9 to 70.5 kg / m. severe obesity was defined using the bmi cutoff of the world health organization (who) (40 kg / m). written informed consent the project was approved by the ethics committees of laval university cardiology and pulmonology institute and laval university medical research center. within a few days before or after the surgery, measurements of total adiposity and body fat distribution were performed in most women (n = 150). classification of normal weight, overweight, and obese women was performed using bmi cutoff points of who. total body fat mass and lean body mass were determined using dual - energy x - ray absorptiometry. abdominal subcutaneous and visceral adipose tissue cross - sectional area measures were obtained at the l4-l5 vertebrae level using a light speed 1.1 ct scanner (general electric medical systems, milwaukee, wi) (20,21). plasma vldl were isolated by ultracentrifugation, and the hdl fraction was obtained after precipitation of ldl in the infranatant with heparin and mncl2. the hdl2 and hdl3 subfractions were obtained after further precipitation of hdl2 with dextran sulfate. cholesterol and triglyceride levels were measured in plasma and lipoprotein fractions by enzymatic methods with a technicon ra-500 analyzer (bayer, etobicoke, canada) as previously described (22). apolipoprotein (apo) b concentrations were measured in plasma, vldl, or ldl fractions by the rocket immunoelectrophoretic method of laurell (20). glucose was measured using the glucose oxidase method and insulin was quantified by radioimmunoassay (linco research, st. the homeostasis model assessment of insulin resistance (homa - ir) index was calculated using the following formula : fasting insulin (in u / ml) fasting glucose (in mmol / l) 22.5 (23). none of the women recruited from the gynecology unit had a previous diagnosis of diabetes or took antidiabetic drugs. however, monitoring fasting plasma glycemia on the morning of surgery revealed that 30 women had impaired fasting glycemia (> 6.1 mmol / l). within 15 min after the beginning of the surgical procedure, paired samples of subcutaneous and omental adipose tissue were respectively collected at the site of surgical incision and the greater omentum. a portion of the tissue sample was digested 45 min at 37c in krebs - ringer - henseleit buffer supplemented with 350 units / ml of type 1 collagenase according to a modified version of the rodbell method (21,24). mature adipocyte suspensions were then washed three times using krebs - ringer - henseleit buffer. mean adipocyte diameter for each adipose tissue sample was calculated from 250 individual measurements using scion image software (frederick, md). sd and skewness of adipocyte size distributions were not independently associated to metabolic alterations and were not included in subsequent analyses. the number of subcutaneous and visceral adipocytes at the l4-l5 vertebrae level was estimated by dividing abdominal subcutaneous and visceral adipose tissue area, respectively, by subcutaneous and omental mean cross - sectional adipocyte surface. nonlinear curve fit of the association between subcutaneous or omental adipocyte size and bmi was performed using a one - phase exponential equation in prism graphpad software. an independent linear regression analysis was performed to predict adipocyte size in each compartment separately using measures of body composition (bmi, fat mass, and lean body mass) and fat distribution (total, subcutaneous, and visceral adipose tissue area at l4-l5 vertebrae) as well as age, menopausal status, and hormone replacement therapy. in models with models were first selected based on the mallows ' cp statistic and further improved using the adjusted r and the predicted residual sum of squares (press). for each model, the study sample was stratified in two subgroups according to whether women had a positive (hypertrophy) or negative (hyperplasia) residual. in each fat depot model, adiposity and metabolic measures between subgroups variables that were not normally distributed based on a significant shapiro - wilk test (p 1.69 mmol / l) using measures of adipocyte size and number in each adipose tissue depot. the model was adjusted for body composition and fat distribution variables as well as age and menopausal status. odds ratios (ors) for each independent variable were computed for a 10% increase. subcutaneous and omental adipocyte size was measured in a sample of 207 lean to severely obese women. the curve - linear relationship between bmi and adipocyte size in each depot adipocyte size increased along with bmi and tended to reach a plateau at 130 m in severely obese women. taken as a whole, omental adipocytes were smaller than subcutaneous adipocytes but tended to reach similar sizes in severely obese women (bmi > 40 kg / m). subsequent analyses were performed in lean to moderately obese women undergoing gynecological surgery, since the relation between adipocyte size and adiposity became nonlinear in severely obese women. subcutaneous and omental adipocyte diameter according to bmi in women undergoing abdominal gynecologic surgery (n = 190) and biliopancreatic diversion (n = 17). women were aged 47.7 5.5 years (range 3068.3 years) with a mean bmi of 28.5 8.8 kg / m (range 17.6 70.5 kg / m). mean subcutaneous and omental adipocyte sizes were 101.5 15.4 m and 85.2 18.6 m, respectively. associations between regional adipocyte size and lipid profile measures were computed in a subsample of women for which data on body composition, fat distribution, and lipid profile were available (n = 150). characteristics of these women are shown in table 1. according to mean bmi and body composition measures, we performed linear regression models to predict subcutaneous and omental adipocyte sizes using all body composition and fat distribution variables available (table 2). based on this analysis, lean body mass as well as subcutaneous and visceral adipose tissue area explained 30.8% of the variance in subcutaneous adipocyte size (model 1). lean body mass, bmi, and visceral adipose tissue area explained 54.3% of the variance in omental adipocyte size (model 2). physical and metabolic characteristics of the subsample of 150 women linear regression analyses predicting omental and subcutaneous adipocyte size in women regression models included bmi, body fat mass, lean body mass, subcutaneous adipose tissue area, visceral adipose tissue area, age, menopausal status, and hormone replacement therapy. models were selected based on the mallows cp, adjusted r, and press statistics ; colinearity among independent variables was assessed through inflation and condition index statistics. a large percentage of the variance in adipocyte size remained unexplained despite considering body composition and fat distribution measures. to assess the impact of adipocyte size variation on the metabolic profile independent of body composition and fat distribution, we stratified the study sample according to the difference between measured and predicted adipocyte size in each model. women with larger adipocytes than predicted by the regression model (positive residual) were identified as hypertrophic, whereas women with smaller adipocytes than predicted by the model were identified as hyperplasic (negative residual) in the corresponding adipose tissue compartment. by design, stratifications performed using each model generated subgroups of women characterized by identical body composition and fat distribution values. figure 2 shows adipocyte size and number in each adipose tissue compartment according to the presence of hypertrophy or hyperplasia in subcutaneous (left) or omental (right) adipose tissue. women characterized by hypertrophic subcutaneous adipocytes had fewer but larger subcutaneous adipocytes than women with hyperplasic subcutaneous adipocytes (p < 0.0001). women characterized by subcutaneous adipocyte hypertrophy also had slightly larger omental adipocytes, but their cell number remained similar to that of women characterized by subcutaneous adipocyte hyperplasia (p < 0.05). similarly, larger but fewer omental adipocytes were observed in women characterized by omental adipocyte hypertrophy compared with women characterized by omental adipocyte hyperplasia (p < 0.0001). although the number of subcutaneous adipocytes was similar in both groups derived from model 2, slightly larger subcutaneous adipocytes were observed in women characterized by omental adipocyte hypertrophy (p < 0.05). adipocyte diameter (a) and adipocyte number at the l4-l5 vertebrae level (b) are shown for subcutaneous (sc ; model 1 ; n = 150) and omental (om ; model 2 ; n = 150) adipose tissue cellularity stratifications. values are mean sem. p < 0.05 ; p < 0.01. no significant difference in blood lipids was observed between women characterized by subcutaneous adipocyte hypertrophy versus subcutaneous adipocyte hyperplasia (fig. women characterized by omental adipocyte hypertrophy had higher levels of plasma triglycerides, higher vldl triglycerides, and higher vldl cholesterol levels compared with women characterized by omental adipocyte hyperplasia (fig. the ratio of total cholesterol to hdl cholesterol was significantly higher in women characterized by omental adipocyte hypertrophy compared with women characterized by omental adipocyte hyperplasia (fig. 3d, right). increased triglyceride and cholesterol content in the vldl fraction may suggest increased particle size (10.89 3.85 vs. 9.27 3.97 mmol of lipids / g of apob, p < 0.01) rather than increased particle number as estimated by vldl - apob (0.13 0.06 vs. 0.12 0.07 g / l, ns). lipid profile in women characterized by hypertrophic or hyperplasic subcutaneous (model 1 ; n = 150) and omental (model 2 ; n = 150) adipocytes. overnight fast values of plasma triglyceride (a) (mmol / l), vldl triglyceride (b) (mmol / l), vldl cholesterol (c) (mmol / l), and total - to - hdl cholesterol (d) are shown. subdivision of women in three subgroups according to the sd of the linear regression model residuals generated results that are comparable with the two - group analyses. specifically, women with adipocyte sizes inside of 1 sd around the mean showed an intermediate phenotype for blood lipid levels compared with women above or below 1 sd of the regression residuals (data not shown). no significant difference was observed in fasting glucose (5.75 0.65 vs. 5.60 0.59, ns) and insulin levels (10.15 5.93 vs. 8.88 5.05, ns) as well as in homa - ir index (2.68 1.71 vs. 2.34 1.60, ns) among women characterized by omental adipocyte hypertrophy or hyperplasia. however, when the linear regression analyses were performed using a single measure of total adiposity (i.e., fat mass) to predict adipocyte size, we observed a trend for higher fasting glucose (5.75 0.64 vs. 5.57 0.59, p = 0.09), significantly higher fasting insulin levels (10.9 6.3 vs. 7.8 3.8, p < 0.005), as well as a higher homa - ir index (2. < 0.001) in women characterized by subcutaneous adipocyte hypertrophy compared with women characterized by subcutaneous adipocyte hyperplasia. to confirm results of the regression analyses, subgroups of women matched for fat mass, visceral adipose tissue area, and subcutaneous adipocyte size but with either small or large omental adipocytes were compared (table 3). by design, both subgroups had a similar body composition and fat distribution. although subcutaneous adipocyte size and number were comparable, women with large omental adipocytes had a lower number of visceral adipocytes. women with large omental adipocytes had higher levels of plasma triglycerides, of vldl triglycerides, and of vldl cholesterol. a trend for lower hdl cholesterol the lower hdl cholesterol value was explained by significantly lower cholesterol content in hdl2, but not in hdl3 lipoproteins, in women with larger omental adipocytes. moreover, the total cholesterol - to - hdl - cholesterol ratio tended to be higher in women with large omental adipocytes compared with women with small omental adipocytes. fasting glucose and insulin concentrations as well as the homa - ir index were similar between women with large versus small omental adipocytes (table 3). characteristics of women matched for visceral adipose tissue area and subcutaneous adipocyte size but with either small or large omental adipocytes (n = 54) the risk of hypertriglyceridemia, defined by triglyceride levels greater than 1.69 mmol / l in women, was assessed using logistic regression analyses (table 4). when adipocyte size and number in each fat depot were included in the model, the risk of hypertriglyceridemia was associated with adipose tissue cellularity measures of the visceral, but not the subcutaneous, fat compartment. indeed, the risk of hypertriglyceridemia was significantly higher in women with 10% larger omental adipocytes (or 2.08 [95% ci 1.483.04 ]) and with 10% more adipocytes in visceral adipose tissue (or 1.16 [1.031.30 ]). after adjustment for bmi, body fat mass, lean body mass, subcutaneous adipose tissue area, visceral adipose tissue area, age, and menopausal status, omental adipocyte size (adjusted or 4.06 [1.929.97 ]) and number (adjusted or 1.55 [1.122.29 ]) remained independent predictors of hypertriglyceridemia in women. mmol / l logistic regression model was adjusted for body fat mass (ns), subcutaneous adipose tissue area (ns), visceral adipose tissue area (p = 0.07), free fat mass (ns), bmi (ns), age (ns), menopausal status (ns), and hormone replacement therapy (ns) ; n = 150. we designed this study to assess whether subcutaneous and omental adipose tissue cellularity measures were related to metabolic alterations independent of body composition and fat distribution in women. using linear regression analyses, women were subdivided in two groups having lower - than - predicted (hyperplasia) or higher - than - predicted (hypertrophy) adipocyte sizes in subcutaneous and omental adipose tissue. women characterized by omental adipocyte hypertrophy presented a deleterious lipid profile compared with women characterized by omental adipocyte hyperplasia. these alterations in the lipid profile were, by design, independent of differences in body composition and fat distribution. in contrast, lipid profiles were similar in women characterized by subcutaneous adipocyte hypertrophy versus subcutaneous adipocyte hyperplasia. in logistic regression analyses including body composition and fat distribution measures, we estimated that a 10% enlargement of omental adipocytes increased the risk of hypertriglyceridemia by more than fourfold, whereas enlarged subcutaneous adipocyte size failed to significantly alter the risk of hypertriglyceridemia in women. to a lower extent, a 10% increase in the number of visceral, but not subcutaneous, adipocytes multiplied the risk of hypertriglyceridemia in women by 1.55-fold. these results suggest that omental fat cell size and number predict lipid profile alterations independent of body composition and fat distribution in women. despite the fact that total and visceral adiposity are known modulators of the lipid profile, we show here for the first time that the extent of omental adipocyte hypertrophy may influence this relation. previous studies reported that subcutaneous adipocyte hypertrophy was associated with alterations in glucose homeostasis (1,5,911). (1) on adipocyte hypertrophy and adipocyte turnover showed that individuals characterized by larger subcutaneous adipocytes had higher fasting insulin levels and homa - ir index. in the current study, all measures of body composition and fat distribution as well as age and menopausal status were used to prepare fully adjusted predictive models for subcutaneous and omental adipocyte size. using these models, we failed to observe alterations in glucose homeostasis measures in women characterized by adipocyte hypertrophy in subcutaneous or omental adipose tissue. on the other hand, linear regression analyses predicting adipocyte size performed using only one measure of total adiposity (e.g., fat mass) revealed glucose homeostasis alterations in women characterized by subcutaneous adipocyte hypertrophy compared with women characterized by subcutaneous adipocyte hyperplasia. these results suggest that associations of glucose homeostasis with subcutaneous adipocyte hypertrophy, as observed by others (1,5,911), may arise from differences in abdominal fat distribution rather than differences in subcutaneous adipose tissue cellularity. lack of statistical adjustment for fat distribution measures in previous studies may explain discrepancies regarding glucose homeostasis alterations in women characterized by hypertrophic subcutaneous adipocytes (1,5,911). to our knowledge, we are the first to report an independent association between omental adipocyte hypertrophy and alterations of the lipid profile in women. with the exception of two studies (10,19), most authors assessed the association between adipocyte size and metabolic parameters using only measures from subcutaneous adipose tissue (1,3,5,11). these studies reported that enlarged subcutaneous adipocytes were associated with insulin resistance independent of obesity itself (1,5,11). in contrast, mundi. (3) observed that insulin and triglyceride levels were slightly better predicted by body composition measures than subcutaneous adipocyte size. therefore, they dismissed the individual effect of subcutaneous adipocyte size on metabolic parameters in men and women (3). subcutaneous and omental adipocyte size measures are closely related (4,25), and part of the effect of omental adipocyte hypertrophy may be indirectly related to subcutaneous adipocyte size. however, lack of visceral adipocyte size measures in most of these studies may explain discrepancies observed between our analysis and previous literature (1,3,5). we corroborate reports showing that body composition and fat distribution measures largely predict subcutaneous and omental adipocyte size (4). however, after adjustment for all anthropometric measures available, interindividual variability observed in subcutaneous and omental adipose tissue was not completely explained. this observation is consistent with the hypothesis that adipocyte size is regulated by factors that are independent of variations in body composition and fat distribution (4). bergman. (26) suggested that fat accumulation occurs primarily in the visceral adipose tissue depot and fat storage then spills over in the subcutaneous adipose tissue depot, whereas others have proposed that subcutaneous fat is a primary compartment (27,28). in any case, lipid uptake and lipolysis rates in each fat depot appear as dynamic processes. (2) observed that hyperplasia is predominant in the subcutaneous adipose tissue, whereas adipocyte hypertrophy is present in both adipose tissue depots. because less than 10% of mature adipocytes are renewed each year in subcutaneous adipose tissue (29), cell hyperplasia may represent a long - term adipose tissue adaptation to face excess energy intake. conversely, the predominance of cell hypertrophy in visceral adipose tissue may suggest that it is a short - term storage compartment. activated receptor- agonist induced hyperplasia has been shown to promote fat redistribution and to reduce visceral fat accumulation (30). these observations show that increasing adipogenic capacity in subcutaneous adipose tissue may attenuate visceral adipocyte hypertrophy and related alterations. although this study was not designed to investigate the mechanisms involved in the association of adipocyte hypertrophy and metabolic alterations, several hypotheses may be put forward. involvement of visceral, but not subcutaneous, adipose tissue cellularity could indirectly reinforce the role of excess free fatty acids (31). indeed, previous studies have reported that large adipocytes showed alterations in lipolysis, insulin sensitivity, and adipokine secretion compared with smaller adipocytes from the same individual (12,13,15,16). although visceral adipose tissue is not believed to be a major source for the circulating free fatty acid pool, the contribution of visceral adipose tissue lipolysis to hepatic free fatty acid delivery is positively associated to visceral fat accumulation (32). increased responsiveness to -adrenergic agonist stimulation (3335) and decreased sensitivity to insulin suppression of lipolysis (36,37) in hypertrophic omental adipocytes could possibly impact splanchnic free fatty acid levels, especially in the visceral obese. increased free fatty acid delivery to the liver was suggested to increased triglyceride - rich vldl production, which is associated with small, dense ldl particles as well as with lower hdl cholesterol levels (38,39). the altered lipid profile observed in women characterized by omental adipocyte hypertrophy is, therefore, consistent with this hypothesis. an altered adipokine secretion pattern, as observed with large adipocytes, may also influence local adipose tissue lipid metabolism and generate abnormal adipose - derived signaling through the portal vein (15). moreover, reduced cellular stability of large adipocytes may increase the risk of cell rupture in hypertrophic adipose tissue (40). chronic, low - grade inflammation provoked by adipocyte death may then contribute to the metabolic alterations associated with obesity (41). results of this study are strengthened by the use of extensive measures of adiposity and body fat distribution together with characterization of subcutaneous and omental adipose tissue cellularity. moreover, the absence of differences in body composition and fat distribution measures between women characterized by hypertrophy and hyperplasia supports the validity and the strength of the methodology used. we suggest that this stratification is adequate to assess metabolic alterations associated to subcutaneous or omental hypertrophy, independent of body composition and fat distribution. however, some limitations in the study design should be acknowledged. the present analyses are based on cross - sectional data, and it is hazardous to conclude cause - and - effect relationships between visceral adipocyte hypertrophy and the development of metabolic alterations in women. the difficulty to recruit lean to moderately obese men undergoing abdominal surgery limits our capacity to perform a comparable study in men. most previous studies (1,3,5,9) included both men and women, but only subcutaneous adipose tissue cellularity was assessed. this study was not designed to assess the involvement of very small adipocytes in metabolic alterations since we used collagenase - digested adipose tissue samples. (42) suggesting that the proportion of very small adipocytes is associated with metabolic alterations. finally, because we only had access to omental and not mesenteric adipose tissue, visceral adipocyte numbers were computed using omental adipocyte surface. we assumed that mean adipocyte sizes in each visceral adipose tissue depot are strongly correlated, as previously observed for the subcutaneous and visceral adipose tissue depots (4). during the revision process of this article, hoffstedt. (43) published a similar study reporting that visceral adipose tissue hypertrophy is associated with dyslipidemia. the recruitment of morbidly obese women and our use of detailed assessment of body fat distribution may account for these discrepancies. in conclusion, our results support the hypothesis that omental adipose tissue cellularity is an important predictor of hypertriglyceridemia in women. we demonstrate that the association between omental adipocyte hypertrophy and lipid profile alterations is independent of differences in subcutaneous adipose tissue cellularity, body composition, and fat distribution in women. | objectivewe assessed whether subcutaneous and omental adipocyte hypertrophy are related to metabolic alterations independent of body composition and fat distribution in women.research design and methodsmean adipocyte diameter of paired subcutaneous and omental adipose tissue samples was obtained in lean to obese women. linear regression models predicting adipocyte size in both adipose tissue depots were computed using body composition and fat distribution measures (n = 150). in a given depot, women with larger adipocytes than predicted by the regression were considered as having adipocyte hypertrophy, whereas women with smaller adipocytes than predicted were considered as having adipocyte hyperplasia.resultswomen characterized by omental adipocyte hypertrophy had higher plasma and vldl triglyceride levels as well as a higher total - to - hdl cholesterol ratio compared with women characterized by omental adipocyte hyperplasia (p < 0.05). conversely, women characterized by subcutaneous adipocyte hypertrophy or hyperplasia showed a similar lipid profile. in logistic regression analyses, a 10% enlargement of omental adipocytes increased the risk of hypertriglyceridemia (adjusted odds ratio [or ] 4.06, p < 0.001) independent of body composition and fat distribution measures. a 10% increase in visceral adipocyte number also raised the risk of hypertriglyceridemia (adjusted or 1.55, p < 0.02). associations between adipocyte size and homeostasis model assessment of insulin resistance were not significant once adjusted for adiposity and body fat distribution.conclusionsthese results suggest that omental, but not subcutaneous, adipocyte hypertrophy is associated with an altered lipid profile independent of body composition and fat distribution in women. |
a 20-year - old incarcerated indigenous australian male, with a history of hepatitis c and intravenous drug abuse, was admitted to the intensive care unit with fulminant hepatic failure following a 5-day history of jaundice, nausea, and vomiting. the patient was opioid dependent but had been previously well with no relevant medical history. investigations at the time of presentation revealed a severe acute hepatitis with marked synthetic dysfunction. his arterial lactate was 5.6 mmol / l, ph was 7.5 and serum ammonia was 132 viral serology revealed positive hepatitis c virus (hcv) antibodies and evidence of immunity to hepatitis b virus. corroborative history from prison medical staff revealed that the patient had thrice injected buprenorphine 1 day prior to the onset of his symptoms. the patient developed life - threatening multiorgan failure as a consequence of the fulminant hepatic failure and met listing criteria for liver transplantation. the patient was managed as per the american association for the study of liver diseases acute liver failure guidelines 1. he was commenced on broad - spectrum antibiotics, antifungal prophylaxis, an n - acetyl - cysteine infusion and continuous veno - venous hemofiltration. the patient survived with supportive intensive care management and was discharged from hospital after 42 days. upon discharge, his liver function was improving with an alt of 278 u / l, inr of 1.2, albumin of 24 g / l and serum bilirubin of 367 mol / l, having peaked at 450 mol / l. buprenorphine is a potent semisynthetic opioid derivative that is prescribed for the treatment of opioid dependence or for analgesic purposes. it undergoes extensive first pass hepatic metabolism utilizing the p450 (cyp 3a4) system 2. acute liver injury from the misuse of sublingual buprenorphine has been described in several case reports and case series. almost all cases of significant hepatocellular injury have been associated with hepatitis c viremia 37. it has been postulated that hcv induces mitochondrial toxicity, leading to more significant liver damage. the spectrum of hepatotoxicity following therapeutic administration, misuse or overdose of buprenorphine ranges from a mild - to - severe hepatitis. the majority of reported cases of intravenous buprenorphine - associated liver damage have been in the context of known or recently detected hepatitis c infection. although intravenous buprenorphine - induced hepatitis is now well recognized, life - threatening fulminant hepatic failure due to this drug has not previously been reported. acute liver injury from intravenous buprenorphine use has been primarily attributed due to the high parenteral doses obtained from crushed sublingual tablets. the mechanism of toxicity is due to inhibition of mitochondrial respiration and fatty acid - b oxidation, leading to atp depletion and hepatocyte necrosis, in rat models 2. most patients who have restarted conventional sublingual doses following an episode of toxicity have not had recurrent liver injury 3,5. the prevalence of hepatitis c virus (hcv) infections is high among opioid - dependent individuals. care needs to be undertaken when prescribing the newer classes of protease inhibitors ; simeprevir, telaprevir, and boceprevir that are metabolized via the p450 system. in addition, the effect of alcohol on the liver contributes to the altered metabolism of buprenorphine via the cyp3a4 enzyme 8. the additive cns respiratory depressant effects of this combination have been fatal in overdose 9. the possibility of intravenous misuse and drug interactions should not prevent clinicians from considering sublingual buprenorphine as a maintenance treatment for heroin addiction. however, caution should be applied in patients with chronic hcv infection and who are at risk of intravenous misuse. | key clinical messagea 20-year - old indigenous australian male was admitted to the intensive care unit with fulminant hepatic failure secondary to intravenous use of buprenorphine, which had been prescribed sublingually for opioid dependence. intravenous buprenorphine - induced hepatitis is well recognized, however, life - threatening fulminant hepatic failure has not previously been reported. |
gemcitabine - induced hemolytic uremic syndrome (hus) is rare but can often be fatal. early detection of hus is critical so that contributing chemotherapeutic agents, such as gemcitabine, can be discontinued. in clinical trials, hus is characterized by renal failure, microangiopathic hemolytic anemia (maha), and thrombocytopenia.1 in addition, the onset of new and uncontrolled hypertension can be diagnostic for hus.2 while a few cases of gemcitabine - induced hus have been reported globally, but only a single korean case of hus in a lung cancer patient is known.3 with the increase in gemcitabine use in pancreatobiliary cancer, early detection of gemcitabine - induced hus is essential. herein, we describe the first case of hus in a pancreatic cancer patient, associated with gemcitabine use in a korean hospital. in june 2011, a 56-year - old male visited a local hospital complaining of dyspepsia and back pain. contrast - enhanced computed tomography (ct) revealed a 4.4 cm mass at the head of his pancreas (fig. he was referred to our hospital, where we found the mass had invaded the main portal vein, common hepatic artery, and stomach. laboratory tests showed serum hemoglobin level of 14.9 g / dl, platelet count of 166,000/l, blood urea nitrogen of 21.3 mg / dl, and serum creatinine level of 1.28 mg / dl. the carcinoembryonic antigen level was 2.34 ng / ml (normal range, 0 to 5 ng / ml) and carbohydrate antigen 19 - 9 was 798 u / ml (normal range, 0 to 37 u / ml). concurrent chemoradiotherapy (ccrt) with original gemcitabine weekly (1,000 mg / m per week, day 1, 8, 15, 22, and 29) and 25 radiation therapy (total radiation dose, 4,500 cgy) was performed from june 30 to august 3, 2011. one month after ccrt, a repeat abdominal ct showed that the pancreatic mass had decreased from 4.4 to 3.0 cm but remained unresectable. on october 21, 2011, gemcitabine therapy was administered weekly (1,000 mg / m per week, day 1, 8, and 15) for 3 out of 4 weeks. a cumulative dose of gemcitabine from june 30, 2011 was 26,250 mg. in february 2012 during week 2 of cycle 5, the patient was admitted for generalized edema and general weakness. he developed acute renal failure and had an elevated serum creatinine level of 2.08 mg / dl (normal range, 0.5 to 1.40 mg / dl). on physical examination, the patient had dyspnea on exertion, lower extremity pitting edema, and a blood pressure of 200/140 mm hg. the patient 's hemoglobin level had decreased to 7.7 g / dl (normal range, 13.0 to 17.0 g / dl) and a platelet count of 87,000/l (normal range, 150,000 to 400,000/l). the reticulocyte count was 11.69% (normal range, 0.5% to 2.31%), with a corrected reticulocyte count of 6.6% and an elevated lactate dehydrogenase (ldh) level of 459 iu / l (normal range, 110 to 247 iu / l). a peripheral blood smear showed macrocytic hypochromic anemia with mild anisopoikilocytosis composed of schistocytes and acanthocytes (fig. 2). there was no evidence of proteinuria or hematuria, but the patient was positive for urine hemoglobin. the patient 's total bilirubin, obtained from an indirect coombs test, was normal and the patient 's haptoglobin level was < 10 mg / dl. the laboratory tests confirmed maha, thrombocytopenia, and acute renal failure - leading to a diagnosis of gemcitabine - induced hus. after 19 plasmapheresis treatments, the patient 's creatinine level decreased to 2.02 mg / dl and he was able to maintain self - urine output. he was transferred to a nursing home for hospice care and has continued with outpatient palliative therapy. hus is characterized by renal failure, thrombocytopenia and maha characterized by a trio of classic symptoms - elevated ldh, low haptoglobin, and schistocytes on the peripheral blood smear. five typical hus cases were first described by gasser. in a pediatric patient with hemorrhagic diarrhea and enterocolitis caused by verotoxin - producing escherichia coli.4 in 1979, a gastric cancer patient developed chemotherapy - related hus associated with mitomycin c (mmc) and 5-fluorouracil.5 the causes of atypical hus include infectious diseases, malignancy, antineoplastic agents such as mmc, antiplatelet agents, pregnancy, hemolytic anemia, elevated liver enzymes, and low platelet syndrome (hellp syndrome), malignant hypertension, systemic lupus erythematosus, and antiphospholipid syndrome. the incidence of gemcitabine - induced hus is low (0.015% to 0.31%), but the mortality rate is as high as 50% in these patients. there have been previous reported cases in patients with nonsmall cell lung cancer, ovarian cancer, and metastatic breast cancer.6 - 8 a review of the literature revealed only 15 patients of gemcitabine - induced hus in pancreatic cancer. these 15 cases are summarized in table 1.1,2,9 - 15 our case is noteworthy because of the first reported case in korea. the mechanism of gemcitabine - induced hus is unclear with hypotheses including microvascular endothelial damage or immunocomplex mediation.9 an increase of von willebrand factor levels in hus suggests a potential role in hus, but further investigation is needed.5 in the literature, the median duration of gemcibatine therapy was 5.8 months, with the majority of patients developing hus within 1 to 2 months of the last gemcitabine infusion.1 the median time between initiation of chemotherapy and onset of gemcitabine - induced hus was 7.4 months.16 gemcitabine - induced hus developed after an estimated median cumulative dose of 20,000 mg / m with a broad range from 2,450 to 48,000 mg / m with no clear dose - response relationship.11 our patient began developing symptoms of peripheral edema and hypertensive emergency after 7 months of therapy and a cumulative dose of 26,250 mg. in patients treated with gemcitabine, hus must be considered when uncontrolled hypertension or worsening preexisting hypertension develops, and it is important to monitor blood pressure to detect any early indication of gemcitabine - induced hus. although there is currently no consensus as to the best treatment for gemcitabine - induced hus, immediate discontinuation of gemcitabine is accepted as the initial step.17 other treatment modalities include corticosteroids, transfusion of fresh frozen plasma, plasmapheresis, or hemodialysis. however, plasmapheresis reportedly had no direct therapeutic effect, which was attributed to discontinuing gemcitabine administration.18 to confirm a diagnosis of hus, a renal biopsy can be performed to show microvascular damage of arterioles and small arteries occluded by eosinophilic hyaline thrombi containing fibrin and platelet aggregates.1 generally, the histopathology of renal tissue in patients with hus shows characteristic thrombotic microangiopathy consisting of thrombi in blood vessels, glomerular mesangiolysis, and widening of the subendothelial space with detachment of endothelial cells from the glomerular basement membrane.4 a renal biopsy was scheduled for the patient in this case but he decided to cancel the procedure due to the high risk of bleeding. it is possible that a diagnosis of hus may be delayed because anemia and thrombocytopenia can also be induced by myelotoxicity secondary to chemotherapy. in particular, an elevated reticulocyte count could differentiate between anemia due to myelotoxicity of gemcitabine and gemcitabine induced - hus.19 furthermore, a sudden decrease in hemoglobin, sudden renal failure, uncontrolled hypertension, pulmonary congestion, peripheral edema, and thrombocytopenia should alert clinicians of the possibility of hus. when hus is suspected, peripheral blood smears should be screened for the presence of fragmented red blood cells and elevated ldh level.20 in conclusion, few cases of gemcitabine - associated hus have been described. with the increase is the use of gemcitabine therapy for pancreatic cancer, it is important to quickly and accurately diagnose gemcitabine - associated hus. this case report demonstrates that a patient could overcome this life - threatening crisis with early clinical detection and immediate discontinuation of gemcitabine. | hemolytic uremic syndrome (hus) is a rare thrombotic complication characterized by a triad of microangiopathic hemolytic anemia, thrombocytopenia, and acute renal failure. hus may be caused by several different conditions, including infection, malignancy, and chemotherapeutic agents, such as mitomycin, cisplatin, and most recently, gemcitabine. the outcome of gemcitabine - induced hus is poor, and the disease has a high mortality rate. this study reports a case of gemcitabine - induced hus in a patient with pancreatic cancer in korea. |
health related quality of life (hrqol) includes not only physical functioning but also emotional and social functioning dimensions. to assess effectiveness of clinical trials, hrqol measuring instruments hrqol instruments assist healthcare services in creating better relationship between patients and their physicians and improving patient 's outcome. in addition, the obtained results help the health system to distribute their assets and facilitate risk assessment. in pediatric field, an instrument to measure hrqol must be able to evaluate cognitive development and viewpoints of both child and parents. it has been shown that using instrument appropriate to age development would make hrqol reports more reliable.[47 ] the pediatric quality of life inventory (pedsql 4.0) that has been invented to determine hrqol is one of the most competent measures in this field. the pedsql 4.0 is designed to integrate the relative merits of generic and disease specific hrqol measuring instruments. adding some constructs and items to the previous pedsql 2.0 and 3.0 matches the pedsql 4.0 generic core scales to dimensions described by who. the pedsql 4.0 generic core scales have been designed to assess child self report for ages 518 and parent proxy report for ages 218. furthermore, disease - specific modules have been invented for chronic conditions and children with cancers by varni. the feasibility, reliability and validity of the original version of the pedsql 4.0 (american english) have been widely demonstrated.[37912 ] considering the capability of pedsql 4.0 in assessing child and parents and its simplicity in applying and scoring, various translations of different languages has been validated.[1321 ] iran has one of the youngest populations in the world. we were the first iranian group obtaining the inventor 's approval (dr. james varni). even though, a persian version of the pedsql 4.0 has been published before our study, according to our best knowledge we carried out translation and validation of the pedsql 4.0 into persian language for children less than 13 years old as the first group. the survey was performed in accordance with the ethical standards of the helsinki declaration. after obtaining the agreement of the main inventor (dr. james varni), approval of isfahan education office license and ethics committee of the research department of isfahan university of medical sciences, 660 children and adolescents and their parents were enrolled. the healthy students aged between 7 - 18 years were chosen from 4 girl and 4 boy - schools among primary and middle schools in isfahan by random cluster method. the 60 children and adolescents with chronic diseases were chosen from patients with malignancy, asthma, beta thalassemia, rheumatologic disorders, immune deficiency disorders and chronic hepatitis referred to private clinics of st. after taking participants consents and providing enough explanation by the project 's staffs, the child and self proxy reports questionnaires were distributed. however, 11 questionnaires were excluded, since they contained more than 50% missing data. five hundred and twenty six questionnaires were eligible and had children and parents proxy reports. out of 526 questionnaires, 239 forms were fully filled by children and 287 forms were completed by the parents. the distribution of the participants according to the age was as follows : (2 - 4 years ; n=30), (5 - 7 years ; n=43), (8 - 12 years ; n=253), (13 - 18 years ; n=180). the forward - backward translation method was applied on 23-items pedsql 4.0 generic core scales, figure 1. pedsql 4.0 generic core scale is a self - administered questionnaire contained 4 subscales including physical, emotional, social and school functions by which different aspects of hrqol are assessed. the child self report assesses children at ages 8 - 12 and 13 - 18 years. the parent proxy report evaluates children between 5 - 7 and 2 - 4 years in addition to the age groups assessed by the child forms. questions of both reports were scored as follows : 0=never a problem ; 1=almost never a problem ; 2=sometimes a problem ; 3=often a problem ; 4=almost always a problem. the items for each of the forms are essentially identical, differing in developmentally appropriate language, or first or third person tense. validation of the persian version of pedsql 4.0 generic core scale followed the recommended guideline, figure 1. process, decision and result according to the international guideline for questioner translation (forward - backward translation) two independent persian versions were made by a general physician fluent in english and an english translator. then, these two drafts were combined by a pediatrician and a community medicine physician. the product of translation process was reviewed and approved by the principal inventor (dr. varni). a pilot study aiming to determine face validity and provide final edition was conducted on 30 students and their parents. our aim was to find items leading to misunderstanding and to edit final version. ultimately, the persian version of pedsql 4.0 generic core scales was the output of the first phase. after obtaining isfahan education office license, a package consisted of a written consent form, a testimonial letter explained how to fill the questioners, contact addresses and phones of main researchers for any possible question or explanation, persian version of pedsql 4.0 parent proxy form and child form were distributed. the children were recruited from 4 girl - schools and 4 boy - schools among primary and middle schools in isfahan by random cluster method. the child forms were filled out by those children whose parents completed the parents proxy forms. sixty children with chronic diseases were recruited from patients referred to private clinics of st. alzahra hospital. the inclusion criteria for these children were the ability of patients and acceptance to fill out the forms. the following statistical methods were applied : the face validity of the persian version of the pedsql 4.0 generic core scale was evaluated by asking the children and their parents in a pilot study about the appearance of the text (its font and size) and the meaning of the sentences. the feasibility of the questionnaire was evaluated by determining the percentage of missing items for each questionnaire. ceiling and floor effects were determined to show whether more than 15% of the participants achieved the highest or lowest score, respectively. the content validity was determined by asking two pediatricians whether the items of the persian version of pedsql 4.0 assessed hrqol and any irrelative item in the inventory. the discriminant validity of the persian version of pedsql 4.0 was determined for 4 subclasses of the questionnaire between healthy children and children with chronic diseases. the intraclass correlation coefficient (icc) was measured to show the agreement between child self - report and parent proxy - report on the pedsql 4.0 subscales : iccs 0.4 as poor to fair agreement ; 0.41 - 0.60 as moderate agreement ; 0.61 - 0.8 as good agreement, and > 0.8 as excellent agreement. in addition, bartlett 's test of sphericity showed that assumption of sphericity was met. exploratory factor analysis (efa), operating principal component analysis and the rotation method (varimax with kaiser normalization) was used to extract factor structure of the pedsql 4.0. confirmatory factor analysis (cfa) was applied to evaluate the fitness of efa extraction model the observed data. the reasonable values of fit indices for cfa were considered as x / df 0.8 as excellent agreement. in addition, bartlett 's test of sphericity showed that assumption of sphericity was met. exploratory factor analysis (efa), operating principal component analysis and the rotation method (varimax with kaiser normalization) was used to extract factor structure of the pedsql 4.0. the extraction method (principal component analysis) and were utilized. confirmatory factor analysis (cfa) was applied to evaluate the fitness of efa extraction model the observed data. the reasonable values of fit indices for cfa were considered as x / df < 5 and root mean square error approximation (rmsea) < 0.08. furthermore, values more than 0.9 were accepted appropriate for comparative fit index (cfi), goodness of fit index (gfi) and adjusted goodness of fit index (agfi). kaiser - meyer - olkin shows the adequacy of sample size and bartlett 's test of sphericity indicates the assumption of sphericity was met five hundred and thirty seven participants out of 660 (81%) returned the questionnaires. eleven out of 537 questionnaires (2%) were omitted because of having more than 50% missing data. five hundred and twenty six questionnaires were eligible and had children and parents proxy reports. out of 526 questionnaires, 236 forms were full filled by children and 290 forms were completed by the parents. forty six percent of healthy children and 52% of children with chronic condition were female. sample characteristics : parent - proxy and child self reports according to health condition to evaluate face validity, 29 children (15 healthy children and 14 children with chronic diseases) and 25 parents (10 parents of healthy children and 15 parents of sick children) were enrolled in a pilot study. twelve healthy children (80%) out of 15 and 57.1% of children with chronic diseases found the meaning of the questions as the researchers expected. eleven healthy participants out of 15 (73.3%) and 71.4% of the children with chronic diseases replied that the appearances of the forms are attractive enough, p=0.99. the p value for the first and second questions between parents groups were 0.18 and 0.13, respectively. these results proved the acceptable face validity of the children self - report and parent proxy - report forms. the feasibility of the forms was evaluated by determining the percentage of missing data and ceiling and floor effects. in healthy children, questions number 18 and 11 with 8 missing, had the highest and questions number 2 and 10 with no missing, had the lowest missing data. total missing data in children self - report forms was 3.6% (7 items in healthy children and 1 item in diseased children ; totally less than 5%). the result was 2.2% in parents - proxy reports (5 items in healthy group and 1 item in diseased group ; totally less than 5%). we did not show any floor effects while ceiling effects were observed from a minimum of 4% in child self - report social functioning to a maximum of 10% in parent proxy - report physical functioning [table 3 ]. the cronbach 's alpha for total scale scores of child - self and parent - proxy reports by applying ceiling and floor effects were 0.73 and 0.9, respectively. the intraclass correlation coefficients (icc) were high for most scales indicating excellent agreement for social functioning and school functioning, good agreement for total score and physical functioning. nonetheless the lowest agreement was identified for emotional functioning scale (icc=0.32, 95% ci : 0.2 - 0.4). item - subscale, item - total and subscale - total score correlations were significant for all items [table 4 ]. floor and ceiling effects of child - self and parent proxy reports item - subscale, item - total, and subscale - total score correlations factor analysis with varimax rotation extracted 7 factors from the pedsql 4.0 self- and parent - proxy reports are shown in table 5. the results of the cfa for 7-factor models showed appropriate fitness of the model for self- and parent - proxy reports [table 6 ]. factor analysis results for child - self reports and parent proxy reports the result of confirmatory factor analysis for the 7 factors model to evaluate discriminant validity, the mean score of each function was compared between two groups separately [table 7 ]. the score for each function was significantly lower in healthy children comparing with children with chronic diseases. in addition, the mean total score for healthy children and children with chronic diseases were 114.1213.48 and 121.625.5 ; respectively, p=0.001. therefore, the pedsql 4.0 persian version has the acceptable discriminant validity for each function and for total score. discriminant validity of the pedsql 4.0 according to health status to evaluate content validity, a pediatrician and a pediatric nephrologist were asked if the items of persian version of pedsql 4.0 assessed hrqol and if there was any irrelative item in the inventory. an appropriate content validity was achieved by kappa analysis that showed 95% and 91% agreement between two physicians for parent - proxy and child - self reports, respectively. spearman coefficient values for parent proxy reports for children 2 - 4 years and 5 - 7 years were 0.3 and 0.73, respectively. the results did not support criterion validity of parent - proxy forms for children 2 - 4 years. the values for children self- and parent - proxy forms of children 8 - 12 years and adolescents 13 - 18 years were more than 0.7. covariance matrices between each function and other functions were less than 0.7 (the least was between social and school functions=0.39 and the greatest value was between physical and social functions=0.58). five hundred and thirty seven participants out of 660 (81%) returned the questionnaires. eleven out of 537 questionnaires (2%) were omitted because of having more than 50% missing data. five hundred and twenty six questionnaires were eligible and had children and parents proxy reports. out of 526 questionnaires, 236 forms were full filled by children and 290 forms were completed by the parents. forty six percent of healthy children and 52% of children with chronic condition were female. sample characteristics : parent - proxy and child self reports according to health condition to evaluate face validity, 29 children (15 healthy children and 14 children with chronic diseases) and 25 parents (10 parents of healthy children and 15 parents of sick children) were enrolled in a pilot study. twelve healthy children (80%) out of 15 and 57.1% of children with chronic diseases found the meaning of the questions as the researchers expected. eleven healthy participants out of 15 (73.3%) and 71.4% of the children with chronic diseases replied that the appearances of the forms are attractive enough, p=0.99. the p value for the first and second questions between parents groups were 0.18 and 0.13, respectively. these results proved the acceptable face validity of the children self - report and parent proxy - report forms. the feasibility of the forms was evaluated by determining the percentage of missing data and ceiling and floor effects. in healthy children, questions number 18 and 11 with 8 missing, had the highest and questions number 2 and 10 with no missing, had the lowest missing data. total missing data in children self - report forms was 3.6% (7 items in healthy children and 1 item in diseased children ; totally less than 5%). the result was 2.2% in parents - proxy reports (5 items in healthy group and 1 item in diseased group ; totally less than 5%). we did not show any floor effects while ceiling effects were observed from a minimum of 4% in child self - report social functioning to a maximum of 10% in parent proxy - report physical functioning [table 3 ]. the cronbach 's alpha for total scale scores of child - self and parent - proxy reports by applying ceiling and floor effects were 0.73 and 0.9, respectively. the intraclass correlation coefficients (icc) were high for most scales indicating excellent agreement for social functioning and school functioning, good agreement for total score and physical functioning. nonetheless the lowest agreement was identified for emotional functioning scale (icc=0.32, 95% ci : 0.2 - 0.4). item - subscale, item - total and subscale - total score correlations were significant for all items [table 4 ]. floor and ceiling effects of child - self and parent proxy reports item - subscale, item - total, and subscale - total score correlations factor analysis with varimax rotation extracted 7 factors from the pedsql 4.0 self- and parent - proxy reports are shown in table 5. the results of the cfa for 7-factor models showed appropriate fitness of the model for self- and parent - proxy reports [table 6 ]. factor analysis results for child - self reports and parent proxy reports the result of confirmatory factor analysis for the 7 factors model to evaluate discriminant validity, the mean score of each function was compared between two groups separately [table 7 ]. the score for each function was significantly lower in healthy children comparing with children with chronic diseases. in addition, the mean total score for healthy children and children with chronic diseases were 114.1213.48 and 121.625.5 ; respectively, p=0.001. therefore, the pedsql 4.0 persian version has the acceptable discriminant validity for each function and for total score. discriminant validity of the pedsql 4.0 according to health status to evaluate content validity, a pediatrician and a pediatric nephrologist were asked if the items of persian version of pedsql 4.0 assessed hrqol and if there was any irrelative item in the inventory. an appropriate content validity was achieved by kappa analysis that showed 95% and 91% agreement between two physicians for parent - proxy and child - self reports, respectively. spearman coefficient values for parent proxy reports for children 2 - 4 years and 5 - 7 years were 0.3 and 0.73, respectively. the results did not support criterion validity of parent - proxy forms for children 2 - 4 years. the values for children self- and parent - proxy forms of children 8 - 12 years and adolescents 13 - 18 years were more than 0.7. covariance matrices between each function and other functions were less than 0.7 (the least was between social and school functions=0.39 and the greatest value was between physical and social functions=0.58). the present study is the first to explain and validate the different aspects and properties of the persian version of pedsql 4.0 generic core scale in children. the results of the study provided enough primary supports for reliability, validity and feasibility of persian version of pedsql 4.0 in toddlers, children and early adolescents self- reports and parent proxy - reports. our results provided enough evidences to support validity, feasibility and reliability of the persian version of pedsql 4.0 generic core scale. at the time of approving the final edition of the manuscript, we realized that another iranian research team has obtained the approval of varni to translate pedsql 4.0 into persian (adolescent 13 - 18 years). however, an important contribution of our paper compared to their work is that, we enrolled a greater sample of chronically ill patients. in addition, the participants of our study were chosen by cluster sampling from different divisions of the city (isfahan). another major contribution of this paper is the fact that, we have studied toddlers and children. we confirmed the face validity of the persian version of pedsql 4.0. the feasibility of our study was approved by missing data analysis (3.6% of children self and 2.2% of parent proxy reports). roizen. demonstrated suitable feasibility by only 9 missing data in 5 - 7 years old range. study spent 5 - 6 min to complete the forms comparing with an average of 5 min in the brazilian study and 5 - 7 min for our participants. the main inventor (dr. varni) achieved cronbach 's alpha=0.89 for child and 0.92 for parent proxy reports with minimal missing data. in our study, the cronbach 's alpha for total scale scores of child - self and parent - proxy reports by applying ceiling and floor effects were 0.73 and 0.9 respectively, the lowest agreement was identified for emotional functioning scale (icc=0.32, 95% ci : 0.2 - 0.4). reported cronbach 's alpha between 0.6 and 0.9 for all dimensions, showing appropriate internal consistency. demonstrated that the finnish version had excellent cronbachs alpha values for the total scale score (0.91 and 0.88 for child self and parent proxy report, respectively). construct validity was assessed using exploratory factor analysis and by exploring the inter - correlations between and among the 4 pedsql subscales for adolescents and their parents. the reliability coefficient for the persian questionnaire was comparable with other studies[6715192227 ] and more than argentinean - spanish version. however, persian version of pedsql 4.0 did not show acceptable reliability for assessing healthy 8 - 12 years old children. it might be due to the fact that children aged 8 - 12 years face physical and psychological aspects of puberty. therefore, more studies are required to assess discriminant validity of each aspect of hrqol. similar to the argentinean - spanish version, the persian version did not show acceptable reliability for 2 - 4 years old toddlers proxy - report. this can be associated to the following reasons : poor communication skills of toddlers results in poor assessment of children for parents.none of school items were completed for toddlers. poor communication skills of toddlers results in poor assessment of children for parents. none of school items were completed for toddlers. other studies also confirm this observation.[67151922 ] probably because physical function is more objective than other items and it is easier to assess. however, utilizing hrqol measurement in pediatric healthcare settings facilitates patient - physician communication, improves patient / parent satisfaction, identifies hidden morbidities, and assists in clinical decision - making. we confirmed that persian version of pedsql 4.0 generic core scales is valid and reliable. firstly, we enrolled a greater sample of chronically ill patients compared to similar studies. secondly, we are the first to validate pedsql 4.0 on iranian toddlers and children. however, another study is yet to be conducted to address cultural specifications of iranian society, especially for pubertal age. we also suggest another study with larger sample size for toddlers while omitting school items. | introduction : to evaluate the reliability, validity and feasibility of the persian version of the pediatric quality of life inventory (pedsql 4.0 4.0) generic core scales in iranian healthy students ages 7 - 15 and chronically ill children ages 2-18.methods:we followed the translation methodology proposed by developer to validate persian version of pedsql 4.0 4.0 generic core scales for children. six hundred and sixty children and adolescents and their parents were enrolled. sample of 160 healthy students were chosen by random cluster method between 4 regions of isfahan education offices and 60 chronically ill children were recruited from st. alzahra hospital private clinics. the questionnaires were fulfilled by the participants.results:the persian version of pedsql 4.0 4.0 generic core scales discriminated between healthy and chronically ill children (healthy students mean score was 12.3 better than chronically ill children, p<0.001). cronbachs alpha internal consistency values exceeded 0.7 for children self reports and proxy reports of children 5 - 7 years old and 13 - 18 years old. reliability of proxy reports for 2 - 4 years old was much lower than 0.7. although, proxy reports for chronically ill children 8 - 12 years old was more than 0.7, these reports for healthy children with same age group was slightly lower than 0.7. constructive, criterion face and content validity were acceptable. in addition, the persian version of pedsql 4.0 4.0 generic core scales was feasible and easy to complete.conclusion:results showed that persian version of pedsql 4.0 4.0 generic core scales is valid and acceptable for pediatric health researches. it is necessary to alternate scoring for 2 - 4 years old questionnaire and to find a way to increase reliability for healthy children aged 8 - 12 years especially, according to iranian culture. |
the use of female human resources is highlighted as an important factor in the aging korean society, especially as married women have started entering the labour market. in addition, a wide range of fields utilize female labour and the distribution ratio of women in each field is increasing, with female participation in social and economic activities reaching 50.2% in 2013, 51.3% in 2014 and 51.8% in 20151. office jobs in particular are experiencing a greater influx of female resources than any other area, accompanied by the risk of various diseases due to poor working posture and environments, long - term use of computers, task over - load, mental stress, etc2. the stress that female office workers experience in the male - centred organizational culture in korea seems to have more significant implications3. in general, moderate stress plays a positive role in physical and mental vitality while achievement motivation elicits feelings of tension. however, undue stress caused by task over - load might have negative effects on the mind and body, for example, personal anxiety, reduced concentration, loss of drive to work, stress - related diseases, etc. it might also have direct or indirect negative effects on organizational performance, for example, occurrence of negligent accidents, expansion of healthcare expenses, a decrease in productivity and loss of employment4. thus, a stress therapy program with easy applicability during busy business hours can improve the health of female office workers. one of the easily applicable stress therapy options, in terms of time and space, is massage. not only does it have therapeutic effects on various bodily functions including the nervous, musculoskeletal, cardiovascular and integumentary systems, it is also recognized as a type of aerohydropathy as it helps to promote local and general circulation, increases immunologic function, optimizes natural healing ability and helps maintain body balance and harmony. massage is the oldest method used to resolve tiredness in the mind and body, and can treat and prevent illness5. massage on the neck muscles extends the range of cervical spine rotation7 and improves neck flexor muscles8. in addition, massage after total knee arthroplasty decreases pain and increases the diarthrodial range9. in the current study, in addition to the extension of diarthrodial range, it influences stress hormones (epinephrine and norepinephrine), which can trigger the activation of sympathetic nerves such as an increase in heart rate, contraction of blood vessels and muscle tension, as well as circulation throughout the body via blood10. thus, massage has been confirmed to have positive effects on changes in hormones11, 12, blood pressure13, 14 and heart rate15. this study was performed to provide a theoretical rationale for the application of massage as stress therapy in female office workers. with the goal of improving health and quality of life, this study is based on the analysis of the effects of massage on stress hormones, including blood pressure and heart rate. the target population of this study was female office workers ; 2049 year old female office workers at 3 institutions located in city g were randomly recruited as an accessible population. 11 were placed in experimental (exp) group (i), 11 in experimental (exp) group (ii) and 12 in the control group ; their physical features are provided in table 1table 1.physical characteristics of the subjectsage (years)height (cm)weight (kg)exp (i) (n=11)42.6 2.0158.1 3.159.2 4.5exp (ii) (n=11)43.1 1.8158.1 2.359.1 1.2cont (n=12)42.6 1.7158.0 1.258.9 1.2exp : experimental group, cont : control group. exp : experimental group, cont : control group this study was approved by the ethics committee at dongguk university of korea and written consent to participate in the study was obtained from all of the subjects in accordance with the declaration of helsinki. scalp massage was performed for 15 minutes / session for exp (i) and 25 minutes / session for exp (ii), twice a week, for a total of 20 times over 10 weeks in both groups. the scalp massage room was maintained at a humidity of 4060% and a temperature of 2627c. the investigator trained one sub - investigator with scalp care experience on the necessity and purpose of this study in order to maintain the accuracy and validity of experimental procedures. the sub - investigator performed the scalp massage under the supervision of the investigator. details of the program are presented in table 2table 2.scalp massage program for 10 weeksweeksordertimecontentsfrequencyexp (i)exp (ii)110warm - up3 min2 minstretching before massagemain program19 min11 min. massage in shoulder (3 times)2 times /week compression by a thumb compression by a wrist grasp and squeeze by a finger champi compression under down [ed : unclear] massage in neck grasp and pull down in the muscle of neck massage under the larynx massage in the occiput massage in a zig - zag wave exercise flip by a finger effeurage by a finger tapotement by a finger pull up a hair grasp up by head rolling and massage a circle in the temples of one s head massage in a ear massage a hole in a facecool - down3 min2 minstretching after massage. blood collection to measure the levels of stress hormones (epinephrine, norepinephrine and cortisol) was performed in the subjects following at least 12 hours of fasting. the collected blood was centrifuged for 10 minutes at 3,000 rpm using a centrifuge (model hc-16a, korea) ; the post - centrifugal plasma was stored at 73 c and referred to laboratory n for hemanalysis. blood collection was conducted before the first scalp massage and at 9 am the following day. blood pressure was measured before the first scalp massage and after 10 weeks, for a total of two times, using a mercury sphygmomanometer (omron hem-770a, japan). the mean value of the 2 measurements was used as the average blood pressure. the subjects heart rates were also measured before the program and after 10 weeks at a stable status using the sport tester pe 3000 (polar electro co. finland) ; each measurement was conducted for 5 minutes and the mean value of the 2 measurements was used as the heart rate. the primary effect and interaction effect on the time interval and between groups were evaluated using a two - way repeated anova, and a post - hoc test was conducted using the duncan test. a scalp massage was performed for 15 minutes / session for exp (i) and 25 minutes / session for exp (ii), for 10 weeks. there was no significant difference in the time interval and interactions between groups for epinephrine (p>0.05). however, there were significant differences in the time interval and interactions between groups for norepinephrine and cortisol. in an assay of primary effects caused by the time interval, significant differences were found among epinephrine, norepinephrine and cortisol, leading to a meaningful difference in exp (ii) for epinephrine in the post - hoc test as well as in exp (i) and exp (ii) for norepinephrine and cortisol with prior and post - hoc tests. significant differences between groups were also found for norepinephrine and cortisol, leading to meaningful differences between exp (i) and the control group and between exp (ii) and the control group for both norepinephrine and cortisol (table 3table 3.the influence of stress hormones according to group by 10 weeks, before and aftervariablespre massagepost massageexp (i)exp (ii)contexp (i)exp (ii)contepinephrine263.8 63.1254.0 7.9262.9 60.1212.4 37.0222.6 33.1254.1 35.4norepinephrine751.6 92.5680.5 110.2739.1 117.8450.0 127.9499.5 126.2655.1 189.3cortisol23.4 2.724.2 3.824.6 2.916.3 2.015.3 2.223.9 1.8values are expressed as mean sd, p0.05). additionally, no significant difference in the primary effect caused by time intervals and between groups was found (p>0.05) (table 4). there are two different neuroendocrine systems associated with stress : one is the sympathetic system - adrenal medulla axis involved in the secretion of epinephrine and norepinephrine and the other is the hypothalamus - pituitary - adrenal medulla (hpa) axis involved in the secretion of cortisol16. the physiological effect of massage in general can be divided into a relaxation effect and a stimulation effect. the relaxation effect involves hypothalamic reactions associated with the decline of sympathetic system activity and an increase in parasympathetic system activity. there are two types of stimulation effects : one is reflexive and the other is mechanical. the reflex effect is refreshing and relaxing due to delivering stimulation at the cutaneous peripheral nerve to the cerebrum. peripheral cutaneous stimulation promotes circulation through stimulation of the parasympathetic nerve, relaxation of muscles and extension of capillary vessels. ultimately, massage reduces sympathetic nerve activity while increasing parasympathetic nerve activity17. in this study, 15-minute and 25-minute scalp massages had a significant effect on norepinephrine and cortisol while the 25-minute scalp massage had a significant effect on epinephrine. this suggests that a scalp massage decreases the activation of the sympathetic nerve while increasing the activation of the parasympathetic nerve, resulting in a decrease in the secretion of norepinephrine and cortisol, or in other words, stabilization of hormone levels. however, only in the case of the 25-minute scalp massage was there a significant difference in the level of epinephrine. this seems to be caused by either personal sensitivity differences and time variance18 or as a result of personal differences between subjects and time variance. in addition, the study demonstrated that the levels of epinephrine were unpredictable and could increase in uncontrolled noise environments19. a previous study demonstrated that the low frequency (lf) sympathetic nerve activity marker significantly decreased whereas the high frequency (hf) parasympathetic nerve activity marker significantly increased after 15 minutes of scalp massage20. another study demonstrated the effects of a 20 minute scalp massage on norepinephrine and cortisol21. the results of our study were in agreement with these existing studies, suggesting that a>15 minute scalp massage has a positive effect on stress hormones. the results also suggested that stress increases cardiac output and blood pressure by influencing the sympathetic nerve, resulting in the increase of peripheral blood pressure by epinephrine, leading to the relaxation of blood vessels and increased blood flow rate in the liver, heart muscles and brain tissues. in addition, norepinephrine is known to cause vasoconstriction, increase blood pressure, general peripheral resistance22, and pulse due to rising blood cortisol concentration levels during mental stress. there was a significant decrease in both systolic and diastolic blood pressure after 15 and 25 minute scalp massages, which seems to be a result of the stabilization of hormone levels caused by an increase in the activation of sympathetic nerves and parasympathetic nerves and the subsequent decrease in the concentration of norepinephrine and cortisol hormone. this is probably because the scalp massage promoted blood circulation, resulting in a positive effect on the neck muscles and the relaxation of blood vessels7, 8. after performing scalp massage in adults at the normotensive or prehypertensive stage additionally, 4 weeks of massage at work was shown to significantly decrease strain and blood pressure14. under the control of the autonomic nervous system, the heart rate increases when the reaction of the sympathetic nerve system is accentuated. in this study, the heart rate did not seem to decrease significantly despite a decrease in the level of norepinephrine and cortisol hormones controlling the sympathetic nerve system. however, the heart rate showed a tendency to drop slightly. in a previous study involving carotid sinus massage in aged subjects, blood pressure and heart rate were shown to decrease24 and a significant decrease in heart rate was confirmed after 4560 minutes of muscle massage in middle aged men and female performed at different intensities25. in a study using massage (at moderate and mild intensity) and a vibrator, massage treatment at moderate intensity was shown to significantly reduce the heart rate26. these differences from the existing study might be because massage in this study was not performed frequently (twice a week) and the intensity was relatively mild. in general, only the heart rate did not show a significant difference and the scalp massage had positive effects in terms of hormones and blood pressure. we recommend active utilization of a scalp massage program with no spatial or time limits in female office workers, as the use of a scalp massage lasting 15 minutes or less as a stress control program could have a positive effect. furthermore, better results are expected in future studies if the intensity of the massage is varied while considering the physical condition of the subjects. | [purpose ] a scalp massage was conducted on female office workers divided into a 15 minute group and 25 minute group and its effect on stress hormone, blood pressure and heart rate was analyzed in order to provide a theoretical rationale to apply scalp massage as stress therapy. [subjects and methods ] a scalp massage was applied to 34 female office workers twice a week for a total of 10 weeks ; the subjects were classified into 15 min., 25 min. and control groups, and their stress hormone levels, blood pressure and heart rate were evaluated. [results ] significant differences in norepinephrine, cortisol and blood pressure (sbp & dbp) were found in terms of interaction by time interval and between groups. [conclusion ] as a result of applying scalp massage to female office workers for 15 and 25 minutes, positive effects were observed on stress hormone, blood pressure and heart rate. therefore, scalp massage can be used for stress control with no spatial or time limit. |
chitin is plentiful in the structural coatings of fungi, insects, and parasitic nematodes, but it is not produced in mammals. chitin, the linear polymer of -1,4 linked -n - acetyl - glucosamine (glcnac), is the most abundant biopolymer in marine ecosystems and, after cellulose, the second most copious polysaccharide in nature. since chitin seems not to be made in mammals, it was initially assumed that chitinases would also be absent, being restricted to species that do contain the polymer. however, the host defense against chitin - containing pathogens includes the production of chitinases. chitinases are endo--1,4 linked -n - acetyl - glucosaminidase, and recently, two distinct chitinases have been identified in humans, chitotriosidase (chit) expressed in phagocytes, and an acidic mammalian chitinase (amcase) expressed in gastrointestinal tract, in lung, and conjunctiva. amcase, 50 kda protein, is a member of the glycosyl hydrolase 18 family (ec 3.2.1.14) and has structural similarity (figure 1) with other glycosyl hydrolase 18 family members, including the lectins ym1 and ym2, which are expressed in macrophages, human cartilage glycoprotein-39 (hc - gp39), and chitotriosidase (muzzarelli, 2008). chit and amcase show chitinase enzymatic activity, whereas other mammalian chitinases do not possess this activity. characteristically, the resistance to acidic ph distinguishes amcase from chit, which has its optimum at ph 6. on the contrary, amcase has its optimum at ph 2 and 4 in rodents and humans, respectively ; amcase is expressed in different tissues such as stomach, lung, and salivary glands, and appears to be associated with inflammatory diseases (chou., 2006). chit, has been shown to be expressed together with lysozyme in the human lacrimal gland (hall., 2008). this chitinase produced by macrophages and neutrophils also has a role in innate immunity (van eijk., 2005). since this chitinase was considered more active in the control of chitin - containing pathogens, the presence of measurable chit activity in tears appears to support the protection of the eye in both humans and mice, where the chitotriosidase (chit) gene is evolutionarily conserved (gianfrancesco and musumeci, 2004). in this context, it is interesting to note that chit, unlike bacterial chitinases, does not appear to have any mucolytic activity (sanders., 2007), and therefore the lacrimal film is not modified in its structure, maintaining the integrity of visual function. moreover, hall. (2008) found that recombinant chit does not inhibit bacterial growth and does not synergize with lysozyme on the growth of gram - positive and gram - negative bacteria, and the antimicrobial activity appears to be limited only to fungi. amcase appears to play a key role as first responder of the immune system (ramanathan., 2006), both in the epithelial cells lining the nasal mucosa and other contiguous conjunctival cells. when the epithelial cells are stimulated against non - existent parasites through a mechanism that induces an il-13 response, they can produce chitinases, which represent a marker of both innate immunity and inflammatory response. in addition to the role in innate immune response, the modulation of amcase expression by specific inhibitors suggests a new pathway for controlling ocular inflammation. this article seeks to provide a succinct review of ocular inflammation, a rapidly changing area of visual science, focusing on the role of chitinase. one of the most important issues in chitinase biology relates to our almost complete lack of understanding of the functions of these strongly conserved, and therefore presumably biologically important, enzymes in human subjects. chitinases have been studied most intensely in lower life forms, where they are produced in significant quantities by hosts defending against infections with chitin - containing organisms. this can be appropriately considered part of the innate immune response against a chitin - containing pathogen. amcase contains a 30-kda n - terminal catalytic domain that can hydrolyse chitin, and it has recently been identified both in human gastrointestinal tract and lung. chitinase production can also play a key role in the life cycle of chitin - containing fungi and parasites, where they control growth and molting, and can be used by pathogens to invade or exploit chitin - containing structures to complete their lifecycle in the host. amcase represents a product of genes on mouse chromosome3 and human chromosome 1 at p13.1p21.3. the enzyme is acid stable, with a ph optimum of 2.0, and its appears to be adapted to function in the extreme environment of the stomach where it may play a role in defense and/or digestion of chitin - containing organisms. 2011) have demonstrated that amcase is active with appropriate ph profile showing that its activity at ph 2.0 was fourfold the activity obtained at higher ph (5.2 and 7.5). this enzyme might also have non - chitin - related biologic effector properties because an amcase - like protein has been reported to possess fibroblast growth - promoting activity (guoping., 1997), and amcase has been implicated in host defenses and food processing (muzzarelli, 2008). amcase is induced at sites of inflammation (e.g., parasitic infections) and remodeling. this raises the possibility that this enzyme plays active roles in human antiparasite and antiinfective defense and repair responses. probably amcase plays a role as sentinel that trigger responses to parasites, infections, and/or antigen challenge, acting directly as chemotactic agents or indirectly by inducing other chemokines that attract eosinophils and t cells to sites of parasitic infection, in other words amcase maybe modulate tissue inflammation, immunity, and remodeling (zhu., 2004). a role for amcase in asthma pathophysiology has been suggested following the demonstration that amcase expression is increased in the lungs of ovalbumin sensitized and challenged mice that develop airway hyperresponsiveness, compared with control animals challenged with phosphate - buffered saline (zhu., 2004). in this animal model of asthma, amcase is expressed in both airway epithelial cells and alveolar macrophages. elevated expression of amcase was also observed in lung tissue from asthmatic patients when compared with normal subjects (zhu., 2004). amcase is produced in human epithelial cells of lower airways and conjunctiva via a th2-specific, il-13-dependent pathway and seems to be associated with asthma and allergic ocular pathologies. amcase has also been proposed as a potential therapeutic target in th2-mediated inflammation (donnelly and barnes, 2004). in fact, the inhibition of amcase activity by the chitinase inhibitor allosamidin decreased the number of inflammatory cells in the bronchoalveolar lavage fluid of ovalbumin sensitized and challenged mice, and reduced asthma symptoms (zhu., 2004. interestingly, the same response was obtained using antisera against amcase given in the airways by aerosol (zhu., 2004). the understanding of the role of amcase in allergic disease is only at its beginning and many issues open new possibilities for its control using specific inhibitors of amcase activity or modulating its expression. in patients with vernal keratoconjunctivitis (vkc) and with seasonal allergic conjunctivitis (sac) the level of amcase activity in the tears was found significantly elevated when compare to healthy controls and the highest levels were found in vkc (musumeci., 2008). when rna was extracted by conjunctival epithelial cells of these patients, quantitative real time pcr measurement confirmed that mrna expression correlates with tear amcase activity and the expression was significantly higher in vkc and sac. also receiver operating characteristic (roc) analysis demonstrated that the sensitivity and specificity of amcase measurement were 100%, addressing the use of amcase assay in the biochemical diagnosis of vkc and sac (musumeci., 2008).recent studies in rabbits, where a uveitis was induced by lps injection into the eye s anterior chamber, confirmed that increased amcase activity was measurable in tears and that epithelial cells of conjunctiva express specific mrna (bucolo., 2008).further, it was previously demonstrated in experimental model of mouse asthma, the inflammatory reaction induced by lps was controlled by the chitinase inhibitor and glucocorticoids, instilled at 3 h interval in conjunctival sac. in dry eye syndrome, another non - allergic ocular pathology, an increased amcase activity was documented and the specific mrna expressed by epithelial conjunctival cells (musumeci., 2009). in this pathology the eye inflammation can be ascribed to a common mechanism mediated by amcase, via a th2-specific, il-13 dependent way. in summary, amcase may be considered an important mediator in the pathogenesis of th2 inflammation eye s diseases, suggesting its potential diagnostic and therapeutic utility. inhibition of amcase, decreasing the inflammation in the mice that overexpressed il-13, suggests a loop in the modulatory effect of amcase in allergic asthma as well in allergic conjunctivitis. in fact, amcase enzyme inhibition with allosamidin does not alter the level of expression of il-13 receptors, but inhibits il-13 from stimulating the expression of several chemotactic factors, typical of allergic inflammation and asthma (romagnani, 2002 ; zhu., 2002). we showed that, in endotoxin - induced uveitis in rabbit, the production of amcase in tears was modulated by glucocorticoids treatment and the inhibition of activity by allosamidin, a chitinase inhibitor, caused an effective reduction of the inflammation clinical score (bucolo., 2008). these data are in accordance with the findings generated from other laboratories, homer. (2006) observed that expression of amcase is upregulated in response to allergen exposure or il-13-induced inflammation in mouse lung and allosamidin strongly inhibits amcase. 2009) showed that allosamidin and demethyl allosamidin, showed a potent anti - inflammatory activity in allergic mouse models, even though the mechanism of action remains unclear. as mentioned before, we found (musumeci., 2008) an overexpression of amcase mrna in epithelial conjunctival cells during vkc and sac, suggesting that the activity of amcase could be used as a differentiate parameter for diagnosis (amcase activity higher in vkc than in sac). in these studies, it was clear that amcase did not directly induce a th2 cytokine response, but mediated or modulated the effector response of il-13 (a cytokine produced by th2 cells).moreover, it seems that amcase activity is required for the increased production of the chemoattractants of the inflammatory cells such as monocyte chemoattractant proteins (mcps) and eotaxins. from these considerations, it seems evident that the expression of amcase and the th2-induced inflammation are driven by a sequential mechanism that is attenuated by inhibitors of chitinase (figure 2). increases in acidic mammalian chitinase levels are a potentially important downstream effect of il-13 stimulation in th2-oriented immune responses to pathogens and parasites amcase seems to be a key mediator of il-13-induced responses in th2-driven immune - inflammatory eye diseases with a lofty implication in ocular allergy conditions as well as in dry eye syndrome. however, the immuno - regulatory function of amcase is not completely clear so far, therefore translational research may be warranted in order to elucidate that. the authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest. | chitinases have an important role in the defense of organisms against chitin - containing parasites. an acidic mammalian chitinase (amcase) has been detected in epithelial cells in lung tissue samples taken from patients with asthma as well as in conjunctival epithelium of patients with inflammatory ocular diseases. particularly, elevated amcase activity has been observed in ocular tissues of patients with vernal keratoconjunctivitis, seasonal allergic conjunctivitis, and in patients affected by dry eye syndrome. this enzyme is induced via a th2-specific, il-13-dependent pathway. amcase may thus be a key mediator of il-13-induced responses in th2-driven inflammatory ocular diseases. |
small cell lung cancer (sclc) is highly malignant neoplasm, derived from neuroendocrine cells. it represents approximately 15% of all bronchial carcinomas, and this percentage is tending to decrease recently. in most cases, sclc arises in the larger airways and grows rapidly, becoming quite large. it also has propensity to metastasize widely throughout the body at an early stage in its clinical course. tonsillar metastasis from sclc is extremely rare, and clinically apparent cases are even less common. idiopathic pulmonary fibrosis (ipf) is a chronic, progressive form of interstitial lung disease with poor prognosis and is a clinical term of usual interstitial pneumonia of unknown cause. it has been reported to be associated with increased risk of lung cancer. in a study, the incidence of lung cancer was increased 7-fold in the ipf group compared with healthy subjects. although the features of the lung cancer with ipf are similar to the general features of lung cancer, sclc is not common in fibrotic area of ipf. the present case report describes sclc in ipf - associated lesion and its tonsillar metastasis, which is rarely seen. a 77-year - old man admitted to our hospital with 1-month history of cough and dyspnea. he had personal history of pulmonary tuberculosis 1 year ago. on physical examination of the thorax, inspiratory dry crackles were heard on both lower lung fields. in observation of oral cavity, a large oval mass composed of soft tissue was detected in his throat. the mass was arising from the right palatine tonsil and extending across the midline of the oropharynx (figure 1a). (a) in physical examination, a large oval mass composed of soft tissue arose from the right palatine tonsil and extended across the midline of the oropharynx. (b) whole - body magnetic resonance revealed intraluminal protruding mass in the right peritonsillar region with heterogeneous enhancement, suggesting malignancy of palatine tonsil. high - resolution computed tomography of chest showed 2 masses in the left lower lobe, 1 mass in the right upper lobe, and multiple enlarged mediastinal lymph nodes of the lung (figure 2). one of the left lower lobe masses was 4.4 4.0 cm sized in superior and lateral segments, and the other was 5.7 3.7 cm sized with fibrosis in subpleural region. also, there was typical honeycomb appearance with traction bronchiectasis and ground - glass opacity pattern, predominantly in subpleural areas of both lower lobes. under suspicion of lung cancer and usual interstitial pneumonia that is pathological equivalent to ipf, further workup was started to confirm the diagnosis. high - resolution computed tomography showed 1 mass in the right upper lobe (a), 2 masses in the left lower lobe (b), and honeycomb appearance in subpleural area of both lower lobe (c). percutaneous transthoracic needle biopsy for lung mass and punch biopsy for tonsillar lesion were performed. both tumors were composed of nests of small, round, or oval cells with little cytoplasm and hyperchromatic nuclei. the cells from both tumors were positive for cd56, a glycoprotein expressed on the surface of neurons and neuroendocrine tumors as well known that it is the neural cell adhesion molecule (ncam). in addition, the cells showed positive staining for synaptophysin and chromogranin a, although the intensity was weaker than cd56, and it was more distinct in lung mass than that in tonsillar mass. based on these pathologic findings and the known fact that sclc belongs to the neuroendocrine lineage of lung cancer, the masses were diagnosed as sclc and tonsillar metastasis (figure 3). representative h&e sections of lung mass (a) and tonsillar mass (b) revealed nests of small, round, or oval cells with little cytoplasm and hyperchromatic nuclei in both lesions. in immunohistochemical staining for cd 56 of lung mass (c) and tonsillar mass (d), cd = cluster of differentiation, sclc = small cell lung cancer. we performed systemic evaluation using whole - body magnetic resonance imaging, which showed a mass indicating brain metastasis in body portion of the right corpus callosum and a 2 2.8 cm - sized, intraluminal protruding mass in the right peritonsillar region with heterogeneous enhancement, suggesting malignancy of palatine tonsil (figure 1b). we suspected that the patient has ipf based on the clinical findings and radiological patterns. there was no history of exposure to any toxic materials and no clinical symptoms of connective tissue diseases. bronchioloalveolar lavage fluid analysis exhibited the percentage of alveolar macrophage, lymphocytes, neutrophil, and eosinophil was 77%, 3%, 15%, and 5%, respectively. chemotherapy with irinotecan and carboplatin for sclc and a standard medication of steroid and acetylcysteine for ipf were applied. however, in following - up, he expired due to respiratory failure by an acute exacerbation of ipf 3 months after the diagnosis. susceptible organs include the liver, abdominal lymph node, bone, brain, adrenal gland, skin, kidney, and pancreas. a few cases of tonsillar metastasis of sclc have been reported. the palatine tonsil is a rare site in which metastatic tumor deposit is found. according to a study, only 12 tumors (0.8%) the metastatic tumors include carcinoma of the breast, stomach, renal cell carcinoma, seminoma, melanoma, and carcinoma of rectum. furthermore, in a review of 76 cases of primary neoplasm complicated by tonsillar metastasis, only 12 were found to be due to carcinoma of the bronchus. among these 12 metastatic cases, evidence for the metastasis to other tissues was found in 10 of the 12 cases. in all reported cases of tonsillar metastasis of sclc, the metastasis developed following presentations : the mean interval of time between development of the primary bronchogenic cancer and the tonsillar metastasis was 8 months, and the mean time interval between appearance of the tonsillar metastasis and death was 5 months. in our present case, most patients with tonsillar metastasis are symptomatic, such as difficulty in breathing, sore throat, irritable cough, dysphagia, otalgia, and swallowing pain accompanied by a foreign body - like sensation. however, in our case, the patient with tonsillar metastasis of sclc did not show any of these symptoms. ipf, which is the most frequent type of interstitial lung diseases, has been reported as an independent risk factor for lung cancer by epidemiological studies. recent studies have reported that alteration of genes like fragile histidine triad (fhit) gene is associated with lung cancer and ipf, supporting that lung cancer can be a result of the occurrence of atypical or dysplastic epithelial changes in fibrosis. ipf is significantly related to the development of lung cancer in peripheral location of the lower lobes. adenocarcinoma and squamous cell carcinoma are the most common histological findings among lung cancer patients with ipf. however, sclc, which is a central disease, shows exceptional tendency to occur in the ipf - nonassociated and nonfibrotic lesion. our case is interesting in the point that sclc has developed from ipf - associated fibrotic lesion of left lower lobe. considering our case presented here, physicians should include metastatic sclc as the differential diagnosis for a single tonsillar mass, although the incidence is very low. in addition, this case shows that sclc can be developed in ipf - associated fibrotic lesion, indicating that physicians should consider a possible association between sclc and lung fibrosis such as ipf. institutional review board of chonbuk national university hospital has stated that it is not necessary to achieve irb approval for this case report, and this report requires obtaining patient consent because this study is dealt with only the patient 's medical record and related images, retrospectively. written informed consent of this case report and accompanying images was obtained from the patient for the publication. | abstractsmall cell lung cancer (sclc) metastasizes widely, but palatine tonsil is an extremely unusual site for metastasis. idiopathic pulmonary fibrosis (ipf) is associated with increased risk of lung cancer. however, the most common histological findings among patients of lung cancer with ipf are known as non - sclc such as adenocarcinoma and squamous cell carcinoma. in addition, the majority of them are located in ipf - associated fibrotic peripheral lesions.a 77-year - old man visited for 1-month persistent cough and dyspnea, with inspiratory dry crackles on both lower lung fields and a large oval mass in his throat. chest computed tomography revealed 2 masses in the left lower lobe, 1 mass in the right upper lobe, and multiple enlarged mediastinal lymph nodes of the lung accompanying with ipf, which were diagnosed as sclc pathologically. very interestingly, the tonsillar mass was also confirmed as the metastatic lesion of sclc. chemotherapy for sclc and medical treatment for ipf were applied. however, in following - up, he expired due to respiratory failure by an acute exacerbation of ipf 3 months after the diagnosis.in this current report, we describe, for the first time, a case of tonsillar metastasis of sclc with ipf detected simultaneously in a 77-year - old man. |
almost all countries, irrespective of economic status, are currently experiencing an obesity epidemic.13 residence area, occupation, and lifestyle significantly modify the body fat composition, and thus affect the prevalence of obesity.4,5 as a country undergoing economic transition, the people s republic of china has its own characteristics, such as the rapid economic growth and huge rural urban disparity, leading to the corresponding disparities in work and lifestyle. whether or not these disparities in the residence area, occupation, and lifestyle increase the risk of obesity in the people s republic of china is a new issue and worthy of further study. obesity is a fundamental pathology of cardiometabolic abnormalities, such as hypertension, type 2 diabetes, and dyslipidemia.6,7 in the people s republic of china, obesity contributes to an increased risk for cardiovascular disorders, the leading cause of death, especially in rural areas.8 therefore, early prevention, identification, and treatment of obesity are recognized as a major public health issue, emphasizing the importance of developing methods and criteria for establishing an accurate diagnosis.9 the excess and abnormal distribution of body fat are the primary defining characteristics of obesity, and various alternative methods of the measurement of obesity, such as the body mass index (bmi) and waist circumference (wc), have been proposed and applied in clinical practice as easy to use and noninvasive. the bmi is often used to reflect total body fat, whereas the wc is used as a surrogate marker of body fat centralization. in recent years, it has been proposed that central rather than overall obesity can serve as an important identifier of cardiometabolic abnormalities.10 accordingly, the wc has been suggested to be a more useful tool in measuring abdominal fat.11 nevertheless, the wc has been criticized for not taking into account differences in the height and hip circumference (hc), thus the waist - to - hip ratio (whr) and waist - to - height ratio (whtr) have emerged as new parameters of body fat, and have been reported to be more closely associated with cardiometabolic abnormalities.12 in addition, another study has provided evidence that the waist - to - hip - to - height ratio (whhr) is the best variable of identifying cardiometabolic abnormalities ; however, it has also been emphasized that the whhr needs to be tested in more diverse populations.13 recently, one study has confirmed that the body adiposity index (bai) is a useful measure of body fat,14 while another study has developed a body shape index (absi), which can play a significant role as a preferable identifier of cardiometabolic abnormalities.15 it is unclear, however, if these new indices of body adiposity better identify cardiometabolic abnormalities than the bmi and wc, and thus the best way to measure body fat has become increasingly controversial. most studies examining the risk of adverse health associated with obesity have been based on data from the united states or european countries, with little data available on populations in the people s republic of china. several studies have demonstrated that many asians with lower bmi levels have higher amounts of body fat, as well as a high prevalence of cardiometabolic abnormalities, thus the limitation of extrapolating anthropometric criteria established with westerners to asian populations has been underscored.16 the applicability of anthropometric criteria to chinese may differ from the current recommendations by other countries due to racial differences in body build and fat distribution,17 thus to apply ethnically - appropriate anthropometric indices for evaluating cardiometabolic abnormalities is of great importance. in view of all these considerations, the aims of the current study were to evaluate the following in a chinese community - dwelling population : 1) the prevalence of obesity and cardiometabolic abnormalities ; 2) the influence of residence area, occupation, smoking, and alcohol consumption on anthropometric indices ; 3) the relationship between anthropometric indices and cardiometabolic abnormalities ; and 4) the appropriate cut - off values for identifying cardiometabolic abnormalities. this community - based survey was conducted in 5,116 participants through a large health check - up program in beijing, people s republic of china. all of the participants were permanent residents of han origin and 18 years of age. a stratified cluster sampling design was used in this survey. in the first stage of sampling, three districts (fengtai, shijingshan, and daxing) were selected from 18 districts in beijing. in the second stage of sampling, two urban communities were selected from two districts (fengtai and shijingshan), and two residential villages were selected from another district (daxing). in the third stage of sampling, the participants were selected from these communities and villages to represent the overall beijing population. two hundred forty - eight subjects with missing data for essential variables were excluded from the analysis. the final study population comprised 4,868 participants. a questionnaire survey that included questions about residence area, occupation, lifestyle, and histories of hypertension, type 2 diabetes, high triglyceride level, high low - density lipoprotein - cholesterol (ldl - c) level, and low high - density lipoprotein - cholesterol (hdl - c) level was administered using a face - to - face counseling method. the questionnaire survey, anthropometric and blood pressure measurements were all performed by physicians in the chinese people s liberation army general hospital. knowledge work mainly depended on mental activities, while manual work mainly entailed physical activities. cigarette smoking was defined as having smoked at least 100 cigarettes in one s lifetime.18 alcohol drinking was defined as the consumption of at least 30 grams of alcohol per week for 1 year.18 physical examination was performed the morning following an overnight fast of at least 12 hours. height was measured in centimeters using a wall - mounted measuring tape, and weight was measured in kilograms using a digital scale. wc was measured on standing subjects with a soft tape midway between the lowest rib and the iliac crest, and hc was measured over the widest part of the gluteal region. the anthropometric indices were calculated using the following equations : blood pressures were measured according to a standard protocol using the right arm of the participants with a calibrated desktop sphygmomanometer (yuwell medical equipment & supply co., ltd., jiangsu, people s republic of china) between 8 am and 10 am, after the participants had been in a seated position for 5 minutes.19 systolic blood pressure (sbp) and diastolic blood pressure (dbp) were recorded at the first and fifth korotkoff sounds, respectively. reported sbp and dbp were the averages of two separate measurements at an interval of more than 1 minute. samples of venous blood were drawn by venipuncture between 8 am and 10 am from fasting participants who had been resting for at least 5 minutes. blood samples were routinely stored at 4c, and delivered to the central laboratory in the department of biochemistry, chinese people s liberation army general hospital, on the same day. serum concentrations of fasting blood glucose, triglyceride, ldl - c, and hdl - c were measured by a technician, blinded to clinical data, using enzymatic assays (roche products ltd basel, switzerland) on a fully automatic biochemical autoanalyzer (cobas c6000 ; roche products ltd). the standard oral glucose tolerance test was completed after the fasting blood samples were collected. participants were given a standard 75 g glucose solution, and blood samples were drawn 120 minutes after the glucose load to measure glucose concentrations. overweight, general obesity, and central obesity were defined respectively, as a bmi 24 kg / m, bmi 28 kg / m, and wc 85 cm for men and 80 cm for women, according to the guidelines on prevention and control of overweight and obesity in chinese adults.20 a subject was considered to have hypertension when sbp 140 mmhg, dbp 90 mmhg, and/or taking an antihypertensive medication.21 type 2 diabetes was defined as follows : fasting blood glucose 7.0 mmol / l, postprandial blood glucose 11.1 mmol / l, and/or taking a hypoglycemic medication or insulin.22 dyslipidemia, including elevated triglyceride level, elevated ldl - c level, and reduced hdl - c level, was diagnosed when the subject had triglyceride level 1.70 mmol / l, ldl - c level 3.37 mmol / l, and hdl - c level < 1.04 mmol / l.23 cardiometabolic abnormalities 1 indicated that the participant had one or more abnormalities (hypertension, type 2 diabetes, and dyslipidemia), whereas cardiometabolic abnormalities 2 indicated that the participant had two or more abnormalities (hypertension, type 2 diabetes, and dyslipidemia). continuous variables were described using the mean and standard deviation for variables with a normal distribution and the median and interquartile range for nonnormally distributed variables. the bivariate association was assessed by student s t - test for continuous variables (normal distribution), mann whitney u test for continuous variables (abnormal distribution), and chisquare analysis for categorical variables. the independent association, after adjusting for age, was evaluated by multivariate regression analyses. the receiver operating characteristic curve and area under the curve (auc) were calculated to compare the power of all anthropometric indices to identify cardiometabolic abnormalities and indicate the corresponding optimal cut - off values. the statistical analyses were two - sided at the 0.05 significance level and performed with spss version 17 (ibm corporation, armonk, ny, usa). this community - based survey was conducted in 5,116 participants through a large health check - up program in beijing, people s republic of china. all of the participants were permanent residents of han origin and 18 years of age. a stratified cluster sampling design was used in this survey. in the first stage of sampling, three districts (fengtai, shijingshan, and daxing) were selected from 18 districts in beijing. in the second stage of sampling, two urban communities were selected from two districts (fengtai and shijingshan), and two residential villages were selected from another district (daxing). in the third stage of sampling, the participants were selected from these communities and villages to represent the overall beijing population. two hundred forty - eight subjects with missing data for essential variables were excluded from the analysis. a questionnaire survey that included questions about residence area, occupation, lifestyle, and histories of hypertension, type 2 diabetes, high triglyceride level, high low - density lipoprotein - cholesterol (ldl - c) level, and low high - density lipoprotein - cholesterol (hdl - c) level was administered using a face - to - face counseling method. the questionnaire survey, anthropometric and blood pressure measurements were all performed by physicians in the chinese people s liberation army general hospital. knowledge work mainly depended on mental activities, while manual work mainly entailed physical activities. cigarette smoking was defined as having smoked at least 100 cigarettes in one s lifetime.18 alcohol drinking was defined as the consumption of at least 30 grams of alcohol per week for 1 year.18 physical examination was performed the morning following an overnight fast of at least 12 hours. height was measured in centimeters using a wall - mounted measuring tape, and weight was measured in kilograms using a digital scale. wc was measured on standing subjects with a soft tape midway between the lowest rib and the iliac crest, and hc was measured over the widest part of the gluteal region. blood pressures were measured according to a standard protocol using the right arm of the participants with a calibrated desktop sphygmomanometer (yuwell medical equipment & supply co., ltd., jiangsu, people s republic of china) between 8 am and 10 am, after the participants had been in a seated position for 5 minutes.19 systolic blood pressure (sbp) and diastolic blood pressure (dbp) were recorded at the first and fifth korotkoff sounds, respectively. reported sbp and dbp were the averages of two separate measurements at an interval of more than 1 minute. samples of venous blood were drawn by venipuncture between 8 am and 10 am from fasting participants who had been resting for at least 5 minutes. blood samples were routinely stored at 4c, and delivered to the central laboratory in the department of biochemistry, chinese people s liberation army general hospital, on the same day. serum concentrations of fasting blood glucose, triglyceride, ldl - c, and hdl - c were measured by a technician, blinded to clinical data, using enzymatic assays (roche products ltd basel, switzerland) on a fully automatic biochemical autoanalyzer (cobas c6000 ; roche products ltd). the standard oral glucose tolerance test was completed after the fasting blood samples were collected. participants were given a standard 75 g glucose solution, and blood samples were drawn 120 minutes after the glucose load to measure glucose concentrations. overweight, general obesity, and central obesity were defined respectively, as a bmi 24 kg / m, bmi 28 kg / m, and wc 85 cm for men and 80 cm for women, according to the guidelines on prevention and control of overweight and obesity in chinese adults.20 a subject was considered to have hypertension when sbp 140 mmhg, dbp 90 mmhg, and/or taking an antihypertensive medication.21 type 2 diabetes was defined as follows : fasting blood glucose 7.0 mmol / l, postprandial blood glucose 11.1 mmol / l, and/or taking a hypoglycemic medication or insulin.22 dyslipidemia, including elevated triglyceride level, elevated ldl - c level, and reduced hdl - c level, was diagnosed when the subject had triglyceride level 1.70 mmol / l, ldl - c level 3.37 mmol / l, and hdl - c level < 1.04 mmol / l.23 cardiometabolic abnormalities 1 indicated that the participant had one or more abnormalities (hypertension, type 2 diabetes, and dyslipidemia), whereas cardiometabolic abnormalities 2 indicated that the participant had two or more abnormalities (hypertension, type 2 diabetes, and dyslipidemia). continuous variables were described using the mean and standard deviation for variables with a normal distribution and the median and interquartile range for nonnormally distributed variables. the bivariate association was assessed by student s t - test for continuous variables (normal distribution), mann whitney u test for continuous variables (abnormal distribution), and chisquare analysis for categorical variables. the independent association, after adjusting for age, was evaluated by multivariate regression analyses. the receiver operating characteristic curve and area under the curve (auc) were calculated to compare the power of all anthropometric indices to identify cardiometabolic abnormalities and indicate the corresponding optimal cut - off values. the statistical analyses were two - sided at the 0.05 significance level and performed with spss version 17 (ibm corporation, armonk, ny, usa). the basic characteristics of all subjects, stratified by sex, are listed in table 1. in total, 2,323 men and 2,545 women were included in this study. the ages of all participants ranged from 1896 years, with mean and median ages of 51.38 (14.96) and 52.00 (40.0063.00) years, respectively. of the study participants, 17.9% of men and 23.5% of women resided in rural areas, and 45.6% of men and 20.4% of women were engaged in knowledge work. there were more men who lived in urban areas and did knowledge work than women. further, 54.8% of men were smokers and 56% consumed alcohol, while 8.7% of women were smokers and 4.4% consumed alcohol. the median bmi levels in both sexes exceeded chinese overweight standard ; 67.1% of men and 65.2% of women were overweight according to the bmi standard. moreover, 22.2% of men and 28.1% of women were considered to be generally obese, with 65.99% of men and 65.97% of women having central obesity. the prevalence of hypertension, type 2 diabetes, elevated triglyceride level, elevated ldl - c level, and reduced hdl - c level was 37.6%, 10.7%, 37.4%, 25.9%, and 25.1% for the total population, respectively ; women had a higher prevalence of elevated ldl - c level and reduced hdl - c level, and a lower prevalence of elevated triglyceride level than men. there was no difference in the prevalence of hypertension and type 2 diabetes between the two sexes. the likelihood that women had 2 cardiometabolic abnormalities was higher than men ; however, no difference existed between women and men with respect to 1 cardiometabolic abnormalities. table 2 summarizes the association of residence area, occupation, smoking, and alcohol consumption with anthropometric indices, stratified by sex. the residents of rural areas or manual workers had moderately or significantly higher anthropometric indices than inhabitants of urban areas or knowledge workers in both sexes, except that there was no significant difference in the bmi between rural and urban men. the anthropometric indices were significantly higher for men who smoked compared with men who did not smoke, except for the bmi and bai. women who smoked only had higher whr and absi than women who did not smoke in all anthropometric indices. table 3 shows the association between the anthropometric indices and different cardiometabolic abnormalities, stratified by sex. all of the anthropometric indices for the male and female participants who had elevated triglyceride level, reduced hdl - c level, and elevated ldl - c level were significantly higher than the participants who did not have these abnormalities. except there was no significant difference in absi between the hypertensive and non - hypertensive women, as well as a moderate difference in bai between women with and without type 2 diabetes, the men and women with hypertension and type 2 diabetes had significantly higher anthropometric indices than those without hypertension and type 2 diabetes. the participants with 1 or 2 cardiometabolic abnormalities in the higher anthropometric indices group outnumbered the participants in the lower anthropometric indices group, except for the absi in women. table 4 displays the auc and cut - off values of all anthropometric indices for identifying different cardiometabolic abnormalities. among all anthropometric indices, whtr had the highest auc values in both sexes for identifying the risk of hypertension and elevated ldl - c level. the cut - off value of whtr for hypertension was 0.53 for both sexes, and the cut - off values for elevated ldl - c level were 0.53 for men and 0.52 for women. additionally, the index with the highest auc values for both sexes to assess the risk of elevated triglyceride level and reduced hdl - c level was wc ; for men, the cut - off values were 87.50 and 89.95 cm, respectively, and for women, the cut - off values were 85.25 and 82.75 cm, respectively. with respect to type 2 diabetes, the auc value of absi was the highest among all anthropometric indices for men (cut - off value : 0.079), while that of the whr was the highest for women (cut - off value : 0.86). the whtr best identified the participants with 1 cardiometabolic abnormalities (cut - off values : 0.51 for men and 0.53 for women), and 2 cardiometabolic abnormalities (cut - off value : 0.54 for both sexes) in all anthropometric indices. alarming increase in adiposity has been identified in many developed and developing countries.13 the current study demonstrates a serious public health problem in the people s republic of china ; specifically, approximately two - thirds of adults are overweight based on the bmi standard. the prevalence of overall obesity is 22.2% in men and 28.1% in women, and that of abdominal obesity is 65.99% for men and 65.97% for women. as a country undergoing economic transition, the people s republic of china has its own characteristics, such as the rapid economic growth and huge rural urban disparity, leading to the corresponding disparities in work and lifestyle. above all of these disparities may have a significant responsibility for such a high prevalence of obesity and overweight in the people s republic of china. the results from the current study illustrate that the residents of rural areas have moderately or significantly higher anthropometric indices than residents of urban areas in both sexes, except that there is no significant difference in bmi between rural and urban men. the relationship of smoking and alcohol consumption with anthropometric indices remains uncertain, especially in a chinese community - dwelling population. some studies have shown that smokers exhibit higher body weight than nonsmokers, whereas other studies have reported the opposite result. smoking not only increases energy expenditure and suppresses appetite, but also is a risk factor for chronic diseases, which may lead to weight loss;24 however, central weight gain as an effect of smoking may be more common and often associated with other adverse behaviors, such as overeating, irregular eating time, and lack of exercise.25 a population - based study involving 21,828 british men and women showed that smokers had a lower mean bmi compared with nonsmokers, but had a higher whr.26 in the current study, the tendency that smoking adds weight is observed for all anthropometric indices reflecting central adiposity, and there is no distinction in bmi and bai, which are representative of general obesity, confirming that cigarette smoking appears to result in a metabolically - adverse fat distribution pattern rather than a simple increase or loss of weight.26 it is widely believed that alcohol energy adds to total energy intake. meanwhile, alcohol induces increased food intake and prolonged meal duration;27 however, an inverse correlation between anthropometric indices and alcohol consumption has been shown in another study.28 the explanation for this paradoxical phenomenon involving alcohol is that the energy from alcohol has a low biological value.29 in the current study, drinking has no influence on the anthropometric indices in both sexes. the result may be the overall expression of the diverse influence of drinking on anthropometric indices. the roles of overweight and obesity, especially abdominal adiposity, in the development of cardiometabolic abnormalities have drawn considerable attention.6,7 in the people s republic of china, obese adults are often at increased risk of cardiovascular disorders, the leading cause of death, especially in rural areas.8 various methods measuring overall and central adiposity have been developed to assess adiposity, and more studies have been designed to systematically and rigorously evaluate their accuracy in identifying cardiometabolic abnormalities. some investigators have reported that wc is more sensitive and indicative of cardiometabolic abnormalities than bmi,11 or that wc and bmi are both suitable measures to identify adults with cardiometabolic abnormalities.30 nevertheless, data from other studies have suggested that whr and whtr are superior in identifying cardiometabolic abnormalities,12 while other investigators have shown that when each of the anthropometric measures is used, whhr emerges as the best identifier of cardiometabolic abnormalities.13 recently, one study has demonstrated that the bai is a better indicator of body mass,14 whereas another study has developed the absi that successfully expresses the excess risk from obesity in a convenient form.15 however, it has yet to be clarified if the new indices of body adiposity, such as the bai and absi, are more useful identifiers of cardiometabolic abnormalities than other indices. there has been considerable heterogeneity in the results from studies that have analyzed the association between cardiometabolic abnormalities and anthropometric indices, and no consensus as to which anthropometric measure best identifies cardiometabolic abnormalities. asians are predisposed to visceral obesity, with higher amounts of body fat corresponding to lower bmi levels than westerners, and their health risks associated with obesity occur at lower bmi levels, suggesting that the heterogeneity may be attributable to difference in the ethnicity distributions of participants.16,17 for chinese adults, the current study indicates that the anthropometric indices have unequal power in the risk assessment of different abnormalities. in other words we observe that the whtr is superior to other anthropometric indices in risk assessment for hypertension and elevated ldl - c level in both sexes. the wc has a better performance than other anthropometric indices in its association with elevated triglyceride level and reduced hdl - c level. the anthropometric indices better identifying type 2 diabetes are absi in males and whr in females. compared with other anthropometric indices, the whtr has stronger ability for identifying participants with greater than one or two cardiometabolic abnormalities, illustrating that whtr has the ability to reflect the compound risk of different cardiometabolic abnormalities. in addition, the cut - off points of anthropometric indices for identifying cardiometabolic abnormalities are derived from studies in the us or european countries, and thus may not be applicable to other ethnic groups. to solve the problem, the cut - off values of anthropometric indices for chinese are obtained in the current study as well. to apply the best anthropometric indices combined with the indigenous cut - off values may provide the greatest convenience for preventing cardiometabolic abnormalities. moreover, it may be worth noting the importance of collecting longitudinal data so we can relate changes in anthropometric indices over time with changes in cardiometabolic abnormalities over time. the current study demonstrates that being overweight is very common for both sexes in the people s republic of china, as are general and central obesity, and the residents of rural areas, manual workers, and smokers have significantly higher anthropometric indices, warning us that overweight and obesity are serious threats to the public health and social security of the people s republic of china. meanwhile, whtr is superior at identifying individuals with hypertension and elevated ldl - c level, wc has a greater ability of identifying elevated triglyceride level and reduced hdl - c level, and the anthropometric indices that are considered to be more indicative of type 2 diabetes are absi in males and whr in females, verifying that the identifying power of each anthropometric index varies for different cardiometabolic abnormalities. more importantly, the ability of the whtr to identify the participants with greater than one or two cardiometabolic abnormalities is superior to other anthropometric indices, showing that whtr has the ability to reflect the compound risk of different cardiometabolic abnormalities and the greatest potential to be widely applied in clinical practice. | objectivethe study aimed to investigate the prevalence of overweight, obesity, and cardiometabolic abnormalities, the influence of residence area, occupation, and lifestyle on new anthropometric indices, and the relationship between anthropometric indices and cardiometabolic abnormalities in a chinese community - dwelling population.methodsthe study included 4,868 residents through a large health check - up program in beijing.resultsoverall obesity existed in 22.2% of men and 28.1% of women. 67.1% of men and 65.2% of women were overweight. 65.99% of men and 65.97% of women had central obesity. residents of rural areas, manual workers, and smokers had significantly higher anthropometric indices. the power of each anthropometric index varied for identifying different cardiometabolic abnormalities, and the ability of the waist - to - height ratio to identify participants with greater than one or two cardiometabolic abnormalities was optimal. the appropriate cut - off values of all anthropometric indices for cardiometabolic abnormalities were obtained.conclusionoverweight is common for both sexes in the people s republic of china, as are general and central obesity. residents of rural areas, manual workers, and smokers have significantly higher anthropometric indices. waist - to - height ratio has the ability to reflect the compound risk of different cardiometabolic abnormalities and the greatest potential to be widely applied in clinical practice. |
isolated pauci - immune pulmonary capillaritis (ipipc) is a rare disorder of unknown etiology. it is characterized by small vessel vasculitis confined to lungs, and systemic manifestations are typically absent. it is considered as a subset of microscopic polyangiitis (mpa) but is mostly sero - negative for anti - neutrophilic cytoplasmic antibodies (anca)., the disorder was fully characterized by jenning. since then only sporadic case reports of the disorder are available in the english literature. we are reporting such a case which, to the best of our knowledge and belief, is the first case report of the disorder in indian medical literature. rd, a 28-year - old housewife from narnaul, haryana, was referred to the outpatient department of respiratory medicine on 23.08.2013 with low grade fever, cough, hemoptysis and shortness of breath (sob) for about 40 days duration. she was a multi - para with normal vaginal deliveries, last being 11 years back. prior to this, she reported to a private practitioner with sudden onset of intermittent fever, cough and sob for which she was put on oral amoxiclav along with symptomatic treatment but after a bout of severe hemoptysis, she was rushed to a general hospital on 18.07.2013 where she was investigated. her x - rays [figure 1a and b ], contrast enhanced computerized tomography (cect) and high - resolution computerized tomography (hrct) thorax [figure 2a and b ] revealed diffuse bilateral cavitating nodular lesions and consolidation in right lower lobe along with small pleural effusion but without any remarkable changes in the mediastinal lymph nodes or the vasculature. echocardiography revealed trivial tricuspid regurgitation without any clot or vegetation, normal pericardium and an ejection fraction of 60%. she was kept on intravenous (iv) meropenem and levofloxacin along with other symptomatic treatment but the patient continued to deteriorate and she was referred to this hospital. laboratory investigations before admission (a) x - ray chest pa view dated 18.07.2013 showing diffuse bilateral necrotizing nodular lesions, more on right than left and heterogeneous opacity in right lower lobe (b) x - ray chest pa view dated 24.07.2013 showing appearance of new nodular shadows in right upper and mid zones and in left lower zone (a) ct thorax dated 24.07.2013 showing diffuse necrotizing nodular lesions (b) topogram dated 24.07.2013 showing bilateral diffuse thick - walled necrotizing nodular lesions with consolidation and small right sided pleural effusion she was an average built, poorly nourished female having pale and toxic look. her body temperature was 98f, pulse ; 160/minute, blood pressure ; 90/60 mm of hg and respiratory rate ; 30/minute. respiratory system examination revealed decreased breath sounds in the right mid and lower chest along with diffuse crepitations and occasional rhonchi. ear, nose, throat and eye checkup did not reveal any abnormality as was the examination of other organ systems. a repeat x - ray chest pa view and ct thorax on admission were remarkably similar to her earlier x - rays and ct thorax [figures 1a and b and 2a and b ]. total leucocyte count (tlc) was 11,370/mm with polymorphs ; 90%, lymphocytes ; 7%, eosinophils ; 1%, monocytes ; 1% and basophils ; 1%. platelet count was 2.48 lacs / cmm but hemoglobin (hb) was low i.e. 9.1 gm%. her sputum samples were negative for pyogenic organisms, acid fast bacilli or fungi. considering necrotizing pneumonia, she was continued on antibiotics and symptomatic treatment. the patient was also given 2 units of fresh blood during the hospital stay and oxygen by face mask, but she continued to have cough, hemoptysis and weight loss. repeat hemogram was unremarkable except that hemoglobin (hb) was further lowered to 7.1 gm%. a repeat serology was negative for angiotensin - converting enzyme (ace), anti - nuclear antibodies (ana), rheumatoid factor (rf), cytoplasmic antineutrophilic cytoplasmic antibodies (c - anca), antiglomerular basement membrane (anti - gbm) antibodies and anti - aspergillous antibodies but positive for c - reactive proteins (crp) and weekly positive for perinuclear antineutrophilic cytoplasmic antibodies (p - anca) by indirect immune - fluorescence assay (iifa). bronchoscopy revealed normal architecture of the airways on either side, but active bleeding was seen from the right lower lobe bronchus. an attempt was made to stop bleeding by instilling frequent aliquots of adrenaline mixed with cold saline but hemostasis could not be achieved so the plan for obtaining lung tissue by transbronchial route was shelved. a provisional diagnosis of ipipc was made on the basis of the overall clinical, radiological and laboratory findings. the patient was started on pulse therapy with methylprednisolone (1 g iv for 3 days) along with cyclophosphamide (15 mg per kg body weight iv on day 1). on this, the patient improved and was discharged on iron therapy with the advice to report for repeat pulse therapies every fortnightly. in due course, cough and sob declined, hemoptysis stopped, hb levels increased to 11.5 g% and x - ray chest pa view showed remarkable clearance [figure 3 ]. patient is still under follow up and is asymptomatic by the end of the 6 cycle. x - ray chest pa view dated 28.10.13 showing resolution of lesions as compared to figure 1b brow suggested that falling hematocrit with or without hemoptysis, unexplained bilateral diffuse alveolar infiltrates in x - ray chest and persistently bloody return in serially aspirated bronchoalveolar lavage (bal) favored the diagnosis of dah. later, cordier. opined that infections, hematological disorders, hemolysis or hemorrhage elsewhere should be ruled out prior to make the diagnosis of dah. based on the presence or absence of systemic symptoms, dah has been classified into two broad groups, one with systemic manifestations of vasculitis and the other without it. the latter is further classified as (a) anti - gbm disease in limited pulmonary form (positive anti - gbm antibody with linear deposits in the lung tissue), (b) pulmonary - limited mpa (positive for p - anca and mpo - anca), (c) isolated pauci - immune pulmonary capillaritis (anca negative neutrophilic pulmonary capillaritis) and (d) idiopathic pulmonary hemosiderosis (acute, sub - acute or chronic bland dah in the absence vasculitis in young smokers). only few case reports of ipipc with varied presentations are available in the medical literature. her thoracoscopic lung biopsy showed capillary destruction and neutrophilic infiltration of the lung interstitium / air spaces but direct evidence for dah was lacking in this case. all had symptoms related to of lower respiratory tract, six of them also having acute respiratory failure and 4 of them needed mechanical ventilation. all were anca negative but direct immune - fluorescence assay (ifa) of lung tissue was positive in 3. monteiro. reported a case of ipipc in a 27-year - old female. this patient used to consume cocaine and presented with hemoptysis and progressive dyspnea of short duration. serum tests for infectious diseases, collagen disorders and vasculitis were negative and urine examination was normal. our patient presented with hemoptysis, cough, shortness of breath and low grade fever for about 40 days duration.. she never smoked or consumed alcohol and did not have a significant illness in the past. respiratory system examination on auscultation revealed diffuse crepitations and occasional rhonchi but other system examination revealed nothing abnormal. her x - ray chest pa view and hrct thorax showed diffuse bilateral cavitating nodular lesions of varying sizes and patchy consolidation. meanwhile, she was continued on antibiotics but she did not respond and showed clinical and radiological deterioration. transbronchial lung biopsy could not be undertaken due to continued bleeding in spite of repeated installation of adrenaline mixed saline. her serology was negative for ana, rf, c - anca, anti gbm, mpo - anca and anti - aspergillous antibodies but was positive for c - reactive proteins and weekly positive for p - anca. thus necrotizing pneumonia and other infections (tuberculosis, fungal infections etc.), systemic vasculitis disorders (collagen vascular disorders, wg, mpa and css) and vasculitis without systemic manifestations (pulmonary renal syndromes and pulmonary limited mpa) as the cause of dah were ruled out in our case. absence of smoking history, presence of frank hemoptysis (rather than blend hemorrhage), persistent bloody aspirates in bal and the presence of necrotizing lesions in her x - ray chest did not favor the diagnosis of idiopathic pulmonary hemosiderosis either. successful response to pulse therapy with methylprednisolone and cyclophosphamide strongly supported the diagnosis of ipipc in our patient. were also of the opinion that although biopsy remained the key to diagnosis of ipipc yet a confident diagnosis may occasionally be made without tissue biopsy on the basis of clinic - radiological features. it is suggested that ipipc, though a rare disease, should be kept in mind and considered in differential diagnosis of dah in an anca - negative young patient in the absence of systemic manifestations, more so when infections, hematological disorders, hemolysis or hemorrhage and collagen vascular diseases have been ruled out. | a young house wife presented with low grade fever, cough, haemoptysis and sob of unknown aetiology for 40 days duration. respiratory system examination revealed diffuse crepts and rhonchi. other organ system examination did not reveal any abnormality. x - ray chest pa view and ct thorax showed diffuse bilateral necrotising nodular lesions of various sizes with small pleural effusion. she also had low resting oxygen saturation with falling haematocrit. her serum was week positive for p - anca and negative for mpo - anca. bronchoscopy revealed continuous bloody aspirates. we could not isolate any organisms in any of the specimens from her and she was unresponsive to any of the antibiotics either. based on the clinical, laboratory data, radiological features and positive outcome to pulse therapy of methylprednisolone and cyclophosphamide, she was diagnosed as a case of ipipc. |
when oxygen supply to the fetus is significantly disrupted, tissue oxygenation deprivation develops, acids begin to accumulate and acidemia develops. umbilical cord blood gas measurement in comparison with the fetal scalp ph monitoring could better detect a hypoxic baby. in 1952, virginia apgar, proposed a scoring system to assess the condition of newborns during the first minutes of life, and to evaluate anesthetic and obstetrical practices. she proposed five objectives and easily - measured clinical signs : heart rate, respiratory effort, muscle tone, reflex irritability, and color. it has been used to assess asphyxia, predict neurological damage and vitality of a neonate during the first minute of life. umbilical cord blood gas assessment seems to be the most objective determination of the fetal metabolic condition at the time of birth [47 ]. manganaro and colleagues suggested that the 5 minute apgar score is useful for immediate clinical assessment and care of the neonate. they found 5 minute apgar score had a high concordance with metabolic acidemia, but anyaegbunam and colleagues ' study revealed that in 20.7% of neonates delivered had an abnormal ph (less than 7.20) and normal apgar. the present study was carried out to determine the correlation between umbilical cord ph and apgar score in high - risk pregnant mother. this was a prospective cross sectional study conducted in a teaching hospital during february 2004 to september 2005. the study was approved by the research ethics committee of babol university of medical sciences. study population consisted of mothers who came with labor pain according to the sample size. at the time of admission, they were assigned to high or low risk group according to whether or not they had any perinatal risk factor that categorized them as high - risk pregnancy. high - risk pregnancy was defined as a pregnant mother who is at risk to deliver a neonate with birth asphyxia according to the definition by american academy of pediatrics. all normal vaginal and cesarean section (c / s) deliveries included in this study were chosen in accordance with this definition. all mothers who delivered a baby with a major congenital anomaly or had intra uterine fetal death (iufd) were excluded from the study. immediately after delivery, umbilical cords were clamped on both ends and an arterial blood sample was collected anaerobically in a pre - heparinized insulin syringe. ph, base excess, carbon dioxide pressure (pco2), po2 and hco3 were measured at 37c by ph and gas analyzer (avl, compact3, australia). apgar score was assessed by a trained physician at 1 and 5 minute after birth. in case of an apgar score less than 8, advanced resuscitation means that a baby required positive pressure ventilation, chest compression and/or drugs administration. all resuscitated babies were transferred to neonatal intensive care unit or newborn services for post resuscitation care. demographic data like gestational age, birth weight, apgar score, need for resuscitation and/or newborn ward admissions were collected by a questionnaire in both groups. sample size calculated 96 mother - fetal pair for each group (if we consider 1-=0.95, r=0.25 and 1-=0.80). for each high - risk mother, a low risk mother was selected as control. student 's t - test, the mann - whitney and test were used for analysis. during a 6-month period, 96 mother - fetus pairs, 49 in high risk, and 47 in low risk group participated in the study. frequency of perinatal risk factors in high - risk groups is shown in table 2. demographic characteristics of neonates of high and low risk groups nicu= neonatal intensive care unit the frequency of perinatal risk factor in high - risk pregnant mothers af = amniotic fluid ; fhr= fetal heart rate ; iugr = intra uterine growth retardation ; c / s= cesarean section the most common perinatal risk factors accompanied with low umbilical artery ph were prolonged rupture0 of membranes, breech presentation, and meconium stained amniotic fluid. the apgar score at 1 and 5 minutes in high - risk patients were significantly lower than in the low risk group (table 3). ph values in mothers were significantly lower than those in low risk counterpart (p<0.001). apgar score at 1, 5, 10 and 15 minutes after birth in high and low risk group (mean sd) umbilical arterial blood ph, base excess and gas values in high and low risk mothers immediately after birth (mean sd) according to kendal correlation coefficient, there was no significant correlation between apgar score at 1 and 5 minute and umbilical cord ph in low risk group (r=0.212, p=0.1), whereas in high - risk group, a significant correlation between apgar score at 1 and 5 minute and umbilical cord ph was found (r=0.01, p=0.036 and r=0.176, p=0.146 respectively). in the current study, we studied correlation of umbilical cord measures of acidosis at birth and the presence of risk factors in pregnancy. the meansd value of umbilical ph in high - risk group was lower than that in low risk group. there was also a significant relation between umbilical cord ph and low apgar score with the incidence of selective neonatal outcomes like nicu admission and need for advanced resuscitation. other authors reported similar short - term outcomes for hospital based patients ' populations giving birth at term [11, 12 ]. meansd for umbilical cord ph, nahco3 and pco2 in neonates delivered to a high - risk pregnant mother differ significantly with those in low risk mothers but mean (sd) values for po2 and base excess did not differ between the two groups. this finding may emphasize the importance of the latter values on prediction of the occurrence of neonatal outcome. in fact metabolic and respiratory acidosis and hypoxia may jeopardize the baby more than a respiratory acidosis alone [13, 14 ]. apgar score at 1st and 5th minute in high - risk group was lower than that in the low risk group. but there was no significant difference in apgar score at 10th and 15th minute between the two groups. this finding emphasizes the impact of the perinatal risk factors on the immediate general condition of the babies at birth and also the effect of immediate intervention on the improvement of the neonatal condition. in our study, there was no significant correlation between apgar score at 1 and 5 minute and the umbilical artery ph in low risk group. these two parameters, apgar score at first minute and umbilical cord ph, behave independently. if immediate intervention and resuscitation commence, there would not be enough time to develop persistent hypoxemia and acidosis. on the other hand, in high - risk group both the apgar score and umbilical cord ph should be employed to interpret the actual condition of the neonate. socol and colleagues showed that neonates with an apgar score less than or equal to 3 at five minutes and a complicated clinical course were more likely to have lower umbilical cord arterial ph measurements and higher base deficit values than did their counterparts with an uncomplicated clinical course. vargas - origel found a significant difference in the apgar score at one and five minute, as well as in umbilical cord ph between asphyxiated babies and control group. in a newly published study, locatelli and colleagues evaluated the predictors of umbilical artery acidemia in term infants and found that evidence of acidemia is present in only 38% of term babies with low apgar score and it is predominantly associated with intrauterine vascular disease like preeclampsia, abruptio placenta, birth weight less than 10 percentile and placental vascular pathologies. although apgar score provides a convenient short hand for the status of the newborn, it is often incorrectly used as a correlate of neonatal acidosis. in fact, only a minority of neonates with low apgar scores at 5th minute have cord evidence of metabolic acidosis. in order to evaluate how often low 5 minute apgar score at term is associated with asphyxia, hogan and their colleagues studied 183 term infants with apgar score below 7 and concluded that in the absence of malformations, the majority of apgar scores below 4 and at least half of scores 4 - 6 could be attributed to birth asphyxia. signs of hypoxia usually appeared during labor but they were present at admission in 38% of cases with apgar score below four. the most common risk factor found in high - risk group in our study was emergency c / s. in a local study, no relation was found between the mode of delivery and umbilical cord blood gases. in another local study there was a significant correlation between c / s, low apgar score and acidemia. emergency c / s can result in acidemia because of the underlying condition that necessitates the emergency c / s. our study highlights a correlation between the presence of perinatal risk factors and umbilical cord ph in high - risk mothers. so we recommend assessing the umbilical cord ph in any mother who has a perinatal risk factor in her history or physical examination. | objectivethe apgar score as a proven useful tool for rapid assessment of the neonate is often poorly correlated with other indicators of intrapartum neonatal well - being. this study was carried out to determine the correlation between umbilical cord ph and apgar score in high - risk pregnancies.methodsthis is a prospective cross - sectional, analytic study performed on 96 mother - fetal pairs during 2004 - 2005 at shahid yahyanejad hospital, which is affiliated to babol university of medical sciences. apgar score at 1 and 5 minutes after birth was taken and an umbilical cord blood gas analysis was done immediately after birth in both groups. mothers came with a labor pain and were divided into high - risk and low risk if they have had any perinatal risk factors. other data like gestational age, birth weight, need for resuscitation and admission to the newborn ward or neonatal intensive care unit was gathered by a questionnaire for comparison between the two groups. p - value less than 0.05 was considered being significant.findingsthe gestational age and birth weight were the same in high - risk and low risk mothers. mean umbilical artery blood ph in high - risk mothers was significantly lower than in low risk mothers (p=0.004). mean apgar scores at 1 and 5 minutes were significantly lower in high - risk mothers than in low risk mothers (p<0.05). according to the kendal correlation coefficient there was no significant correlation between apgar score at 1 and 5 minutes and umbilical cord ph in low risk group (r=0.212, p=0.1). but in high - risk group there was significant correlation between apgar score at 1st and 5th minute and the umbilical cord ph (r=0.01, p=0.036 and r=0.176, p=0.146, respectively).conclusioncombination of apgar score and umbilical cord ph measurement in high - risk pregnant mother could better detect jeopardized baby. |
vaccination has been one of the most effective measures to control infectious diseases in the 20th century. most of the viral (mumps, rubella, measles, varicella and rotavirus) and bacterial (tetanus, diphteria, haemophilus influenzae, streptococcus pneumoniae, meningococcus group c) diseases traditionally affecting childhood all over the world are now preventable with vaccines (givs 20062015 at http://www.who.int/vaccinesdocuments). besides paediatric and adult population, vaccination has been successfully extended to adolescents, elderly, immunocompromised subjects and pregnant women. although therapeutic vaccines targeting ongoing chronic infectious diseases have not yet been developed, effective vaccines exist that prevent chronic infections by human papilloma virus (hpv) and hepatitis b virus (hbv), which in some cases cause cancer. in summary vaccination has been and continues to represent a success story (plotkin., 2008 ; levine., 2009). however, there is still a long list of viruses [e.g. hepatitis c virus (hcv), human immunodeficiency virus (hiv), dengue, respiratory syncytial virus (rsv) and cytomegalovirus (cmv) ] ; parasites (e.g. plasmodium, leishmania, schistosoma, trypanosoma) and bacteria [e.g. mycobacterium tuberculosis (tb), group a streptococcus (gas), group b streptococcus (gbs), staphylococcus aureus, meningococcus group b (menb), shigella, pathogenic escherichia coli ] that are not preventable by vaccination, and infectious diseases are still a major cause of death and disability worldwide. thirtythree million people are currently infected by hiv that causes 1.8 million deaths per year (who global summary of aids epidemic : http://www.unaids.org/globalreport). malaria and tuberculosis together kill almost 3 million people every year (who report 2010 : http://www.who.int). the total number of deaths increases even more if we include those caused by vaccine preventable infections in areas of the world where vaccines are not available to the population. it is estimated that over 2.5 million of children under 5 years of age die every year as a result of vaccine preventable infections (givs 20062015). in the future, the increased availability of existing vaccines and the development of new vaccines can have a tremendous impact in reducing the mortality and disability caused by infectious diseases worldwide. traditional vaccines based on inactivated or attenuated pathogens or on purified pathogen subunits, such as toxins, proteins and polysaccharides, have been very efficient in preventing infections of pathogens with low degree of antigen variability. these vaccines work mainly by eliciting functional antibodies that can (i) neutralize viral invasion, (ii) neutralize bacterial toxins and (iii) induce opsonophagocytosis or complementmediated killing of bacteria (germain, 2010). it has also been possible to develop effective vaccines to prevent pathogens that have a moderate degree of antigen variability and exist in multiple strains. the problem has been solved by developing multivalent vaccines, which combine multiple antigens, each directed against one strain or serotype, in the same formulation. four different viruslike particles that elicit neutralizing antibodies have been assembled in one hpv vaccine (pomfret., 2011). in the case of bacterial pathogens, polysaccharides derived from the capsule of various strains have been conjugated to protein carriers and mixed in the same vaccine. multivalent glycoconjugate vaccines have been developed for pneumococcus (7valent ; 10valent and more recently a 13valent vaccine) (prymula and schuerman, 2009 ; duggan, 2010) and for menigococcus (two 4valent vaccines have been licensed for a, c, w and y serotypes) (pace, 2009). a glycoconjugate strategy could not be adopted for meningococcus serotype b because its capsular polysaccharide is similar to a self antigen. in this case traditional techniques were unable to identify a universal vaccine against all menb strains, and a novel approach based on genomic information called reverse vaccinology has been applied as described in detail in the next paragraph (giuliani., 2006). traditional vaccines have been also able to prevent diseases caused by pathogens, such as influenza (flu), that are not only present in different strains or clades, but change the antigenic target of neutralizing antibodies (haemagglutinin, ha) at every season. in this case the problem has been solved by producing a trivalent vaccine and by changing the composition of the vaccine every year. however a flu vaccine with increased breadth would greatly help the manufacturing process and increase vaccine efficacy. some investigators have suggested various strategies to develop a universal flu vaccine, capable of preventing infection by all existing flu strains (nabel and fauci, 2010 ; dormitzer., 2011). a universal flu vaccine would be ideal to prevent, or at least mitigate, the risk of pandemic outbreaks, which are originated by unpredictable flu strains of animal origin. until today vaccines failed to prevent infections by pathogens characterized by a high mutation rate that are able to modify the target antigens and evade the antibody response during the course of the same infection. in addition, vaccines have been very inefficient in preventing infections that are not controlled by antibodies but by cellular immunity. the challenge for the future is even higher if we consider that the infections caused by some of the most variable pathogens, such as hiv and hcv, are probably not preventable only by antibodies but require a combination of humoral and cellular responses (mcelrath and haynes, 2010). the success of future vaccines against highly variable pathogens depends on the ability to induce a universal b cell response, characterized by the production of functional antibodies that can crossreact with multiple variants of the same antigen. however, in some cases a universal antibody response may not be enough for protection and must be complemented by the ability to elicit efficient cd4 and cd8 responses (sallusto., 2010). in this case, similarly to antibody assays (neutralization, opsonophagocytosis, bactericidal killing) that have been used as correlate of protection for vaccines that work through b cells, new cellular assays to evaluate vaccine efficacy must be developed. it has been highlighted by several experts in the field that if we want to understand the cellular correlate of vaccine protection it is not sufficient to count the frequencies of antigenspecific t cells in response to vaccination but it is important to assess their quality : the cytokine that they make, their differentiation state (central memory, effector memory, effector t cells) and the receptors expressed on the surfaces that may predict their localization in case of infection (germain, 2010 ; sallusto., 2010). the following paragraphs will focus on the strategies that have been recently employed to cope with bacterial and viral diversity and on the technologies that can be applied to the vaccines of the future to prevent highly variable pathogens. we will then describe new technologies including adjuvants, delivery systems and viral vectors that can be used to shape the cellular and humoral immune response to vaccination. finally we will review new system biology approaches to better understand human correlates of vaccine efficacy. a vaccine discovery process called reverse vaccinology has been efficiently adopted for bacterial pathogens that have a high degree of antigen variability and circulate in multiple strains (sette and rappuoli, 2010). in summary, genomic information of multiple strains has been used to select surfaceexposed antigens that are more conserved. in silico selected antigens the antigens that gave the best bactericidal antibody responses or in the absence of a correlate of protection the best survival rate in animal challenge studies have been selected for prototype vaccines. this approach has been adopted for the development of a vaccine that was able to prevent infection by most men b strains in mice (giuliani., 2006). the preclinical results were confirmed by clinical trials in adults and infants and a licence application has been recently submitted in europe (findlow., 2010 ; snape., 2010). the same approach has been adopted for the development of a vaccine able to protect all circulating strains of gbs (maione., 2005). in most of the cases the antigens identified by reverse vaccinology were unknown and their characterization has often helped to understand the biology of the pathogen. for example, a protective antigen identified by reverse vaccinology in gas was found to be the component of a previously unknown piluslike structure mediating bacterial adhesion to host cells (lauer., 2005). in the future, reverse vaccinology can be applied to generate universal proteinbased vaccines against variable pathogens such as gas, s. pneumoniae and pathogenic e. coli strains. reverse vaccinology is not applicable to the development of vaccines against viruses with high mutation rate. in contrast, it has been shown that the interrogation of human b cell response to infection can lead to the identification of crossreactive protective epitopes that may represent structural determinants of broadly protective vaccines. this process has been called analytical vaccinology and is based on the recent development of several methods of generating human monoclonal antibodies from human blood samples (reviewed in sallusto., 2010). one approach that has been used to analyse the human b cell compartment was to select total memory b cells from infected or vaccinated subjects, immortalize them through a new efficient method and screen them for antibody functional assays (traggiai., 2004). alternatively it was possible to select plasma cells induced by vaccination (wrammert., 2008) or antigenspecific memory b cells (scheid., 2009) the antibodies were then expressed in heterologous cells and used for functional assays (liao., 2009). through these approaches it has been possible to analyse the human antibody repertoire generated by infection of variable pathogens such as flu and hiv. in both cases broadly neutralizing human monoclonal antibodies have been identified and the epitopes that they recognize have been mapped. flucrossneutralizing antibodies recognize a conserved region in the stem of the ha protein (ekiert. these antibodies do not prevent ha binding to receptors, but interfere with conformational changes in ha that are required for membrane fusion (sui., 2009). in the case of hiv, extremely efficient monoclonal antibodies crossneutralizing 91% of primary isolates tested in the form of pseudoviruses bind a conserved epitope in the cd4 binding site of the env protein (wu., 2010). the same approach has been used for the identification of human monoclonal antibodies that neutralize human cmv infection. in this case the analytical vaccinology has shown that protective neutralizing epitopes are generated on a pentameric protein complex which represents a new candidate for vaccine development (macagno., 2010). the identification of conserved epitopes in hiv and flu that are target of broadly neutralizing antibodies is only the first step for the development of universal vaccines. a lot of work needs to be done to rationally design and express the target antigens (e.g. ha or env) in a form that stabilizes crossreactive epitopes and makes them sufficiently immunogenic. the only way to achieve this goal is to have a deep structural knowledge of the candidate antigens (fig. 1). human samples from vaccinated or convalescent subjects are used to isolate crossreactive monoclonal antibodies through in vitro functional assays (viral neutralization or bacterial killing). structural vaccinology methods are applied to express and stabilize the antigens in the protective conformation. structural information can also be exploited to generate chimeric proteins which contain protective epitopes deriving from multiple variants of the same antigen. this approach has been recently used for a novel menb candidate vaccine that was rationally designed by assembling epitopes from three different variants of menb factor hbinding protein (scarselli., 2011). the identification of protective epitopes and the production of rationally designed antigens that induce crossneutralizing antibody are not enough to produce an effective vaccine. the antigens need to be formulated with antigen delivery systems and immunomodulators or must be inserted in viral or nucleic acid expression vectors. it has been well known for a long time that adjuvants can increase the amount, the quality and the duration of the antibody responses to vaccination. empirically derived adjuvants such as aluminium salts have been used in human vaccines for more than 70 years. however, in the last decade the knowledge of the human innate immune system has rapidly progressed leading to the identification of novel classes of molecules [tolllike receptors (tlrs), riglike receptors (rlrs), nodlike receptors (nlrs), ctype lectins ] that are validated targets for the development of new immunomodulators (reviewed in this issue by carlos gutzman., in press). in parallel, a significant amount of work has been done to advance antigen delivery systems including liposomes, virus like particles and nanoparticles, which can be used to formulate antigens and immunomodulators in the same vaccine allowing codelivery (bachmann and jennings, 2010). despite the universally acknowledged potential of vaccine adjuvants, only two compounds are licensed for human use in the usa (alum and the tlr4 agonist mpl) and few others in europe (virosomes and the oil in water emulsions mf59 and as03) (coffman., 2010 ; mpl adsorption to an alumhpv vaccine promoted higher neutralizing antibody titres and increased memory b cell frequencies (giannini., 2006). the addition of oil in water emulsions mf59 and as03 to h5 pandemic flu vaccines was required for protection (seroconversion) and promoted crossreactivity versus various heterologous h5 clades (banzhoff., 2009 ; 2009). besides antibodies, mf59 enhanced both h5specific cd4 and memory b cell responses to vaccination (galli. 2009a, b). interestingly, the increased breadth of the response to h5 induced by mf59 was associated with the recognition of an increased number of epitopes on ha (khurana., 2010). these data suggest that adjuvants can change not only the amount but also the quality of the humoral responses to vaccination, allowing for a better coverage of less immunodominant epitopes that may be very important for crossprotection. many new adjuvants, in isolation or in combination, are in clinical trials with very promising results. one example is as01, a combination of liposomes, mpl and qs21 that was used in a malaria phase ii clinical trial that achieved for the first time 3050% protection (cohen., 2010). another promising adjuvant in clinical development is the tlr9 agonist cpg oligonucleotide (steinhagen., 2010). in addition, many other tlr agonists, such as polyi : c, flagellin, lipoproteins, have already been tested in humans in combination with various antigen and delivery systems (steinhagen., 2010). it is easy to predict that in the future more immunomodulators and antigen delivery systems will be available for use in human vaccines. the use of these compounds in various combinations will greatly help to rationally design vaccines with distinct properties, with the aim of generating protective adaptive immune responses. however the induction of cd8 by subunit vaccines, which relies on crosspresentation mechanisms, is generally very poor. some adjuvants, such as the saponinbased iscoms have shown the ability to increase crosspresentation in animals and human studies (drane. however in humans iscoms produced only partial cd8 responses (drane., 2009). probably the best approach to induce mhcimediated immune responses is to use nucleic acid or viral vectors, which mediate the expression of the selected vaccine antigens directly in the cytoplasm of target cells (liu, 2010) (reviewed in this issue by oliver elbert, in press). several dnas and replicating or nonreplicating viral vectors have been tested in clinical trials until now ; however, none of them has been licensed for human use. one very wellknown example of a nonreplicating vector is the adenovirusderived ad5 vector that was used for the step hiv trial and induced strong cd8 responses in all subjects that did not have preexisting ad5 antibodies (mcelrath., unfortunately, the trial showed that a good cd8 response did not correlate with prevention of hiv infection, suggesting that probably a more balanced immune response including both cd4 and cd8 t cells and crossneutralizing antibodies is required (buchbinder., 2008 ; mcelrath., 2008 ; mcelrath and haynes, 2010). one way to obtain multifunctional responses is to use prime boost regimens, in which the priming with an expression vector targeting cd4 and cd8 responses is followed by a boost with a subunit vaccine, which improves antibody production. subjects have been primed with a canarypox vector encoding gag, env and protease and boosted with a gp120 subunit vaccine. the rv144 trial showed for the first time 31% efficacy against hiv1 acquisition (rerksngarm., 2009). these results are very encouraging and can be considered as a proof of concept that an hiv vaccine strategy can be applied. however, more work needs to be done to optimize the vaccine by using more efficient expression vectors and a rationally designed env antigen eliciting crossneutralizing antibodies formulated in adequate immunomodulators and delivery systems. although the rv144 trial has shown the benefit of a primeboost strategy for a preventive vaccine against hiv, it is still not clear which are the effector mechanisms elicited by the vaccine that protected some of the subjects from hiv infection. for difficult intracellular pathogens such as hiv and hcv, protection probably arises from the integration of different effector mechanisms of the appropriate quality. therefore, correlates of protection are not easily measured by simple antibody or t cell assays, but require more complex readouts. with the progress of genomics it is possible to generate a lot of highthroughput data from human blood samples including : rna expression profiles measured through dna microarrays ; protein expression profiles measured by mass spectrometry ; and analysis of genomic polymorphism measured by deep sequencing. systems biology is required to integrate genomic data with the data obtained from the same subjects through classical immunological assays for antibodies (neutralization, bactericidal activity, elisa) or for t cell characterization (elispot ; facsintracellular staining ; proliferation, cytotoxic activity) (pulendran., 2010) (reviewed in this issue by david klatzmann, in press). in addition, more innovative immunological assays are now available, such as antibody repertoire analysis or single cell phosphospecific flow cytometry (krutzik., 2011) that allow to evaluate in detail the quality of humoral and cellular responses to vaccination. systems biology approaches have been already successfully applied to humans vaccinated with the yellow fever vaccine yf17d (gaucher., 2008 ; these studies have shown that it is possible to predict the efficacy of the vaccine by measuring the transcriptome of pbmcs few hours after vaccination. interestingly, the predictive innate immune signatures were not obvious, involving genes, such as eif2aka that had not been associated with the generation of adaptive responses before. the new biological information discovered by systems biology approaches helped to better understand the mechanism of action of the vaccine and can now be exploited to rationally design improved vaccine adjuvants targeting the protective genes or pathways (pulendran., 2010). however there are still many infectious diseases that cause millions of deaths every year worldwide. however there is great hope for the future that is based not only on new technologies available for vaccine development, but also in an increased ability to interrogate the human immune responses and integrate complex readouts through computational methods. we think that vaccinology of the future will be less empirical than it used to be and will learn more from human immunology. adjuvants and expression vectors will be selected based on the type of humoral and cellular immune responses which correlate with protection. a more rational vaccine design will hopefully allow the development of effective preventive vaccines for all remaining difficult targets including hiv, malaria and tuberculosis. | summaryconventional vaccines have been extremely successful in preventing infections by pathogens expressing relatively conserved antigens through antibodymediated effector mechanisms. thanks to vaccination some diseases have been eradicated and mortality due to infectious diseases has been significantly reduced. however, there are still many infections that are not preventable with vaccination, which represent a major cause of mortality worldwide. some of these infections are caused by pathogens with a high degree of antigen variability that can not be controlled only by antibodies, but require a mix of humoral and cellular immune responses. novel technologies for antigen discovery, expression and formulation allow now for the development of vaccines that can better cope with pathogen diversity and trigger multifunctional immune responses. in addition, the application of new genomic assays and systems biology approaches in human immunology can help to better identify vaccine correlates of protection. the availability of novel vaccine technologies, together with the knowledge of the distinct human immune responses that are required to prevent different types of infection, should help to rationally design effective vaccines where conventional approaches have failed. |
suicide is an important health challengeand one of the main leading causes of premature death worldwide (1, 2). suicide rate is among the top three causes of death in people aged 1544 yr old (3). the rate of suicide ideation is reported to be highest among elderly (1). completed suicide is 5% to 10% of suicide attempts including one attempt in every 3 and one death from suicide in every 40 sec (5). according to the report of iranian ministry of health and medical education in 2004, suicide is the 13 cause of death (6) and the second cause of death from external causes of morbidity(7), the rate of suicide attempt is 3 times in women, while, the rate of completed suicide is 4 times more common in men (8). among iranian population at 2012, 3216 suicide attempts occurred including 6 suicide attempts per 100,000 populations with the highest rate in the second decade of life (9, 10). classification is defined as an approach to determine a class for a new object applied using different methods such as data mining (machine learning) techniques (11). decision tree (dt), k - nearest neighbor, logistic regression (lr), naive bayes, c4.5, support vector machine (svm) and linear classifier are among conventional classification methods (1214). classification methodsinclude two main steps : first, a training sample of the dataset is determined randomly to find the model and the second step tests the resulted model (12). according to the kind of dataset, different methods result in different accuracy of prediction. the comparison among the methods can be applied using different criteria such as area under curve, which measures the accuracy of the prediction (13). among different classification methods, lr is the most popular predicting the presence or absence of an attribute using covariates. however, dt is preferable when there are predetermined set of attributes, the response is discrete and disjunctive and graphical results are required (15). artificial neural network (ann)as a non - linear, flexible, and general tool is capable of dealing with any sort of arbitrary function. support vector machine (svm)is a kind of generalized linear models with a classification decision according to the value of the linear combination of features(16, 17). this study aimed to determine factors putting people at a higher risk of completed suicide using different classification methods including lr, dt, ann and svm. we used the dataset of a study conducted to predict risk factors of completed suicide in hamadan province, the west of iran, in 2010 (18). the dataset was based on a large population survey conducted in 2010 where all cases of suicide occurring in hamadan province from apr 2008 to mar 2010 were enrolled. the presence of missing values was 17% in the dataset ; therefore, expectation maximization (em) algorithm was used for imputation. for this purpose, parameters in the equations imputed the missing values (expectation), then, parameters were updated using all observations including the imputed ones (maximization). this procedure ended at the convergence (19). to assess the fatal suicide, which is a binary variable ; using several risk factors, several classification methods were performed. the data needed to be divided into two sub - sets where training sample of the dataset finds the model and the testing sample tests the resulted model. the result derived from the learning sample (70% of cases) was then evaluated by utilizing the test sample (30% of cases). the applied methods were compared using sensitivity (se), specificity (sp), positive predicted value (ppv), negative predicted value (npv), accuracy (acc), and the area under curve (auc). logistic regression : lr is one of the most common applied classification methods in medical data analysis. the model can be written as : log(1)=+i=1kixi in this model, the xi s are the covariates to classify the response and the i s are the regression coefficients. the log it, log(1), indicates the odds ratio of classifying the response in category one than zero. artificial neural network : this method is an information - processing tool based on human brain performance. among different ann models, multilayer perceptron (mlp) is the most common used method, which includes layers as input, output, and hidden with nodes in each layer. an activation function transforms the data in each layer to the latter one by introducing a degree of non - linearity. input layer consist of all risk factors affecting the result of suicide, here including 6 variables. the response variable is shown in the output layer with two nodes as the possible outcomes for suicide attempts. to find the best performance of the network, a complicated nonlinear mapping between input and output layers is found using the number of nodes determined empirically in the hidden layer (20). support vector machine : a mapping function whether a classification or regression function is used in svms. to classify the result of suicide, a non - linear kernel function is used in order to transform the input data to a high - dimensional space where the input data can be separated as well. radial basis function (rbf) kernel consists of two parameters trading off misclassification of training sample against simplicity of the decision surface (cost parameter) and to evaluate the influence degree of a training sample. choosing the kernel function as well as the parameters, acclaims svm as a flexible method, which the ability of the user can make the results more appealing. using maximum - margin hyper planes, the classes will be best separated in the data. by contrasting two parallel hyper planes on each side of the separating hyper plane, the minimum generalization error will be achieved when the distance between the hyperplanes takes place (21). the dt can be applied when the aim of the research is to identify or discriminate high - risk subjects. the direction begins at the node and extends to the leaf, which connects the features. the tree is a disjunction of these connections and these disjunctions separate the branch population into sets with the same likelihood of events. at each stage, the graphical feature presentation makes ease of interpretation and allowing to different alternatives (15). to check the adequacy of the models, indices such as sensitivity, specificity, diagnostic accuracy (da), positive predictive value (ppv), negative predictive value (npv), and the area under curve (auc) were calculated using the observed data as the gold standard. the cochran - q test was used to check differences in proportion among methods. to assess the association between the observed and predicted values several statistics were measures such as coefficient, contingency coefficient, and kendall tau - b. the dt can be applied when the aim of the research is to identify or discriminate high - risk subjects. the direction begins at the node and extends to the leaf, which connects the features. the tree is a disjunction of these connections and these disjunctions separate the branch population into sets with the same likelihood of events. at each stage, the graphical feature presentation makes ease of interpretation and allowing to different alternatives (15). to check the adequacy of the models, indices such as sensitivity, specificity, diagnostic accuracy (da), positive predictive value (ppv), negative predictive value (npv), and the area under curve (auc) were calculated using the observed data as the gold standard. the cochran - q test was used to check differences in proportion among methods. to assess the association between the observed and predicted values several statistics were measures such as coefficient, contingency coefficient, and kendall tau - b. of 5414 people who attempted suicide 50.8% were male, 53.7% were married, 92.8% had no history of suicide, 47.3% and aged between 20 to 29 yr, 8.4% (457 subjects) died of suicide. the mean age of subjects was 26.3 yr (25.3 yr in females and 27.3 yr in males) ranged from 10 to 90 yr. to identify the risk factors affecting completed suicide, lr, svm, dt and ann were performed to the data. the test and train samples were composed of 1626 (30%) and 3788 (70%) cases, respectively. the factors such as gender, job, age, education, marital status, and history of suicide attempt were considered as the explanatory variables for the performed methods. completed suicide was significantly associated with gender (p<0.0001) and age (p<0.05) in the lr model. those aged between 2029 yr old was 3.14 times more likely to die from suicide than those aged 1019 yr (table 1). logistic regression model results among several ann models, the best model included one hidden layer and six hidden nodes. hyperbolic tangent and softmax were the activation functions for hidden and output layers, respectively. the importance of the variables is shown in fig. 1 presented by scores using sensitivity analysis. the normalized importance of the variables in decision tree and artificial neural network to perform the svm model, gaussian radial basis function was used as the best non - linear kernel function for classifying the successful attempts. this method showed a kernel parameter (sigma) of 0.24, a cost parameter of 5, and 2178 support vectors as the estimated parameters of the kernel function. in training sample, the weight assigned to the svm method was 11 for completed suicide and one for suicide attempt., the probability of completed suicide is presented according to the condition mentioned in its corresponding branch (fig. the classification tree with the probabilities of success for suicide attempts in each node a comparison of sensitivity, specificity, positive probability value, negative probability value, accuracy and the area under curve for training and testing sets of classification methods are shown in table 2 and fig. cochran - q test resulted in differences between proportions in different methods (p<0.001). multiple comparison adjusted for significance level was performed using mcnemar test which showed a significant difference in proportions of any two methods (p<0.001). to evaluate the association of the method predictions and observed value of suicide attempts, coefficient, contingency coefficient, and kendall tau - b were performed which resulted in the best performance of svm in comparison to others (table 3). the area under curve for the performed methods comparison of classification techniques lr : logistic regression, dt : decision tree, ann : artificial neural network, svm : support vector machine the association of performed methods with observed values lr : logistic regression, dt : decision tree, ann : artificial neural network, svm : support vector machine in this study, gender was recognized as a significant risk factor for predicting completed suicide so did for age, and educational level in different applied methods. despite not being significant, marital status and history of suicide were the less important variables predicting completed attempts in dt and ann. in a study, furthermore, age of 2130 yr was associated with the highest rate of completed suicide. classifying educational level into three categories (low - intermediate - high), the intermediate educated cases associated with the highest ratio of completed suicide. moreover, they showed that married cases were more prone to die comparing to the single people (22). in other study, men selected high - risk methods of suicide and suicide related mortality rate was higher in men (23). the rate of completed suicide was 2.5 in males compared to females. moreover, age, occupation, marital status, and educational levelwere reported as significant risk factor for completed suicide (24). age was affecting significant variable for suicide with the highest age - specific suicide after 45 yr old in both japan and south korea (25). marital status was a significant risk factor resulted from the odds of completed suicide 2.77 for married cases (26). this study showed that among four statistically different classification methods including svm, lr, dt and ann for this data, svm had the best performance in classifying the risk factors associated with completed suicide. in spite of the least sensitivity in the testing sample and presence of unbalanced data (8.3% fatality in training sample), svm had the outperformance among mentioned methods and indicated the highest association between svm predicted and the observed values as well as the highest accuracy. although, the assigned weight for the training sample was the best choice among all other assignments, the testing sample did not result in the same shape as the training sample because of different rates for fatal to non - fatal suicide attempts. to compare seven classification methods based on sample size and type of attributes, a sufficient number of records dt, svm, k - nearest neighborhood and c4.5 obtained a higher area under curve than lr, naive bayes, and linear classifier (27). in another study, several classification methods including linear discriminant analysis, logistic regression, neural networks, support vector machines, classification trees and random forests were used to predict the dementia. despite of the highest specificity and lowest sensitivity of svm, this method had the highest accuracy among all different methods (28). the functioning of svm based methods against ann assessed in a study of analytical chemistry. they recommended that the svm - based approach for practical application according to the robustness (29). conducting an empirical comparison between svm and ann, for classifying document - level sentiment, ann showed a better statistically significant prediction comparing to svm, even on the context of unbalanced data (30). finding predictive models for pre - operative diagnosis of rotator cuff tear, ann and lr were compared. the study resulted in a higher predictive accuracy of ann than lr (31). in a study, ann and dt an outperformance for ann was found compared to dt where the mean absolute errors for the former were less than the latter one (32). to classify the magnetic resonance imaging data in alzheimer s disease, different classification methods dt, ann, svm and orthogonal projections of latent structures (opls) were compared. although there was no statistical difference among several methods, svm and opls outperformed slightly than dt and ann (33). in a study that assessed differences between svm and lr, concluded that svm achieves a better performance in comparison to lr when fewer variables are included (34). to determine statistically the sex from craniometrists, three different methods lr, svm and linear discriminant analysis were compared. the study showed a better reliability existed for males than females using all the methods while the results for svm had to be developed. moreover, they found that lr was much more feasible than svm according to the choice about the kernel function and the parameters (35). predicting the hospital mortality in critically ill patients with hematological malignancies, svm and lr the comparing results were not statistically significant even though lr was resulted in a better predictive accuracy comparing to svm. moreover, to predict the model using svm, only 4 variables were needed, whereas this number was 7 and 8 for lr (36). despite its limitations such as missingness in the data imputed, this study compared four different methods suggesting svm as the best classifier model, which may help the policymakers determining suicide risk factors. svm had a better performance in classifying risk factors of completed suicide than other classification methods including dt, k - nearest neighbor, lr, naive bayes, c4.5, svm, and linear classifier. the flexibility of this method according to several choices for parameters and kernel function can make it as the first choice method for classification of such data. ethical issues (including plagiarism, informed consent, misconduct, data fabrication and/or falsification, double publication and/or submission, redundancy, etc.) have been completely observed by the authors. | background : we aimed to assess the high - risk group for suicide using different classification methods includinglogistic regression (lr), decision tree (dt), artificial neural network (ann), and support vector machine (svm).methods : we used the dataset of a study conducted to predict risk factors of completed suicide in hamadan province, the west of iran, in 2010. to evaluate the high - risk groups for suicide, lr, svm, dt and ann were performed. the applied methods were compared using sensitivity, specificity, positive predicted value, negative predicted value, accuracy and the area under curve. cochran - q test was implied to check differences in proportion among methods. to assess the association between the observed and predicted values, coefficient, contingency coefficient, and kendall tau - b were calculated.results:gender, age, and job were the most important risk factors for fatal suicide attempts in common for four methods. svm method showed the highest accuracy 0.68 and 0.67 for training and testing sample, respectively. however, this method resulted in the highest specificity (0.67 for training and 0.68 for testing sample) and the highest sensitivity for training sample (0.85), but the lowest sensitivity for the testing sample (0.53). cochran - q test resulted in differences between proportions in different methods (p<0.001). the association of svm predictions and observed values, coefficient, contingency coefficient, and kendall tau - b were 0.239, 0.232 and 0.239, respectively.conclusion:svm had the best performance to classify fatal suicide attempts comparing to dt, lr and ann. |
amyotrophic lateral sclerosis (als) is a common progressive, neurodegenerative disorder of the voluntary motor system that affects motor neurons in the cerebral cortex, brain stem, and spinal cord. the latter rapidly leads to pulmonary compromise requiring mechanical ventilation and represents the major cause of mortality. eighty - four percent of als patients die of respiratory complications and respiratory insufficiency in 2 - 3 years after diagnosis. on a retrospective chart review, it was found that 2.7% of patients with als had respiratory symptoms as their first symptom and that the average survival period of these patients was only 2 months [3, 4 ]. hence, reduced ventilation results, in part, from progressive motor neuron degeneration, which leads to respiratory muscle weakness. respiratory muscle weakness is defined as the inability of the respiratory muscles to generate normal levels of pressure and air flow during inspiration and expiration. this leads to respiratory insufficiency, which is defined as inadequate pulmonary ventilation to the point that gas exchange is impaired, resulting in carbon dioxide retention, hypoxemia, and respiratory failure [5, 6 ]. als patients are able to withstand respiratory muscle weakness by using other muscles at the initial stage, but the symptoms progress gradually to respiratory insufficiency and eventual respiratory failure. in the majority of cases, death is related to respiratory events. the time of progression and the degree of respiratory muscle weakness are, therefore, important prognostic factors for als patients. hypoventilation in als patients is due to respiratory muscle weakness and is associated with poor survival, cognitive impairment, and a poor quality of life ; this has led to an increase in concern about the respiratory problems faced by als patients and a dramatic increase of relevant knowledge. several studies have shown that treatment of als patients with noninvasive positive pressure ventilation (nippv) for respiratory insufficiency appears to prolong life expectancy and improves the quality of life ; this is possibly attributable to the slower rate of decline of pulmonary function in these patients. these recent studies have demonstrated reduced mortality in als patients with respiratory complications and prolonged the average survival period through an increase in the use of respiratory assisting devices for managing respiratory problems. however, there is no effective treatment for respiratory dysfunction in als patients so far. it has been shown in a recent study to be a safe and potentially effective intervention in patients with dyspnea, which is a major symptom of chronic obstructive pulmonary disease (copd). acupuncture has also been shown to be effective for symptoms in an animal model of als [9, 10 ]. korean sa - am acupuncture methods with lung tonification effects were chosen for this study after a review of the korean traditional literature. k - alsfrs - r scores, which are the main assessment tools of als, were used to analyze the relationship between the status of als patients and respiratory physiology parameters. the aim of this study was to report changes in etco2, spo2, rr, and pulse rate, after sa - am acupuncture treatment on als patients. this study was conducted at the wonkwang university als clinic from january through july, 2012. eighteen eligible als patients were selected out of all als patients admitted during that period. this study was approved by the institutional review board (irb), and written informed consent was obtained from all participants. the inclusion criteria were as follows : patients who (1) satisfied el escorial criteria and were diagnosed with als by emg, (2) signed a consent, (3) cooperated with this study, (4) had not exercised within the previous 24 h, (5) had not smoked, drank alcohol, coffee, or green tea within the previous 8 h, (6) had eaten at least 1 h prior to testing, and (7) were not menstruating. the exclusion criteria were as follows : patients who (1) needed intensive care for respiratory insufficiency, (2) were not able to give basic information owing to severe bulbar palsy, (3) had heart disease of ischemic or other etiology, (4) had endocrine disorders such as thyroid disease, (5) had renal diseases such as chronic renal failure, (6) had fever, (7) had a seizure disorder, (8) had mental illness, (9) were addicted to drugs such as alcohol, nicotine, or caffeine, and (10) were considered not eligible for this study at the discretion of the researcher. capnography & pulse oximetry (nonin medical, japan) was chosen because it is easy to handle and is a useful measuring tool for observing changes in a patient 's respiratory condition. this study measured end - tidal carbon dioxide (etco2), peripheral oxygen saturation (spo2), respiratory rate (rr), and pulse rate. the amyotrophic lateral sclerosis functional rating scale (alsfrs) is a validated, questionnaire - based scale that measures physical functional status in terms of the ability to carry out activities of daily living in patients with als. it has been used in clinical trials, as well as in clinical practice, because of its ease of use and its correlation both with objective measures of disease status and levels of disability. the score reflects the deterioration of function in the natural course of als, but may have a lower sensitivity in advanced disease stages. the scale was developed primarily to assess outcomes in pharmaceutical clinical trials and does not rely upon physical examinations or instruments [11, 13 ]. an initial imbalance within the scale that minimized the importance of respiratory function was rectified by a revision (als functioning rating scale, revised [alsfrs - r ]) to incorporate respiratory symptoms and assess the need for ventilation. the k - alsfrs - r was made to reflect its domestic applicability ; a preliminary experiment reported that it had high reliability and validity. it consists of 4 sections (verbal, detailed motor, gross motor, and respiration function) and 12 subsections ; the maximum possible score is 48 points. the greater the decrease in muscle function, the lower the score ; in other words, higher scores reflect better patient status. experimental procedure was as follows : (1) subjects were advised to rest and not to undertake strenuous exercise before measurement, (2) respiratory parameters (etco2, spo2, rr, and pulse rate) were measured using capnography and oximetry for 15 min, (3) sa - am acupuncture was conducted at specific acupoints using 0.25 40 mm, sterile, disposable acupuncture needles made of stainless steel (dongbang acupuncture inc. the depth of insertion at each point was predetermined to be within the normal range of 820 mm, depending on the location of the point. the sp3 and lu9 acupoint needles were electrically stimulated at 100 hz with the clinician adjusting the intensity so that the patients felt an uncomfortable sensation that was not painful ; (4) needles were kept in place for 15 min while measuring etco2, spo2, rr, and pulse rates simultaneously. each patient received acupuncture treatment twice a day for 5 days (figure 1). acupuncture points sp3, lu9, ht8, and lu10 (figure 2) were selected on both sides of the body according to sa - am five - element acupuncture method found in the traditional korean medical literature. these sets of acupuncture points are used for the tonification of lung functions. certified practitioners with at least 6 years of experience in traditional korean medicine, and an additional 3 years of clinical experience, performed the acupuncture treatment according to the who standard acupuncture point locations for the western pacific region. according to the korean sa - am acupuncture literature, sp3 (figure 2(a)) and lu9 (figure 2(b)) are used for the tonification of lung - qi, while ht8 (figure 2(c)) and lu10 (figure 2(d)) are used to clear lung fire. the selection of acupuncture points was based on the who standard acupuncture point guidelines. statistical analysis using spss version 20.0 for windows was used to compare changes in etco2, spo2, rr, and pulse rate. the relationship between k - alsfrs - r on one side and etco2, spo2, rr, and pulse rate on the other was analyzed with pearson 's correlation analysis. to compare the difference between parameters of respiratory function before and after acupuncture stimulation, a paired t - test was conducted using the mean value over a period of 15 min. a significance level of p 0.05) and respiratory rate (p = 0.180 > 0.05) after acupuncture stimulation. however, there was a significant difference between spo2 before and after acupuncture stimulation, with an increase from 95.42% to 95.58% (p = 0.002 < 0.05). there was a significant change in pulse rate after acupuncture stimulation, with a decrease from 82.49 bpm to 80.08 bpm (p < 0.001) (table 4, figure 5). als is a fatal and progressive neurodegenerative disease, leading to muscle weakness, paralysis, and death by respiratory failure. although respiratory failure is generally the cause of death in als, little is known about the treatment and control of respiratory problems. therefore, this pilot study was conducted to study the effect of acupuncture treatment on parameters of respiratory function with the help of capnography and oximetry to set up guidelines for preventing and managing respiratory problems in als. it reports several causes of wei symptoms and suggests a treatment, which is focused on making the digestive system healthy. therefore, als patients have received acupuncture treatment specifically at the spleen meridian (sp) and stomach meridian (st) in almost all case studies [19, 20 ]. sa - am acupuncture, an original and traditional korean acupuncture method, elicits a strong pain response when applied on the upper and lower extremities. sa - am acupuncture is widely used in korean clinical practice because it is considered safe and effective ; unfortunately, there has been little research on its use in als. according to this theory, the 4 causes of imbalances in meridian energies are deficiency, excess, cold, and heat. since there are 12 meridians with 4 possible imbalances, there are a total of 48 preestablished acupuncture prescriptions, each with 5 transport points to use to restore these imbalances. the 5 transport points are important acupoints of meridian that control the 5-phase qi of viscera and bowels. the acupoints of sp3, lu9, ht8, and lu10, which are referred to as lung tonification prescriptions, were selected as intensive acupuncture treatment in this study because respiratory muscle weakness and respiratory symptoms such as sputum production and shallow respiration are thought to be related to lung dysfunction. reported that shortness of breath improved by more than 50% after acupuncture treatment at ht8 and lu10 points on als patients. jiang. reported that electroacupuncture could be an effective anti - inflammatory treatment for the respiratory impairment that occurs in animal models of als. however, almost all of these studies were conducted with small sample sizes, and the patients received not only acupuncture but also various traditional treatments. moreover, some of these studies were performed on animal models of als and not on human patients. it is significant that the present study was conducted to observe the effect of acupuncture, with more patients recruited than ever before. it is known that etco2 exceeds 42 mm hg when severe bronchial obstruction exists. therefore, the control of etco2 was considered an appropriate marker of an improvement of respiratory function and the alleviation of respiratory symptoms. hypoxemia is defined as an spo2 equal to or less than 93%, lasting for 15 s or longer ; severe hypoxemia kills cells and suppresses mental activity, resulting in a comatose state and a reduction in the ability of muscles to perform work. dyspnea and hyperpnea are the most important symptoms of hypoxemia. a pulse rate of 60100 beats / min and respiratory rate of 1620 breaths / min a number of studies have shown impaired cardiac autonomic control in patients with als together with parasympathetic dysfunction and sympathetic predominance. disturbances in autonomic cardiac control in respirator - dependent patients with als may significantly influence survival and may lead to hypertensive crisis, circulatory collapse, and sudden death. observing and regulating etco2, spo2, respiratory rate, and pulse rate can, therefore, be an important process of care for als patients. analysis of k - alsfrs - r score and parameters of respiratory function measured before and after acupuncture treatment showed that patients with high scores in k - alsfrs - r had a greater difference in pulse rate before and after acupuncture stimulation. patients who recorded high scores in k - alsfrs - r showed a decrease in the values of etco2, rr, and pulse rate and an increase in the values of spo2 after acupuncture stimulation. overall results showed a high correlation with k - alsfrs - r score and the therapeutic result of sa - am acupuncture. there is rapid nerve degeneration as als progresses, therefore ; acupuncture stimulation did not affect patients who were affected more severely. it is possible that sa - am acupuncture would be more effective in the early stages of als. there was a significant change in pulse and spo2 after lung tonification by sa - am acupuncture. unfortunately, etco2 and rr showed no statistically significant changes despite a decrease at the time of measurement. throughout this study, the results showed sa - am lung tonification to be more effective in controlling inspiration rather than expiration. all patients who received sa - am lung tonification showed a decrease in pulse rate ; this suggests that sa - am acupuncture treatment may play an important role in stabilizing sympathetic nerves and regulating the autonomic nervous system. however, acupuncture treatment achieved only a slight improvement in respiratory parameters ; even then, it may help als patients to maintain respiratory function and retard the progression of respiratory muscle weakness. there were no side effects during the study ; nevertheless, the study had its limitations. als is a progressive and incurable disease, so it was not possible to distinguish between the treatment group and the nontreatment group for ethical reasons. it was also impossible to measure the effect of acupuncture treatment over long periods because these patients experience difficulty in breathing in the supine position. research on the role of acupuncture in alleviating respiratory symptoms in als patients provides basic data for preventing respiratory complications, which generally lead to death in als patients. ongoing research with the development and validation of new acupuncture treatment should continue in order to extend the life of als patients and improve their quality of life. sa - am acupuncture led to a statistically significant difference in pulse rate and spo2 after acupuncture stimulation. patients in the earlier stages of the disease with high k - alsfrs - r scores responded better to acupuncture treatment than did patients with lower k - alsfrs - r scores. this study needs to be taken further with a larger sample size to obtain more valuable and meaningful data. | respiratory dysfunction and complications are the most common causes of death in amyotrophic lateral sclerosis. this is a pilot study to observe the changes in respiratory physiology parameters after sa - am acupuncture treatment. eighteen als patients received sa - am acupuncture treatment twice a day for 5 days. the etco2, spo2, rr, and pulse rate were measured for 15 min before and during treatment, using capnography and oximetry. correlation of k - alsfrs - r scores against measured parameters showed that patients who had high k - alsfrs - r scores had greater changes in pulse rate after acupuncture stimulation ; they also showed a decrease in etco2, rr, and pulse rate and an increase in spo2. a comparison of the mean values of these different parameters before and after sa - am acupuncture stimulation revealed statistically significant differences (p < 0.05) in spo2 and pulse rate, but none in etco2 and rr. sa - am acupuncture treatment on als patients seems to be more effective in the early stages of the disease. in light of increased spo2 values, sa - am acupuncture appears to have a greater effect on inspiration rather than on expiration. as a pilot study of acupuncture on als patients, this study could be used as a basis for future research. |
technological advances in industrial societies have led to sedentary lifestyles at home and at work. inactivity and the excess intake of calorie - rich foods are the main culprits in the twin epidemics of obesity and type 2 diabetes - associated cardiovascular disease, disproportionately affecting women, especially those of african descent. skeletal muscle, the major site of glucose disposal, increases with body weight with steeper slopes of the regression lines in black women compared with white women and is associated with decreased insulin sensitivity. on the other hand, chronic elevation in insulin levels as a result of deceased insulin sensitivity may have direct effects on skeletal muscle mass by stimulating contractile protein synthesis, as has been shown in animal models, possibly due to structural homology with insulin - like growth factor. increased muscle mass -- especially in the lower extremities -- might be adaptive for accommodating an increased body mass during ambulation. increased muscle mass however, may be maladaptive if muscle function is compromised by lipid - induced interference with mitochondrial oxidative metabolism, with greater imbalance between myofibrillar contractile units and energy supply for activation. we addressed this hypothesis by : 1) analyzing cross - sectional insulin and muscle mass data from overweight women entering a worksite wellness program, and 2) determining changes in insulin and body composition following 6 months of participation in the diet and exercise program. as black women are reported to have greater skeletal muscle mass than white women in relation to body size, as a secondary analysis we examined racial differences in associations between extremity muscle mass and insulin. overweight (body mass index [bmi ] 25 to 30 kg / m) and obese (bmi 30 kg / m) non - diabetic (fasting glucose 0.790, p<0.001). despite increased total extremity muscle mass with greater obesity, both exercise duration (r= 0.360, p<0.001) and vo2 peak (r= 0.382, p<0.001) were inversely associated with total extremity muscle mass. total extremity muscle masses correlated positively with fasting insulin (figure 2) independent of age and height. significant correlations were also noted for lower (r= 0.328, p < 0.001) and upper (r=0.310, p < 0.001) extremity muscle masses analyzed separately. when fat mass was introduced into the multivariable regression model, the independence of insulin as a predictor of total extremity muscle mass for those with igf-1 measurements, there were no significant associations between igf-1 and total extremity muscle mass (r= 0.016, p=0.883). to evaluate the relationship between insulin and muscle mass prospectively, data were analyzed from 132 subjects in this cohort who completed the 6 month weight loss and exercise program. the group that completed the program did not differ from those who did not complete the program (n=65) in racial distribution (p=0.547). however, the group that did not complete the program tended to be younger (4410 vs. 4711 years, p=0.079) and were significantly more obese when entering the study (bmi 35.46.7 vs. 33.26.0 kg / m, p=0.016). additionally, those who did not complete the program trended to have higher fasting insulin values (8.97.3 vs. 6.85.5 u / ml, p=0.069). significant reductions in weight (2.54.3 kg, p<0.001) and fat mass (1.93.5 kg, p<0.001), and improvement in exercise duration (+ 4665 seconds, p<0.001) and vo2 peak (+ 1.53.2 ml o2/kg / min, p<0.001) were determined for subjects completing the program, although the range of measurements indicated increases in weight and fat mass in some subjects (table 1). reduction in fasting insulin from baseline to completion of the program was significantly associated with reductions in total extremity muscle mass (figure 3), largely due to correlation for change in lower extremity mass analyzed separately (r = 0.194, p=0.026). correlation between change in insulin and change in upper extremity mass was not significant (p=0.452). to consider the confounding effects of hormonal status and hormonal treatments on insulin and muscle mass, we analyzed primary outcomes data separately on 103 premenopausal subjects who were not on hormonal therapy. at baseline, higher level of fasting insulin was associated with greater total extremity muscle mass (r= 0.304, p=0.002). similarly, reduction in fasting insulin for those who completed the 6 month weight loss and exercise program was significantly associated with a reduction in total extremity muscle mass (r= 0.297, p=0.021). thus, baseline and outcomes with respect to insulin and muscle mass for premenopausal women not on hormonal treatment were similar to findings in the entire cohort. multivariable regression analysis with total extremity muscle mass as the dependent variable showed that change in insulin was a predictor of change in total extremity muscle mass (= 0.053, p=0.038), independent of changes in fat mass, exercise duration or vo2 peak(table 2). reduction in extremity muscle masses was not associated with diminished exercise performance by exercise duration (r= 0.176, p=0.045) or by vo2 peak (r=0.138, p=0.117). the principal finding of our study is that total extremity muscle mass was significantly associated with insulin levels in overweight or obese women, independent of age, body size or fat mass. this relation was present for load - bearing lower extremities as well as non - load - bearing upper extremities when analyzed separately. correlations were similar for black women compared with white women, with too few subjects in other racial or ethnic groups for comparison. in order to determine an independent contribution of insulin to muscle mass prospectively, subjects participated in a 6-month weight loss and exercise program at our institution, with the expected reduction in weight and fat mass and improvement in exercise performance achieved by the majority of participants who completed the program. reduction in fat and muscle masses achieved by our participants were not associated with improvement in insulin sensitivity by the minimal model, consistent with findings in women participating in the targeted risk reduction intervention through defined exercise (strride) trial despite reduction in thigh intermuscular adipose tissue. we found that reduction in insulin concentrations was significantly associated with reduction in total (primarily lower) extremity muscle mass, independent of fat mass loss and improvement in exercise performance and vo2 peak, and without interaction by race. thus, our data support the contribution of chronic hyperinsulinemia to increased extremity muscle mass in overweight or obese non - diabetic women that is partially reversible with reduction in insulin concentrations associated with weight loss and improved exercise performance. increases in extremity muscle mass and muscle have been reported in diabetics compared with controls, especially notable in older individuals, who in the non - diabetic state commonly lose muscle mass with aging (sarcopenia). thus, in the health, aging, and body composition (health abc) study 485 type 2 diabetics (including 212 women) aged 7079 years underwent dxa determination of extremity muscle mass, which was found to be 510% greater for both upper and lower extremities (for both men and women) compared with 2,132 non - diabetic controls of the same age range. albu reported that skeletal muscle mass measured by magnetic resonance imaging increased with body weight, with steeper slopes of the regression lines in black women compared with white women. increased skeletal muscle mass associated with obesity has been considered to be a result of generalized increase in body mass (including fat and bone). in addition, increased lower extremity skeletal muscle mass may compensate for increased fat mass in order to accommodate standing and ambulation. we considered possible effects of insulin on muscle as an additional contributor to muscle mass in our subjects, a hypothesis supported by animal studies. in humans, an increase in myosin heavy - chain mrna was noted after 3 hours of insulin infusion. in addition to stimulatory effects of insulin on glucose transport via irs-1/pi3-k / akt signaling, insulin also activates protein synthesis in skeletal muscle through signaling downstream of the mammalian target of rapamycin (mtor) associated with igf-1 receptor activation, especially following muscle loading in animal models. framingham investigators recently reported inverse correlations of igf-1 concentrations with age, bmi and insulin resistance by the homeostatic model assessment in men and women. we also found an inverse association of igf-1 levels with age, with a trend to an inverse association with fat mass in our subjects who underwent this testing, but not with insulin sensitivity assessed by the minimal model. thus, we find no evidence to suggest resistance to igf-1 signaling analogous to resistance to insulin signaling in obesity with a compensatory rise in insulin concentrations, and no reason to suspect that igf-1 is responsible for increases in extremity muscle mass measured in our subjects. our data are consistent with the possibility that with increasing obesity - associated insulin resistance, higher chronic insulin levels may stimulate muscle hypertrophy through increased protein synthesis. animal data suggest, however, that with increasing severity of obesity, resistance to insulin may ultimately develop in the mtor pathway, which may compromise further skeletal muscle hypertrophy in response to fat mass load, and even lead to muscle loss. our data further suggest that increased extremity muscle mass in overweight or obese non - diabetic women is partially reversible with reduction in insulin concentrations. the increase in muscle mass associated with obesity may not translate into improved exercise performance, in contrast to increases in muscle mass associated with strength and endurance training. investigators in the health abc study reported muscle strength per unit cross sectional area in upper and lower extremities was lower in older men and women with diabetes than those without diabetes despite greater extremity muscle masses. increased inter - and intramuscular fat -- reported to be greater in women (especially of african descent) than men --may not only contribute to insulin resistance but also compromise muscle bioenergetics. petersen reported that lipid content in the soleus muscle (measured by h magnetic resonance spectroscopy) was 80% higher and rates of mitochondrial phosphorylation (measured by p magnetic resonance spectroscopy) were 30% lower in insulin - resistant offspring of type 2 diabetic patients compared with insulin - sensitive control subjects. an additional effect of insulin that may contribute to poor exercise performance despite increased muscle mass is the experimental evidence in the rat model that insulin changes fiber composition in skeletal muscle to more fast - twitch, type iib fibers, which in humans is associated with low capillary and mitochondrial density and low oxidative capacity. muscle biopsy studies in insulin - resistant first - degree relatives of patients with adult - onset diabetes and in obese women suggest the same phenomenon. a predominance of type iib fibers may be deleterious to endurance - type activity necessary to improve exercise fitness with sufficient energy expenditure to achieve weight loss. in our study, there was a significant improvement in exercise performance for the total cohort. whether reduced insulin in obese subjects following weight loss has favorable effects on muscle fiber type conducive to endurance exercise is unknown and deserves further study. a limitation of this study is the cross - sectional design for some analyses that limits conclusions regarding cause and effect. in particular, the time - course of onset of increases in extremity muscle mass relative to increases in fat mass, development of insulin resistance and elevation in insulin levels can not be determined from our study. nonetheless, loss of muscle mass (including lower extremity muscle mass) with weight loss reported by others supports the compensatory nature of increased muscle mass. our data suggest that reduction in insulin might also lead to a reduced muscle mass, independent of fat loss, but without compromise to exercise performance. another potential limitation is that muscle mass determination in the extremities by dxa scanning might not accurately measure skeletal muscle. muscle mass measured in the thigh by dxa strongly correlated with skeletal muscle measured in the thigh by multi - slice ct (r=0.96) in a study by levine., but with a systematic overestimate of thigh muscle mass averaging 12%, partly explained by inclusion of skin mass as muscle mass in dxa measurements but not in ct muscle measurements. we are not aware of correlations between dxa measures of extremity muscle mass and either ct or mri measures of skeletal muscle in extreme obesity (bmi 40 kg / m), however, which comprised approximately 15% of our cohort. in conclusion, fasting hyperinsulinemia in overweight women is associated with increased total extremity muscle mass, which is partially reversible with reduction in insulin independent of fat mass loss or improvement in exercise performance and peak vo2 or with significant increase in insulin sensitivity, and consistent with stimulatory effects of insulin on skeletal muscle mass. | obesity disproportionately affects women, especially those of african descent, and is associated with increases in both fat and muscle masses. although increased extremity muscle mass may be compensatory to fat mass load, we propose that elevated insulin levels resulting from diminished insulin sensitivity may additionally contribute to extremity muscle mass in overweight or obese women. the following measurements were performed in 197 non - diabetic women (57% black, 35% white ; age 4611 years [meansd ], bmi range 25.0 to 57.7 kg / m2) : dual - energy x - ray absorptiometry for fat and extremity muscle masses ; exercise performance by duration and peak oxygen consumption (vo2 peak) during graded treadmill exercise ; fasting insulin and in 183 subjects insulin sensitivity index (si) calculated from the minimal model. si (range 0.5 to 14.1 liter / mu1min1) was negatively, and fasting insulin (range 1.9 to 35.6 u / ml) positively, associated with extremity muscle mass (both p<0.001), independent of age and height. sixty - seven percent of women completed 6 months of participation in a weight loss and exercise program : we found a significant association between reduction in fasting insulin and a decrease in extremity muscle mass (p=0.038), independent of reduction in fat mass or improvement in exercise performance by vo2 peak and exercise duration, and without association with change in si or interaction by race. thus, hyperinsulinemia in overweight or obese women is associated with increased extremity muscle mass, which is partially reversible with reduction in fasting insulin concentration, consistent with stimulatory effects of insulin on skeletal muscle. |
target of rapamycin (mtor) is an evolutionarily conserved serine / threonine kinase that regulates cell growth and division in response to energy levels, growth signals, and nutrients. control of mtor activity is critical for the cell since its dysregulation leads to cancer, metabolic disease, and diabetes. in cells, mtor exists as two structurally distinct complexes termed mtor complex 1 (mtorc1) and mtor complex 2 (mtorc2), each one with specificity for different sets of effectors. mtorc1 couples energy and nutrient abundance to cell growth and proliferation by balancing anabolic (protein synthesis and nutrient storage) and catabolic (autophagy and utilization of energy stores) processes. although different cellular locations for mtorc1 and mtorc2 have been reported (these studies were recently summarized in an excellent review by betz and hall), there is a consensus that the localization of mtorc1 to lysosomes is critical for its ability to sense and respond to variations in the levels of amino acids. when amino acid levels are high, mtorc1 is recruited to the lysosomal surface, where it is activated by the guanosine-5-triphosphate (gtp)-loaded form of the small gtpase rheb (ras homolog enriched in brain). the amino acid - dependent translocation of mtor to the lysosome requires active rag gtpases and a ragulator, a pentameric protein complex that anchors the rag gtpases to lysosomes [6 - 8 ]. the rag proteins function as heterodimers in which the active complex consists of gtp - bound raga or b complexed with guanosine diphosphate (gdp)-bound ragc or d. it has been proposed that amino acids trigger the gtp loading of raga / b proteins, thus promoting the binding to raptor and assembly of an activated mtorc1 complex. however, it is important to mention that this model was recently challenged by a study suggesting that the activation of mtorc1 is not dependent on the rag gtp charging. therefore, further studies will be required to solve this apparent discrepancy and fully characterize the mechanism of mtorc1 translocation to lysosomes upon amino acid stimulation. meanwhile, the activity of rheb is regulated by a complex consisting of tuberous sclerosis complex 1 (tsc1), tsc2, and tbc1 domain family member 7 (tbc1d7). this complex also localizes to lysosomes and functions as a gtpase - activating protein (gap) that inhibits the activity of rheb. in the presence of growth factors or insulin, tsc releases its inhibitory activity on rheb, thus allowing the activation of mtorc1. it is important to keep in mind that lysosomes do not simply serve as platforms for the proper assembly of the mtorc1 regulatory pathway. in the last few years, it has become apparent that the activities of mtorc1 and lysosomes are intimately interconnected. the level of amino acids inside the lumen of lysosomes directly modulates mtorc1 activity through the vacuolar h - adenosine triphosphatase (v - atpase). in addition, recent evidence suggests that mtorc1 may play a crucial role in lysosomal function by regulating biogenesis, distribution, and activity of lysosomes (figure 1). under nutrient - rich conditions (left panel), active mtorc1 promotes the retention of tfeb and tfe3 in the cytosol as well as inhibition of the ulk1/2 complex and the atp - sensitive na channel. in addition, lysosomes move toward the periphery of the cell. in contrast, under starvation conditions (right panel), the inactivation of mtorc1 leads to rapid translocation of tfeb and tfe3 to the nucleus (1), induction of lysosomal biogenesis (2), autophagy activation (3), and changes in lysosomal membrane potential (5). inactivation of mtorc1 might also be required to facilitate the fusion between autophagomes and lysosomes (4). after prolonged periods of starvation, reactivation of mtorc1 is critical to induce autophagic lysosomal reformation (6). abbreviations : mtorc1, mechanistic target of rapamycin (serine / threonine kinase) complex 1 ; rheb, ras homolog enriched in brain ; tfe3, transcription factor binding to ighm enhancer 3 ; tfeb, transcription factor eb ; ulk, uncoordinated 51-like kinase ; v - atpase, vacuolar h - adenosine triphosphatase ; zkscan3, zinc finger with krab and scan domains 3. however, recent evidence revealed that cells constantly monitor lysosomal function and possess the ability to increase the number and activity of lysosomes in response to their energetic or degradative needs. the transcription factor eb (tfeb), a member of the basic helix - loop - helix leucine zipper family of transcription factors, promotes expression of many lysosomal proteins and is considered a master regulator of lysosomal biogenesis. upon activation, tfeb binds directly to a 10-base pair motif (gtcacgtgac), known as coordinated lysosomal expression and regulation (clear) element, enriched in the promoter regions of numerous lysosomal genes, thus promoting their transcription. interestingly, the activation of tfeb is regulated by mtorc1 [16 - 18 ]. mtorc1-dependent phosphorylation of tfeb in serine 211 (s211) is particularly important, as this residue mediates the interaction between tfeb and the cytosolic chaperone 14 - 3 - 3 and causes retention of tfeb in the cytosol. following starvation and consequent inactivation of mtorc1, the tfeb/14 - 3 - 3 complex dissociates, allowing rapid transport of tfeb to the nucleus and tfeb - regulated expression of lysosomal genes. active tfeb also upregulates the expression of critical regulators of the autophagic process, including several proteins implicated in the formation of autophagosomes as well as proteins required for the fusion between autophagosomes and lysosomes. therefore, tfeb contributes to synchronize the two major cellular degradative systems : autophagy and lysosomes. the ability of mtorc1 to block autophagy initiation was well established but was thought to occur at the protein level. mtorc1 directly phosphorylates and inhibits key components of the uncoordinated 51-like kinase 1/2 (ulk1/ulk2) complex, required for autophagy initiation. however, the above - mentioned findings indicate that, by promoting the sequestration of tfeb in the cytosol, mtorc1 also modulates autophagy at the transcriptional level. finally, tfeb induces expression of critical regulators of lipid metabolism that facilitate the use of energy stores during prolonged periods of starvation and this is a role well conserved during evolution [22 - 24 ]. phosphorylation of tfeb by mtorc1 requires the recruitment of tfeb to lysosomes, the compartment where active mtorc1 resides. rags also bring follicullin (flcn) and tsc to lysosomes [13,26 - 28 ]. flcn functions as a ragc / d gap and is thought to be critical for mtorc1 reactivation when cells change from starvation to nutrient - rich conditions. therefore, rags play an important role as docking sites for recruiting mtorc1, mtorc1 effectors (tfeb), and mtorc1 regulators (flcn and tsc) to the lysosomal surface in a nutrient - dependent manner. interestingly, tfeb (as well as tfe3 ; see below) induces the expression of flcn, suggesting the presence of a regulatory loop in which tfeb may contribute to mtorc1 reactivation and therefore its own inhibition. tfeb belongs to the mitf / tfe family of transcription factors that includes mitf, tfe3, and transcription factor ec (tfec). tfe3 was previously implicated in the development of osteoclasts, activation of the immune system, and control of the allergic response. however, recent evidence showed that tfe3 also induces autophagy and lysosomal biogenesis in starved cells by binding to the clear elements present in the promoter regions of autophagic and lysosomal genes. at the same time, the transcription factor zkscan3 functions as a negative regulator of lysosomal biogenesis and autophagy in fed cells. interestingly, the nuclear localization of both, tfe3 and zkscan3, is directly regulated by mtorc1. overall, the emerging picture reveals a close collaboration between mtorc1 and several master regulators of autophagy and lysosomal biogenesis to facilitate an efficient response to the varying energetic demands of the cell. to maintain an efficient autophagic flux during starvation conditions, cells must couple formation and degradation of autophagosomes. for this, synthesis of autophagosomes ideally should be linked to increased autophagosome - lysosome fusion and increased lysosomal degradative activity. zhou and colleagues compared lysosomal function between fed and starved cells by measuring lysosomal acidification, cathepsin activity, and rate of proteolysis. they found that the activity of lysosomes was significantly augmented in starved cells, a factor that is probably critical to ensure efficient degradation of the autophagic content. increased lysosomal activity under starvation conditions required the inactivation of mtorc1, translocation of tfeb to the nucleus, and fusion of autophagosomes with lysosomes. early investigations revealed that the inhibition of protein synthesis by cycloheximide (chx) resulted in diminished lysosomal function. since chx causes a dramatic increase in the amount of intracellular amino acids, it is possible that the observed effect was due to chx - mediated activation of mtorc1. the requirement of autophagosome - lysosome fusion for efficient lysosomal degradation is very intriguing and suggests that autophagosomes might contribute important regulators of lysosomal activity. this idea is supported by the finding that tfeb - mediated lysosomal exocytosis is significantly reduced in autophagy - defective cells. the current model suggests that mtorc1 regulates lysosomal function by directly preventing autophagy and tfeb activation. under starvation, inactivation of mtorc1 in addition, tfeb is free to translocate to the nucleus and upregulate the expression of critical regulators that further enhance autophagic flux. tfeb also activates expression of genes that directly increase lysosomal activity, including several subunits of the v - atpase, lysosomal hydrolases, and receptors required for the delivery of those hydrolases to lysosomes. in addition, the inactivation of mtorc1 seems to be required to allow the fusion between autophagosomes and lysosomes. choi and colleagues (2012) recently identified a synthetic compound, mhy1485, which inhibits lysosomal fusion during starvation - induced autophagy by directly binding and activating mtorc1. mtorc1 might also regulate lysosomal function by directly modulating the activity of key lysosomal proteins. the v - atpase is a multisubunit proton pump composed of a v1 complex that catalyzes atp hydrolysis and a transmembrane vo complex that rotated upon atp hydrolysis. the best - characterized role of v - atpase is to pump protons inside the lumen of the lysosome, thus promoting lysosomal acidification. however, recent evidence suggests a novel function of v - atpase in nutrient sensing. when the level of amino acids in the lumen of lysosomes is low, the v - atpase interacts with the ragulator and prevents the activation of rag gtpases. in contrast, when amino acids are abundant, the v - atpase undergoes conformational changes that release the guanine nucleotide exchange factor (gef) activity of ragulator, leading to the activation of rag heterodimers and the recruitment of mtorc1 to lysosomes. although it is still undetermined whether the v - atpase is a direct sensor of amino acid levels or additional proteins are implicated in this process, it is clear that v - atpase plays a critical role linking amino acid levels to mtorc1 activation. open questions that remain unanswered include how the structural rearrangements undergone by the v - atpase in different nutrient conditions affect lysosomal acidification and whether mtorc1 directly modulates v - atpase activity as part of a regulatory feedback loop. in this regard, it was recently shown that mtorc1 is involved in controlling v - atpase assembly in dendritic cells. another interesting possibility was recently suggested by cang and colleagues. under nutrient- and atp - rich conditions, mtorc1 binds and inhibits the activity of an endolysosomal atp - sensitive na channel formed by the two - pore channels tpc1 and tpc2. when the levels of atp are reduced, something that occurs under starvation and other stress conditions like hypoxia, ischemia, or hyperosmotic stress, mtorc1 redistributes to the cytosol, allowing opening of the channel and the release of na and other ions from the lumen of the lysosome to the cytosol. the depolarization of the lysosomal membrane may potentially affect many different lysosomal parameters, including the luminal ph, fusion of lysosomes with autophagosomes, or transport of nutrients. in the presence of amino acids and growth factor, lysosomes tend to localize closer to the plasma membrane. it has been suggested that lysosomal positioning may have an important impact on mtor activity by regulating the proximity of the kinase to upstream signals. in contrast, the retrograde transport of lysosomes to the perinuclear area under starvation conditions is thought to be critical to facilitate fusion with autophagosomes. the rubinsztein group recently used different approaches to alter the distribution of lysosomes within cells. as expected, the localization of lysosomes in the periphery correlated with increased mtor activity, whereas the inhibition of lysosomal scattering resulted in diminished mtor activity and, consequently, increased number of autophagosomes. the rubinsztein group observed that, under starvation conditions, certain proteins implicated in the anterograde movement of lysosomes, such as kinesin kif2 (kinesin heavy chain member 2) and arl8b (adp - ribosylation factor - like 8b), dissociate from lysosomal membranes, thus facilitating the movement of lysosomes towards the center of the cell. the mechanism by which nutrients regulate lysosomal distribution is currently unknown, but, as mentioned in the previous section, it is possible that nutrient - dependent changes in lysosomal membrane potential or mtorc1 activity regulate the association of lysosomes with microtubules or specific motors. the catabolic activity of autophagy not only is critical to ensure cell survival when nutrients are limited but also regulates autophagy termination and lysosomal homeostasis. the lenardo group found that, after long periods of starvation, mtorc1 is reactivated. this is probably due to the degradation of the autophagosome content and consequent increase in the level of amino acids inside lysosomes. in agreement with this idea, for example, the depletion of spinster, a late endosomal / lysosomal sugar transporter, prevented mtorc1 reactivation and caused the accumulation of enlarged autolysosomes. the role of spinster in lysosomal homeostasis was further supported by a recent study showing that this protein regulates fertility and fat content of lipid droplets in caenorhabditis elegans by modulating lysosomal function and morphology. importantly, mtorc1 reactivation is required for the formation of nascent lysosomes from autophagosomes, a process known as autophagic lysosomal reformation (alr). in this process, long and stable tubules emanate from autolysosomes and eventually pinch off to form nascent lysosomes. over time, these proto - lysosomes become acidic and acquire degradative capacity, thus becoming mature lysosomes. the mechanism that orchestrates alr is very complex and requires the synthesis and accumulation of specific phosphatidylinositols in certain regions of the reformation tubules as well as recruitment of clathrin and clathrin adaptors ap2 and ap4. although it is still unclear how mtorc1 reactivation promotes alr, these findings reveal an important role of mtorc1 in the recycling of critical lysosomal components for which synthesis and transport are energetically demanding. reactivation of mtorc1 may also halt or at least slow down autophagy, thus preventing autophagic cell death. the participation of mtorc1 in alr may seem contradictory with its inhibitory role in lysosomal biogenesis and function. however, these observations probably reflect a broader role of mtorc1 in coordinating autophagy termination. it is also worth mentioning that alr was observed when autophagy was induced by serum and glutamine starvation. further studies will have to determine whether alr also occurs under other forms of starvation (for example, glucose or amino acid starvation). torc1 has been shown to mediate vacuolar fission (but not fusion). under nutrient restriction, torc1 inactivation could alter the fusion - fission equilibrium, resulting in an increase in the size and a reduction in the number of vacuolar (lysosomal) structures, as it has been observed in both yeast and mammalian cells. internalization of pathogens or live cells by phagocytosis or entosis, respectively, results in the formation of large macroendocytic vacuoles that fuse with lysosomes. upon degradation, nutrients and vacuolar components are recycled back to the lysosomal network by the mtorc1-regulated fission of the phagosome / entotic vesicle. although this recycling closely resembles alr, it is important to point out that there are important differences between the two processes. the two most remarkable are that the vesicles produced by phagosome and entotic vacuole fission contain luminal components and that inhibitors of mtorc1 do not completely block tubulation (although the vesicle shrinkage is significantly delayed). mtorc1 stimulates important anabolic processes, such as protein synthesis and the accumulation of energy stores, whereas lysosomes are critical mediators of catabolism. therefore, it is not surprising that the activities of mtorc1 and lysosomes are deeply interconnected. in the last few years, we have begun discerning the molecular mechanism that governs this connection. the emerging picture points to lysosomes as key regulators of nutrient signaling and energy homeostasis. however, many exciting questions still await clarification, including how the whole cell adapts to starvation conditions, the identification of novel regulators of the lysosomal / mtorc1 pathway, and the interplay between nutrient sensing and disease. | lysosomes are key cellular organelles that play a crucial role in catabolism by degrading extracellular and intracellular material. it is, therefore, very intriguing that mtorc1 (mechanistic target of rapamycin complex 1), a major promoter of anabolic processes, localizes in its active form to the surface of lysosomes. in recent years, many exciting observations have revealed a tightly regulated crosstalk between mtorc1 activity and lysosomal function. these findings highlight the complex regulatory network that modulates energy metabolism in cells. |
there are several medical and surgical [2, 3 ] treatment options for obstructive sleep apnea (osa). patients who use continuous positive airway pressure (cpap) devices have been shown to have nasal obstruction as a common complaint (estimated prevalence : 2545%) [46 ]. as described by poiseuille 's law, airflow resistance is proportional to the length and is inversely proportion to the radius to the fourth power. because the radius is such an important variable, small changes, such as a 10% increase in the cross - sectional area of the nasal cavity airway, can result in a 21% increase in airflow. although surgery on the nose has not been shown to dramatically improve osa, it can improve cpap device use. a recent systematic review and meta - analysis also demonstrated that isolated nasal surgery reduces cpap device therapeutic treatment pressures by 2 - 3 centimeters of water pressure (cwp). therefore, surgically increasing the size of the nasal airway decreases nasal resistance and reduces cpap device pressure requirements. however, to our knowledge, for patients who have not undergone nasal surgery it is unknown whether patients with smaller turbinates have lower cpap therapeutic treatment pressure requirements when compared to patients with larger turbinates. a recently published systematic review did not identify any study in the international literature that used inferior turbinate size as a variable in mathematical equations to predict cpap. for this study we hypothesized that, in patients who have not had nasal surgery, large turbinates would require higher cpap therapeutic treatment pressures than small inferior turbinates. because it has previously been shown that nasal surgery can reduce cpap therapeutic treatment pressures, we planned to exclude patients with prior nasal surgery in order to remove this confounding variable. the objective of this study is to evaluate the effect of turbinate sizes on the cpap titration based therapeutic treatment pressures (in centimeters of water pressure) for patients with osa who have not previously undergone nasal surgery. the stanford hospital and clinics institutional review board was contacted and written approval was granted prior to commencing this study. inclusion criteria are as follows : (1) stanford sleep medicine clinic patients who had a nasal examination and underwent an attended in - lab cpap titration study and (2) the nasal examination needed to include nasal septal deviation severity and inferior turbinate grades for the left and right sides separately. exclusion criteria are as follows : (1) patients who have undergone nasal surgery. the american academy of sleep medicine (aasm) manual for the scoring of sleep and associated events was used by stanford and outside institutions. the stanford hypopnea scoring criteria included 10 seconds with 30% reduction in airflow measured by the nasal flow transducer associated with a 3% desaturation and/or an electroencephalogram arousal as described in the aasm scoring manual 2013, version 2.0.2. in order to fully evaluate the effect of inferior turbinate size, a tool (inferior turbinate classification system, grades 1 to 4) this inferior turbinate classification system provides a method for grading the amount of airway space that the anterior aspect of the inferior turbinate occupies relative to the total available airway space and is summarized as follows : grade 1 is 025% of the total airway space, grade 2 is 2650% of the total airway space, grade 3 is 5175% of the total airway space, and grade 4 is 76100% of the total airway space ; see figure 1. the nasal obstruction symptom evaluation (nose) scale was used to evaluate nasal obstruction and a patient with a score > 40 was considered to have nasal obstruction. the data was cataloged using microsoft excel 2013 (redmond, wa, usa). the ibm statistical package for social sciences (spss) software version 20 (armonk, new york, usa) was used for statistical analyses. the patient data was analyzed by calculating the means, standard deviations (m sd), and 95% confidence intervals [95% ci ]. one - way analysis of variance (anova) was used to evaluate ordinal and nominal data ; spearman 's rank correlation coefficient (rs) was used for continuous data measures. the rs was selected for correlating variables because it is less sensitive to strong outliers and it can also be used for calculating correlation coefficients for both continuous and discrete variables. the standard recommendations for rs strengths were used : 0.00.19 = very weak, 0.200.39 = weak, 0.400.59 = moderate, 0.600.79 = strong, and 0.801.0 = very strong. variables evaluated included the cpap titration data, age, and body mass index (bmi) in kilograms per meter squared (kg / m), race / ethnicity, apnea - hypopnea index (ahi), oxygen desaturation index (odi), lowest oxygen saturation (lsat), inferior turbinate size, nasal septal deviation severity, and other physical exam findings. for cpap titration pressures, if a fixed pressure was prescribed, that value was used and if pressure ranges were prescribed, then the average of the pressure range was calculated and used as the cpap therapeutic treatment pressure for analysis purposes. the mean standard deviation (m sd) for age was 54.6 22.4 years and for body mass index was 28.5 5.9 kg / m. table 1 provides demographic information for the patients to include age, ahi, bmi, odi, lsat, nose scale scores, race information, nasal septal deviation severity, inferior turbinate size, and mask type. the spearman 's rank correlation coefficient (rs) between cpap therapeutic treatment pressures and several variables were calculated and were weakly correlated (age rs = 0.29, nasal obstruction rs = 0.30), moderately correlated (body mass index rs = 0.42 and lowest oxygen saturation rs = 0.47), or strongly correlated (apnea - hypopnea index rs = 0.60 and oxygen desaturation index (rs = 0.62)). no statistical significance was found with one - way analysis of variance (anova) between cpap therapeutic treatment pressures and inferior turbinate size (right turbinates p value = 0.2012, left turbinate p value = 0.3064), nasal septal deviation (p value = 0.4979), or mask type (p value = 0.5136) ; see table 2. the m sd for therapeutic cpap for grade 1 (five patients) : 12.8 2.5 cwp, grades > 1 to 2 (eleven patients) : 11.5 1.6 cwp, grades > 2 to 3 (twenty - one patients) : 11.3 1.8 cwp, and grades > 3 to 4 (eight patients) : 12.2 2.9 cwp, with a one - way anova p value of 0.4599 ; see table 3. mean diagnostic cpap titration based treatment pressure by inferior turbinate size (grades 14) was evaluated with multivariate analysis with the standard least squares linear regression model with an r = 0.08, p value = 0.9953 consistent with no association to very weak association ; see figure 2. among a subgroup analysis of patients without nasal obstruction (as evaluated by a nose scale score of 40 out of 100 or less, n = 34 patients) the m sd for age was 56.0 23.8 years, for body mass index was 28.4 6.6 kg / m, and for inferior turbinate size was 2.47 0.80. the m sd for cpap therapeutic treatment pressures for all 34 patients was 11.8 2.2 cwp, for grade 1 (four patients) : 13.3 2.7 cwp, grades > 1 to 2 (seven patients) : 11.6 1.7 cwp, grades > 2 to 3 (eighteen) : 11.5 1.8 cwp, and grades > 3 to 4 (five patients) 12.1 3.5 cwp, with a one - way anova p value of 0.5213 ; see table 3. for patients with complaints of nasal obstruction (11 patients) the m sd for age was 62.5 15.4 years, for body mass index was 29.2 3.5 kg / m, and for inferior turbinate size was 2.3 0.9. the m sd for cpap therapeutic treatment pressures for all 11 patients was 11.1 1.3 cwp, for grade 1 (one patient) : 11 cwp, grades > 1 to 2 (four patients) : 11.3 1.5 cwp, grades > 2 to 3 (three patients) : 10.0 0.0 cwp, and grades > 3 to 4 (three patients) 12.5 0.7 cwp, with a one - way anova p value of 0.4722 ; see table 3. there were three categories in the subanalysis for mask type : unknown mask types (7 patients), nasal masks (27 patients), and oronasal masks (11 patients). the m sd for cpap therapeutic treatment pressures for the seven patients with an unknown mask type was 11.3 1.7 cwp ; the m sd turbinate sizes were 2.66 0.30. the m sd for cpap therapeutic treatment pressures for twenty - seven patients with nasal masks was 11.8 2.3 cwp ; the m sd turbinate sizes were 2.41 0.96. for nasal masks, the one - way anova p value of 0.9217, see table 3. the m sd for cpap therapeutic treatment pressures for eleven patients with oronasal masks was 11.2 1.5 cwp ; the m sd for turbinate sizes was 2.45 0.82. for oronasal masks, the one - way anova p value of 0.2732, see table 3.. first, cpap therapeutic treatment pressures do not seem to be influenced by inferior turbinate sizes in patients who have not undergone nasal surgery. it has been shown that patients who have undergone nasal surgery will have a decrease in cpap therapeutic treatment pressures by approximately 2 - 3 centimeters of water pressure ; therefore, patients with prior nasal surgery were intentionally excluded from the study in order to eliminate this variable as a confounder. the mean diagnostic cpap and inferior turbinate sizes (grades 14) were evaluated with multivariate analysis with the standard least squares linear regression model with an r = 0.08, p value = 0.9953 consistent with no association to very weak association. there was a weak correlation between cpap therapeutic treatment pressures and nasal obstruction using the nose scale questionnaire (rs was 0.21, two - tailed p value 0.57, not statistically significant) and a very weak correlation for patients without nasal obstruction using the nose scale questionnaire (rs was 0.05, two - tailed p value 0.78, not statistically significant). given the lack of an association of the inferior turbinate sizes, nasal septal deviation severity, and nasal obstruction overall, these findings suggest that simply observing nasal abnormalities in a patient may not warrant surgery if they do not have complaints of nasal obstruction. another finding was that lowest oxygen saturation rs = 0.47 and body mass index (rs = 0.42) were moderately correlated. the moderate correlation with bmi is not unexpected as it is logical that a larger person would require more pressure than a thin person given the additional mass in the upper airway and in the abdomen. two variables, apnea - hypopnea index (rs = 0.60) and oxygen desaturation index (rs = 0.62), were strongly correlated, which is a logical finding given that a cpap titration is intended to reduce arousals and to improve oxygenation. additional research is needed in order to evaluate whether cpap therapeutic treatment pressures are truly independent of inferior turbinate sizes. as a retrospective case series utilizing chart review we would like to encourage researchers to incorporate and use the inferior turbinate classification system, grades 14 as it is a tool which has high intra- and interrater reliability. by using the classification system, the influence that the inferior turbinate sizes have as related to nasal obstruction and cpap can be more accurately ascertained. furthermore, despite the lack of an association between cpap therapeutic treatment pressures and inferior turbinate sizes in patients without nasal surgery we would still recommend that patients with nasal obstruction and large turbinates undergo turbinoplasties as several studies have demonstrated a quality of life benefit and improvement in cpap use and decreased cpap in patients who have undergone nasal surgery. additionally, to our knowledge, this study is the first to evaluate the association between inferior turbinate sizes and therapeutic cpap ; therefore, we caution against making generalizations. in order to increase the level of evidence once several studies have been published, a systematic review and meta - analysis would more accurately answer the question using statistical analysis with random effects modeling. first, we are limited to the constraints which are shared by all retrospective studies in that only the previously collected data could be utilized and analyzed ; therefore, if there are missing data, then patients may have had to be excluded solely based on the lack of documentation. second, in this study we did not review cpap device pressures for patients who had previous nasal surgery ; however, this was done intentionally given that a meta - analysis of eighteen studies demonstrated a reduction by 2 - 3 centimeters of water pressure after isolated nasal surgeries. third, given that we did not have rhinomanometry nor acoustic rhinometry, we were not able to evaluate the relationship between the data from these tools and the inferior turbinate sizes and nasal function as it relates to cpap ; future studies could evaluate these relationships. lastly, there was no rigid or flexible endoscopy performed in the assessment of these patients as the sleep medicine clinics do not have them available ; however, each patient underwent a nasal examination by the first author who is a board - certified otolaryngologist. in this study, cpap titration based therapeutic treatment pressures were not found to be associated with inferior turbinate sizes ; however, the cpap therapeutic treatment pressures were strongly correlated with apnea - hypopnea index and oxygen desaturation index. | objective. to evaluate the effect of turbinate sizes on the titrated continuous positive airway pressure (cpap) therapeutic treatment pressures for patients with obstructive sleep apnea (osa) who have not had nasal surgery. study design. retrospective case series. methods. a chart review was performed for 250 consecutive patients. results. 45 patients met inclusion criteria. the mean standard deviation (m sd) for age was 54.6 22.4 years and for body mass index was 28.5 5.9 kg / m2. the spearman 's rank correlation coefficient (rs) between cpap therapeutic treatment pressures and several variables were calculated and were weakly correlated (age rs = 0.29, nasal obstruction rs = 0.30), moderately correlated (body mass index rs = 0.42 and lowest oxygen saturation rs = 0.47), or strongly correlated (apnea - hypopnea index rs = 0.60 and oxygen desaturation index (rs = 0.62)). no statistical significance was found with one - way analysis of variance (anova) between cpap therapeutic treatment pressures and inferior turbinate size (right turbinates p value = 0.2012, left turbinate p value = 0.3064), nasal septal deviation (p value = 0.4979), or mask type (p value = 0.5136). conclusion. in this study, cpap titration based therapeutic treatment pressures were not found to be associated with inferior turbinate sizes ; however, the cpap therapeutic treatment pressures were strongly correlated with apnea - hypopnea index and oxygen desaturation index. |
in november 2009, a bat was found on the ground in bokeloh, lower saxony, germany (522513.99n ; 92331.56e). it was given mealworms and water ad libitum supplemented with minerals and vitamins. in february 2010, the bat began to act aggressively, directly approaching any moving object, vigorously trying to bite, and screaming ferociously. this agitated stage lasted for 7 days and was followed by general weakness, lethargy, and paralysis. after the first 3 days of the clinical course the bat was submitted for testing, and rabies diagnosis was performed by using immunohistochemical analysis. lyssavirus antigen was detected in numerous neurons of the cerebral cortex, cerebellum, and especially the nucleus funiculi lateralis and the nucleus olivaris of the medulla (figure 1). organs other than the central nervous system, e.g., the salivary glands, did not contain lyssavirus antigen. after the first cell passage in rabies tissue culture infection test (9), virus was isolated from brain tissue. antigenic typing performed with a panel of 10 antinucleocapsid monoclonal antibodies (10) clearly differentiated the isolated virus from all other tested lyssavirus species (table). immunohistochemical analysis of brain of natterer s bat for lyssavirus antigen by using the avidin biotin complex method. cytoplasmic granular - to - diffuse staining for rabies antigen is visible in the perikarya and neuronal processus. b) medulla and neurons of the nucleus funiculi lateralis showing strong cytoplasmic staining for rabies antigen. bblv, bokeloh bat lyssavirus ; rabv, rabies virus ; lbv, lagos bat virus ; mokv, mokola virus ; duvv, duvenhage virus ; eblv, european bat lyssavirus ; ablv, australian bat lyssavirus ; + + +, strongly positive ;, negative. results of discriminatory reverse transcription pcr results for eblv-1 and eblv-2 (11,12) were negative, and only a generic reverse transcription pcr (13) yielded a 605-bp amplification product similar to that of the positive control. the nucleotide sequence was determined by using standard methods (primers and protocols are available upon request). sequence analysis of the nucleoprotein gene performed with mega version 4.0 software (www.megasoftware.net/mega4/mega.html) showed that bblv differed from all other published lyssavirus sequences with the highest nucleotide identity to khuv (80%), followed by arav (79%), eblv-2 (79%), australia bat lyssavirus (77%), eblv-1 (77%), irkut virus (76%), shimoni virus (76%), rabv (73 - 75%) and duvenhage virus (75%). lagos bat virus (72 - 74%), mokola virus (72%), and west caucasian bat virus cbv (72%) showed the highest divergence to bblv. also, phylogenetic analysis based on concatenated n - p - m - g - l nucleotide sequences showed that bblv is most closely related to khuv, followed by eblv-2 (figure 2). phylogenetic tree inferred from concatenated n - p - m - g - l sequences of bat lyssaviruses. the neighbor - joining method (kimura 2-parameter) was used as implemented in mega4 software (www.megasoftware.net). rabv, rabies virus ; ablv, australian bat lyssavirus ; arav, aravan virus ; khuv, khujand virus ; bblv, bokeloh bat lyssavirus ; european bat lyssavirus ; irkv, irkut virus ; duvv, duvenhage virus ; shibv, shimoni bat virus ; lbv, lagos bat virus ; mokv, mokola virus ; wcbv, west caucasian bat virus. we report the discovery of a lyssavirus (designated as bblv) from a natterer s bat that died with rabies - like clinical signs. initially, a distinctive pattern in the reaction with a panel of antinucleocapsid monoclonal antibodies indicated the presence of an antigenically atypical isolate. the differentiation from other lyssavirus species was confirmed by phylogenetic analysis (figure 2). bblv is pathogenic because it caused a fatal disease in the natterer s bat that was similar to the clinical picture of rabies seen in other bats. viral antigen was present in many locations of the brain (figure 1) but surprisingly not in the salivary glands. since the exact date of infection is unknown, the incubation period can only be estimated as > 4 months. whether the natterer s bat is the natural reservoir species of bblv or whether it was bats (eblv-2 from m. daubentonii, m. dasycneme, khuv from m. mystacinus, and arav from m. blythii) indicating that myotis spp. if one considers the history of bat rabies in europe, it seems unlikely that bblv had spread from a distant origin into central europe or that the bat itself was translocated over long distances. the fact that bblv has been identified only in 1 bat is puzzling, considering the relatively high level of surveillance in germany. also, of 63 natterer s bats tested during 19992010 in a retrospective study, none tested positive for rabies (t. mller. germany is the only country where several bat species other than serotine bats, i.e., pipistrellus nathusii, pipistrellus pipistrellus, and plecotus auritus, have been found infected with eblv-1 during this study (t. mller. data), and this is the second discovery in recent years of a new lyssavirus species through routine passive bat rabies surveillance. also, in 2007 a daubenton s bat found on the ground was taken to a rehabilitation center, where it died and subsequently tested positive for eblv-2 (7). in both cases, the person who took care of the animal had completed the full preexposure vaccination, as a required risk - mitigating measure. an encounter with a bblv - infected natterer s bat could lead to a fatal outcome because bat lyssaviruses have caused several human cases of infection (4). in europe, species conservation and research require the handling of bats by bat workers. during 20002010, a total of 37,140 handlings were recorded for the natterer s bat (bat marking centre, saxon state office for environment and geology, dresden, germany), underlining the need for adequate prophylaxis for bat handlers. if one considers the close phylogenetic relationship between bblv and eblv-2 humans who receive rabies prophylaxis however, recent studies of the antigenic relationships of lyssaviruses have shown the difficulty of interpreting antigenic differences by using sequences alone (14). thus, in vitro and in vivo cross - neutralization and protection studies with current anti - rabv vaccines are urgently required for assessing the public health risk posed by this new lyssavirus. | a virus isolated from a natterer s bat (myotis nattererii) in germany was differentiated from other lyssaviruses on the basis of the reaction pattern of a panel of monoclonal antibodies. phylogenetic analysis supported the assumption that the isolated virus, bokeloh bat lyssavirus, may represent a new member of the genus lyssavirus. |
however, its administration is associated with problems, such as the adjustments needed to maintain an effective blood concentration. in addition, dietary restrictions are necessary, because vitamin k levels influence warfarin s efficacy. in recent years, novel oral anticoagulants (noacs) those have demonstrated encouraging outcomes, as well as effects comparable to those of warfarin. noacs are increasingly being used for primary and secondary prevention with respect to cardiogenic embolisms. with the aging of society, elderly patients suffering from atrial fibrillation make up a growing population. as a result, the prevention of cardiogenic embolisms is becoming a major issue for social and health economics. since warfarin has a long half - life, controlling intracranial bleeding is very difficult. there are some reports that the incidence of intracranial bleeding after noac administration is not as frequent as in patients treated with warfarin. other reports indicate that the clinical course following intracranial hemorrhage is better in noac patients than in warfarin patients. we investigated the progress and prognosis of cases at our institute in which conservative therapy was selected, with intracranial bleeding as a complication during noac administration. noacs were administered to 313 patients (dabigatran : 173, rivaroxaban : 140) at our institute between 2011 and july 2014. all patients were diagnosed with non - valvular atrial fibrillation and noac medication was started for the prevention of cardiogenic embolization. patients were allocated to dabigatran or rivaroxaban on the basis of their application time, medication compliance, and other factors. random assignment was not performed. among these patients, an 85-year - old man with a history of high - blood pressure, diabetes, and episodic atrial fibrillation (cardiac pacemaker) was staying at a health and welfare institution for the elderly because of dementia. he was taking a hypoglycemic agent, -inhibitor, anti - dementia drug, aspirin (100 mg per day), and dabigatran (220 mg per day). he was transported to our institute after the occurrence of left hemiparesis. on initial examination, his level of consciousness was 10 points on the japan coma scale (jcs 10) and pupil diameters were equal at 2.5 mm bilaterally ; however, right conjugate deviation was observed. paralysis was observed in the left upper and lower limbs, and he scored 3/5 on a manual muscle test. his renal function indicated a creatinine clearance (ccr) of 49.6 ml / min. an evaluation of the risk factors for cerebral stroke gave 3 points on the congestive heart failure, hypertension, age, diabetes mellitus, and stroke / tia (chads2) scale, and 4 points on the congestive heart failure, hypertension, age, diabetes mellitus, stroke / tia, vascular disease, age and sex category (cha2ds2-vasc) scale, making him eligible for anticoagulant therapy. at the same time, his risk factors for bleeding during anticoagulant therapy produced a hypertension, abnormal renal / liver function, stroke, bleeding history or predisposition, labile inr, elderly, drug / alcohol use (has - bled) score of 2 points. mild midline shift was observed on a head computed tomography (ct) scan as a complication of subcortical bleeding in the right frontal lobe (figure 1afigure 1case 1 : the patient s head computed tomography on admission (a) and 7 days later (b).). anticoagulant therapy (dabigatran) and antiplatelet therapy (aspirin) were discontinued and conservative therapy was applied. the level of consciousness shifted to jcs 3 without aggravation of the cerebral hemorrhage (figure 1b). the patient was transferred to a nursing home 10 months after the onset of cerebral hemorrhage and the discontinuation of anticoagulant therapy. case 1 : the patient s head computed tomography on admission (a) and 7 days later (b). an 81-year - old man with a history of cerebral infarction, diabetes, and atrial fibrillation was treated with insulin and rivaroxaban (10 mg per day). a residual disability caused by cerebral infarction left him requiring total assistance in his day - to - day living. he was admitted because of torso tilt and vomiting. on initial examination, his consciousness level was jcs 3 with no apparent immobility of the pupils, deviation, or paralysis of the four limbs. regarding risk factors for cerebral stroke, his chads2 score was 4 points and cha2ds2-vasc was 5 points. at the same time, the has - bled score was 2 points, with the risk of bleeding during anticoagulant therapy determined as moderate. a head ct scan indicated a left cerebellar hemorrhage (figure 2afigure 2case 2 : the patient s head computed tomography on admission (a) and 5 days later (b).). anticoagulant therapy the patient was transferred to a geriatric health services facility 3 months after the onset of cerebellar hemorrhage while still receiving no anticoagulant therapy. case 2 : the patient s head computed tomography on admission (a) and 5 days later (b). an 85-year - old man with a history of high - blood pressure, diabetes, and episodic atrial fibrillation (cardiac pacemaker) was staying at a health and welfare institution for the elderly because of dementia. he was taking a hypoglycemic agent, -inhibitor, anti - dementia drug, aspirin (100 mg per day), and dabigatran (220 mg per day). he was transported to our institute after the occurrence of left hemiparesis. on initial examination, his level of consciousness was 10 points on the japan coma scale (jcs 10) and pupil diameters were equal at 2.5 mm bilaterally ; however, right conjugate deviation was observed. paralysis was observed in the left upper and lower limbs, and he scored 3/5 on a manual muscle test. his renal function indicated a creatinine clearance (ccr) of 49.6 ml / min. an evaluation of the risk factors for cerebral stroke gave 3 points on the congestive heart failure, hypertension, age, diabetes mellitus, and stroke / tia (chads2) scale, and 4 points on the congestive heart failure, hypertension, age, diabetes mellitus, stroke / tia, vascular disease, age and sex category (cha2ds2-vasc) scale, making him eligible for anticoagulant therapy. at the same time, his risk factors for bleeding during anticoagulant therapy produced a hypertension, abnormal renal / liver function, stroke, bleeding history or predisposition, labile inr, elderly, drug / alcohol use (has - bled) score of 2 points. mild midline shift was observed on a head computed tomography (ct) scan as a complication of subcortical bleeding in the right frontal lobe (figure 1afigure 1case 1 : the patient s head computed tomography on admission (a) and 7 days later (b).). anticoagulant therapy (dabigatran) and antiplatelet therapy (aspirin) were discontinued and conservative therapy was applied. the level of consciousness shifted to jcs 3 without aggravation of the cerebral hemorrhage (figure 1b). the patient was transferred to a nursing home 10 months after the onset of cerebral hemorrhage and the discontinuation of anticoagulant therapy. case 1 : the patient s head computed tomography on admission (a) and 7 days later (b). an 81-year - old man with a history of cerebral infarction, diabetes, and atrial fibrillation was treated with insulin and rivaroxaban (10 mg per day). a residual disability caused by cerebral infarction left him requiring total assistance in his day - to - day living. he was admitted because of torso tilt and vomiting. on initial examination, his consciousness level was jcs 3 with no apparent immobility of the pupils, deviation, or paralysis of the four limbs. regarding risk factors for cerebral stroke, his chads2 score was 4 points and cha2ds2-vasc was 5 points. at the same time, the has - bled score was 2 points, with the risk of bleeding during anticoagulant therapy determined as moderate. a head ct scan indicated a left cerebellar hemorrhage (figure 2afigure 2case 2 : the patient s head computed tomography on admission (a) and 5 days later (b).). the patient was transferred to a geriatric health services facility 3 months after the onset of cerebellar hemorrhage while still receiving no anticoagulant therapy. case 2 : the patient s head computed tomography on admission (a) and 5 days later (b). among the 313 patients treated with noac at our institute between 2011 and july 2014 (dabigatran : 173, rivaroxaban : 140) the average age was 74.6 years, with 96 patients aged up to 70 years and 217 aged 71 years or older. regarding chads2, 54 patients scored 01 point, while 259 scored 2 points or more. the cha2ds2-vasc score was 01 point in 24 cases and 2 points or more in 289 cases. serious complications requiring discontinuation of anticoagulant therapy occurred in 9 cases, of which intracranial bleeding was observed in 2. in addition, recurrence of cerebral infarction was observed in 8 cases. the intracranial bleeding involved hemorrhage in the cerebral parenchyma (1 case of cerebral hemorrhage in the cerebrum and 1 case of cerebellar hemorrhage), with no cases of subdural bleeding. conservative therapy was selected in both intracranial hemorrhage cases ; no aggravation of hematoma was observed as a result of the antihypertensive therapy, the administration of a hemostatic drug, and the discontinuation of anticoagulants. i have described above two cases in which the administration of anticoagulants was indicated (chads2 and cha2ds2-vasc scores of 2 points or more) and intracranial hemorrhage was observed as a complication during noac administration. the bleeding risk in both cases was moderate (has - bled score 2 points). however, no increase in the quantity of hematoma or aggravation of the abnormal neurological findings were observed as a result of conservative therapy. whereas the half - life of warfarin in serum is considered to be 20 to 60 hours, the half - lives of 1217 hours for dabigatran and 59 hours for rivaroxaban mean that the anticoagulant effect of noac is likely to disappear within a very short period of time following withdrawal. in addition, in noac the mechanism of coagulation inhibition and hemostasis differs from that of warfarin, being limited to a single cause (figure 3figure 3mechanism of hemostasis and targets of warfarin and novel oral anticoagulants.). furthermore, the brain system is rich in tissue factor, which contributes to the activation of the extrinsic clotting system due to factor vii. unlike warfarin, the microenvironment of the hemorrhaged brain system contains abundant active factor vii in patients taking noac. therefore, the rapid functional restoration of the coagulation and hemostasis mechanisms occurs in parallel with the decline in blood concentration once the noac intake is stopped. thus, the increase in hematoma size can be controlled by rapid withdrawal alone, even after the onset of intracranial bleeding. a comparative outcome analysis has indicated that noac have a lower incidence of complications such as massive bleeding or intracranial bleeding. substantial anticoagulant effects in preventing cerebral infarction and systemic embolisms have been reported,. although intracranial bleeding during anticoagulant therapy is likely to become serious, no sudden increase in the hematoma is observed, because of the short half - lives of noac in blood compared to that of warfarin. in cases with a high anticoagulant therapy recommendation score (chads2 and cha2ds2-vasc) currently, in japan, the suitability of drugs is limited to inhibiting the occurrence of ischemic cerebral stroke and systemic embolisms in patients with non - valvular atrial fibrillation. based on our results and a previous report, noac are more likely than warfarin to inhibit an increase in severity at the onset of hemorrhagic complications. the has - bled score is usually used for the prediction of the risk of intracranial hemorrhage during anticoagulation therapy. however, a moderate has - bled score has not been established as a prognostic factor. the relationship between has - bled score and hematoma size or prognosis needs to be investigated by accumulating more patient cases and performing a statistical analysis., the number of elderly patients developing atrial fibrillation as underlying disease is expected to increase. accordingly, for the purposes of primary and secondary prevention, the expansion of the range of diseases suitable for anticoagulant therapy can not be avoided. against such a social background, it has been predicted that cases of intracranial bleeding during anticoagulant therapy will continue to increase in the future. although the use of noac may be limited by social and health economic backgrounds, along with the limitation of pharmaceutical registration, the types of disease suitable for anticoagulant treatment are likely to increase. since noac have the advantage of not worsening the severity of serious complications, they may be useful in the management of patients with atrial fibrillation. | objective : oral anticoagulants are widely administered to patients with atrial fibrillation in order to prevent the onset of cardiogenic embolisms. however, intracranial bleeding during anticoagulant therapy often leads to fatal outcomes. accordingly, the use of novel oral anticoagulants (noacs), which less frequently have intracranial bleeding as a complication, is expanding. a nationwide survey of intracranial bleeding and its prognosis in japan reported that intracranial bleeding of advanced severity was not common after noac administration. in this report, two cases from our institute are presented.patients : case 1 was an 85-year - old man with a right frontal lobe hemorrhage while under dabigatran therapy. case 2 was an 81-year - old man who had cerebellar hemorrhage while under rivaroxaban therapy.result : in both patients, the clinical course progressed without aggravation of bleeding or neurological abnormalities once anticoagulant therapy was discontinued.conclusion : these observations suggest that intracranial hemorrhage during noac therapy is easily controlled by discontinuation of the drug. noac administration may therefore be appropriate despite the risk of such severe complications. further case studies that include a subgroup analysis with respect to each noac or patient background will be required to establish appropriate guidelines for the prevention of cardiogenic embolisms in patients with atrial fibrillation. |
many people are not aware of the harmful effects of radiofrequency waves (rf) and their role in cancer and other serious risks. scientific evidence suggests that cancer is not only linked to mobile phone radiation and that other factors also may be involved in its development. most mobile operators use from radiofrequency waves in the range up 300 mhz to 3 ghz that can be harmful for human health (1). many scientific studies have been done on the radiobiological effects of rf waves, and most of them have reported the rare relationship between rf exposure and risks posed by mobile phones on the body in the last 15 years (2). however, they could lead to increased body temperature, especially in the head and neck, who have a low threshold dose and increases the probability of injury if there is long - term exposure to these waves. when using mobile phones, various factors should be considered, such as the duration, location, and method of use, in order to reduce the possible effects of exposure to radiation in the rf. because of the risk of mutation and sexual trauma and to prevent infertility due to the effect on male sexual cells, the mobile phone should be away from the waist. in this study, we measured the emitted dose from a radiofrequency device at different frequencies using a scintillation detector. the results of this study and international commission of non ionization radiation protection (icnirp) reports show that people who use cell phones more than 50 minutes a day face early dementia or other thermal damage due to the burning of glucose in the brain (3). currently, the ranges of subjects related to occupational and public exposure of electromagnetic fields have been increased. due to the possibility of adverse effects on human health, performance study of these devices there are two types of electromagnetic fields based on the frequency range, i.e., extremely low frequency (elf) and very low frequency (vlf) fields. elf and vlf fields frequency ranges are between 3 and 300 hz and 3 and 300 khz, respectively. due to the nature of static electricity, electric and magnetic fields at these frequencies operate separately from each other and will be measured in the different circumstances (5). electric and magnetic fields can be produced by carrying of electric current at any wiring or equipment, such as overhead or underground power lines, home wiring, medical equipment, and electronic devices (6). the most important factors that influence human exposure are power density of the frequency generator source, distance from the source, type and thickness of the exposed beams, and frequency (penetrated depth) of the rf input signal to the body (7). the high voltage power supply (hvps) that was used in this study was made based on the push - pull topology (48). it was designed and built for the single photon emission computed tomography(spect) system, and its performance was evaluted through simulation as well as experiment. the nominal frequency of shvps is about 31 khz that is achieved in half of the duty cycle. some module specifications, such as the maximum intensity of the magnetic field (bmax), output voltage (vo), output current (io), maximum switching frequency (fmax), reference voltage (vr), and bias voltage are bmax = 200 mt, vo = 2000 v, io = 20 ma, fmax = 60 khz, vr = 5.5 v, and vc = 12 v, respectively. figure 2 shows the prototype model of the constructed module. in switched mode power supplies, switching frequency interference results in a sudden flow of electric and magnetic fields. thus, to reduce the effects of interference, electromagnetic interference (emi) filters and rf shielding are used. the body exposure that is caused by rf waves of electric and magnetic fields, which varies with the intensity of the output signal usually energy measurement of transmitted rf signals that enters the body is not exactly possible. for quantification of the exposure effects on the human body, the term used is dosimetry (dosimetric quantities). other quantities related to the exposure of tissue include the electric field strength, the induced current in the body, and the rate of energy absorbed by the external field. the dosimetry role is to evaluate the amount of the induced electric fields in the body. also, the dosimetry term often is used to connect the body and the biological effects resulting from energy absorption of electromagnetic waves. to measure the absorbed energy in the body by rf wave that was emitted from the hv module, an alnor dosimeter was used in this study. usually, the exposure limit is expressed by specific absorption rate (sar), which is a measureable metric. sar is the time rate at which electromagnetic energy is absorbed in a biological tissue and can be expressed as (9, 10) : where is specific conductivity s m 1, erms is the effective amount of field in tissue, and is the density of the radiation - exposed tissue, which, for the head is 1.027 (gr cm). because the head is the most important member of the body, it is necessary to express the above parameters for this body part. for head limp at room temperature is 1.79 (s m), erms for this module at 1.5 m distance from the dosimeter (on the head) is 1500 (v m and density of it is 1.027 (gr cm). the penetration depth of the rf wave enters into biological tissues and the frequency of the emitted wave are inversely related. the electric field due to the rf input signal to the body will decrease after a distance from its original amount (griffiths, 1989), known as the skin depth (figure 3). the skin depth of each tissue type organ depends on the electrical permittivity and conductivity. where is the angular frequency,,, and are magnetic permeability of materials, conductivity (s m), and permeability, respectively. usually, in tissues has essentially the same value as in free space, 4 10 (h m). the relationship between skin depth of rf waves and frequency is shown in figure 4. the high voltage power supply (hvps) that was used in this study was made based on the push - pull topology (48). it was designed and built for the single photon emission computed tomography(spect) system, and its performance was evaluted through simulation as well as experiment. the nominal frequency of shvps is about 31 khz that is achieved in half of the duty cycle. some module specifications, such as the maximum intensity of the magnetic field (bmax), output voltage (vo), output current (io), maximum switching frequency (fmax), reference voltage (vr), and bias voltage are bmax = 200 mt, vo = 2000 v, io = 20 ma, fmax = 60 khz, vr = 5.5 v, and vc = 12 v, respectively. figure 2 shows the prototype model of the constructed module. in switched mode power supplies, switching frequency interference results in a sudden flow of electric and magnetic fields. thus, to reduce the effects of interference, electromagnetic interference (emi) filters and rf shielding are used. the body exposure that is caused by rf waves of electric and magnetic fields, which varies with the intensity of the output signal was determined. usually energy measurement of transmitted rf signals that enters the body is not exactly possible. for quantification of the exposure effects on the human body, the term used is dosimetry (dosimetric quantities). other quantities related to the exposure of tissue include the electric field strength, the induced current in the body, and the rate of energy absorbed by the external field. the dosimetry role is to evaluate the amount of the induced electric fields in the body. also, the dosimetry term often is used to connect the body and the biological effects resulting from energy absorption of electromagnetic waves. to measure the absorbed energy in the body by rf wave that was emitted from the hv module, an alnor dosimeter was used in this study. usually, the exposure limit is expressed by specific absorption rate (sar), which is a measureable metric. sar is the time rate at which electromagnetic energy is absorbed in a biological tissue and can be expressed as (9, 10) : where is specific conductivity s m 1, erms is the effective amount of field in tissue, and is the density of the radiation - exposed tissue, which, for the head is 1.027 (gr cm). because the head is the most important member of the body, it is necessary to express the above parameters for this body part. for head limp at room temperature is 1.79 (s m), erms for this module at 1.5 m distance from the dosimeter (on the head) is 1500 (v m and density of it is 1.027 (gr cm). the penetration depth of the rf wave enters into biological tissues and the frequency of the emitted wave are inversely related. the electric field due to the rf input signal to the body will decrease after a distance from its original amount (griffiths, 1989), known as the skin depth (figure 3). the skin depth of each tissue type organ depends on the electrical permittivity and conductivity. where is the angular frequency,,, and are magnetic permeability of materials, conductivity (s m), and permeability, respectively. usually, in tissues has essentially the same value as in free space, 4 10 (h m). the relationship between skin depth of rf waves and frequency is shown in figure 4. in table 1, the maximum allowable working with high frequency sources, according us federal communication commission (fcc), is shown. according to the first row of table 1, at the frequencies of 15 to 60 khz, the maximum work time for an operator should not be more than 6 min. to determine the intensity of magnetic field exposure limit values are effective values (rms) in this study, the value of b at 25 khz switching frequency is 2.4 mt. figure 4 demonstrated the dose rate received to dosimeter, at a distance of 1.5 meters from the module. the row data for calculation cumulative dose and relationship between the penetration depths of rf wave entrances to body with the frequency of emitted wave at different distances from module can be collected. as is clear from the results, the penetration depth of the wave rf and the frequency of the emitted wave is inversely : (fs). numerous epidemiological studies, the association between public and occupational exposure, particularly exposure to elf fields and the risk of cancer, including leukemia, brain tumors and breast cancer has shown (8). an internal em field is induced in the tissue when a biological tissue is exposed to rf waves (11). according to american conference of governmental industrial hygienists (acgih) and icnirp reports (7), the normal occupational exposure limit, for the whole body, not to exceed 60 mt or 600 g. occupational exposure limit values of the magnetic flux density of the magnetic field with a frequency range 30 khz and less than it, refers to a value that if employees are frequently faced with it, does not have any negative effect on their health. to determine the intensity of magnetic field exposure limit values are effective values (rms) occupational exposures in the range of extremely low frequency (elf) 1 hz to 300 hz, the amount shall not exceed the ceiling provided in the link below : in the above equation, the level of exposure by tesla (t) and f is the frequency in hz. occupational exposures in the frequency range of 300hz to 30 khz (including audio frequency band [vf ] from 300 hz to 3 khz and very low frequency band [vlf ] is the 3 khz to 30 khz) shall not exceed the ceiling 0.2 mt. exposure limit values for frequency of 300hz to 30 khz, including the body and is also part of the body. at frequency of 35 khz (nominal module frequency) maximum value of factor is equal to 19.9 m in the middle of the skull bone (1 to 7 mm) is much less, but the rf energy can penetrate into the brain in the area of the bone in place leaves. ncrp-86 reporting threshold of biological effects caused by occupational exposure to rf radiation absorption to 0.4 (w kg) determined according to the amount of 39.21 (w kg) of this module is to measure the duration of the ongoing work in the once the device should not exceed 15 min. also, in switching power supplies, there are long lines of communication between circuits components can reduce the effectiveness of high - frequency filters that this will lead to an increase in exposure. in general, considering that mental and psychological effects of these fields has been reported in small quantities, in order to control the possible harmful effects rf fields as possible from exposure. in this field, through the reduction of working time with a beam and the distance of the field with equipment distance and the training of personnel exposed to radiation, is prevented. | introductionpublic and occupational exposure to electromagnetic fields due to the growing trend of electronic devices may cause adverse effects on human health. this paper describes the risk of mutation and sexual trauma and infertility in masculine sexual cell by mobile phone radiations.methodsin this study, we measured the emitted dose from a radiofrequency device, such as switching high voltage at different frequencies using a scintillation detector. the switching high voltage power supply (hvps) was built for the single photon emission computed tomography (spect) system. for radiation dosimetry, we used an alnor scintillator that can measure gamma radiation. the simulation was performed by matlab software, and data from the international commission on non - ionizing radiation protection (icnirp) were used to verify the simulation.resultswe investigated the risks that result from the waves, according to a report by international commission on non ionizing radiation protection (icnirp), to every organ of the body is defined by the beam and electromagnetic radiation from this electronic device on people. the results showed that the maximum personal dose over a 15-min period working at the mentioned hvps did not exceed 0.31 sv / h (with an aluminum shield). so, according to other sources of radiation, continuous working time of the system should not be more than 10 hours. finally, a characteristic curve for secure working with modules at different frequencies was reported. the rf input signal to the body for maximum penetration depth () and electromagnetic energy absorption rate (sar) of biological tissue were obtained for each tissue.conclusionthe results of this study and international commission of non ionization radiation protection (icnirp) reports showed the people who spend more than 50 minutes a day using a cell phone could have early dementia or other thermal damage due to the burning of glucose in the brain. |
the transforming growth factor - beta (tgf) was discovered more than two decades ago and was isolated as a secreted factor from sarcoma virus - infected cells [13 ]. tgf was shown to transiently confer on normal fibroblasts phenotypic properties of transformed cells, as demonstrated by their acquired ability to grow in soft agar in an anchorage - independent manner. since then, more than 40 different family members have been identified, including the activin / inhibin subfamily, the bone morphogenetic proteins (bmps), nodal, myostatin, and the mullerian inhibitory substance (mis) [47 ]. as for the tgf subfamily, three distinct isoforms have been identified (tgf-1, -2, -3), each encoded by a different gene [4, 810 ]. of the three different types of tgf, which share around 70% homology within their sequence, tgf-1 has been the most studied isoform and will hereinafter be referred to as tgf. the active tgf molecule is a homodimer stabilized by hydrophobic interactions strengthened by a disulfide bond. this active form of tgf is synthesized from a large inactive precursor molecule, called latent tgf. as shown in figure 1, latent tgf is composed of a tgf dimer in a noncovalent complex with the tgf propeptide or latency - associated peptide (lap) that remains bound to tgf after secretion, retaining tgf in an inactive form and the latent tgf-binding protein (ltbp) which is linked to lap by a disulfide bond. this precursor molecule is stored in the extracellular matrix that acts as a reservoir for tgf. the activation of the tgf precursor is controlled by multiple processes, such as proteolytic enzymatic activity (furins, plasmin, calpain, etc.) but also acid, alkali, and heat - induced proteolysis. moreover, tgf can be activated by glycosidases, thrombospondin, and by some therapeutic molecules, such as antiestrogens and retinoic acid [13, 14 ]. the mature tgf is a homodimeric protein composed of two monomeric subunits linked by a single disulfide bond strengthened by some hydrophobic interactions. in 1982, massagu. identified a 60 kda high - affinity cell surface receptor (type i receptor) for tgf. subsequently, using affinity cross - linking approaches, other tgf receptors were discovered and identified (type ii and type iii receptors). following identification of its specific receptors, tgf was shown to control and modulate a plethora of biological effects, ranging from cell growth and differentiation, embryogenesis, hormonal synthesis and secretion, immunity, reproduction, bone formation, tissue remodeling and repair, and erythropoiesis, among others [5, 6, 8, 17, 18 ]. tgf and its receptors are widely expressed in all tissues and tgf signal transduction pathways play a major role in human diseases. indeed, while loss of function has been implicated in hyperproliferative disorders, tumor formation, inflammation, and autoimmune diseases, gain of function leads to immunosuppression and tumor metastasis [6, 9, 19, 20 ]. thus, tgf plays a dual role in human cancers, acting both as a tumor suppressor and as a promoter of tumor metastasis. the tumor suppressive effects of tgf, which include inhibition of cell proliferation, induction of apoptosis, and inhibition of cell immortalization, are observed in normal cells and early carcinomas. conversely, the tumor promoting effects of this growth factor, which include induction of epithelial - mesenchymal transition (emt), cell adhesion, migration, invasion, chemoattraction, and tumor metastasis, are more specifically observed in aggressive and invasive tumors [6, 2125 ]. in addition, as tumors grow and progress, they generally produce and secrete a large amount of autocrine tgf that is then released in the tumor vicinity. these increased tgf levels not only affect the tumor cells themselves but also the surrounding stroma by inhibiting cell adhesion, inducing immunosuppression and angiogenesis, and by promoting the degradation of the extracellular matrix, further contributing to the metastatic process. thus, the dual role played by tgf and particularly its prometastatic effects make it an attractive target for the development of novel therapies aimed at specifically blocking the pro - metastatic arm of its signaling pathway. tgf ligands interact with a complex of two transmembrane serine / threonine kinase receptors [5, 8, 27 ]. signaling starts with ligand binding to the extracellular domain of the type ii tgf receptor (trii), a constitutively autophosphorylated serine / threonine kinase receptor (figure 1). of the three tgf isoforms, tgf2 has the lowest affinity for the type ii receptor. as such, tgf2 requires binding to an accessory receptor (type iii receptor / betaglycan) first to efficiently bind trii. tgf binding to trii is followed by the recruitment of the type i into the complex receptor and its transphosphorylation by the trii kinase domain. tgf interacts with three distinct type i receptors, including the activin - like - kinase 1 (alk1), alk2 or alk5. of note, alk5 is the predominant form expressed in epithelial cells and is commonly referred as tri, being the preferred partner for tgf. given the dimeric nature of mature tgf, the resulting receptor complex is in fact a tetramer, composed of two molecules of trii associated with two molecules of type i receptor. phosphorylation of the type i receptors occurs mainly in the juxtamembrane region of the intracellular domain of the receptor, called the gs domain as it is rich in glycine and serine residues [5, 7 ]. the penultimate residue in the gs domain of the type i receptor, adjacent to the kinase domain, is always a threonine or a glutamine residue. mutation of this residue to aspartate or glutamate confers elevated kinase activity on the receptor in vitro and constitutive signaling activity in the cell, allowing the type i receptor to fully transmit signals in the absence of ligand or type ii receptor. the activated type i receptor is the main component of the tgf receptor complex and controls various downstream signaling pathways, including the canonical smad - dependent pathway as well as non - smad signaling mechanisms. the activated type i receptor recruits and phosphorylates the smad proteins, the main known effector molecules for these serine kinase receptors. the two receptor - regulated smads (r - smads), smad2 and smad3, are phosphorylated by the tgf and activin type i receptors (alk5 and alk4, resp.) on their c - terminal serine residues (sxs motif). while activin and tgf share the same r - smad signaling molecules and mostly signal through smad2 and smad3, other members of the tgf superfamily, such as the bmps, signal through distinct r - smad proteins (smad1, 5 and 8) following activation of their specific receptors by ligand binding (figure 1). once phosphorylated, smad2 and smad3 detach from the receptor complex and associate with the common partner smad4 within the cytoplasm [8, 3134 ]. thus, the activated smad complex is heterotrimeric, composed of two phospho - smad2 or phospho - smad3 moieties with a smad4 molecule. the smad complex is then translocated to the nucleus where it acts as a dna site - specific transcriptional regulator. nuclear translocation of the smad complex is regulated by both importin - dependent and -independent mechanisms [36, 37 ]. smad proteins recognize the dna sequence cagac, termed the smad binding element (sbe), as well as some gc rich sequences, but their affinity for dna is low. in order to achieve a high - affinity dna binding, the smads associate with various dna binding partners. the smads and associated cofactors bind in concert with their respective cognate recognition sites on dna, thus ensuring specific selection of the targeted gene promoters and of the tgf-mediated transcription response. these co - factors may be functionally expressed in different cell types thus providing another basis for tissue and cell type - specific functions for tgf ligands [40, 41 ]. furthermore, the smad complex associates with transcriptional coactivators or corepressors, resulting in the induction or repression, respectively, of a given tgf-smad target gene [42, 43 ]. while the smad pathway represents the canonical signaling pathway for tgf ligands, other intracellular signaling cascades have been shown to be activated in response to these ligands (figure 2). in particular, the stress - activated kinases p38 and jnk (jun n - terminal kinase) have been shown to be induced by tgf ligands and synergize with smad signaling to lead to apoptosis and epithelial - mesenchymal transition (emt) [4449 ]. the p38 kinase pathway also plays an important role downstream of activin signaling and was shown to be required for activin - mediated cell growth arrest in breast cancer and activin - mediated inhibition of human pit-1 gene expression in pituitary tumors. tgf can also signal through the mitogen activated protein kinase (mapk) pathway by activating the extracellular - signal - regulated kinases 1 and 2 (erk1 and erk2), further leading to the induction of emt [45, 50, 51 ]. rho gtpases have been shown to relay the tgf signals leading to cytoskeleton reorganization, cell motility, and invasion, through activation of rhoa, cdc42, and rac [52, 53 ]. finally, tgf was also shown to signal through the mtor and the phosphoinositide 3-kinase (pi3 k)/akt pathway to regulate cell growth inhibition and induction of emt [55, 56 ]. although mechanistically simple, the tgf / smad signaling cascade is regulated by multiple autoregulatory mechanisms that exist to maintain tightly regulated tgf-induced responses. the first tgf/smad inhibitory pathway that has been described involves a smad family member, smad7. the inhibitory smad7 functions through a negative feedback loop mechanism to terminate signaling by sterically preventing access of smad2/3 to the kinase domain of the type i receptor [57, 58 ]. in addition, smad7 also recruits protein phosphatases and ubiquitin ligases (smurf1/2) to the activated tgf receptor, further contributing to the termination of tgf signaling [5962 ]. the tgf receptors can be constitutively internalized by clathrin - independent or -dependent mechanisms, through the recruitment of endocytic adaptors like ap-2 and arrestins to the tgf receptor [64, 65 ]. at the smad level, nuclear translocated smad2 and smad3 can be subject to ubiquitin - proteasome - mediated degradation or dephosphorylation, leading to termination of signaling in both cases [6669 ]. cross - talk from other signaling pathways, such as epidermal growth factor (egf) and oncogenic ras signaling, which interfere with the nuclear translocation of the smads, can further act to negatively regulate smad - dependent signal transduction [7072 ]. while phosphorylation of the c - terminal mh2 domain of the smad by the type i receptor leads to activation of the r - smads, phosphorylation of the linker domain by various nonreceptor intracellular kinases inhibits smad signaling. as shown in figure 2, such kinases include the mapk kinases [70, 73 ], calcium - calmodulin - dependent protein kinase ii, cyclin - dependent kinase cdk2/4, casein kinase, protein kinase c, and g protein - coupled receptor kinase 2 (grk2). these kinases specifically target the linker region of the r - smads on multiple distinct serine and threonine residues, leading to termination of smad signaling. thus, the linker domain appears as a primary site for negative regulation of smad signaling. in the case of grk2, the kinase itself is regulated by tgf signaling and acts in a negative feedback loop. because grk2 plays a central role in modulating g - protein coupled receptor signaling, we also found tgf-induced grk2 expression antagonizes angiotensin ii - regulated vascular smooth muscle cell proliferation and migration. grk2 physically interacts with the mh1 and mh2 domains of the receptor - regulated smads and phosphorylates their linker region on a specific single serine / threonine residue. grk2-induced smad phosphorylation then leads to complete inhibition of tgf-induced smad activation, nuclear translocation, and target gene expression and inhibits the tgf antiproliferative and pro - apoptotic responses. thus, grk2 appears as a novel tgf antagonist that strongly inhibits cell growth arrest and apoptosis in both normal and cancer cells. interestingly, mutating the grk2 phosphorylation site within the smad linker domain to an aspartate residue to mimic a constitutively phosphorylated smad generates dominant negative smads that efficiently inhibit tgf responses. the most well - characterized effects of tgf are its tumor suppressor effects in epithelial, endothelial, myeloid, and lymphoid cell types. expression of the tgf type ii receptor (trii) in breast cancer cells prevents tumor formation, while inactivating mutations or overexpression of a dominant negative form of the receptor abolish tgf tumor suppressive effects and increase tumorigenicity [8284 ]. moreover, low expression levels of trii are correlated with more advanced and aggressive tumor stages, suggesting that the tgf signaling pathway acts as a tumor suppressor in the early stages of tumor development. as shown in figure 3, tgf exerts strong cytostatic effects on most of its target tissues by inhibiting cell cycle in the g1 phase. in addition, tgf induces apoptosis and prevents cell immortalization in numerous target tissues [4, 24, 48, 86 ]. cell cycle progression is controlled by intracellular protein kinases, called cyclin dependent kinases (cdks). once activated and associated to their regulatory cyclin subunits, the cdks induce gene transcription of a number of cell cycle regulators (dna polymerases, oncogenes, etc.), allowing for cell cycle progression from g1 to s phase. as shown in figure 3, tgf induces cell cycle arrest in g1 by inducing the expression of small inhibitory molecules, the cyclin - dependent kinase inhibitors (cdkis) p15 and/or p21, which in turn inhibit specific cdk activity. tgf-induced gene transcription of p15 and p21 is mediated by smad association with specific transcription factors, such as foxo forkhead and sp1 [90, 91 ]. p15 interacts with either cdk4 or cdk6 or with cdk4-cyclin d or cdk6-cyclin d complexes, while p21 interacts with cdk2-cyclin a or cdk2-cyclin e complexes. tgf-induced p15 expression leads to p15 binding to cdk4 and cdk6, blocking their association with their regulatory cyclins, thereby inhibiting their function and inducing g1 arrest. moreover, p15 binding to the cyclin d - cdk4/6 complexes also displaces p21 or the related p27 from these complexes, thus allowing these proteins to bind and inactivate cdk2-cyclin a and cdk2-cyclin e complexes [92, 93 ]. in addition, tgf represses the expression of growth promoting factors such as the oncogene c - myc [78, 94 ], and the i d family of helix - loop - helix transcription factors (id1, id2, and id3) [9597 ]. these proteins regulate angiogenesis, cell growth, and differentiation and are often upregulated in human cancer [95, 96, 98 ]. thus, inhibition of their expression by tgf largely contributes to this growth factor 's antiproliferative effect. tgf-mediated c - myc downregulation is mediated by a transcriptional regulatory complex including smad3, smad4, the repressor e2f4/5, and p107 and directly leads to cell growth arrest. interestingly, expression of the two cdkis p15 and p21 is normally restrained by the binding of c - myc and the zinc - finger protein miz1, in the proximal region of their promoters [100, 101 ]. inhibition of c - myc expression by tgf thus further contributes to increased expression of p15 and p21 and induction of g1 arrest. the i d proteins interact with the retinoblastoma tumor suppressive protein (prb) to promote cell proliferation and have been implicated in promoting tumorigenesis [96, 98 ]. id1 also delays cellular senescence in primary mammalian cells through inhibition of the cell cycle regulatory protein p16. tgf inhibits id1 expression in a smad3-dependent manner through induction of the activating transcription factor-3 (atf-3), a well - known id1 repressor. as c - myc binds the id2 gene promoter and activates id2 gene expression, its downregulation by tgf also leads to inhibition of id2 gene transcription [96, 97 ]. in ovarian cancer cells, we also found the engulfment protein gulp to act as a key regulator of tgf-mediated growth inhibition. finally, tgf was also shown to specifically inhibit expression of the tyrosine phosphatase cdc25a in normal mammary epithelial cells, by means of a smad3/e2f4/5/p130 inhibitory complex. thus, tgf-mediated inhibition of cdc25a expression allows for sustained cdk4/6 phosphorylation on their inhibitory sites and further induces cell cycle arrest (figure 3(a)). recent work from our laboratory has also revealed a role for the tumor suppressor menin downstream of tgf/smad signaling in pituitary adenoma cells [5, 104, 105 ]. pituitary adenomas are common monoclonal neoplasms accounting for approximately twenty percent of primary intracranial tumors with prolactin - secreting pituitary adenomas (prolactinomas) and are the most common form of pituitary tumors in humans [106, 107 ]. they are associated with very high levels of the hormone prolactin, exhibit increased tumor growth, and they give rise to severe endocrine disorders, including amenorrhea, infertility issues associated with galactorrhea in females, and impotence in males [106108 ]. multiple endocrine neoplasia type 1 (men1) is an autosomal dominant disorder characterized by endocrine tumours of the parathyroid, pancreatic islets, and anterior pituitary, particularly prolactinomas [5, 109 ]. we found menin to physically interact with smad3 in somatolactotrope cells and showed that inactivating menin expression antagonizes tgf signaling. we found that menin suppresses tgf-induced transcriptional activity by inhibiting the binding of the smads to dna. the role of menin is not restricted to tgf as we also found menin to be required for activin signaling in pituitary cells. results from our laboratory indicate that activin negatively regulates prolactin gene expression through reduction of pit-1 expression in a smad- and menin - dependent manner and that menin is required for activin - induced cell growth inhibition in somatolactotrope cells, highlighting a critical role for activin in mediating pituitary cell growth and pit-1/prolactin gene expression through the smads and menin [5, 110 ]. tgf stimulates cell death in various target tissues and these effects have been particularly well documented in the various epithelium, liver, and immune system [111116 ]. however, the molecular mechanisms and signaling pathways underlying these pro - apoptotic effects of tgf remain largely uncharacterized. several apoptotic regulators have been implicated downstream of the tgf signaling pathway, often in a cell- or tissue - specific manner. for instance, in hepatocarcinomas, the daxx adaptor protein couples the tgf signaling pathway to the cell death machinery through its interaction with the type ii tgf receptor (trii). interaction of the daxx protein to trii leads to its stabilization and further activation of the jnk and fas - mediated apoptotic pathways. in liver cancer, tgf also induces gene expression of the death - associated protein kinase dapk, which promotes cell death, in a smad - dependent manner, thereby linking the smad proteins to the mitochondrial proapoptotic processes. the tgf-inducible early - response gene (tieg1) is a krppel - like zinc finger transcription factor that mediates apoptosis in pancreatic epithelial cells. another mitochondrial protein that has been shown to mediate some of the tgf responses is the septin - like protein arts (apoptosis - related protein in the tgf signaling pathway), which can potentiate apoptosis induced by tgf, even in cells resistant to tgf-mediated cell death. the stress - activated protein kinase / c - jun n - terminal kinase (sapk / jnk) signaling pathway also plays a critical role in mediating tgf, through smad interaction with the activator protein ap1 [121, 122 ]. tgf causes smad- and the sapk / p38-dependent transcriptional induction of the pro - apoptotic bcl-2 family members bmf and bim, which in turn activate the pro - apoptotic factor bax that induces mitochondrial release of cytochrome c and activation of the apoptosome, leading to caspase - dependent apoptosis in hepatocytes and b - lymphocytes [123, 124 ]. conversely, the anti - apoptotic proteins bcl - xl and bcl-2 have been demonstrated to be down - regulated by tgf in various cell types [125128 ]. each of these signaling events eventually couples tgf to the cell death machinery, leading to changes of expression, localization, and activation of members of the bcl-2 family and caspases. interestingly, the pro - apoptotic effects of tgf are particularly strong in immune cells. work from our laboratory showed that in lymphocytes, both tgf and activin induce the expression of the src homology 2 domain - containing 5 inositol phosphatase (ship), leading to immune cell death. tgf-induced ship expression is smad - dependent and results in intracellular changes in the pool of phospholipids. upon tgf stimulation, the increased ship expression leads to decreased levels of second messenger pip3 (phosphatidyl inositol triphosphate), further contributing to inhibition of the akt survival pathway and resulting in cell death in both b and t lymphocytes. tgf also antagonizes survival signaling by inhibiting expression of the prosurvival protein survivin through the physical interaction of smad3 with akt, leading to apoptosis in colon cancer [129131 ]. in prostate epithelial cells, inhibition of survivin by tgf is smad - dependent and involves recruitment of a prb / e2f4 repressive complex to the survivin promoter. finally, we recently uncovered a central mechanism by which tgf induces apoptosis in both normal and cancer cells of various origins. indeed, we found tgf to increase expression of the transcription factor e2f1, further leading to the formation and binding of a transcriptionally active e2f1-prb - p / caf complex on multiple tgf pro - apoptotic target gene promoters, thereby activating their transcription and highlighting e2f1 as a central mediator of the tgf apoptotic program (figure 3(b)). normal cells are only able to replicate a defined number of times, called the hayflick limit, after which cells enter senescence and die. this limited number of replication cycles is due to the progressive shortening of the ends of the chromosomes, called telomeres, as dna polymerases fail to completely replicate genetic material at each cell division. as a result, after a number of cell divisions, the length of the telomeres shortens to a critical point, eventually leading to chromosome instability, senescence, and cell death. interestingly, cancer cells are not subjected to this limitation and thus achieve immortalization, due to the reactivation of an enzymatic program, the telomerase activity. in fact, elevated telomerase activity is so commonly observed in cancer cells that it is being used as a prognostic marker for cancer. telomerase is a reverse transcriptase that adds telomeric dna repeats at the end of chromosomes, thereby preventing their shortening. interestingly, tgf regulates the levels of human telomerase reverse transcriptase (htert), the protein component of the telomerase enzyme, by repressing its expression in normal and cancer cells [48, 135, 136 ]. this tgf-mediated repression of telomerase is smad3-specific and requires the transcription factor e2f1 as well as the stress - activated kinase and histone deacetylase activities (figure 3(c)). in summary, tgf acts as a tumor suppressor, acting through three different signaling arms (cell cycle inhibition, induction of apoptosis, and prevention of cell immortalization) and it is by the combined effects of these three separate signaling axes that tgf exerts its potent tumor suppressive effects in most cell types and tissues. the role of tgf as a potent tumor suppressor is further highlighted by the fact that many inactivating mutations in tgf receptors and smad genes have been found to be an underlying cause for human cancer [4, 9, 24, 137 ]. multiple genetic and epigenetic alterations of the tgf signaling pathway components have been reported to inhibit tgf tumor suppressive effects, thereby favoring tumor development. these are often found in human cancers of various origin (table 1) [4, 6, 9, 24, 137 ] and clearly illustrate the critical role played by the tgf signaling pathway in preventing tumor formation. mutations in either alleles of the tgf type ii receptor (t rii), leading to the formation of truncated or kinase inactive mutant forms of the receptor, are frequently found in colorectal, gastric, biliary, pulmonary, ovarian, esophageal, head and neck cancers, and gliomas [137, 138 ]. they also occur in other types of tumors, such as those of the endometrium, pancreas, liver, and breast cancers, though with a lower frequency. these inactivating mutations of trii are more frequently observed in tumors with microsatellite instability, due to mutations in mismatch repair genes. the type i tgf receptor (tri) also often harbors frameshift and missense mutations in ovarian, breast, esophageal, pancreatic, and head and neck cancers. epigenetic alterations of the tgf receptor genes, such as promoter hypermethylation or altered / defective expression of the transcription factors that regulate their expression, also lead to decreased receptor expression and inefficient receptor activity. like the tgf receptors, the genes coding for the smad proteins are often mutated or deleted in human cancers. smad mutations are due to loss of chromosome regions, deletions, frameshift mutations, nonsense, and missense mutations. of the three domains that compose the smad molecules, it is within the carboxy - terminal mh2 domain of the smads that these mutations preferentially occur [137, 140 ]. these mutations are mostly found in smad2 and smad4 and either prevent complex formation with the smad partners or block activation of smad - mediated gene transcription [139, 140 ]. as shown in table 1, the tumor suppressor smad4, also known as dpc4 (deleted in pancreatic cancer), is particularly affected by these genetic alterations, being mutated or deleted in no less than half of human pancreatic cancer, where it was originally characterized. since then, mutations in the smad4 gene have been characterized in other type of tumors (table 1). mutations in the smad2 gene also have a relatively high occurrence in lung, liver, and colorectal cancers [137, 142 ], while the rate of mutation in the smad3 gene is much lower. in fact, to date there are only few examples of such defects in smad3 expression, found in some gastric cancers and certain types of leukemia. finally, inhibition of smad expression in human tumors can also result from gene amplification of smad transcriptional repressors. in particular, the two smad repressors snon and ski are often found activated in in human cancers, highlighting their potent oncogenic properties. similarly, overexpression of the inhibitory smad family member, smad7 has been reported in several types of human cancers, including pancreatic, endometrial, and thyroid follicular tumors, resulting in inhibition of tgf/smad signaling. besides the known mutations in the tgf receptors and canonical smad pathway, other types of genetic alterations have also been reported to affect tgf signaling and tumor formation. for instance, oncogenic activation of the ras - raf - mapk pathway and c - jun nh2-terminal kinase in hepatocellular carcinoma has been reported to induce phosphorylation of the smad3 linker domain by mapk, further preventing c - terminal phosphorylation of the smad by the tri kinase domain and inhibiting tgf cytostatic effects. moreover, epigenetic alterations of other signaling components can also favor the tgf pro - metastatic effects. indeed, hypomethylation of the platelet - derived growth factor (pdgf) gene promotes glioblastoma cell proliferation in response to tgf. epigenetic downregulation of human disabled homolog 2 (dab2) also switches tgf from a tumor suppressor to a tumor promoter in head and neck carcinomas. finally, the sine oculis homeobox homolog 1 (six1) homeoprotein was also shown to induce human mammary carcinoma cells to undergo epithelial - mesenchymal transition and metastasis in mice through increasing tgf signaling. as described in the previous sections, the tumor suppressive role of tgf has been well described in multiple target tissues. interestingly, while tgf acts as a tumor suppressor in normal cells and early carcinoma, its cytostatic effects are often lost during the progression of the disease. indeed, as indicated above, many different human tumors are either resistant to the tgf cytostatic effects, due to genetic and epigenetic modifications in the tgf signaling components, or become resistant, due to the activation of prooncogenic signaling pathways (mapk, pi3 k, ras, c - myc), which then simply override any growth inhibitory signaling pathways, including tgf/smad [9, 20, 72, 151 ]. meanwhile, other tgf responses prevail, unrelated to the tgf cytostatic effects, which favor tumor progression and metastasis [6, 9, 20, 151 ]. these pro - metastatic effects of tgf were best characterized in breast cancer, the most common form of cancer in women in north america. in 2012, 1.8 million new cases of cancer will be diagnosed in the us and canada alone, including 250,000 new cases of breast cancer. for 2012 alone, the breast cancer - related death toll is estimated at 45,000 individuals in the us / canada (http://www.cancer.org/, http://www.cancer.ca/). there are several classifications of breast tumors, depending on the tumor size, the presence of tumor cells outside the primary site either in the lymph nodes or distant metastatic sites, and the growing speed rate of the cancer cells (established from biopsies). while the rate of remission and overall survival are high in the case of localized primary tumors, they are dramatically lower for metastatic tumors that have propagated to distant sites. the growth of these tumors is independent of hormonal and human epidermal growth factor receptor 2 (her2) levels. as a result, they usually are insensitive to hormone - based therapies (tamoxifen, aromatase inhibitors) and therapies targeting the her2 receptor (trastuzumab, herceptin). these aggressive metastatic tumors are responsible for the large majority of breast cancer - related deaths. although typically associated with the tgf cytostatic responses, the smad proteins are also critical for tgf-mediated tumor metastasis. indeed, expression of dominant negative smad3 or expression of a mutant form of the tgf type i receptor that fails to recruit the smad proteins, in human mammary epithelial cells, significantly diminished their ability to colonize the lungs [154, 155 ]. moreover, overexpression of the inhibitory smad7 impaired mammary carcinoma cell invasion. finally, gene silencing of the common partner smad4 in mda - mb231 invasive breast cancer cells impaired their ability to form osteolytic lesions and the formation of bone metastasis [157, 158 ]. as shown in figure 4, tgf plays a major role in promoting breast cancer migration, invasion, and metastasis by acting at various levels : (a) on the stroma and neighboring cells surrounding the tumor and (b) directly on the cancer cells themselves. these pro - metastatic responses of tgf include the ability to remodel the surrounding extracellular matrix (ecm), through stimulation of matrix metalloproteinase (mmp) expression and modulation of the plasminogen activation system, resulting in tgf-mediated matrix degradation and, consequently, an increasing release of stored tgf from the ecm that acts as a tgf reservoir. indeed, in many types of cancer, increased production of tgf correlates with higher tumor grade [159, 160 ]. increased tgf expression is observed in breast tumors and correlates with the aggressiveness and advanced stage of the tumor. interestingly, in mammary carcinoma patients, immunocytochemical analysis revealed that secreted tgf strongly localizes to the advancing edges of the primary tumor and to lymph node metastases [161, 162 ]. this tumor - derived tgf can exert autocrine effects, that is, effects on the tumor cells themselves, as well as paracrine effects on components of the tumor milieu such as stromal fibroblasts, endothelial cells, and immune cells. increased secretion of tgf affects and stimulates angiogenesis, contributes to myofibroblast differentiation and causes local and systemic immunosuppression, further contributing to tumor progression and metastasis [9, 20, 137, 151 ]. as outlined above, cancer cells themselves respond to and are affected by the increased tgf levels, which leads to phenotypic change of the cancer cells from epithelial to mesenchymal (emt transition), loss of polarity and adhesion, associated with increased migration and invasion cell properties, as well as increased chemoattraction to distant tissues (e.g., bone), thereby favoring and inducing metastatic development [9, 20 ]. thus, the tumor - permissive effects of tgf provide for a unique therapeutic opportunity in that specifically blocking this signaling network may interrupt mechanisms that are essential for tumor metastasis. however, it is important to note that because of the dual role played by tgf, acting as both tumor suppressor and tumor promoter, elucidation of the molecular events and components leading to both arms of tgf signaling will be critical to further design therapeutic strategies aimed at specifically blocking tgf-mediated tumor metastasis without affecting the tumor suppressive effects of this growth factor. elevated tgf levels released from the ecm reservoir exert a profound effect on the immune system. indeed, as mentioned above, tgf acts as a potent inducer of apoptosis in immune cells. through up - regulation of the lipid phosphatase ship and subsequent inhibition of the pi3kinase / akt survival pathway tgf directly targets cytotoxic t cell functions during tumor evasion of immune surveillance by suppressing production of cytolytic factors (pore - forming protein perforin, caspase - activating factors granzymes a and b, and pro - apoptotic cytokines fas - ligand and interferon). tgf also inhibits expression and activity of interleukin-2 and its receptors and blocks t lymphocyte stimulation by dendritic cells during an immune response. tgf inhibits proliferation and differentiation of t lymphocytes, lymphokine - activated killer cells, natural killer cells (nk), neutrophils, macrophages, and b cells. finally, tgf also decreases tumor cell surface immunogenicity by inhibiting expression of major histocompatibility complex class ii antigens, through a smad3-dependent mechanism [167169 ]. these immunosuppressive effects of tgf are best illustrated by the blockade of the tgf signaling cascade through overexpression of a dominant - negative receptor (dn - trii), which restored both cd8 and cd4 mediated immune response. thus, the increased concentration of released tgf in the tumor vicinity dramatically contributes to the tumor progression process, as local and systemic immunosuppression induced by tgf allows the tumor to escape host immunosurveillance [4, 6, 137 ]. the process of angiogenesis is essential to tumor growth as it allows blood vessels to deliver nutrients and oxygen to the tumor cells, and allows cancer cells that have detached from the primary tumor to reach and intravasate into the blood system. the tgf signaling pathway is a potent inducer of fibrosis and angiogenesis in vivo and promotes chemoattraction of angiogenic cytokine - secreting monocytes. tgf signaling in endothelial cells is rather complex as these cells express two tgf type i receptors (alk1 and alk5). the classic alk5-mediated pathway leads to smad2/3 activation, resulting in vessel maturation and angiogenic resolution, while alk1-mediated signaling antagonizes tgf/alk5 responses by inducing smad1/5 and generates transcriptional responses that are linked to angiogenesis [174177 ]. the tgf effects in angiogenesis are best illustrated by the multiple germline mutation mice models (ligand, receptors (i, ii and iii) and smad1/5) that all lead to vascular and endothelial cell defects [178185 ]. increased expression of tgf correlates with increased microvessel density and with poor prognosis in various tumor types, such as breast cancer and nonsmall cell lung carcinoma [186, 187 ]. tgf stimulates the expression of angiogenic factors such as vascular endothelial growth factor (vegf) and connective - tissue growth factors (ctgf) in epithelial cells and fibroblasts [188, 189 ]. tgf-induced expression, secretion, and activity of mmps also contribute to the dissolution of mature vessels around the tumor and the release of endothelial cells from the basement membrane, allowing them to further migrate and invade. moreover, tgf represses expression of angiopoietin-1, a critical factor in maintaining vessel integrity, in fibroblasts thereby contributing to the permeable properties of tumor - associated blood vessels. many recent studies have focused on the emerging role of tumor - stroma interactions, which are essential for supporting tumor progression. myofibroblasts, also known as cancer - associated fibroblasts (cafs), are mesenchymal cells harboring characteristics from both fibroblasts and smooth muscle cells. these cells can secrete numerous cytokines, growth factors, and ecm components and have the ability to substantially promote tumorigenesis and their appearance precedes the invasive stage of cancer. in a coimplantation breast tumor xenograft model, resident human mammary fibroblasts progressively converted into caf myofibroblasts during the course of tumor progression. during this process, they displayed increased autocrine signaling loops, mediated by tgf and sdf-1 cytokines, which acted in both auto - stimulatory and cross - communicating fashions. these autocrine - signaling loops initiated and maintained the differentiation of fibroblasts into myofibroblasts and the concurrent tumor - promoting phenotype. tgf also significantly increased the percent of myofibroblasts and invasion rate in caf cultures and increased the production and secretion of urokinase - type plasminogen activator (upa) by human breast myofibroblasts. thus, tgf induces the generation and maturation of myofibroblasts from precursor fibroblasts which, in turn, stimulate invasion of the tumor cells through secretion of proliferative, proinvasive and proangiogenic factors. as summarized in figure 4, tgf plays an important role in promoting tumor growth and development as well as tumor metastasis by allowing tumor cells to survive, detach and migrate away from the primary tumor to invade the surrounding tumor environment and metastasize to distant organs. the increased tgf levels produced by the tumor cells also contribute to the formation of a favorable microenvironment for tumor growth and spread by acting directly on the tumor cells themselves. tumor - produced tgf stimulates emt, cell migration, and invasion and promotes chemoattraction of the tumor cell towards distant organs (figure 4). the emt process characterizes the differentiation of highly organized and tightly connected networks of epithelial cells into disorganized and mobile mesenchymal cells with stem cell - like properties. the emt process involves a loss of cell - to - cell contact and the acquisition of fibroblastic characteristics by the epithelial cells as well as the acquisition of migratory and invasive properties by the cancer cells [196, 197 ]. emt drives and governs morphogenesis by inducing the differentiation of the epithelium into mesenchymal cell types to generate the different embryonic territories. the emt process is characterized by the dissolution of epithelial tight junctions and basolateral adherens junctions, resulting in the loss of epithelial cell polarity. this is best exemplified by the loss of epithelial gene expression (e - cadherin, zo-1, occludin, claudin, cytokeratins 8, 18, and 19, desmoplakin) and the induction of more mesenchymal markers (n - cadherin, vimentin, fibronectin, tenascin - c, and vitronectin). during emt, the actin cytoskeleton is also reorganized from a cortical adherens - associated location into actin stress fibers anchored to focal adhesion complexes that contribute to the formation of filopodia and promote cell migration. indeed, down - regulation of e - cadherin allows for the release of -catenin leading to increased expression of c - myc, cyclin d1, and mmp7, thereby promoting the invasive behavior of the cells. moreover, during emt, increased secretion of extracellular proteases and reduced expression of ecm proteins further contribute to cancer cell invasion. emt is under the control of several key transcription factors that regulate expression of mesenchymal markers and repression of epithelial genes. these include the zinc - finger proteins snail and slug, the basic helix - loop - helix factor twist, the zinc - finger / homeodomain proteins zeb-1 and -2, as well as the forkhead factor foxc3, which are all regulated and under the control of tgf. tgf induces reversible emt in both normal and cancer contexts [199, 200 ]. reciprocally, blocking tgf signaling by overexpression of a dominant - negative trii efficiently prevents skin squamous cancer cells from undergoing emt in vivo. interestingly, the tumor cells located at the invasion front, which contain high levels of tgf, show enhanced emt features. smad - dependent activation of transcription leads to expression of the emt regulatory factors, such as snail, slug, zeb-2, and twist through induction of the expression of high - mobility group a2 (hmga2) protein, resulting in repression of e - cadherin expression and dissociation of desmosomes. phosphorylation of the cell polarity protein par6 by trii also leads to the dissolution of cell junction complexes. while smad - dependent tgf-induced emt is enhanced by ras signaling, other smad - independent tgf downstream signaling pathways, including the ras / pi3 k [206208 ], rhoa, mtor [56, 209 ], erk mapk [50, 210212 ], and p38 stress - activated kinase pathways also contribute to tgf-induced emt. micrornas (small noncoding rnas) also play an important role in the regulation and maintenance of emt downstream of tgf [6, 213 ]. micrornas (mirnas) have eluded researchers for decades, stealthily regulating many of the major biological processes in eukaryotic cells. mirnas regulate gene expression posttranscriptionally by guiding the rna - induced silencing complex (risc) to their cognate site of the 3untranslated region (3utr) of the target mrna. while mirnas represent only 1% of all human genes, over a third of the transcriptome is regulated by these mirnas. individual mirnas can regulate hundreds of genes directly and thousands indirectly [215, 216 ]. by controlling and regulating the expression of so many genes, it clearly became apparent that mirnas play a central and critical role in the pathogenesis of human diseases, including cancer [217222 ]. moreover, about half of the mirna encoding genes are located in chromosomal regions that are being altered during tumorigenesis. numerous mirna signatures have been characterized in human cancers and implicated in the tumorigenic process [6, 224228 ]. while some mirnas exert their effects as classical oncogenes or tumor suppressors, others act in the advanced stages of the disease by promoting cancer progression and tumor metastasis [230233 ]. tgf represses expression of mir-200 leading to increased levels of the mir-200 target, zeb2/sip1. as zeb2/sip1 acts as the main repressor of e - cadherin expression, tgf-mediated down - regulation of mir-200 leads to decreased e - cadherin levels and emt in breast, pancreatic, and colorectal cancer. in turn, zeb2/sip1 targets tgf and mir-200 transcription in a feedforward loop which stabilizes emt. by inducing emt and modifying the cell phenotype, tgf alters cell - to - cell contact and communication as well as the adhesive, migratory and invasive properties of the tumor cells (figure 4). tgf-induced emt leads to changes in the expression profiles of adhesion molecules that profoundly diminish cell - to - cell and cell - to - substrate adhesion. these inhibitory effects on cell adhesion promote the detachment of the cancer cells from the primary tumor and their dissemination throughout the stroma. for instance, in the skin, melanocytes are tightly connected to keratinocytes through surface expression of e - cadherin. in melanoma, tgf-induced emt leads to downregulation of e - cadherin and alters the communication between keratinocytes and melanocytes and further allows melanoma cells to attach and communicate with fibroblasts from the stroma and endothelial cells, thereby favoring their propagation throughout the derma. in osteosarcomas, tgf inhibits cell adhesion to the substrate laminin, by down - regulating expression of the laminin receptor, 3 1 integrin. interestingly, tgf specifically inhibits cell interaction with laminin, as receptors for other substrates, such as collagen (2 1 integrin) and fibronectin (5 1 integrin) are not affected by tgf. a direct consequence of emt is the acquisition of migratory and invasive properties by the cancer cells. as mentioned above, tgf regulates the expression of several transcription factors such as hmga2, snail, slug, and twist during the emt process. expression of hmga2, snail or twist alone can induce emt and increase cell migration. overexpression of constitutively active tri restores cellular motility through the activation of pi3 k / akt and mapk pathways. in order to migrate, these events are coordinated by rho - family gtpases which are themselves activated by tgf. although the induction of rho signaling by tgf is not fully understood, it has recently been shown that rhoa activator is up - regulated by tgf in a smad4-dependent manner. while tgf exerts an important role in breast cancer progression as a pro - metastatic factor, notably through enhancement of cell migration, it is becoming clear that micrornas also play a crucial role in the mediation of these effects. for instance, mir-155 (which is regulated by tgf targets rhoa, thus directly contributing to epithelial plasticity. interestingly, we recently found tgf-mediated regulation of several micrornas to be critical for tgf-induced cell migration [242, 243 ]. in particular, our results highlight a novel signaling route whereby tgf silences expression of the microrna mir-584, further leading to actin re - arrangement and breast cancer cell migration. tgf down - regulation of mir-584 in breast cancer cells leads to increased expression of its downstream target, the actin - binding protein phactr1, resulting in enhanced cellular migration (figure 4). accordingly, over - expressing mir-584 or knocking down expression of the target phactr1 resulted in a drastic reorganization of the actin cytoskeleton and impaired tgf-induced cell migration. in addition to its promigratory role, tgf also contributes to the ecm remodeling and invasiveness of the cells by increasing the expression of metalloproteinases and the generation of plasmin which, in turn, contributes to the release of stored tgf from the ecm, further increasing cell invasion. the increased tgf levels allow cancer cells to cross through the ecm to reach distant metastatic sites. in invasive hepatocellular carcinomas, tgf induces transcriptional expression of 31-integrin, a key player in basement membrane invasion [245, 246 ]. moreover, tgf has been shown to inhibit expression of tissue inhibitor of metalloproteinase 3, timp3, further contributing to hepatocellular carcinoma cell invasion. though often associated together, tgf-induced emt can be dissociated from tgf-induced invasion and metastasis. indeed, using a skin carcinogenesis mouse model, it was found that tgf-mediated emt requires a functional tgf type ii receptor (trii), whereas tgf-mediated tumor invasion is associated with reduced trii signaling in tumor epithelia. we and others have demonstrated the mir-181 family of micrornas to be up - regulated by tgf and activin, a closely related tgf family member [243, 247, 249 ]. in hepatocellular carcinoma, we also found mir-181 to be a downstream regulator of activin / tgf-induced cellular migration and invasion in breast cancer (figure 4). as a critical regulator of tumor cell migration and invasion and breast cancer progression in vitro, mir-181 could potentially be an important therapeutic target. finally, a recent study from our laboratory identified p21cip (p21), a member of the core cell cycle machinery, as a key regulator of tgf-mediated breast cancer cell migration and invasion (figure 4). we found p21 expression to correlate with poor overall and distant metastasis - free survival in breast cancer patients. furthermore, using in vivo xenograft animal models, we found p21 to be essential for local tumor invasion. p21 interacts with smad3 and the acetyltransferase p / caf to regulate smad acetylation and transcriptional activity, as well as gene transcription of downstream tgf-induced pro - metastatic genes [250, 251 ]. our data also showed a significant association between tgf/smad3 signaling, p21, and p / caf expression with lymph node positivity, making them potential useful prognosis markers for lymph node metastasis. together these findings highlight an important role for the p21-p / caf - smad3 signaling axis in promoting breast cancer cell migration and invasion at the transcriptional level, and support the notion of a direct oncogenic role for p21 in the progression of breast cancer to a metastatic disease. while tgf directly contributes to local invasion, this is only the first event in a multistep process which will eventually lead to the formation and establishment of secondary tumors. as shown in figure 4, tgf also contributes to the establishment of metastasis by contributing to the growth of these secondary lesions. tumor cells that have migrated through and invaded the matrix can penetrate blood vessels, through a mechanism called intravasation. once in the circulation, tumor cells will then disseminate to distant sites and organs by exiting blood vessels (extravasation) and form new colonies in a new favorable distant microenvironment. tgf stimulates the secretion of osteolytic cytokines, which further contribute to the metastatic process by digesting the bone matrix. these events are smad - dependent as the formation of osteolytic lesions in mice by breast cancer, melanoma, and renal carcinoma cells can be blocked by overexpressing the inhibitory smad7 or a dominant negative tgf receptor [254256 ]. tgf stimulates the secretion of the parathyroid hormone related protein (pthrp) which promotes the differentiation of osteoclast precursors and bone resorption and induces expression of the bone homing receptor c - x - c chemokine receptor type 4 (cxcr4). association of the stroma - derived factor-1 (sdf-1) ligand to its receptor cxcr4 promotes chemoattraction of the breast cancer cells to the bone secondary sites [4, 258 ]. tgf also induces the expression of the interleukin proteins il-1, il-6, il-11, and connective tissue growth factor (ctgf), leading to osteoclastic differentiation and angiogenesis, and further contributing to bone resorption and the formation of osteolytic lesions [95, 158, 259261 ]. tgf-mediated il-11 and pthrp by the cancer cells leads to increased expression of receptor activator of nuclear factor kappa - b ligand (rankl) at the osteoclast cell surface, further enhancing progenitor cell differentiation into osteoclasts and bone demineralization [159, 160 ]. in addition to the development of bone osteolytic lesions, tgf also contributes to the development of metastases by directing cells to specific tissues and by enhancing the extravasation of breast cancer cells into the lung parenchyma, by inducing expression of cyclooxygenase-2 (cox2), epidermal growth factor receptor (egfr), and angiopoietin - like 4 (angptl4), and promoting development of lung metastasis. some of these genes (cox2 and egfr) have also been associated with brain metastasis. mammary gland growth and differentiation is a complex process regulated by steroids, polypeptide hormones, and growth factors, among which tgf and the hormone prolactin play major roles. while prolactin is required for lobuloalveolar formation and functional differentiation of mammary epithelial cells, tgf exerts an opposite effect, inducing apoptosis during mammary gland involution and inhibiting milk protein expression [265, 266 ]. prolactin signaling is mediated by the interaction of its specific receptor with the intracellular tyrosine kinase jak2 [267273 ]. once phosphorylated, tyrosine residues on both jak2 and the prolactin receptor create docking sites for the recruitment and activation of the transcription factor stat5 which will then activate gene transcription of target genes such as those encoding milk proteins and cell growth regulators [274276 ]. the importance of stat5 in mammary gland development is further highlighted by the stat5a knockout mouse which show no lobuloalveolar development during pregnancy and a complete absence of lactation. interestingly, an elegant study from the ali laboratory, revealed prolactin to also act as a suppressor of metastasis in breast cancer. indeed, prolactin and jak2 were shown to play a critical role in regulating epithelial - mesenchymal transition. activation of the prolactin / jak2 signaling pathway in mesenchymal - like breast cancer cells suppressed their mesenchymal properties and reduced their invasive behavior while blocking prolactin autocrine function in epithelial - like breast cancer cells induced mesenchymal - like phenotypic changes and enhanced their invasive capacity. interestingly, blocking prolactin signaling led to activation of the two major prometastatic pathways, the mitogen - activated protein kinase and the tgf/smad signaling pathways, highlighting prolactin as a critical regulator of epithelial plasticity and defining a new role for prolactin as an invasion suppressor hormone in breast cancer. tgf is expressed in each phase of postnatal mammary gland development and all three isoforms of tgf are up - regulated during mammary gland involution [265, 280, 281 ]. tgf inhibits alveolar formation, milk protein synthesis and induces apoptosis during involution of the mammary gland [282284 ], suggesting that tgf signaling may also antagonize prolactin - induced signals in mammary cells [285, 286 ]. in a recent study, we identified a novel antagonistic crosstalk mechanism by which tgf/smad signaling inhibits prolactin signaling and stat5-mediated gene transcription and mammary epithelial cell differentiation, by preventing stat5 binding to its coactivator cbp. these studies indicate that the prolactin and tgf signaling cascades oppose their effects not only to regulate differentiation of mammary epithelial cells and lactation but also to modulate tumor formation and breast cancer metastasis [278, 287 ]. as outlined in this paper, tgf tumor suppressive effects are often lost in aggressive tumors, while tumor promoting and pro - invasive responses remain and prevail, leading to the development of distant metastases. moreover, since tgf expression is increased in many cancers and correlates with the stage of the tumor [159, 160 ], blocking the tgf signaling pathway may provide for a unique therapeutic opportunity against tumor metastasis. as such, several approaches to develop new therapeutic tools that would interfere with the tgf pathway have been undertaken in recent years (figure 5) [6, 159, 160, 288291 ]. blocking antibodies against specific tgf isoforms, such as tgf-1 (metelimumab or cat-192 ; cambridge antibody technology and genzyme) and tgf-2 (lerdelimumab or cat-152 or trabio ; cambridge antibody technology) [292294 ] however, both proved unsuccessful in clinical trials and were subsequently discontinued [295298 ] (reviewed in). as all tgf isoforms influence tumors, pan - tgf antibodies are also being developed, as they may prove more efficient than isoform - specific antibodies [6, 288, 291, 299302 ]. targeting tgf signaling with soluble receptors has also been investigated, using exogenous expression of a soluble trii [303, 304 ], soluble recombinant triii (betaglycan), decorin, a proteoglycan induced by tgf [306308 ] or even a chimeric soluble receptor was constructed by fusing the extracellular domain of trii to the fc regions of human immunoglobulin igg1 (fc : trii or sr2f) [307, 308 ]. synthetic short peptides derived from tgf receptors that block tgf binding to its receptors are also an interesting avenue. one such promising candidate, p144 (digna biotech), inhibits tgf signaling and collagen type i synthesis in cardiac fibroblasts and potentially prevents myocardial fibrosis in hypertensive rats. antisense oligonucleotides (aso) are 1325-nt single - stranded nucleic acids, chemically modified or not, that are complementary to the target mrna [6, 310, 311 ]. the compound ap 11014, currently in advanced preclinical studies, specifically targets the tgf-1 mrna and has been shown to significantly reduce tgf-1 secretion in multiple cancer cell lines, impeding tgf-1-induced immunosuppression [6, 312, 313 ]. another compound, known as ap 12009 (or trabedersen ; antisense pharma), was designed to target the tgf-2 mrna and showed some efficacy in pancreatic cancer and glioblastoma [6, 302, 314317 ]. the development of tgf receptor kinase inhibitors has primarily focused on tri, due to extensive knowledge of the effect of this receptor on smad phosphorylation and since targeting tri would not disrupt potential tri - independent pathways initiated by trii [23, 296 ]. multiple tri kinase inhibitor compounds have been developed and tested and showed various levels of efficacy (reviewed in). some were tested in xenograft models in breast and non - small cell lung cancers and showed tumor growth delay in vivo. oral administration of ly2157299 with advanced malignancies was determined to be safe and well tolerated in a phase i clinical trial. moreover, sd-093 strongly inhibits the tgf-induced motility and invasiveness of pancreatic carcinoma cells and tgf-induced emt in mammary epithelial cells [321, 322 ]. sd-208 and sx-007 efficiently inhibit tgf-induced migration and invasion and prolonged survival in murine glioma tumors [323, 324 ]. sb-431542, a selective inhibitor of smad3 phosphorylation by tri, inhibits tgf-induced fibronectin and type i collagen synthesis in renal epithelial carcinoma cells. ly2109761 inhibits both smad - dependent and -independent tgf responses and attenuates tgf-induced cell migration, invasion, and tumorigenicity in colon adenocarcinoma and decreases liver metastases and prolonged survival in a murine pancreatic cancer model. one of the main concerns in targeted therapy is the off - target effects. because tgf exerts a dual role in cancer, targeted therapy to block tgf signaling raises a major concern. indeed, blocking the tgf pro - metastatic effects will only be beneficial if the therapy does not affect the tumor suppressive arm of tgf signaling. in a neu - driven breast cancer model, constitutively active tri increased the latency of the primary mammary tumor but also increased pulmonary metastasis whereas a dominant negative trii decreased the latency of the primary tumor but also significantly decreased the number of lung metastases. this finding indicates that although tgf acts as a tumor suppressor on the primary tumor, it may act on the ability of the breast cancer cells to extravasate from lung vessels to the parenchyma. although loss of trii correlates with poor prognosis in esophageal cancer and renal carcinoma, it also correlates with better survival rate in colon cancer and gastric cancer, clearly indicating that the role of trii in carcinogenesis may be stage and tissue specific. these potential risks give rise to the necessity to identify patients in which the pro - metastatic arm of the tgf signaling pathway is predominant. assessing the levels of tgf in the serum or the tumor has been studied as a tool for screening patients, showing a correlation between high serum levels and tumor progression and metastasis. many efforts have been made to target tgf in cancer due to its crucial role in cancer progression. several strategies have been developed and some molecules have shown encouraging and promising results. however, these strategies still have very important challenges to overcome. alternatively, new strategies aimed at specifically targeting the pro - metastatic arm of the tgf signaling pathway may prove more useful and safer. for this, identifying the downstream signaling components and elucidating the molecular mechanisms by which this growth factor promotes cell migration, invasion, emt, and metastasis will be critical for establishing successful specific therapies. for instance, rna interference approaches specifically targeting intracellular downstream molecules relaying these tumor promoting effects of tgf could prove useful. we recently found the cell cycle regulator, p21 to play a central role in the mediation of tgf-mediated local tumor cell invasion. using in vitro approaches and in vivo xenograft animal models, we found that blocking p21 expression with specific shrnas and sirnas could significantly alter the tgf tumor promoting effects, without affecting cell growth or tumor formation. thus, designing therapeutic strategies aiming at knocking down p21 expression in breast cancer patients may prove useful to prevent or circumvent the metastatic disease in this tissue (figure 5). indeed, while the tgf pro - metastatic effects have been relatively well characterized in breast cancer, the role of tgf in tumor cell invasion and metastasis in other tissues remain elusive. for instance, in melanoma, which is the leading cause of death due to cancer in young adults from 25 to 30 years old, the tgf effects on tumor progression remain unclear. while some reports suggested that tgf signaling could promote the metastatic potential of the cells [150, 217 ], other studies, including recent work from our laboratory, indicate that tgf potently inhibits cell migration and invasion of melanoma cells issued from various patients with different clinical backgrounds [333, 334 ]. thus, in this particular context, clinical strategies aiming at mimicking the tgf antiproliferative, antimigratory and anti - invasive effects may prove beneficial for the treatment of melanomas at different stages of their progression, including primary and metastatic tumors. tgf plays a major role in regulating cancer formation and progression. while acting as a tumor suppressor in normal cells and early carcinomas, tgf switches roles to in fact understanding and elucidating the intracellular and molecular mechanisms that trigger the tgf tumorigenic effects will thus be critical for the development of novel anticancer therapies based on the use of tgf antagonists. combined research from academia and industry has led to the development of such new therapeutic tools, some of which have demonstrated promising results. although the available tgf antagonists tested so far have shown some relative efficacy in different types of cancer, their use may also be limited. indeed, even though several antagonists are currently being tested in clinical trials, their long - term efficiency and potential adverse side effects remain to be determined. in particular, it is difficult to predict whether blocking all tgf effects, as with the current strategies, will allow for sufficient blockage of the pro - metastatic arm of the tgf pathway without affecting the tumor suppressive arm, thereby giving rise to spontaneous tumors elsewhere in the organism. in that respect, it will be vitally important to focus efforts on the development of novel strategies aimed at specifically manipulating the downstream signaling components of the tgf tumor promoting effects, as they may prove more effective and safer in the long run. | the transforming growth factor - beta (tgf) superfamily encompasses widespread and evolutionarily conserved polypeptide growth factors that regulate and orchestrate growth and differentiation in all cell types and tissues. while they regulate asymmetric cell division and cell fate determination during early development and embryogenesis, tgf family members play a major regulatory role in hormonal and immune responses, cell growth, cell death and cell immortalization, bone formation, tissue remodeling and repair, and erythropoiesis throughout adult life. the biological and physiological functions of tgf, the founding member of this family, and its receptors are of central importance to human diseases, particularly cancer. by regulating cell growth, death, and immortalization, tgf signaling pathways exert tumor suppressor effects in normal cells and early carcinomas. thus, it is not surprising that a high number of human tumors arise due to mutations or deletions in the genes coding for the various tgf signaling components. as tumors develop and progress, these protective and cytostatic effects of tgf are often lost. tgf signaling then switches to promote cancer progression, invasion, and tumor metastasis. the molecular mechanisms underlying this dual role of tgf in human cancer will be discussed in depth in this paper, and it will highlight the challenge and importance of developing novel therapeutic strategies specifically aimed at blocking the prometastatic arm of the tgf signaling pathway without affecting its tumor suppressive effects. |
intersphincteric resection (isr) is a procedure that aims to preserve anal function in patients with very low rectal cancer. since the introduction of laparoscopic rectal surgery for rectal cancer first described laparoscopic isr (lap isr) in 2000, and a number of cases have since been reported. we present our clinical experience with lap isr performed using needlescopic instruments in two patients with very low rectal cancer. the first port for the camera is placed at the umbilicus using the open technique. two 5-mm ports for the operator are inserted through the right upper and lower quadrants, and the upper port is placed where a diverting stoma would be created. two needlescopic forceps (endo - relief hope denshi co., chiba, japan) for the assistant are inserted into the left upper, and lower quadrants, such that a total of 5 ports are placed [figure 1 ]. the port placement (schema and external view) the endo - relief is a new type of needlescopic forceps. the tip of this new forceps has the same shape and size as that of the conventional 5-mm forceps ; however, the new forceps type has a shaft measuring 2.4 mm in diameter. during the operation, this needlescopic forceps is assembled from five parts, including the tip which has the same size and shape as the conventional 5-mm forceps and a shaft measuring 2.4 mm in diameter assembling and use this forceps as follows. the shaft guide plus (hope denshi co., chiba, japan) is directly inserted into the abdominal cavity through the abdominal wall.the top of the guide is passed through a 5-mm port extra - corporeally, and the inner needle is removed.the forceps end is connected to the shaft guide outer sheath.the handle parts are finally assembled, and the forceps top is then intra - corporeally pulled through a 5-mm port [figure 3 ]. the shaft guide plus (hope denshi co., chiba, japan) is directly inserted into the abdominal cavity through the abdominal wall. the top of the guide is passed through a 5-mm port extra - corporeally, and the inner needle is removed. the handle parts are finally assembled, and the forceps top is then intra - corporeally pulled through a 5-mm port [figure 3 ]. the shaft guide plus (hope denshi co., chiba, japan) is directly inserted into the abdominal cavity through the abdominal wall (a). the top of the guide is passed through a 5-mm port extra - corporeally, and the inner needle is removed (b). the forceps end (arrow) is connected to the shaft guide outer sheath (c). the handle parts are finally assembled, and the forceps top is intra - corporeally pulled through a 5-mm port (d) after dissection of the rectosigmoid mesentery at the level of the sacral promontory [figure 4 ], the lymph nodes around the inferior mesenteric artery (i m a) are dissected using the electrocautery device and laparoscopic coagulating shears. the i m a is divided above the left colic artery, after ligation with the clip, and the inferior mesenteric vein is divided at the same level. colonic mobilisation is performed using a medial to lateral approach, to preserve the ureter and the gonadal vessels. the mesorectum is carefully mobilized into the pelvis to preserve the hypogastric nerves and pelvic plexus. the needlescopic forceps inserted at the left upper quadrant is pulled cranially with the band or the gauze tied around the rectum circumferentially, and total mesorectal excision is carried down to the levator ani muscles. the puborectalis and the hiatal ligament are dissected to the intersphincteric plane, which is about 2 cm distal from the top of the hiatal ligament. the rectosigmoid mesentery is grasped with the end - relief, and then dissected using the electrocautery device (a). the puborectalis and the anococcygeal ligaments (hiatal ligament) are dissected to the intersphincteric plane, which is about 2 cm distal from the top of the anococcygeal ligament (arrow) (b) in the transanal dissection, the application of a self - holding retractor (lone star retractor cooper surgical inc., a circular incision of the mucosa and the internal anal sphincter is performed at the dentate line. a loop ileostomy is created at the right upper port site, and the drainage tube is inserted into the pelvis. after the operation, the only abdominal incisions are limited to the stoma site and the port sites [figure 5 ]. one patient was a 60-year - old woman with a body mass index (bmi) of 22.5. colonoscopy revealed a rectal tumour and the distal border was located 3.5 cm distal from the anal verge [figure 6 ]. colonoscopy revealed a rectal tumour and the distal border was located 4.5 cm distal from the anal verge [figure 6 ]. operative times were 384 min and 406 min, and blood losses were 10 ml and 55 ml, respectively. there were no intraoperative complications, and we were able to perform this procedure without the need for insertion of an additional port or changing the needlescopic forceps to conventional 5-mm forceps. the postoperative hospital stays were 14 days and 20 days, respectively, because both patients needed to acquire sufficient knowledge of stoma management. numbers of harvested lymph nodes were 7 and 16, respectively. however, there was no lymph node metastasis in either case. colonoscopy reveals a rectal tumour located 3.5 cm distal from the anal verge in a 60-year - old woman (a). a rectal tumour is located 4.5 cm distal from the anal verge in a 61-year - old man (b) intersphincteric resection is a sphincter - preserving procedure developed with the aim of avoiding the need for a permanent stoma in cases with very low rectal cancer, and was introduced by schiessel. with the spread of laparoscopic surgery to colon cancer lap isr was first described by watanabe. in 2000. since then, the feasibility of lap isr, in terms of complications, outcomes and anal functions, has been reported. however, the sample sizes in these reports are relatively small, and only three reports collected > 100 cases having undergone lap isr. first, this technique is often performed for low rectal cancer at stage 0/1 or clinical t1/t2. second, the lap isr procedure requires greater skill than laparoscopic low anterior resection. with lap isr procedures, it is not necessary to create an extraction site, because the resected specimen is extracted through the anus. furthermore, in many cases, a diverting ileostomy is created to avoid any potential risk for anastomotic leakage. in the recent literature, a diverting stoma was created in 88 - 100% of cases, the exception being the 11% described by park. the stoma site can be used for single - incision laparoscopic surgery (sils) placement to reduce the number of ports. reported that they created the sils port in the right lower abdomen, where a diverting stoma would be created, and the other two ports (10 mm port just above the umbilicus, 5 mm port at the left side of the abdomen). two ports were inserted through the sils port, such that 4 ports in total were placed. the concept of reduced port surgery includes not only sils, but also needlescopic surgery (ns). ns has been introduced for benign abdominal diseases, such as cholecystitis with gallbladder stones and appendicitis. in colorectal cancer surgery, ns is more advantageous as the natural orifice, that is, the rectum or the vagina, can be used for specimen extraction. however, ns is adapted for isr procedures in order to avoid the creation of an extraction site, because the surgical specimen is extracted through the anus. moreover, ns does not require expert techniques, as compared with the sils procedure. therefore, we apply needlescopic instruments with the end relief when performing lap isr. during the operation, there is no tissue damage due to grasping the mesorectum or rectum with the end relief. because the needlescopic forceps tip has the same shape and size as those of a conventional 5-mm forceps, the risk of tissue injury with grasping and lifting the end relief may have a particular advantage for laparoscopic colorectal surgery, because this procedure involves lifting tissues to maintain the operating field and grasping the large intestine. furthermore, ns can more easily and safely be applied to laparoscopic colorectal surgery than needlescopic forceps in which both the needle shaft and tip are small. the skin damage is only 2.4 mm in diameter, with the passage of the 2.4 mm shaft through the abdominal wall, because it can be used without a trocar. although the end relief is strong enough for grasping tissues, the shaft can easily be slightly bent, when heavy tissues such as the mesorectum are lifted up. therefore, we remain mindful of the direction of forceps implementation, in order to avoid bending the shaft. laparoscopic isr is suitable for reduced port surgery, particularly ns, single port surgery, or mixed surgery, because the surgical specimen is extracted through the anus without creating a new extraction site. our lap isr procedure using needlescopic instruments is a feasible procedure for minimally invasive surgery. | intersphincteric resection (isr) is a procedure designed to preserve anal function in cases with very low rectal cancer. we report our clinical experience with laparoscopic isr (lap isr) performed using needlescopic instruments. first, a camera port is created at the umbilicus. two 5-mm ports are then inserted at the right upper and lower quadrants. two needlescopic forceps (endo - relief hope denshi co., chiba, japan) are inserted at the left upper and lower quadrants. we then perform the following procedures ; ligation of the inferior mesenteric artery and vein, total mesorectal excision and dissection of the intersphincteric space. after the transanal intersphincteric dissection, the specimen is extracted through the anus and a hand sewn coloanal anastomosis is performed. the covering ileostomy is finally created at the right upper port. we performed lap isr using needlescopic forceps in two patients with very low rectal cancer. in both cases, we were able to perform this procedure without insertion of an additional port or to change the needlescopic forceps to conventional 5-mm forceps. lap isr with needlescopic instruments is a feasible procedure for minimally invasive surgery. |
it is well known that the main phenomenon of hypoxemia in acute lung injury / acute respiratory distress syndrome (ali / ards) is the high shunt fraction caused by the nonaerated areas of the lungs. during the disease process, the volume of extravascular lung water and the lung weight increase and promote the collapse of peripheral airways and lung parenchyma, mainly in the gravitation - dependent lung regions (fig. the relationship between the nonaerated, poorly aerated, normally aerated and hyperinflated lung regions depends on the degree of heterogeneity of the ali / ards and the net result of the interaction of the pressure applied to the lung parenchyma (airway pressure / end expiratory pressure) and chest wall mechanics, as illustrated in the report by henzler and colleagues appearing in this issue of critical care. the most important force is not the airway pressure or tidal volume itself but the stress and strain that this airway pressure / tidal volume generates and the duration of these stresses and strains. at the bedside, the rough equivalent of stress is transpulmonary pressure, and the rough equivalent of the strain is tidal volume / end expiratory lung volume. this modern and complex mechanical ventilatory approach of ali / ards recruitment maneuvers and positive end - expiratory pressure (peep)/tidal ventilation titration is a meshwork of interdependent but heterogeneously affected lung subunits that are behave according to different and multiple pressure volume envelopes of the respiratory system during mechanical ventilation, which in some cases can be represented by respiratory mechanics (depending on the heterogeneity and etiology of the ali / ards and the net results of the mechanical configuration of the respiratory system and the applied inspiratory / expiratory pressure along the mechanical ventilatory support duration). in 1998, a brazilian prospective, randomized and controlled trial of mechanical ventilation in patients with ards demonstrated that a lung protective ventilation strategy that used recruitment maneuvers (a continuous positive airway pressure of 35 to 45 cmh2o) for 40 s with a higher peep set 2 cmh2o above the lower inflection point of the pressure volume curve of the respiratory system and tidal volumes less than 6 ml / kg was associated with a 28-day intensive care survival rate of 62%. this contrasted with a survival rate of only 29% with conventional ventilation (the lowest peep necessary for acceptable oxygenation with a tidal volume of 12 ml / kg without recruitment maneuvers - number necessary to treat = 3, p < 0.001). in a post hoc analysis, the same group stratified the 53 patients of the trial into quartiles according to peep levels and analyzed the 28-day survival rate. a peep of more than 12 cmh2o, and particularly greater than 16 cmh2o, was significantly correlated with an improved survival rate in these ards patients. ranieri and colleagues corroborated these results by demonstrating that a ventilation strategy involving higher peep / low tidal volume significantly decreased bronchoalveolar lavage and systemic blood levels of tumor necrosis factor-, il-8 and il-6 compared with low peep / high tidal volume ventilation. more recently, the same brazilian group showed that when an almost full recruitment is achieved and maintained by means of sufficient applied peep levels (in ards patients this is about 18 to 26 cmh2o of peep), a partial arterial oxygen tension plus partial arterial co2 tension of more than 400 mmhg at a fraction of inspired oxygen of 100% is well correlated with less than 5% of lung collapse as shown on a thoracic computed tomography (ct) scan, ensuring more homogeneous ventilation (fig. recruitment maneuvers, peep and tidal ventilation titration in ali / ards exert varied effects on airway caliber, the ventilation : perfusion ratio distribution, cardiac output and many as yet incompletely understood effects on the macromechanical and micromechanical properties of the diseased lung parenchyma [6 - 8 ]. the history of mechanical ventilation in previous breaths and the applied peep level strongly determine the working envelope in the present breath and the chances of promoting intratidal recruitment during mechanical ventilation in ards patients. overdistension and the opening and closing of alveoli during tidal ventilation are important issues in ventilator - induced lung injury. airspace collapse as shown by a thoracic ct scan is associated with hypoxemia in early ali / ards and can be reversed with a maximum lung recruitment strategy that can be applied to critically ill patients and may lead to 2 better pulmonary function at hospital discharge. so, careful studies of the mechanical, gas - exchange and hemodynamic consequences of mechanical ventilatory support in the experimental and clinical critical care settings of ali / ards are still necessary for a better understanding of the extremely complex issues involved in improving the prognosis of this still life - threatening clinical problem. more intriguing are the recent results showing that dead space fractions were elevated early in the course of ards patients and that the dead space fraction is an independent risk factor for death. corroborating these results are the observations that ali / ards patients who had a decreased partial arterial co2 tension during a prone - position protocol had improved survival compared with the nonresponders. so, respiratory mechanics, gas exchange and hemodynamic parameters as well as medical treatment for the etiology of ali / ards (for example viral infections, bacterial infections, pancreatitis or gastric aspiration) are important issues that have to be kept in the mind of the critical care physicians when treating a patient with ards in the intensive care unit. ali = acute lung injury ; ards = acute respiratory distress syndrome ; ct = computed tomography ; il = interleukin ; peep = positive end - expiratory pressure. thoracic tomography of two different models of acute lung injury / acute respiratory distress syndrome (ards). (a) computed tomography (ct) scan of pigs after saline lung lavage before and after recruitment maneuvers with 45 cmh2o of pressure, maintaining a positive end - expiratory pressure (peep) of 10 cmh2o, showing some redistribution of ventilation. (b) ct scan of acute respiratory distress syndrome patients before and after a recruitment maneuver with 60 cmh2o maximal inspiratory pressure maintaining peep values of 20 and 25 cmh2o. | recruitment maneuvers and positive end - expiratory pressure (peep)/tidal ventilation titration in acute lung injury / acute respiratory distress syndrome (ali / ards) are the cornerstone of mechanical ventilatory support. the net result of these possible adjustments in ventilatory parameters is the interaction of the pressure applied in the respiratory system (airway pressure / end expiratory pressure) counterbalanced by chest wall configuration / abdominal pressure along the mechanical ventilatory support duration. refinements in the ventilatory adjustments in ali / ards are necessary for minimizing the biotrauma in this still life - threatening clinical problem. |
fast quantum - chemical methods are indispensable for computationally demanding calculations of electronic properties of large molecules. in the 1970s and 1980s, semiempirical quantum - chemical (sqc) methods were the workhorse in computational studies of ground - state properties. since the 1990s, they have largely been replaced in such studies by ab initio and density functional theory (dft) approaches, which are typically slower by at least 3 orders of magnitude but also generally more accurate and robust. however, even nowadays, sqc methods often remain the only practical choice when treating huge molecules or very large numbers of molecules or when performing extensive molecular dynamics (md) simulations, e.g., in the context of quantum mechanics / molecular mechanics (qm / mm) studies on large biomolecular systems. examples of recent sqc applications include the calculation of electronic properties used in 3d - qsar models (3d - quantitative structure activity relationship), qm - based computer - aided drug design, the study of band gaps, uv / vis spectra, and charge - transfer processes in systems relevant for molecular nanoelectronics and organic photovoltaics, the investigation of local properties used to understand electron and hole transport mechanisms in transistors, gas - phase md simulations of electron impact mass spectra, ground - state qm / mm md simulations of enzymes, excited - state nonadiabatic dynamics simulations of organic chromophores, and studies of organic and enzymatic reactions in solution. most of the widely used sqc methods are variants of the mndo model which is based on the nddo (neglect of diatomic differential overlap) integral approximation. these mndo - type methods include am1, pm3, mndo / d, am1, rm1, pddg / mndo and pddg / pm3, pm6, and pm7 (see the cited references for details and the meaning of the acronyms). a common feature of these approaches is that they attempt to improve the accuracy within the confines of the given electronic structure model, mainly by adding modifications to the core repulsion functions and by performing a more thorough and extensive parametrization. prime examples of this strategy are the pmx methods (pm3, pm6, and pm7). all mndo - based methods solve the hartree fock secular equations as if the basis set were orthogonal, without explicitly accounting for the terms arising in the ab initio treatment when transforming the fock matrix from the original nonorthogonal to an orthogonal basis. this neglect of orthogonalization terms gives rise to several qualitative deficiencies of standard sqc methods that can not always be eliminated simply by parametrization. it has been shown that reintroduction of the overlap matrix into the secular equations can indeed significantly improve the accuracy of the mndo method. in the orthogonalization models (omx) developed in our group, orthogonalization corrections are included into the fock matrix to different extent (along with other interactions of similar size), which leads to the following general - purpose sqc methods : om1, om2, and om3. the theoretical formalism, the optimized parameters, and the initial validation of the omx methods are described in detail in a companion article. both the mndo - type and omx methods formally neglect dispersion which causes their poor performance in systems where dispersion interactions play an important role. in recent years, a number of empirical dispersion corrections have been developed and successfully applied for dft methods, e.g., the d2 and d3 corrections proposed by grimme. the same types of dispersion corrections can also be combined with the omx methods, with no change in the omx parameters and only minor adjustment of the d2 and d3 parameters. the resulting omx - dn methods provide a much improved treatment of noncovalent interactions. their formalism, parameters, and initial validation are described in detail in ref (42). among the mndo - type methods, the latest variant (pm7) incorporates explicit dispersion corrections in its definition, and unlike in the case of the omx - dn methods, its parameters were optimized with these corrections included. given the severe approximations in the formalism of sqc methods and the presence of empirically determined parameters, it is essential to validate these methods carefully against reliable experimental data and/or accurate high - level theoretical results. some validation studies are available for omx methods, which cover both ground - state and excited - state properties. they indicate that the omx methods outperform other sqc methods in most cases. some of this validation work also offers comparisons between sqc methods and standard dft approaches showing that the former may approach or sometimes even exceed the accuracy of dft results for ground - state properties of organic molecules. considering the extensive recent benchmark exercises for ab initio and dft methods, it is obvious that a more comprehensive validation of the omx and omx - dn methods is required to establish their reliability and to allow for a more detailed assessment of their accuracy compared with other sqc methods. the objective of this article is to provide such validation for ground - state properties. we cover standard mndo - type sqc methods (mndo, am1, pm3, pm6, and pm7), omx methods (om1, om2, and om3), and dispersion - corrected omx - dn methods (om2 and om3 with d2 and d3 corrections as well as d3 t corrections with additional three - body terms). the omx and omx - dn methods are fully specified in ref (42). in addition to our own validation sets, we use benchmark sets mostly taken from the available ab initio and dft literature. we include from these sets only those reference molecules that contain the elements h, c, n, o and/or f, for the simple reason that the omx methods have up to now only been parametrized for these elements. we address the following ground - state properties : heats of formation, bond lengths, bond angles, dihedral angles, relative energies, reaction energies, dissociation energies, atomization energies, proton affinities, activation barriers, vertical and adiabatic ionization potentials (ips), adiabatic electron affinities (eas), dipole moments, noncovalent interaction energies, and geometries of noncovalent complexes. overall, the benchmarks contain 13035 original or derived reference data. the results from the benchmarking will help to establish the accuracy of different sqc methods in different areas and can also serve as a guide for choosing the most appropriate sqc method in specific application projects. all mndo, am1, pm3, omx, and omx - dn calculations were carried out with our locally modified mndo2005 program. standard conventions were used in all calculations. for the sake of documentation, we specify some of these standard options here. the convergence criterion for the scf energy was set to 10 ev ; in addition, the diagonal elements of the density matrix were required to be converged to 10 in mndo2005. doublet and triplet states were treated with the restricted open - shell half - electron approach, and higher spin states were described using unrestricted hartree we did not use molecular mechanics corrections for peptides in pm6 or pm7, and we did not apply any cutoffs for the three - center orthogonalization corrections in the omx and omx - dn methods. geometry optimizations were considered converged when the gradient norm became smaller than 0.01 kcal/(mol). in mndo2005 the bfgs optimization algorithm was used by default ; in difficult cases, we also applied eigenvector following, with the hessian being computed numerically by one - sided finite differences of the gradient (at the first step and every 10 following steps) and with a minimum trust radius of 0.00001. in mopac the eigenvector following algorithm was used throughout ; in most cases the full hessian matrix was constructed and recalculated every 10 steps using numerical single - sided derivatives of the gradient. frequently mopac stopped when the heat of formation remained essentially constant in subsequent cycles ; in these cases the gradient norm was usually smaller than 0.1 kcal/(mol). it is well - known that sqc methods generally predict the heat of formation of the proton with very large errors. therefore, in line with common semiempirical practice, we used the experimental value of 367.171 kcal / mol for the heat of formation of the proton at 298 k to calculate the proton affinities included in the gmtkn30-chnof and ce345-chnof databases (see later text). the reference molecules in the various benchmark sets are generally still quite small by semiempirical standards. therefore, they were computed using single - cpu serial versions of the mndo2005 and mopac programs. for large - scale applications, parallel versions of mndo2005 are available using shared - memory and distributed - memory message - passing parallelization, as well as a hybrid version using graphics processing units (gpus). similarly, there are shared - memory and gpu - parallelized versions of mopac. to illustrate the scope of sqc calculations, we note that the hybrid gpu version of mndo2005 has been used to compute water clusters of up to 5400 atoms and to optimize the geometries of a large series of representative proteins. calculations with d3 and d3 t dispersion corrections were done using our interface of the mndo2005 program to the dft - d3 stand - alone program by grimme (versions 3.0 rev 2 and 3.1 rev 0). default cutoffs (95 au for two - body terms and 40 au for coordination numbers) were used with version 3.0 and increased cutoffs (95 au for two - body terms, coordination numbers, and three - body terms) with version 3.1. in this section we define the benchmark sets used presently to evaluate different sqc methods. we divide these sets into two groups : those designed to benchmark ground - state properties in general (ovs7-chnof, g2g3-chnof, w4 - 11-chnof, gmtkn30-chnof, ce345-chnof, pddg, pm7-chnof, and c7h10o2) and those specifically designed to benchmark noncovalent interactions (a24-chnof, s66, s66a8, jsch-2005-chnof, s7l, s30l - chnof, and af6). we note that this division is somewhat arbitrary as some reference molecules and complexes from the first group also feature noncovalent interactions and may thus appear again in the second group. all data sets and subsets from the first and second groups are listed in tables 1 and 2, respectively. most of them are taken from the literature and described in the same terms as in the published work. included are all molecules from the original data sets that contain only the elements c, h, n, o, and/or f, for which omx parameters are available ; all other molecules are ignored. detailed numerical results on all individual reference molecules from all benchmark sets are compiled in the supporting information (si) which also cites the origin of the published reference data. in the following two sections, we will focus on statistical evaluations of these results for all sets and subsets (with more than two molecules). data in these databases were normally corrected to represent total energies without any zero - point vibrational energies and thermal corrections ; this is also true for the expt and theor values given for the subsets ; theor usually means best theoretical estimate. intermolecular distances were obtained from mp2/cc - pvtz geometries by interpolating estimated ccsd(t)/cbs energies along dissociation curves. reference geometries were obtained by optimizing intermolecular distances at the ccsd(t)/ha - cc - pvdz level using monomer geometries optimized at the b3lyp level with large basis sets. following semiempirical tradition, all sqc methods tested presently were parametrized with regard to heats of formation. their performance can thus be evaluated in a straightforward manner when reliable reference data for heats of formation are available either from experiment or from high - level ab initio calculations. this is the case for the chno and fluor data sets used during the parametrization of the omx methods and for the ovs7-chnof, g2g3-chnof, pddg, and pm7-chnof sets used in the present benchmarking. performance evaluation of atomization enthalpies at 298 k is also straightforward for the c7h10o2 set as reference values can be easily obtained from the reference ab initio enthalpies at 298 k. most of the entries in the other databases of tables 1 and 2 represent relative energies, for example isomerization energies, interaction energies, binding energies, reaction energies, barrier heights, proton affinities, ionization potentials, and electron affinities. the corresponding ab initio reference data are generally obtained from energy differences, whereas the respective semiempirical values are differences of heats of formation (i.e., enthalpies at 298 k) that implicitly include zero - point vibrational energies (zpves) and thermal enthalpic corrections (from 0 to 298 k). these corrections are normally quite similar for related molecules since they tend to be bond - specific and transferable, and hence they will cancel to a large extent when comparing related systems. as in many semiempirical studies over the past decades, we will thus mostly ignore the distinction between relative energies and heats of formation in the following, and consider the semiempirical values to be directly comparable with ab initio relative energies. this line of reasoning obviously breaks down when it comes to atomization energies, because there is no longer any cancellation. for a realistic comparison between ab initio and semiempirical atomization energies, the zpves and thermal enthalpic corrections this is computationally demanding at an accurate ab initio level, and therefore we computed both these corrections at the sqc level using the harmonic - oscillator and rigid - rotor approximations. to illustrate the necessity of including these corrections, we compare the om2 results for propane with experimental and ab initio w4 reference data : the experimental heat of formation at 298 k (25.0 kcal / mol) is well - reproduced by a straight om2 calculation (24.4 kcal / mol at the ab initio reference geometry), whereas the w4 atomization energy (1007.9 kcal / mol) is well - reproduced by om2 (1005.6 kcal / mol) only after applying the corrections described previously, which are far from negligible (51.8 kcal / mol). the distinction between relative energies and heats of formation is also relevant for the reaction energies in other fragmentations, e.g., bond dissociation reactions : while the corresponding zpve and thermal enthalpic corrections are much smaller than for atomization energies, they are still non - negligible. in the next section, we will apply these sqc corrections to all benchmarks for atomization energies (especially w4 - 11-chnof). we will also investigate their effect on the reaction energies covered by the four subsets of the w4 - 11-chnof set and by those subsets of the gmtkn30-chnof and ce345-chnof sets that address fragmentation reactions. the sqc corrections are combined with single - point sqc results obtained at the ab initio reference geometries. the conversion of semiempirical heats of formation at 298 k to zpve - exclusive relative energies at 0 k is described in detail in the supporting information. the omx methods have been parametrized using the relatively small chno and fluor training sets (140 and 48 molecules, respectively). for these sets, the omx results are generally found to be superior to the results from other sqc methods, as documented in detail in a companion article. various omx parametrization runs starting from different initial values were often found to produce different parameter sets that gave results of similar quality for the molecules in the training set ; therefore we performed further tests on several validation sets to help identify the most suitable parameters. the main validation sets are collected in the ovs7-chnof benchmark which consists of seven subsets. detailed numerical results are given in the si (tables s9s14), while statistical evaluations in terms of mean absolute errors (maes) are provided in table 3. with regard to heats of formation, om2 and om3 outperform all other sqc methods for six of the seven subsets, namely, radicals71, anions24, cations41, bigmol20, isomers44, and fluorine91 ; the corresponding maes for om2 and om3 are 5.05.6, 8.49.6, ca. 6.9, ca. 5.0, 1.11.8, and 7.27.3 kcal / mol, respectively. in the conformers30 subset, om1 gives the lowest mae (1.8 kcal / mol) followed by pm7 (2.4 kcal / mol) and om2 and om3 (ca. considering relative energies, om2 and om3 again outperform all other sqc methods in the case of the radicals71 and cations41 subsets, and they also give very good results for the isomers44 subset (maes of 0.82.1 kcal / mol). ionization potentials for the radicals71 subset (from koopmans theorem) are best predicted by the three omx methods, with maes of 0.40.5 ev. this is also true for the barriers in the conformers30 subset, for which the maes for the omx methods range between 1.3 and 1.5 kcal / mol. in the fluorine91 subset, bond lengths are well - described by om1, om2, pm3, and pm6, while bond angles are best reproduced by the omx methods. finally, we note that the inclusion of dispersion corrections must deteriorate the accuracy of the omx methods for heats of formation, because the omx parameters were optimized without such corrections. since dispersion effects are always attractive, their subsequent introduction will systematically lower the computed heats of formation and thus increase their errors compared with experiment. on the other hand, dispersion corrections have no or very little effect on ionization potentials and barriers ; they improve the relative energies for the isomers44 subset while leaving those for the radicals71 and cations41 subsets almost unaffected (for further details see the si table s1). the g2 and g3 sets provide well - established accurate reference data that have often been used to validate the performance of ab initio and dft methods. they have been published after the development of om1 and om2. in previous work we have reported an evaluation of the omx methods for the g2 and g3 sets, and we can thus be brief. the statistical results are summarized in table 4 (for detailed numerical results, see si tables s16s18). om2 yields the lowest maes for the heats of formations in g2 and g3 (3.23.4 kcal / mol) and for the relative energies in the alkanes28 subset of the g3 set (0.6 kcal / mol), while om3 gives the lowest mae for the heats of formations in alkanes28 (0.7 kcal / mol). as pointed out before, the performance of om2 and om3 for the g2 and g3 sets is respectable, even when compared with standard dft approaches, and there is no systematic error for alkane chains of increasing length that plagues some standard dft functionals (including b3lyp). again, as in the ovs7-chnof set and for the same reasons, dispersion corrections deteriorate the accuracy of the omx methods for heats of formation in the g2g3-chnof set, while improving relative energies (si table s2). the w4 - 11 benchmark set includes the w4 - 08 subset of the gmtkn30 set (see later discussion) and, in addition, contains numerous further atomization energies (collected in the extended subset tae140) as well as four other subsets with reaction energies (bde99, hat707, isomer20, and sn13 ; see table 1). we reduced the w4 - 11 to the w4 - 11-chnof data set in the usual manner, by eliminating entries for species containing elements other than h, c, n, o, and f. the sqc results from single - point calculations at the reference geometries are compared with the reference data obtained from ab initio total energies without zero - point vibrational corrections. for reasons discussed above, we converted the semiempirical heats of formation to atomization energies at 0 k by applying zpve and thermal enthalpic corrections. we also compared the results for the reaction energies in the subsets calculated with and without such corrections (see table 5 for the statistical evaluation). (corr) means that energies are obtained by removing zpve and thermal corrections from the sqc results (see text). the atomization energies in the tae140 subset are computed with errors somewhat larger than typical errors for heats of formation. these errors are of similar magnitude in the omx and pm7 methods (maes of 4.817.02 kcal / mol, om2 lowest). the maes remain roughly of similar size when multireference cases are excluded (tae_nonmr124, maes of 4.846.75 kcal / mol, om2 lowest). errors in the corrected and uncorrected reaction energies for heavy - atom transfer (hat707 subset), for isomerization (isomer20 subset), and for transformations in the sn13 subset are very close to each other, without any systematic improvement due to the corrections. this provides further justification for the usual practice of directly comparing the semiempirical scf energies with ab initio relative energies in such cases. thus, we will discuss for these sets and similar benchmark sets of this kind only the noncorrected reaction energies calculated from the semiempirical scf energies. on the other hand, removing the zpve and thermal corrections from the semiempirical scf energies systematically improves the computed bond dissociation energies (bde99 subset, mae lowered by ca. this indicates that these corrections are generally relevant for fragmentation reactions, and hence we will discuss them for such transformations in the following. the reference energies for bond dissociation and heavy - atom transfer are best reproduced by om2 (maes of 6.25 and 8.92 kcal / mol) followed by om3 (maes of 7.51 and 9.44 kcal / mol). isomerization energies are computed reasonably well by all sqc methods considered (maes of 7.659.37 kcal / mol, am1 lowest). the reaction energies in the sn13 subset are also well - reproduced, particularly by pm7, om1, and om3 (maes of 3.14, 4.02, and 4.31 kcal / mol, respectively). the largest outliers in the computed atomization energies are found for c2 (om1), isocyanic acid hnco (om2), hnnn (om3), and h2 and n2 (pm7), with errors ranging from 21 to 35 kcal / mol. dispersion corrections have only very little effect on any of the statistical results reported presently (see si table s19). the grimme group published comprehensive collections of validation sets for general main group thermochemistry, kinetics, and noncovalent interactions, to allow for a thorough assessment of the performance of various dft methods. in previous work, we extracted from the initial gmtkn24 database all species containing only the elements h, c, n, and o. for the resulting gmtkn24-hcno database, we performed calculations using the am1, pm6, om1, om2, om3, scc - dftb, b3lyp, and pbe methods and their dispersion - corrected counterparts to compare their accuracy in a comprehensive manner. here we update our previous evaluation of sqc methods by starting from the more recent gmtkn30 database and extracting all species containing only the elements h, c, n, o, and f. the resulting gmtkn30-chnof database differs from the previous gmtkn24-hcno variant in the following aspects. (b) it includes three new subsets isol22, bsr36, and adim6 from gmtkn30 ; the other three new subsets in gmtkn30 contain elements other than h, c, n, o, and f and are therefore disregarded. (c) it also includes the full w4 - 08 subset with multireference (mr) cases, in addition to the previously used w4 - 08womr subset without mr cases. (d) it utilizes updated, more accurate reference data from the gmtkn30 database whenever available (in subsets idisp, s22, bhperi, and pa). specifically, the gmtkn30 reference data for the bhperi subset are now based on w1 rather than cbs - qb3 calculations ; the latter had been employed in gmtkn24-hcno for some reaction barriers. the gmtkn30 reference values for the proton affinities of small molecules in the pa set are now taken from vibrationally back - corrected w1 data rather than from best estimates from vibrationally back - corrected experimental values in gmtkn24-hcno. overall, the gmtkn30-chnof database consists of 24 subsets with 480 reference data that are determined from 806 single - point calculations at the reference geometries. six subsets from the full gmtkn30 database are missing (al2x, nbprc, alk6, rg6, heavy28, and cyconf), because they consist of species containing elements not yet parametrized for the omx methods. detailed numerical results for the gmtkn30-chnof database are documented in the supporting information. generally speaking, our present results support and generalize the conclusions on the performance of sqc methods obtained previously for the gmtkn24-hcno set. overall, the omx and omx - dn methods provide the most accurate results. the maes for the entire gmtkn30-chnof set are lowest for om3 (7.17 kcal / mol) followed by om2 (7.94 kcal / mol), but are substantially higher for the pmx methods (14.4416.49 kcal / mol) and am1 (16.45 kcal / mol). inclusion of dispersion corrections for om2 and om3 changes the overall maes only very slightly (om3-dn, 7.217.26 kcal / mol ; om2-dn, 7.767.91 kcal / mol). compared to the previously reported uncorrected values in ref (52) (w4 - 08womr subset), the semiempirical scf atomization enthalpies at 298 k become much more accurate by correcting to zpve - exclusive atomization energies at 0 k (errors reduced by ca. the omx methods remain the most accurate sqc methods in the w4 - 08womr subset (maes of 4.126.00 kcal / mol). for the full w4 - 08 subset, the maes of the corrected atomization energies are somewhat higher at the omx level (4.197.58 kcal / mol) and still slightly larger for pm6 and pm7 (6.537.95 kcal / mol). (corr) means that energies are obtained by removing zpve and thermal corrections from the sqc results (see text). without mb08 - 165 and c20. upon geometry optimization, some of the artificial molecules adopted structures very different from the reference geometries so that the computed corrections may be less accurate in these cases (see text). the adduct o3 + c2h2 is better described as an open - shell singlet at omx - dn. subset dc9 without c20 bowl / cage isomerization energy. c20 bowl / cage isomerization energy. for some sqc methods reference dissociation energies of four (h2o)20 clusters were updated with more accurate values from ref (132) (see text). (corr) means that energies are obtained by removing zpve and thermal corrections from the sqc results (see text). without mb08 - 165 and c20. upon geometry optimization, some of the artificial molecules adopted structures very different from the reference geometries so that the computed corrections may be less accurate in these cases (see text). the adduct o3 + c2h2 is better described as an open - shell singlet at omx - dn. subset dc9 without c20 bowl / cage isomerization energy. c20 bowl / cage isomerization energy. for some sqc methods reference dissociation energies of four (h2o)20 clusters were updated with more accurate values from ref (132) (see text). in the case of the mb08 - 165 subset, the zpve and thermal corrections reduce the errors for all of the methods except for mndo. this subset is generally most challenging as it was especially designed to test the robustness of computational approaches. it consists of the dissociation energies of randomly generated species with unusual bonding situations to diatomics and hydrides. for some of the artificial molecules in this subset, the geometry optimization (needed to evaluate the zpve and thermal corrections) led to structures very different from the reference geometries (especially with mndo), and hence to corrections that may be less accurate ; the corrected values were included in the statistics also in these cases. the omx methods have the smallest maes for both the uncorrected (19.4622.47 kcal / mol, om1 and om3 lowest) and the corrected (11.0415.03 kcal / mol, om1 lowest) dissociation energies of the mb08 - 165 subset, while the maes for pm6 and pm7 exceed 100 kcal / mol. in the case of the bsr36 subset, going from the scf bond separation enthalpies at 298 k to the corresponding zpve - exclusive energies at 0 k reduces the errors only for the om2, om3, and omx - dn methods. the reason for the increased error with the other methods is the way in which this benchmark is constructed : bond separation energies refer to reactions of hydrocarbons with methane to produce ethane molecules ; thus large numbers of ch4 and c2h6 molecules are involved (up to 22 and 18, respectively), and any errors in the computed zpve and thermal corrections for these two molecules will accumulate. this happens, for instance, in the case of pm7 where the mae for the bsr36 subset drops from 17.40 to 9.08 kcal / mol (similar to the value of 9.63 kcal / mol without corrections) when using the zpve energies of methane and ethane calculated at the w4 level instead of the pm7 values. such problems are not encountered with the om2, om3, and omx - dn methods which give realistic zpve and thermal corrections for methane and ethane. om3 is most accurate for the bsr36 subset, both for the uncorrected (mae of 3.46 kcal / mol) and the corrected (mae of 1.90 kcal / mol) bond separation energies. as noted previously, the cage / bowl isomerization of c20 fullerene is also a very challenging problem for sqc methods : the absolute error in the isomerization energy exceeds 85 kcal / mol for all sqc methods tested here. c20 is part of the dc9 subset with only seven members ; to avoid misleading impressions because of the c20 outlier, we provide maes not only for the complete dc9 subset but also for dc9woc20 (dc9 without c20), as well as the errors for c20. if we remove the most challenging test cases (mb08 - 165 and c20) from the overall statistics, the maes remain lowest for om3 (6.30 kcal / mol) and om2 (6.95 kcal / mol), whereas those for the pmx methods drop considerably (9.6011.26 kcal / mol) ; am1 benefits less (14.65 kcal / mol). inclusion of dispersion corrections for om2 and om3 again affects their maes only slightly. according to the statistical evaluations, the omx and omx - dn methods are most accurate for barrier heights of hydrogen transfers, heavy - atom transfers, nucleophilic substitutions, unimolecular reactions, and association reactions (bh76) and for the corresponding reaction energies (bh76rc) ; and for selected reaction energies in the g2 set (g2rc). the om2, om3, and omx - dn methods also perform best for adiabatic ionization potentials (g21ip) and electron affinities (g21ea) ; for isomerization energies of large organic molecules (isol22) ; and for relative energies of phenylalanyl glycyl glycine tripeptide conformers (pconf) and of sugar conformers (sconf). the lowest maes are provided for proton affinities (subset pa) by om1 followed by om3 and om3-dn ; for systems with intramolecular dispersion interactions (idisp) by om2, om3, and om3-dn ; for interaction energies between n - alkane dimers (adim6) by omx - d3, omx - d3 t, and pm7 ; and for relative energies of alkane conformers (aconf) by omx - dn. pm6 is statistically most accurate for systems with problems related to the dft self - interaction error (sie11) ; for other molecules that are difficult cases for dft (dc9 and dc9woc20) ; for reaction energies and barriers of ozone addition to c2h4 and c2h2 (o3add6) ; and for energies of diels alder reactions (darc). pm7 appears best suited for calculating barrier heights of pericyclic reactions (bhperi) ; isomerization energies of small- and medium - sized organic molecules (iso34) ; and binding energies of noncovalently bound dimers (s22). in the latter case, the omx - dn methods are of similar accuracy (see section 5). benchmark energies for four (h2o)20 clusters used in the water27 subset (binding energies of water, h(h2o)n, and oh(h2o)n clusters) were taken from mp2/cbs calculations. recently more accurate reference energies were obtained at the ccsd(t)/cbs(45) level of theory for those four complexes. when including these updated reference data, the pm7, om2-dn, and om3-d2 methods perform similarly well (maes of 6.206.57 kcal / mol, om2-d3 lowest). interestingly, radical stabilization energies (rse43) are best predicted by the venerable am1 method closely followed by om1. this demonstrates that the most recent scq models and parametrizations may not always be the best for a problem at hand. hence, if possible, sqc methods should generally be calibrated against available experimental or high - level theoretical data for related systems before using them in actual applications. as already discussed earlier the largest outliers (si figure s1 and table s20) are often found for species in the mb08 - 165 subset ; this is the case for pmx, om2, and om2-dn, with especially large errors of more than 100 kcal / mol seen frequently with pmx. in the case of the om3 and om3-dn methods, the largest errors are encountered for the isomerization energy of c20, followed by species in the mb08 - 165 subset. other notable outliers (figure 1) occur for om1 in the water27 subset (four errors larger than 100 kcal / mol) ; for om2 in the pa subset (error of 58 kcal / mol in the proton affinity of molecular hydrogen) ; and for om3 in the o3add6 subset (error of 45 kcal / mol in the energy of adduct formation between o3 and c2h2). according to the statistical evaluations, the omx - dn methods have essentially the same overall accuracy as the standard omx methods. as expected, the dispersion corrections significantly improve the accuracy for systems, in which noncovalent interactions play a significant role. the barriers of pericyclic reactions (bhperi subset) are also slightly improved when using the omx - dn methods, on average by more than 1 kcal / mol. on the other hand, as already noted, inclusion of dispersion corrections (without any change in the standard omx parameters) will lower the total energies systematically and thus tend to increase the errors in the calculated heats of formation. interestingly, the dispersion corrections even deteriorate the results for the idisp subset designed to capture the effects of intramolecular dispersion. apparently, intramolecular noncovalent interactions of this kind are already largely taken into account (on average) by the general parametrization of omx methods such that adding further dispersion interactions is detrimental. overall, the positive and negative effects of including empirical dispersion corrections (while retaining the standard omx parameters) seem to cancel out in the case of the gmtkn30-chnof benchmark set so that the overall maes are similar for the omx and omx - dn methods. the chemistry energetic database with 345 entries (ce345) is a collection of accurate reference data used for the evaluation and parametrization of dft functionals. we note that some of the reference data in the ce345 database are also present in the gmtkn30 database, although the actual reference values may differ slightly because of the use of different methodology for obtaining them. we reduced the ce345 database to the ce345-chnof set by accepting only species containing exclusively the elements h, c, n, o and/or f. therefore, the srmbe13 subset had to be excluded. the mrbe10 and dc9 subsets were also excluded from the error analysis because there are only two surviving species in each subset. the final ce345-chnof set consists of 11 subsets with a total of 187 entries. error distribution of ground - state properties calculated at the omx and pm7 levels for the gmtkn30-chnof benchmark set (excluding the mb08 - 165 subset and the isomerization energy of c20). the subsets are marked with alternating gray and white backgrounds, and their numbers correspond to those in table 6. overall, the lowest maes for the ce345-chnof set are obtained from the omx and omx - dn methods (6.177.91 kcal / mol). as expected, the inclusion of dispersion corrections improves the results for the ncce31/05 subset targeting noncovalent complexation energies ; this is also true for the subsets abde12, hc7/11, and tc13 that contain systems in which dispersion plays a prominent role. (corr) means that energies are obtained by removing zpve and thermal corrections from the sqc results (see text). scf calculations of the nh3f2 complex in the ncce31 subset could not be converged with om2 and om3 ; thus we excluded this complex from the statistics. looking at the statistical evaluation for the individual subsets, atomization energies (after correcting scf atomization enthalpies at 298 k to zpve - exclusive energies at 0 k, mgae109/11) are found to be best reproduced by omx and omx - dn methods (maes of 4.194.98 kcal / mol, om2 and om2-dn lowest). isomerization energies (isol6/11) are best reproduced by om2 and pm7 (mae of ca. we note that this subset contains the six smallest pairs of isomers from the isol22 subset of gmtkn30 ; their isomerization energies were calculated at a more accurate level of theory than those in isol22. (corr) means that energies are obtained by removing zpve and thermal corrections from the sqc results (see text). scf calculations of the nh3f2 complex in the ncce31 subset could not be converged with om2 and om3 ; thus we excluded this complex from the statistics. the subsets for adiabatic ionization potentials (ip21) and electron affinities (ea13/03) overlap with the corresponding gmtkn30-chnof subsets (g21ip and g21ea). in all cases, the om2 and om3 methods perform best (maes of 0.520.57 ev for ip21 and 0.400.42 ev for ea13/03), with substantially larger errors for the other sqc methods. as expected, dispersion corrections have essentially no effect on the om2 and om3 results for these properties. all entries in the subset for proton affinities (pa8/06) also appear in the pa subset of gmtkn30 (albeit with slightly different reference values). as in the pa case, om1 has the lowest statistical error (mae of 4.8 kcal / mol). alkyl bond dissociation energies (abde12) are best reproduced by the omx - dn and omx methods (maes after accounting for zpve and thermal corrections of 7.3210.52 kcal / mol and above 17 kcal / mol for the mndo - type methods), while hydrocarbon chemistry (hc7/11) is best described by om3-dn and pm6 (maes of 3.204.34 and 4.57 kcal / mol, respectively). the thermochemistry of systems (tc13) is best treated by the om2 and om2-dn methods (maes of 2.54 and 2.732.74 kcal / mol, respectively ; the data set includes some proton affinities of polyenes that are part of the pa subset of gmtkn30). the entries in the subsets for hydrogen and non - hydrogen transfer reactions (htbh38/08 and nhtbh38/08) are also present in the bh76 subset of gmtkn30. om2 and om2-dn perform best for the barrier heights to hydrogen transfer followed by om3 and om3-dn (maes of 4.954.96 and 5.996.61 kcal / mol, respectively), while the barrier heights to non - hydrogen transfer are described less well (lowest mae of om1 9.60 kcal / mol). as expected, noncovalent complexation energies (ncce31/05) are best reproduced by the omx - dn methods (maes of 1.081.51 kcal / mol) followed by pm7 (maes of 1.58 kcal / mol). the error distributions of the omx and pm7 methods with regard to the ce345-chnof set are shown in figure 2. they appear similar overall ; however, there are more outliers for pm7 compared with the omx methods and especially with om2. error distribution of ground - state properties calculated at the omx and pm7 levels for the ce345-chnof set. the subsets are marked with alternating gray and white backgrounds, and their numbers correspond to those in table 8. the largest outliers at the omx and omx - dn levels have errors between 40 and 63 kcal / mol in the computed energies. these are the ionization energy of the carbon atom (om1) ; the proton affinity of molecular hydrogen (om2 and om2-dn) ; and the barrier of the hcn hnc isomerization (om3 and om3-dn) ; see the supporting information. the pddg set was used in the development and validation of the pddg / mndo and pddg / pm3 methods. it contains 979 experimental reference data for heats of formation, bond lengths, bond angles, dihedral angles, ionization potentials, and dipole moments of 622 closed - shell molecules (elements h, c, n, ond o). the pddg set is divided in the original work into ch, chn, cho, and chno subsets. in addition, we introduce subsets for bond lengths, bond angles, and dihedral angles according to the elements involved. in our evaluations, the resulting maes for the different properties are summarized in table 10 for the mndo, am1, pmx, and omx methods and in si table s3 for the omx - dn methods. considering heats of formation, the pddg / pm3 method gives the lowest mae for the pddg set (3.2 kcal / mol), which is not too surprising since it was trained on this set. slightly higher are the maes for pm7 (3.34 kcal / mol), om2 (3.55 kcal / mol), and om3 (3.68 kcal / mol), while those for the other sqc methods are above 4.0 kcal / mol. the om2 and om3 methods systematically overestimate the heats of formation in the pddg set (mean signed errors (mses) of 0.92 and 1.08 kcal / mol, respectively), more so than pm7 (mse of 0.49 kcal / mol). error histogram of heats of formation calculated at the omx and pm7 levels of theory for the pddg benchmark set. as noted previously, post - scf dispersion corrections deteriorate the accuracy of heats of formation calculated by sqc methods parametrized at the scf - mo level (compare table 10 and si table s3). this shortcoming can be avoided by simultaneously fitting the parameters of the sqc method and of the dispersion corrections, as has been done in pm7 ; this may contribute to its good performance for the pddg set. considering geometries, all tested sqc methods perform reasonably well for bond lengths (overall maes of 0.0110.018, lowest for pm3 and pm7). except for mndo, this is also true for bond angles (overall maes of 1.902.15, lowest for om3). dihedral angles are described best by the omx methods (overall maes of 5.78.5, lowest for om3). ionization potentials are reproduced best by om2 (mae of 0.37 ev) ; the other sqc methods have somewhat larger errors (0.480.70 ev). the maes in the calculated dipole moments are in the range between 0.24 and 0.38 d ; the am1, pm3, and omx methods provide the best estimates (maes of 0.240.27 d). the pm7-chnof benchmark set was assembled from the online database of reference data that have been used for the development and validation of the pm7 method. it includes all species of the database that consist only of the elements h, c, n, o, and f. it contains 1595 experimental and high - level ab initio reference values for heats of formation, bond lengths, bond angles, ionization potentials, and dipole moments of 1177 neutral and charged, open - shell and closed - shell species. it is divided into the da, ch, chn, cho, chf, and chno subsets. the diatomic da subset consists of the h2, n2, o2 (triplet and singlet), and f2 molecules. in our evaluations, the resulting maes for the different properties are summarized in table 11 for the mndo, am1, pmx, and omx methods and in si table s4 for the omx - dn methods. the pm7 method was trained and validated on this benchmark set, and it is thus not surprising that pm7 shows the best overall accuracy for the heats of formation (mae of 3.78 kcal / mol, mse of 0.18 kcal / mol) ; its error distribution is narrow and symmetric (figure 4). the omx methods tend to overestimate the heats of formation for this set, especially om1 (figure 4). om2 and om3 perform similarly (maes of 4.834.85 kcal / mol). they outperform pm7 in the da and chf subsets and have similar accuracy in the ch subset, while pm7 is superior in the chn, cho, and chno subsets. again, for the same reasons as discussed before, post - scf dispersion corrections deteriorate the accuracy of the heats of formation calculated by the omx methods (compare table 11 and si table s4). error histogram for heats of formation calculated at the omx and pm7 levels of theory for the pm7-chnof benchmark set. concerning the other properties, the conclusions for the pm7-chnof set are similar to those for the pddg set (see preceding discussion). bond lengths and bond angles are generally reproduced reasonably well by all sqc methods, and there are no great differences in the overall accuracy of the computed ionization potentials and dipole moments. for bond lengths, pm7 has the lowest overall mae (0.013) followed by pm3, pm6, om1, and om2 (0.015 in each case). the omx methods have a slight overall advantage over the other sqc methods for bond angles (maes of 1.481.76, om1 lowest), ionization potentials (0.330.53 ev, om2 lowest), and dipole moments (0.220.26 d, om3 lowest). accurate thermochemical calculations at the g4mp2 level of theory were recently performed for a set of 6095 constitutional isomers c7h10o2. they were drawn from the chemical universe database gdb-17 that contains 166.4 billion molecules with up to 17 non - hydrogen atoms. we derived atomization enthalpies at 298 k for these 6095 c7h10o2 isomers from the corresponding g4mp2 enthalpies reported in ref (123). the reported geometries were obtained at the b3lyp/6 - 31g(2df, p) level of theory, which is not considered accurate enough to be used as reference in our present benchmarking. the omx and pmx methods perform similarly, with maes ranging between 6.30 and 8.92 kcal / mol (om2 lowest, closely followed by pm7). dispersion corrections have essentially no effect on the atomization enthalpies for the molecules in the c7h10o2 set (si table s26). upon the request of a reviewer, we have carried out analogous rm1 calculations for all benchmark sets considered in preceding discussion. the statistical evaluations of the rm1 results are given in the supporting information (tables s34s41) along with a brief assessment. overall rm1 tends to be generally more accurate than am1, about as accurate as the pmx methods, and somewhat less accurate than the omx methods. noncovalent interactions are difficult to describe for standard sqc methods which do not include dispersion in their formalism and often also have well - documented problems with hydrogen bonding. in this section, we will focus on the pm7 and omx - dn methods with explicit dispersion corrections, while presenting the results for the other (less accurate) sqc methods only in the supporting information. we selected seven data sets from the literature to benchmark the accuracy of sqc methods for noncovalent interactions. as usual, we excluded in these sets the species containing elements for which omx parameters are not yet available. thus, three of the data sets were reduced : a24 from 24 to 21 entries in a24-chnof, jsch-2005 from 143 to 134 entries in jsch-2005-chnof, and s30l from 30 to 24 entries in s30l - chnof. the remaining four sets were not changed (s66, s66a8, s7l, and af6). we first report results from single - point calculations at the reference geometries for all seven data sets. thereafter we present results from sqc geometry optimizations for the a24-chnof, s66, s7l, and af6 sets, for which high - level reference geometries are available. we note in this context that the s66a8 set is specifically designed for single - point calculations. the reference geometries in the jsch-2005-chnof set are taken from different sources and include experimental, purely theoretical, and combined experimental / theoretical data, with purely theoretical geometries constituting only a minor part ; many of these reference geometries are not minima in the gas phase, which would make it meaningless to compare them with optimized gas - phase sqc geometries. in the validation studies with full geometry optimization, the reason is simple : if a method fails to predict the correct geometry for a noncovalent complex, the computed interaction energy will be meaningless. hence, we primarily examine the most important intermolecular distances and angles for each complex (except for af6 where we check the intramolecular geometry parameters in the folded conformation). the statistical evaluation of the interaction energies obtained from single - point sqc calculations is given in table 13 for the pm7 and omx - dn methods and in si table s5 for all other methods. the omx - dn methods perform best overall for the a24-chnof set (maes of 0.560.69 kcal / mol, om2 lowest), while pm7 has somewhat larger errors (mae of 1.10 kcal / mol). the most problematic case in a24-chnof is the hfhf complex : at the nonoptimized reference geometry, the reference interaction energy is attractive (4.57 kcal / mol) ; om2 and om2-dn give only a rather weak attraction (1.47 to 1.15 kcal / mol), whereas mndo, pm6, pm7, om1, om3, and om3-dn yield a repulsive interaction energy (up to 5.81 kcal / mol in pm7) ; see the discussion in section 5.2. when this complex is disregarded in the statistics, the errors drop further (maes of omx - dn 0.430.49 kcal / mol and pm7 0.64 kcal / mol). considering the subsets, the omx - dn methods outperform pm7 for complexes with dispersion - dominated and mixed electrostatic / dispersion interactions, and the om3-dn methods are best for h - bonded complexes. in the s66 set, the pm7 and omx - d3 methods are of similar accuracy (maes of 0.79 vs 0.82 kcal / mol), whereas the latter are superior in the s66a8 set (maes of 0.78 vs 0.600.62 kcal / mol) ; the omx - d2 treatments are generally somewhat less accurate. in all three sets, the omx - dn methods outperform pm7 for the complexes with dispersion - dominated and mixed interactions. in the jsch-2005-chnof set (interaction energies of dna base pairs and amino acid pairs), the omx - d2 results are again slightly less accurate than the omx - d3 results, and among the latter, om3-d3 has generally the lowest or close - to - lowest errors. om3-d3 also outperforms pm7 for the jsch-2005-chnof set overall (maes of 1.33 vs 1.93 kcal / mol) as well as for two of the subsets (intrastrand and stacked base pairs), whereas the maes are similar in the case of h - bonded base pairs (maes of 2.53 vs 2.62 kcal / mol) and higher for om3-d3 in the case of amino acid pairs (maes of 2.56 vs 2.19 kcal / mol). the largest outliers generally occur for pairs of oppositely charged amino acids : when these six complexes are excluded from the analysis, the errors drop substantially, as can be seen from the maes for om3-d3 and pm7 for the neutral amino acid pairs (maes of 0.49 vs 1.17 kcal / mol) and for the overall set (maes of 1.10 vs 1.83 kcal / mol). evidently, om3-d3 performs best for the jsch-2005-chnof set. without the hfhf complex, two complexes correspond to both h - bonded and charged complexes subsets ; see ref (49). in a similar vein, om3-d3 describes the and interactions for large complexes (s7l set) significantly better than any of the other sqc methods considered. the corresponding mae value for om3-d3 (0.66 kcal / mol) is small on an absolute scale and much lower than that for pm7 (4.91 kcal / mol). the s30l set is an expansion of the s12l set and consists of very large noncovalent complexes, which are deemed useful to check against overfitting to too small molecular cases. the omx - dn methods perform fairly well (maes of 3.596.69 kcal / mol, om3-d3 t lowest). they outperform other sqc methods investigated here and in ref (49), and especially pm7, which strongly overestimates most interaction energies (mae of 15.44 kcal / mol). the omx - dn methods generally describe stacking interactions better than h - bonding interactions ; however, even in the latter case, they are more accurate than pm7, which appears to suffer from an accumulation of errors in the very large noncovalent complexes of the s30l - chnof set. the largest outliers in the omx - dn results are found for positively charged complexes, as previously reported for om2-d3 and om3-d3 in the case of the s12l set. the af6 set provides reference folding energies and enthalpies for a series of alkanes. the pm7 values should be compared to the enthalpies, since pm7 was parametrized against heats of formation. in the case of the omx - dn methods, one may argue that the computed values can be considered as enthalpies or as energies, since the parameters for the underlying omx methods and for the dispersion corrections were calibrated against heats of formation and interaction energies, respectively. to be unbiased, we provide data for both types of comparisons, and the results differ only very slightly (see table 13). the pm7 and omx - dn methods have rather small errors both for the folding enthalpies (maes of 0.201.18 kcal / mol, om2-d2 lowest, pm7 highest) and energies (maes of 0.181.05 kcal / mol, om2-d2 lowest, pm7 highest). however, more important than these statistics is a qualitative finding : only om2-d2, om3-d2, and om3-d3 correctly predict that the hairpin conformation of alkanes becomes more favorable than the linear conformation for 17 1 carbon atoms, in agreement with experiment, while the hairpin conformation is more favorable already for alkanes with less than 16 carbon atoms in the case of pm7 and om2-d3. finally, we briefly compare the effect of the different dispersion corrections applied in the omx - dn calculations. in the large majority of cases, the d3 correction turns out to be superior to the d2 correction, albeit often only by a small margin. the role of three - body corrections is negligible in small complexes (a24-chnof, s66, and s66a8 sets) and still quite small but not entirely negligible in hydrogen - bonded and stacked base pairs (jsch-2005-chnof) and in folded alkanes (af6). as expected, three - body corrections become significant in stacked systems modeling interactions between graphene layers (s7l) and in very large noncovalent complexes (s30l - chnof). since om2-d3 and om3-d3 underestimate the attractive dispersion interactions, the repulsive three - body corrections increase the corresponding errors (on average by 0.7 kcal / mol). on the other hand, om2-d3 and om3-d3 tend to overestimate the binding energies in most of the very large noncovalent complexes, and hence the repulsive three - body corrections decrease the error in this case by 0.310.77 kcal / mol for the s30l set, as already reported earlier for the s12l set (om2-d3 and om3-d3) and for the s30l set (om2-d3). full geometry optimizations were carried out for the a24-chnof, s66, s7l, and af6 sets using all sqc methods examined presently. the ab initio reference geometries were taken from published work at the following levels : ccsd(t)/cbs extrapolation for a24-chnof ; counterpoise - corrected mp2/cc - pvtz for s66 with intermolecular distances obtained by interpolating estimated ccsd(t)/cbs energies along dissociation curves ; intermolecular distances optimized at the ccsd(t)/ha - cc - pvdz level with monomer geometries optimized at the b3lyp level with large basis sets for s7l ; and mp2/cc - pvtz for af6. in the original work on the a24 set, three stacked complexes were not optimized but constrained artificially ; therefore they are not considered here. the statistical evaluation of selected optimized interatomic distances and bond angles is presented in table 14 for the pm7 and omx - dn methods and in si table s6 for the other sqc methods. the corresponding statistical data for the associated interaction energies are documented in the supporting information (tables s7 and s8). we note that some of the noncovalent complexes are present in several data sets, with slightly different reference geometries. we start with some general remarks before discussing individual data sets and complexes. the selected distances and angles mostly refer to intermolecular geometrical parameters, e.g., distances defining the separation between the monomers or the length of hydrogen bonds and angles characterizing the relative orientation of the monomers in a complex. the associated minima on the potential energy surface are usually rather extended and flat, and hence the deviations of the sqc results for the geometrical parameters from the ab initio reference values are expected to be much larger than in the intramolecular case. inspection of table 14 confirms that this is indeed the case : the maes are typically of the order of 0.3 for the intermolecular distances and 10 for the angles. the overall statistics in table 14 for the omx - dn methods indicate that the om3-based results are mostly superior to the om2-based results, the d3 correction normally performs better than the d2 correction, and the three - center dispersion correction usually has no significant influence on the geometries of the rather small complexes considered here (d3 vs d3 t). in an overall assessment, om3-d3 outperforms pm7 for the a24-chnof set (maes of 0.26 vs 0.38 and 8.7 vs 12.9) and the s66 set (maes of 0.32 vs 0.41 and 13.2 vs 15.8), while it shows somewhat larger errors for the s7l set (maes of 0.39 vs 0.33) and the af6 set (maes of 0.17 vs 0.09 and 7.1 vs 6.6). om3-d3 works especially well for the dispersion - dominated complexes in the a24-chnof set (maes of 0.02 and 1.2). we now address individual sets and complexes. in the case of the a24-chnof set, all sqc methods considered give qualitatively reasonable complex geometries in general (realistic shape and intermolecular distances within 1 of the reference values), except for mndo which often fails qualitatively and yields much too large intermolecular separations. a problematic case is the hf dimer : the reference geometry has a linear hydrogen bond (f hf), which is also found with pm7 (but much too long) whereas the omx and omx - dn methods give cyclic hydrogen - bonded structures (si figure s2). if the angle f hf is constrained to the reference value, the hf molecules drift apart upon sqc geometry optimization : the hydrogen bond is too long by 0.160.18 at om2 and om2-dn, by 0.510.56 at om3 and om3-dn, and by 1.041.05 at pm6 and pm7. thus, when calculations are run on the reference geometry, the sqc methods either underestimate the binding energy (om2 and om2-dn) or even predict a repulsive interaction (om3, om3-dn, and especially pm6 and pm7 ; see section 5.1). as discussed previously, the methane dimer is also problematic : om3-d3 gives a qualitatively correct geometry, while other sqc methods fail in this regard. the reference interaction energies are derived in the original work on the a24 set without accounting for the deformation of the monomers (i.e., from single - point calculations on monomers at the geometry of the respective complex). however, for small monomers such as those in the a24 set, these deformation energies are negligible, and the interaction energies computed with and without them are essentially the same for all tested sqc methods (si table s27). in the s66 set, the omx - dn methods again give the lowest maes for the distances and angles, but there are some notable outliers, in particular the benzeneacoh (oh-), h2omeoh, c6h6menh2, peptidepeptide, and t - shaped pyridine dimer complexes (for their optimized reference and sqc structures see si figures s3s7). the pm7 geometries are reasonable for some of these complexes, presumably due to the inclusion of special h - bond correction terms (see si figures s3s7). finally, a severe failure of the om3 and om3-dn methods is that they give symmetric structures for carboxylic acid dimers, in which two hydrogen atoms are equidistant to the oxygen atoms in the corresponding hydrogen bonds. this is found for the acetic acid dimer in the s66 set and for the formic acid dimer in the validation of omx and omx - dn methods. since the monomer moieties in such dimers are strongly distorted, their interaction energies are strongly overestimated (see si table s28 for acetic acid dimer). in the s7l set, pm7 gives overall the best intermolecular distances ; om3-d3 has slightly smaller errors for the hh distances. a specific problem is that pm7 optimizes the parallel displaced naphthalene dimer to the sandwich conformation, contrary to omx - dn. in the af6 set, the overall statistical errors of the pm7 and omx - dn methods are rather similar, with slight advantages for pm7. all sqc methods without explicit dispersion corrections fail qualitatively since mndo, am1, pm6, and the omx methods predict the most favorable conformation to be linear for all alkanes while pm3 predicts the opposite. only the om2-d2 and om3-dn methods give the correct result that the linear conformation dominates for short chain lengths and that the hairpin conformation becomes more favorable for alkanes with 17 1 carbon atoms, in agreement with experiment. at the end of such a large and diverse benchmark study, it is impossible to provide a single unbiased grand error estimate for each sqc method. the chosen benchmark sets strongly differ in their size and in the quality of the reference data, and they address different properties. thus, instead of attempting to provide such grand error estimates, we summarize the results for all subsets and identify the methods which are best for a given subset / property combination and which are most robust in general. this information can be used as a guide for selecting the sqc method that performs best for applications of the kind considered here. tables 15 and 16 present the benchmark results for ground - state properties and noncovalent interactions, respectively ; they list the two top - performing methods with their maes for all data sets and subsets (as described in tables 1 and 2). maes for energies in kcal / mol, for bond lengths in, for angles in degrees, for ionization potentials in ev, and for dipole moments in d. if maes differ by less than ca. 0.1 kcal / mol for energies or 0.01 for bond lengths, the corresponding methods are listed together. mae of dispersion - corrected counterpart (om2-dn or om3-dn) is very close (within 1.0 kcal / mol for energies and within 1.0 for angles) and may actually be slightly lower. without the isomerization energy of c20 ; see text. only the omx - dn and pm7 methods are considered here since the other sqc methods do not adequately describe noncovalent interactions. if maes differ by less than 0.1 kcal / mol for energies or 0.01 for distances or 1.0 for angles, the corresponding methods are listed together. without the hfhf complex single - point calculations on reference geometries. deviations upon optimization with sqc methods. most of the entries in table 15 involve one of the omx or omx - dn methods, which appear to perform best overall for the large variety of ground - state molecules and properties considered presently. as a rule of thumb, the omx and omx - dn methods can thus be recommended as default for ground - state sqc studies, especially when a specific validation against experimental data or high - level calculations is not feasible. common organic molecules are also described with good accuracy by pm6 and pm7 because of their extensive parametrization. this indicates that sqc methods with orthogonalization corrections still have a large potential for improvement when larger data sets are used for their parametrization ; this is one target of our current research. the omx - dn and pm7 methods have similar overall accuracy for noncovalent binding energies and geometries of noncovalent complexes. the omx - dn methods generally outperform pm7 for dispersion - dominated complexes and mixed electrostatic / dispersion interactions, while pm7 is normally superior for hydrogen - bonded complexes ; the latter can be attributed to special hydrogen - bond corrections that are present in pm7 but not in the omx - dn methods. while it has been shown for water clusters that the description of hydrogen bonds can be improved by a special parametrization of omx methods, it would seem more promising to target a complete reparametrization at the omx - dn level that includes noncovalent interaction energies and geometries as reference data. | the semiempirical orthogonalization - corrected omx methods (om1, om2, and om3) go beyond the standard mndo model by including additional interactions in the electronic structure calculation. when augmented with empirical dispersion corrections, the resulting omx - dn approaches offer a fast and robust treatment of noncovalent interactions. here we evaluate the performance of the omx and omx - dn methods for a variety of ground - state properties using a large and diverse collection of benchmark sets from the literature, with a total of 13035 original and derived reference data. extensive comparisons are made with the results from established semiempirical methods (mndo, am1, pm3, pm6, and pm7) that also use the nddo (neglect of diatomic differential overlap) integral approximation. statistical evaluations show that the omx and omx - dn methods outperform the other methods for most of the benchmark sets. |
electroconvulsive therapy (ect) is one of the most efficacious treatment of major depressive disorder (mdd). it is used as a rapid and efficacious treatment in emergent and/or treatment resistant forms of other psychiatric disorders. the cognitive aspects which are influenced immediately after ect are : orientation, information processing, retrograde and anterograde amnesia, visuospatial capacity and word finding. donepezil in a case report study galantamine in study without control group, a glucocorticoid antagonist (cort108997) in another study, thyroid hormones, antihypertensive, poropofol, physosigmine, naloxone, piracetam showed some benefits in preventing ect - induced cognitive impairment ; but many of those have not enough reliability because were not statistically significant, or were case reports, unblanded, uncontrolled or on animals. the other studies did not show efficacy of the used drugs in prevention of memory impairments. considering these non sufficient and controversial data, more structured studies with control group on human samples are necessary. so, we chose the so - called memoral herbal for our this preventive new study, which has never been used for this propse. the memoral herbal capsules, each contains 360 mg of boswellia oleo - gum resin and 36 mg of zimgiber rhizome. the most important constituent of boswellia is gum resin 60%, mosilage 20 - 23% and essence 5 - 9%, which contains a - b - thujon, p - cymen and linanol. boswellia serrata has boswellic acid and acetyl- 11- keto - beta boswellic acid too, which are responsible for many therapeutic effect. the volatile oils in boswellia dilate the vasculator of the brain thus can increase the blood passage through the brain. boswellia extract has been demonstrated by a battery of rigorous tests to have anti - inflammatory effect. the mechanism by which boswellia can improve memory is through its anti - inflammatory effect on the brain. the aim of this study is to compare memoral herbal with placebo in prevention of ect - induced cognitive impairments. this study is a randomized and controlled clinical trial which was done during 9 months of 2011 at noor hospital (isfahan, iran). the sample size was 70 patients, based on 80% power and 95% confidence interval. each consequent patient placed randomly in intervention (memoral) or control (placebo) group, i.e., 35 patients in each group. the inclusion criteria were : 15 - 65ys / o, diagnosis of major depressive disorder (mdd) or bipolar disorder (bd) according to the impression of therapist (academic member) documented in the patient file, prescribed at least 4 ect sessions by the therapist and informed consent for participation. exclusion criteria were : mental retardation (mr) or dementia as documented in hospital file by psychiatrist, discontinuation of ect before fourth sessions, discontinuation of participation before the end of study, and arising delirium during the course of study. the ethical issues of this study was approved by ethical committee of behavioral sciences research center (bsrc) of isfahan university of medical sciences (iums), iran. the study process registered at iranian registering of clinical trials office, with irct i d : irct201102072232n2. the study was illustrated to patients and an informed consent form was signed by each patient or his / her first degree related person who was supporting him / her (in cases the patient was unable to decide). each patient received three capsules (tid) of either memoral or placebo from the day before first ect session until 2 months after the last session of ect. memoral and placebo capsules were produced by goldaru pharmaceutical manufactory (isfahan, iran). the capsules were labeled by only a or b brand and were taken to patients by a nurse. this scale was developed through extension of mini mental status examination (mmse) memory, language and visuospacial ability subscales, and adding verbal fluency subscale to it. it is translated to persian and validated by pouretemad h.r.(2006). its sensitivity and specificity for screening normal persons from mild cognitive impairment (mci) is 93% and 91%, respectively ; for screening alzheimer patients from mci patients is 73% and 93%, respectively, and for screening normal persons from alzheimer patients is 100% and 96%, respectively. it has five subscales : attention and orientation, memory, language, verbal fluency, visuospatial ability. also it has the capacity for scoring based on the past so - called mmse scale. the patients cognitive function was assessed at four occasions : before the first ect session, before the 4 ect session, 1 month and 2 months after the last ect session, by a psychologist. the probable side effects of memoral were also evaluated at those occasions, using a checklist. the findings were analyzed by repeated measure of analysis of variance (anova) and pearson chi - square, using spss18. this study is a randomized and controlled clinical trial which was done during 9 months of 2011 at noor hospital (isfahan, iran). the sample size was 70 patients, based on 80% power and 95% confidence interval. each consequent patient placed randomly in intervention (memoral) or control (placebo) group, i.e., 35 patients in each group. the inclusion criteria were : 15 - 65ys / o, diagnosis of major depressive disorder (mdd) or bipolar disorder (bd) according to the impression of therapist (academic member) documented in the patient file, prescribed at least 4 ect sessions by the therapist and informed consent for participation. exclusion criteria were : mental retardation (mr) or dementia as documented in hospital file by psychiatrist, discontinuation of ect before fourth sessions, discontinuation of participation before the end of study, and arising delirium during the course of study. the ethical issues of this study was approved by ethical committee of behavioral sciences research center (bsrc) of isfahan university of medical sciences (iums), iran. the study process registered at iranian registering of clinical trials office, with irct i d : irct201102072232n2. the study was illustrated to patients and an informed consent form was signed by each patient or his / her first degree related person who was supporting him / her (in cases the patient was unable to decide). each patient received three capsules (tid) of either memoral or placebo from the day before first ect session until 2 months after the last session of ect. memoral and placebo capsules were produced by goldaru pharmaceutical manufactory (isfahan, iran). the capsules were labeled by only a or b brand and were taken to patients by a nurse. this scale was developed through extension of mini mental status examination (mmse) memory, language and visuospacial ability subscales, and adding verbal fluency subscale to it. it is translated to persian and validated by pouretemad h.r.(2006). its sensitivity and specificity for screening normal persons from mild cognitive impairment (mci) is 93% and 91%, respectively ; for screening alzheimer patients from mci patients is 73% and 93%, respectively, and for screening normal persons from alzheimer patients is 100% and 96%, respectively. it has five subscales : attention and orientation, memory, language, verbal fluency, visuospatial ability. also it has the capacity for scoring based on the past so - called mmse scale. the patients cognitive function was assessed at four occasions : before the first ect session, before the 4 ect session, 1 month and 2 months after the last ect session, by a psychologist. the probable side effects of memoral were also evaluated at those occasions, using a checklist. the findings were analyzed by repeated measure of analysis of variance (anova) and pearson chi - square, using spss18. of 70 patients, two patients from memoral group and five patients from placebo group were excluded because of premature discontinuation of ect and discharge from the hospital, causing disruption of participation. so 33 patients in memoral group, and 30 patients in placebo group remained and there was not any significant difference between two groups (p = 0.95). considering educational levels, there was not any significant difference between two groups (p = 0.18) : including 6 (9.5%) patients had primary school studies, 23 (36.5%) guidance school, 24 (38.1%) high school, 10 (15.9%) had university studies. the frequency of patients diagnosis was : 26 (41.3) patients with bd manic or mixed episode, 13 (20.6%) patients bd depressive episode, 24 (38.1%) patients with mdd. there was not any significant difference between memoral and placebo group about diagnoses distribution (p = 0.09). also there was no significant difference between the number of ect sessions in two groups (pv=0.95%) ; included 2 (3.2%) patients received five ect sessions,25 (39.7%) received six sessions, four (6.3%) received seven sessions, 17 (26.9%) received eight sessions, two (3.2%) patients received nine sessions, 8 (12.7%) received 10 sessions, two (3.2%) patients received 11 sessions, two (3.2%) patients received 12 sessions, and one (1.6%) patient received 14 ect sessions. the mean energy of ect used in memoral group was 25.8 joule in memoral group, and 25.9 joule in placebo group (p = 0.98).the mean age of patients in memoral group was 33.8 ys / o, and 32.8 ys / o in placebo group, with no significant difference between them (p = 0.17).we did not find any important side effects that could exclude patients from the study, and there was not any significant difference between two groups for this issue (p = 0.30). there was not any significant difference between two groups between total score of ace of memoral group (76) and placebo group (79.4) before intervention (p = 0.32). also, there was not any significant difference between two groups for the scores of attention and orientation subscales (p = 0.64), memory (p = 0.75), verbal fluency (pv = 0.35), language (p = 0.11), and mmse subscales (p = 0.40) before intervention. table 1 shows the mean sd of the total score of ace and the scores of ace subscales in memoral and placebo groups at four assessment occasions, i.e., before intervention, before the 4 ect session, 1 month and 2 months after the last ect sessions [figure 1 ]. the mean sd of the total score of ace and the scores of ace subscales in memoral and placebo groups at four assessment occasions : before intervention, before 4 ect session, 1 month and 2 months after last ect sessions the mean of the total score of ace in memoral and placebo groups at four assessment occasions : before intervention, before 4 ect session, 1month and 2months after last ect sessions repeated measure of anova showed that the total score of ace and the scores of attention and orientation subscale, memory, verbal fluency and mmse subscales were significantly higher in memoral group than in placebo group after intervention, and also were uprising, unlike placebo group [table 1, figure 2 ]. but the scores of the language, and visuospatial ability subscales did not show any significant difference between two groups, although these were uprising in memoral group, unlike in placebo group [table 1 ]. the paired comparison of each subscales in two groups also showed significant difference at each stage. the mean of the memory subscale score of ace in memoral and placebo groups at four assessment occasions : before intervention, before 4 ect session, 1month and 2 months after last ect sessions this study assessed the efficacy of memoral herbal on prevention of ect - induced cognitive impairments, using ace scale. the total score of ace and the subscales of memory, attention and orientation, verbal fluency and mmse in memoral group were significantly higher than in placebo, and did not declined by ect unlike in placebo group. the noticeable finding was the largest amount of difference for memory subscale in this intervention [table 1, figure 2 ]. this study showed that memoral herbal prevents ect - induced memory impairment more rapidly and more impressively than other drugs used in previous similar studies.[313 ] unlike previous studies[313 ] in this intervention, we assessed cognitive status in more detailed and itemized description. so these data may have more reliability and scrupulosity than previous findings, and may somehow interpret the inconsistency of some previous findings.[1215 ]. the sample size of this study is another precedence of this study over previous ones. this was some deal because of the blinding goals and the independence of researchers from therapists. the future studies on prevention of ect - induced cognitive impairments may comprise memoral herbal with other drugs in a research, and on patients with other diagnoses. our work showed the effective prevention of ect - induct cognitive impairment by using memoral herbal. the memory, attention and orientation, verbal fluency, and mmse of patients showed improvement by memoral use. | background : electroconvulsive therapy (ect) is one of the most efficacious treatment for major depressive disorder (mdd), it is also used as a rapid and efficacious treatment for other psychiatric disorders, especially treatment resistant ones. the cognitive impairment is one of the most important side effects of ect. this study examined the memoral herbal efficacy in prevention of ect - induced memory impairment.methods:in a randomized clinical trial, 70 patients with mood disorders who were candidates for ect enrolled in either memoral or control group, and received either memoral or placebo. the memory was assessed by addenbrook cognitive examination (ace), and the findings were analyzed by anova under spss18.results:the memoral group patients showed significantly higher total ace scores than placebo group (p < 0.001). the scores of attention and orientation, verbal fluency and memory subscales not only never decreased during the study in memoral group, but also increased. there was no significant difference between these scores of memoral and placebo groups for the subscales of language and visuospacial ability.conclusion:the memoral herbal is an efficacious and safe choice in prevention of ect- induced cognitive impairment. |
the off - season period (i.e. transition period) has taken a relevant role across the soccer s world. considering the growing economic importance of soccer competitions and the increase of the number of matches around the world (regardless of the competitive - level), soccer referees have been involved in officiating during most of the year (da silva., subsequently, the transition period is shorter and soccer referees have only few weeks to prepare to the competitive season. given that refereeing is a very demanding activity both physically and physiologically (mallo., 2008), soccer referees must ensure a high level of physical fitness. indeed, field referees (fr) were reported to cover a total distance of 10 - 12 km (krustrup and bangsbo, 2001 ; weston., 2007) and assistant referees (ar) of 6 - 8 km during competitive matches (krustrup., 2002 ; mallo., 2008). furthermore, fr and ar may perform as much as 1,269 and 1,053 activity changes during a match, respectively, with fr undertaking 21.3 - 30.5 sprints at a speed above 25.2 kmh-1 (krustrup and bangsbo, 2001), whereas ar run up to 20 sprints per match (krustrup., 2002). given that match officials must be physically fit to keep up with the game tempo to make appropriate decisions (weston., 2012), referees international and national governing boards require the evaluation of fr and ar s physical fitness at the start of the competitive season with the pass of set limits to be included in the seasonal match appointment list. as a result, the seasonal evolution of the physical fitness of match officials is currently considered as a key methodological issue in modern soccer (castagna., only few studies have examined seasonal variations in physical performance of elite level frs with no research report addressing ar (weston. furthermore, no study has focused on the training transition period or the part of the season that spans from the end of the previous competitive season to the new one. considering that there is no information regarding the physical fitness of elite level match officials during the off - season period, a better understanding of the evolution of their physical fitness during this period would help adjust their training programs. therefore, it would be interesting to determine the effects of this transition period on fr and ar s physical performance. in soccer, several studies focused on the off - season period (miller., 2011 ; (2011) observed that soccer players maintained a relatively high fitness level, while other researchers (amigo., 1998 ; caldwell and peters, 2009) reported a loss of physical performance during the off - season period. as a result, no direct evidence is currently available on how to conduct an effective transition training - period in soccer players and match officials. therefore, the aim of this study was to analyze the effects of the transition period (i.e. june to august) on physical performance variables (i.e. linear straight sprint, change of direction ability and endurance) in national soccer division referees. forty - five spanish soccer referees (29.6 7.8 yr, 73.3 7.6 kg, 175.9 5.6 cm and 23.7 2.1 kgm) that officiated during official soccer matches of the spanish national division during the 2014/2015 season volunteered to participate in this study. participants were classified according to competitive status, fr (n = 23, 30.0 6.7 yr, 73.3 8.1 kg, 176.8 6.1 cm and 23.4 1.8 kgm) and ar (n = 22, 29.2 8.9 yr, 73.3 7.3 kg, 175.0 5.6 cm and 23.9 2.3 kgm). the study was performed in accordance with the declaration of helsinki (2013), was approved by the university of the basque country (upv / ehu) ethics committee and met the ethical standards in sport and exercise science research. the study was designed to determine the effects of a 9 week off - season period on the results of relevant fitness tests evaluating linear sprint, change of direction and intermittent high - intensity performance in fr and ar (castagna. tests were performed at the end of the season (t1, in june) and at the start of the pre - season (t2, in august) at the same venue. during the off - season period, soccer referees trained following the recommendations designed by a professional in sports sciences who worked as a personal trainer for the spanish committee of soccer referees during the competitive season. the training program recommended three weeks of low intensity activities (i.e. walking or cycling) followed by five more weeks focusing on low intensity running (under 70% of the individual maximum heart rate, hrmax) and exercise, (i.e. paddle or swimming). tests were preceded by a standard warm - up consisting of 7 min of slow jogging, followed by progressive sprints and static stretching. referees undertook, in this order, a 30-m straight sprinting test, a modified agility t - test (matf) and the yo - yo intermittent recovery level 1 test (yyir1). the linear straight sprint and the matf eight minutes of standardised recovery for all participants were allowed between the matf and the yyir1. linear straight sprinting test : each referee performed an acceleration test consisting of three maximal sprints of 30 m length (krustrup., 2002), with a 90 s rest period between each sprint. participants were asked to place themselves 0.5 m from the starting point and began when they felt ready. split time at 20 m and the time to cover the 30 m were measured. modified agility test free (matf) : the referees completed the protocol by yanci. all participants performed the test three times with at least three minutes of rest between trials. the total distance covered was 20 m. a photocell (microgate polifemo, bolzano, italy) was used to record the time. participants were asked to begin 0.5 m behind the starting line (point a, figure 1) and sprint forward as fast as possible, touching with one hand and in this order the top of cone b, c, d and b, and finally return to line a. modified agility t - test free (matf) design. yo - yo intermittent recovery level 1 test (yyir1) : the yyir1 consisted of 2 x 20 m runs back and forth between two lines at a progressively increasing speed controlled by audio beeps. when the participants twice failed to reach the corresponding line in time, the distance covered was recorded and represented the test result. each bout was interspersed with a 10 s active rest period consisting of 2 x 5 m of jogging. during the test, the heart rate (hr) was recorded every 1 s using the polar team system (polar electro oy, kempele, finland). the individual maximum hr (hrmax) was determined as the peak value reached during the test. at the end of the test the data were downloaded to a computer and processed using polar precision 2.0 software (polar, kempele, finland). one minute after finishing the yyir1, tympanic temperature (thermoscan 5 irt 4520, braun gmbh, krongerg, germany) was measured (hamilton., 2013). the 0 - 10-point scale (foster., 2001) of the respiratory rate of perceived exertion (rperes) (green., 2009) and the leg muscular rate of perceived exertion (rpemus) (los arcos., 2014) were used to determine the subjective perception of fatigue. vo2max was estimated from the following equation : vo2max (mlminkg) = yyir1 distance (m) 0.0084 + 36.4 (bangsbo., 2008). normal distribution was tested using the kolmogorov - smirnov test and statistical parametric techniques were applied. a t test for paired samples was used to analyze the differences between the end of the season (t1) and the start of the preseason (t2) independently for each group (fr, ar and total sample). both in t1 and t2, a paired t - test for independent samples was used to compare results between fr and ar. the in - between groups (fr and ar) comparison from t1 to t2 effect sizes (d) of above 0.8, between 0.8 and 0.5, between 0.5 and 0.2 and lower than 0.2 were considered as large, moderate, small and trivial, respectively. differences between means were expressed as percentages. statistical significance was set at p 0.05, es = trivial to small) between group differences were detected at t1 for the 30 m test and matf1 (table 1). moderate differences between groups (es = 0.70) for the matf were detected at t2. a pre - to - post loss of performance (p 0.05), in yyir1 performance (es = 0.97, p < 0.05) and vo2max (es = 0.97, p < 0.05) at t2. furthermore, despite a higher reported rperes score in ar, rpemus values were lower than that for fr in t2. only fr showed significantly better results in the distance covered and vo2max in the yo - yo intermittent recovery test (level 1) after the offseason period. therefore, while fr improved their cardiovascular performance, ar tended to obtain worse results in the yo - yo intermittent recovery test (level 1). it would be interesting to implement specific endurance programs especially in ar during this period in order to not decrease the resistance capacity during the off - season period. match officials in soccer have to possess an optimum level of physical fitness at the start of the competitive season to warrant their eligibility to be appointed for championship matches and for this reason during the transition period soccer referees do not stop their physical training completely. even though both fr and ar must complete 20 intervals consisting of 150 m running in 30 s, the former have a recovery time of 30 s to walk 50 m, whilst for the latter, the recovery time is longer (16.6%) (mallo., 2009). it is likely that fr had trained their cardiovascular capacity more during the off - season because generally they are responsible for making key decisions to ensure the proper course of the game and consequently they are required to keep up with the play at the start of the competitive season. however, ar do not need such rigorous physical preparation to develop their refereeing activity in competition. we could conclude that soccer referees decreased acceleration capacity after 9 weeks of the off - season period, suggesting that it would be interesting to analyze whether specific training programs would compensate for this loss of acceleration performance. it could be speculated that implementing specific lower - limb power training throughout the transition period could compensate the loss of acceleration performance in fr and ar. furthermore, ar transition training should focus more on endurance training and/or consider more specific testing of endurance (castagna., given that a strong relationship has been observed between physical performance and match activity (weston., 2009), this study provides coaches with information useful to guide match official preparation during the off - season. | abstractthe evolution of referees physical fitness has been studied over one or several seasons, however, the variation of the physical performance between the end of the competitive season (t1) and the start of the following pre - season (t2) has not been ascertained. therefore, the aim of this study was to analyze the effects of the transition period on physical performance variables (i.e. linear straight sprint, change of direction ability and endurance) in national soccer division referees. forty - five spanish referees volunteered to participate in this study. participants were classified according to competitive status, field referees (fr, n = 23) and assistant referees (ar, n = 22). a loss of performance (p < 0.05) was observed in the 20 and 30 m linear straight sprint between t1 and t2 in both fr (1.64 - 1.56%, d = 0.29 to 0.32) and ar (2.01 - 3.41%, d = 0.33 to 0.60). in t2 the fr significantly improved the distance covered (p < 0.05, 13.11%, d = 0.39) in the yo - yo intermittent recovery test (yyir1). besides, significant differences were observed between fr and ar in the distance covered (p < 0.05, 23.55%, d = 0.97) in the yyir1 test in t2. more research may be necessary to focus on the off - season period in order to implement specific training programs and consequently reduce the loss of sprint ability in field and assistant referees and the decrease in cardiovascular fitness in assistant referees. |
prevalence and risk of overweight were assessed in 232 children who were enrolled in the prospective german gdm offspring study between 1989 and 2000. all children were from singleton births, had a mother with gdm diagnosed according to criteria of the german diabetes association using an oral glucose tolerance test (ogtt) with a 75-g glucose load. women were considered to have gdm if two of three capillary blood glucose values exceeded the following limits : > 5 mmol / l (fasting) before an oral glucose tolerance test, > 10.0 mmol / l after 60 min, and 8.6 mmol / l after 120 min. prevalence of overweight was compared with that of a cohort of ot1d (n = 757) and offspring of nondiabetic mothers (ondm) (n = 431) followed in the babydiab study (6). in both the gdm and babydiab studies, mothers and their offspring were recruited between 1989 and 2000 germany - wide through the same network of collaborating obstetric departments and pediatricians and entered into the study before the offspring reached the age of 3 months. all children were followed with similar follow - up visits until the age of 14 years. in both studies, > 98% of the families were caucasian. in families with more than one offspring participating in the study, data on weight and height were collected at birth and at age 2, 8, and 11 years by physicians. at birth, length was measured on a measuring board and weight on a calibrated scale by health care professionals. weight and height were obtained for 215 children at age 2 years, 89 children at age 8 years, and 74 children at age 11 years. birth weight was adjusted for sex and gestational age and is expressed as a percentile of the german reference population (8). offspring less than the 10th percentile were defined as small for gestational age, those 90th percentile as large for gestational age and those between > 10th and 10th and < 90th percentile as appropriate for gestational age. from age 2 years, height and weight were measured by physicians at regular clinical visits using a standard protocol. weight was measured on a calibrated scale with the subject wearing light clothing, and height was measured with a fixed tape standing barefoot. height, weight, and bmi from age 2 years were expressed as percentiles adjusted for age and sex according to german reference data (9). maternal height and weight in early pregnancy were collected by physicians at the first pregnancy visit, bmi was calculated and categorized as obese (30 kg / m), overweight (25.029.9 kg / m), or normal weight (< 25 kg / m). data on diabetes treatment (categorized as treated by insulin injections or) and maternal smoking behavior during pregnancy (defined as < or 1 cigarette / day) were obtained from a questionnaire at birth. written informed consent since 2003, fasting blood samples were collected for the determination of insulin resistance in ogdm as well as in offspring from the babydiab study. insulin resistance was measured by homeostasis model assessment of insulin resistance (homa - ir). a total of 751 children (including 74 ogdm and 425 ot1d) with data on homa - ir either at age 8 or 11 years were included in this substudy. fasting insulin was determined centrally using an automated immunoassay analyzer (aia 360 ; tosoh, san francisco, ca). the interassay coefficient of variation of the insulin assay is 8.4% at a concentration of 22.6 u / ml and the lower limit of detection is 0.5 u / ml. overweight prevalence in offspring at age 2, 8, and 11 years was compared between groups using the test. logistic regression analyses were performed to test the association of maternal bmi at early pregnancy, birth size, maternal smoking during pregnancy, treatment modality during pregnancy, and breast - feeding duration as univariate covariates with overweight risk in ogdm, giving the odds ratio (or) and 95% ci for overweight for each of the covariates. factors that were significantly associated with overweight risk in the univariate analysis were included as covariates in the multivariate logistic regression analysis. missing data were not considered in the univariate analyses but were included as a separate categorical value for each of the variables included in the multivariate analysis. linear regression was used to compare homa - ir in ogdm with those in ot1d and in ondm adjusted for age and sex and to analyze factors associated with homa - ir in ogdm. for all analyses, two - tailed p < all statistical analyses were performed using spss 17.0 (spss, chicago, il). prevalence of overweight in ogdm was 17.2% at age 2 years, 20.2% at age 8 years, and 31.1% at age 11 years. prevalence of overweight was increased in ogdm compared with ot1d (15.8, 11.0, and 15.8% at age 2, 8, and 11 years, respectively ; p < 0.05, p = 0.03, and p < 0.01, respectively) and with ondm (11.4, 10.3, and 15.5% ; p = 0.7, p = 0.02, and p = 0.005, respectively (fig. 1). prevalence of overweight at 2, 8, and 11 years of age in ogdm (), ot1d (), and ondm (). in ogdm, maternal obesity was a strong predictor of overweight in children at age 2, 8, and 11 years in the univariate and multivariate logistic regression model (tables 1 and 2). the prevalence of overweight in children at 2, 8, and 11 years of age was 24.6, 36.4, and 45.8% in children of obese mothers compared with 9.2, 11.3, and 11.9% of children born to nonobese mothers (p = 0.01, p = 0.02, and p = 0.003, respectively) (fig. birth size, therapy of gdm during pregnancy, and maternal smoking during pregnancy showed inconsistent associations with overweight risk in the offspring (tables 1 and 2). associations of maternal bmi in early pregnancy, birth size, therapy modality, and smoking behavior during pregnancy on overweight - risk (bmi 90th percentile) in ogdm at 2, 8, and 11 years of age : univariate analysis aga, appropriate for gestational age ; lga, large for gestational age ; sga, small for gestational age. associations of maternal obesity in early pregnancy, lga status, and maternal smoking during pregnancy on overweight risk in ogdm at 2, 8, and 11 years of age : multivariate analysis variables included in the multivariate analysis : maternal bmi before pregnancy, birth size, and maternal smoking during pregnancy. prevalence of overweight at 2, 8, and 11 years of age in ogdm in relation to maternal bmi in early pregnancy : bmi < 25.0 kg / m (), bmi 25.029.9 kg / m (), and bmi 30 kg / m (). in the substudy on insulin resistance, ogdm had increased homa - ir compared with that of children of ondm and of ot1d (p = 0.04 and p = 0.03, adjusted for age and sex) (fig. homa - ir was associated with the child 's bmi (p = 0.01, adjusted for age and sex) (supplementary fig. 1a, available in an online appendix at http://care.diabetesjournals.org/cgi/content/full/dc10-0139/dc1) but not with maternal obesity in early pregnancy (supplementary fig. 1b). homa - ir (mean 1 se) at age 11 years in ogdm compared with ot1d and ondm. prevalence of overweight in ogdm was 17.2% at age 2 years, 20.2% at age 8 years, and 31.1% at age 11 years. prevalence of overweight was increased in ogdm compared with ot1d (15.8, 11.0, and 15.8% at age 2, 8, and 11 years, respectively ; p < 0.05, p = 0.03, and p < 0.01, respectively) and with ondm (11.4, 10.3, and 15.5% ; p = 0.7, p = 0.02, and p = 0.005, respectively (fig. 1). prevalence of overweight at 2, 8, and 11 years of age in ogdm (), ot1d (), and ondm (). in ogdm, maternal obesity was a strong predictor of overweight in children at age 2, 8, and 11 years in the univariate and multivariate logistic regression model (tables 1 and 2). the prevalence of overweight in children at 2, 8, and 11 years of age was 24.6, 36.4, and 45.8% in children of obese mothers compared with 9.2, 11.3, and 11.9% of children born to nonobese mothers (p = 0.01, p = 0.02, and p = 0.003, respectively) (fig. birth size, therapy of gdm during pregnancy, and maternal smoking during pregnancy showed inconsistent associations with overweight risk in the offspring (tables 1 and 2). associations of maternal bmi in early pregnancy, birth size, therapy modality, and smoking behavior during pregnancy on overweight - risk (bmi 90th percentile) in ogdm at 2, 8, and 11 years of age : univariate analysis aga, appropriate for gestational age ; lga, large for gestational age ; sga, small for gestational age. associations of maternal obesity in early pregnancy, lga status, and maternal smoking during pregnancy on overweight risk in ogdm at 2, 8, and 11 years of age : multivariate analysis variables included in the multivariate analysis : maternal bmi before pregnancy, birth size, and maternal smoking during pregnancy. prevalence of overweight at 2, 8, and 11 years of age in ogdm in relation to maternal bmi in early pregnancy : bmi < 25.0 kg / m (), bmi 25.029.9 kg / m (), and bmi 30 kg / m (). in the substudy on insulin resistance, ogdm had increased homa - ir compared with that of children of ondm and of ot1d (p = 0.04 and p = 0.03, adjusted for age and sex) (fig. homa - ir was associated with the child 's bmi (p = 0.01, adjusted for age and sex) (supplementary fig. 1a, available in an online appendix at http://care.diabetesjournals.org/cgi/content/full/dc10-0139/dc1) but not with maternal obesity in early pregnancy (supplementary fig. homa - ir (mean 1 se) at age 11 years in ogdm compared with ot1d and ondm. this study shows an increased prevalence of overweight at 2, 8, and 11 years of age in ogdm compared with ot1d and ondm. in ogdm, overweight up to age 11 years was strongly associated with maternal obesity at early pregnancy. birth size of the child, maternal smoking during pregnancy, or treatment modality of gdm were less consistent predictors of overweight in children of mothers with gdm. this gdm offspring study is a prospective study from birth, in which questionnaires addressing in pregnancy - related factors were administered at birth and children 's weight and height were measured regularly by a pediatrician according to standard protocols. recall bias and over- or underestimating of growth that may occur when data are reported by parents are therefore avoided or limited. our results are based on the definition of overweight as bmi percentile 90, which has been proposed by the german association of obesity during childhood and obesity. findings were consistent when overweight was defined as bmi 85 or 95 percentile (data not shown). although not population based, this is the largest prospective study following caucasian children of mothers with gdm from birth in defined follow - up intervals, enabling a longitudinal analysis of factors affecting overweight risk. nevertheless, the numbers of children of mothers with gdm remains relatively low at ages 8 and 11 years. as a consequence, cis for some of the risk estimates are wide, and some associations may change with a larger number of subjects. a small number of studies have examined the impact of maternal diabetes on overweight risk in offspring, differentiating between ogdm and offspring of mothers with preexisting type 1 diabetes (10,11). clausen. (11) reported an increased prevalence of overweight in adults who were from gdm pregnancies (40%) or type 1 diabetes pregnancies (41%) compared with control subjects (24%). (10) found only a minor increase in the prevalence of overweight in ogdm children (30%) compared with control children (23%) and ot1d (23%). the findings of lawlor. support our previous report that ot1d are not at higher risk for overweight compared with ondm (6). however, in the current study, we saw a pronounced difference between ogdm and ot1d, whereas differences are not significant in the study of lawlor. to reconcile this discrepancy, it is noted that our study in germany reported a lower background prevalence of overweight in children compared with the u.s. and that the numbers of ot1d and ogdm are larger in our study. our findings are unlikely to be biased with respect to the comparisons between ogdm and ot1d children because both cohorts were recruited over the same period and from the same region and ethnic background. they are also supported by expected associations of maternal bmi with overweight risk in the ogdm. however, data for potential confounders such as socioeconomic status, dietary habits during childhood, and physical activity were not collected, and we can not exclude the possibility that the associations observed could, in some cases, be due to one or more of these confounding variables. the finding that overweight risk is significantly higher in ogdm compared with ot1d indicates that exposure to hyperglycemia during pregnancy per se can only partly explain the increased prevalence of overweight in these children. maternal obesity in early pregnancy was the strongest predictor of overweight risk at 2, 8, and 11 years in ogdm. others have shown that maternal pregravid bmi is the strongest predictor of childhood obesity independent of maternal glucose status or birth weight (7). it has also been shown that childhood bmi correlates more closely with maternal bmi than does paternal bmi, indicating that in addition to genetic influences an obese intrauterine environment per se may contribute to the higher overweight risk in children of obese mothers (12,13). this finding is strengthened by results from animal studies showing that feeding - induced obesity at conception increases the prevalence of obesity in offspring. programming of obesity in animals was independent of changes in birth weight and was associated with significant changes in metabolic and endocrine parameters and adipose tissue cellularity independent of postnatal caloric intake (13). finally, the possibility that the influence of maternal obesity on childhood overweight could also be due to similar adverse dietary and physical activity behaviors in obese mothers and their offspring should be considered. similarly, the observation that maternal smoking during pregnancy is associated with overweight in the child could further indicate an influence via an unhealthy postnatal environment. the relationship between childhood obesity and both maternal obesity and smoking during pregnancy became more pronounced with age, further giving support to this hypothesis. not unexpectedly given the increased risk for overweight in ogdm, our substudy found higher homa - ir in ogdm than in ondm and also in ot1d. (14) who also reported a relationship between offspring bmi and insulin resistance in a small group of 23 female ogdm. a recently published study found that in offspring of mothers with pregravid obesity fetal insulin resistance strongly correlated with fetal adiposity (15). we could not find a significant association between insulin resistance in the offspring and maternal obesity in early pregnancy, but numbers were relatively small. in summary, our results show that prevalence of overweight and, as a consequence, homa - ir during childhood is higher in ogdm than in ot1d, indicating that maternal diabetes type affects overweight risk. the finding that overweight risk was associated with maternal obesity and to a lesser extent with birth size suggests that a combination of genetic predisposition, fetal overnutrition and lifestyle factors are likely to contribute to childhood growth in ogdm. | objectivegestational diabetes mellitus (gdm) is associated with high birth weight in the offspring. this may lead to overweight and insulin resistance during childhood. the aim of the study was to assess the impact of gdm on overweight risk and insulin resistance in offspring.research design and methodsbmi measurements were collected at age 2, 8, and 11 years from 232 offspring of mothers with gdm (ogdm) and compared with those from 757 offspring of mothers with type 1 diabetes (ot1d) and 431 offspring of nondiabetic mothers (ondm) born between 1989 and 2000. insulin resistance (homeostasis model assessment of insulin resistance [homa - ir ]) was determined at age 8 and 11 years in 751 children (74 ogdm). overweight was defined as bmi percentile 90 ; insulin resistance was defined by homa-ir.resultsoverweight prevalence was increased in ogdm compared with ot1d and to ondm throughout childhood (age 11 years 31.1, 15.8, and 15.5% ; p = 0.005). maternal obesity was an important predictor of overweight risk in children (age 11 years odds ratio 7.0 [95% ci 1.827.7 ] ; p = 0.006) ; birth size and maternal smoking during pregnancy were inconsistently associated with and treatment of gdm during pregnancy did not affect overweight risk. homa - ir was increased in ogdm compared with offspring of ondm mothers (p = 0.01, adjusted for sex and age) and was associated with the child 's bmi (p = 0.004).conclusionsoverweight and insulin resistance in children is increased in ogdm compared with ot1d or ondm. the finding that overweight risk is associated mainly with maternal obesity suggests that familial predisposition contributes to childhood growth in these offspring. |
talc pleurodesis (tp) is a procedure first described by bethune 1 in 1935 as a means to anchor the lung during lobectomy 2. ever since then, tp has been employed worldwide in the management of recurrent pneumothorax and malignant pleural effusion, with a success rate in excess of 90% 3 - 5. tp is performed by instillation of sterile talc into the pleural cavity (chemical pleurodesis), with chemical irritation leading to pleuritis, and ultimately to pleural fibrosis. computed tomography (ct) and f - fluorodeoxy - glucose (fdg) positron - emission tomography (pet) are important imaging techniques for the detection and staging of cancers that, used together, may provide a more accurate tumor staging tool 6. however, tp treatment has been reported to increase fdg uptake in thickened pleura, making it difficult to distinguish between benign inflammatory processes and malignancies, with the potential for false positive results 7 - 16. malignant pleural mesothelioma (mpm) is a thoracic neoplasm originating from the pleural lining, where an increased fdg uptake has been observed 17, and several studies have reported on the use of pet to monitor chemotherapeutic response 18 - 21. in this setting, the concomitant presence of a disease such as mpm, and the application of a procedure like tp, may produce an increase in suv that underestimates the tumor response to chemotherapy. in clinical practice, the distinction of responding patients by those with stable or progressive disease has a prognostic implications and this is confirmed by some reports 22, 23. the distinction of these two categories of disease may allow to recognize patients with better prognosis which may benefit by trimodality therapy by those with a resistant disease where it may be suggested a conservative approach. the aim of this retrospective mpm case series was to analyze tp - induced pleural changes by fdg pet / ct and to verify the concordance between metabolic and radiologic response after chemotherapy. inclusion criteria was biopsy confirmed mpm performed during tp, followed by fdg pet / ct staging. all patients subsequently underwent three or four courses of pemetrexed based chemotherapy before fdg pet / ct restaging. written informed routinary consent was obtained from each patient before each examination, as standard quality of our institution require. all data were acquired on a combined pet / ct in - line system (biograph sensation 16, siemens). patients were instructed to fast for at least 6 h prior to scanning, which started 50 - 60 min after injection of 5.18 mbq fdg per kilogram body weight. the glucose level was checked before radiotracer injection and it always was lower than 160 mg / dl. all patients received a non - enhanced ct scan with 80ma, 120 kv parameters. scans were acquired in the head and neck area, the thorax and the lower abdomen during shallow breathing, and the upper abdomen during non - forced expiration. immediately after ct, the pet scan was performed using an acquisition time of 3 min per bed position. the patient remained in the same supine position on the pet / ct table throughout the procedure, and acquisition times ranged from 18 to 24 min. ct data were used for attenuation correction, and pet images were reconstructed by standard 2d - iterative algorithm (ordered subset expectation maximization). images were derived from an isocontour volume of interest (voi) drawn over the corresponding hemithorax region using a commercially available workstation (siemens esoft 2004). voi margins were checked to cover the whole area of interest in all three planes. the kidneys and the myocardium were spared from analysis by tracing a second voi positioned over the heart or kidneys. the minimal suv within the first voi was set to a level of 2.5 to eliminate any physiological liver uptake, that may have influenced mpm measurements. an example of pet / ct scan with recognized voi is represented in figure 1. all patients were radiologically staged according to brigham and women 's hospital 's revised mpm surgical staging system which considers tumor histology, resectability and nodal status 24. pleural biopsies obtained during tp procedures were analyzed to determine the histologic mpm subtypes 25. ct scans were assessed by a thoracic radiologist, experienced in mpm measurement and unaware of patient outcome ; radiologic response to chemotherapy was assessed using the modified mpm recist criteria 26. qualitative and semiquantitative evaluations of pet / ct images were performed by drawing a voi around the pleural thickness ; suvmax and suvmean depended on the initial dose administered and the body mass index. criteria for therapeutic response assessment were according to the european organization for research and treatment of cancer (eortc) that defined guidelines for the use of pet using fdg. these guidelines stated a 25% reduction in the maximum standardized uptake value (suvmax) as threshold for definition of partial response 27. all analyses were carried out by sas statistical software for windows version 9.1 (sas institute inc., cary, nc, usa). inclusion criteria was biopsy confirmed mpm performed during tp, followed by fdg pet / ct staging. all patients subsequently underwent three or four courses of pemetrexed based chemotherapy before fdg pet / ct restaging. written informed routinary consent was obtained from each patient before each examination, as standard quality of our institution require. all data were acquired on a combined pet / ct in - line system (biograph sensation 16, siemens). patients were instructed to fast for at least 6 h prior to scanning, which started 50 - 60 min after injection of 5.18 mbq fdg per kilogram body weight. the glucose level was checked before radiotracer injection and it always was lower than 160 mg / dl. all patients received a non - enhanced ct scan with 80ma, 120 kv parameters. scans were acquired in the head and neck area, the thorax and the lower abdomen during shallow breathing, and the upper abdomen during non - forced expiration. immediately after ct, the pet scan was performed using an acquisition time of 3 min per bed position. the patient remained in the same supine position on the pet / ct table throughout the procedure, and acquisition times ranged from 18 to 24 min. ct data were used for attenuation correction, and pet images were reconstructed by standard 2d - iterative algorithm (ordered subset expectation maximization). images were derived from an isocontour volume of interest (voi) drawn over the corresponding hemithorax region using a commercially available workstation (siemens esoft 2004). voi margins were checked to cover the whole area of interest in all three planes. the kidneys and the myocardium were spared from analysis by tracing a second voi positioned over the heart or kidneys. the minimal suv within the first voi was set to a level of 2.5 to eliminate any physiological liver uptake, that may have influenced mpm measurements. an example of pet / ct scan with recognized voi is represented in figure 1. all patients were radiologically staged according to brigham and women 's hospital 's revised mpm surgical staging system which considers tumor histology, resectability and nodal status 24. pleural biopsies obtained during tp procedures were analyzed to determine the histologic mpm subtypes 25. ct scans were assessed by a thoracic radiologist, experienced in mpm measurement and unaware of patient outcome ; radiologic response to chemotherapy was assessed using the modified mpm recist criteria 26. qualitative and semiquantitative evaluations of pet / ct images were performed by drawing a voi around the pleural thickness ; suvmax and suvmean depended on the initial dose administered and the body mass index. criteria for therapeutic response assessment were according to the european organization for research and treatment of cancer (eortc) that defined guidelines for the use of pet using fdg. these guidelines stated a 25% reduction in the maximum standardized uptake value (suvmax) as threshold for definition of partial response 27. all analyses were carried out by sas statistical software for windows version 9.1 (sas institute inc., cary, nc, usa). eight patients were included in our study, 6 (75%) males and 2 (25%) females, with a median age of 65 years (range : 54 - 77). histologic examination revealed an epitheliod subtype in 7 (88%) cases and a mixed subtype in 1 (12%), 5 (63%) of which were located on the left side and 3 (37%) on the right. overall, 7 (88%) stage i and 1 (12%) stage ii diseases were reported. all patients underwent pemetrexed - based chemotherapy (3 patients in combination with carboplatin, 4 with cisplatin, and 1 with pemetrexed only) with a median of 3.5 courses (range : 3 - 4) before restaging. after tp treatment, there was a mean interval of 14 days (range : 9 - 22) and 125 days (range : 76 - 162) between fdg pet / ct staging and restaging, respectively, and, on average, 26 days (range : 6 - 46) between the last course of chemotherapy and the fdg pet / ct restaging. suv mean and suv max before and after treatment is summarized in the table 2. the concordance between two kinds of responses was respectively 37.5% (3 out of 8 patients) and 75% (6 out of 8 patients). comparing radiologic and metabolic response, expressed as suvmax and suvmean (see table 2 and table 3), the concordance was 37.5% (3 out of 8 patients) and 75% (6 out of 8 patients), respectively. talc pleurodesis is used to treat patients with persistent pleural effusions or pneumothorax not amenable to other therapies 1 - 5. pathologic studies have shown that granulomatous inflammation occurs in the visceral and parietal pleural immediately after tp, resulting in pleural inflammation. for several years, this has been presumed to be the cause of increased fdg activity, leading to the difficulty in distinguishing between benign and malignant disease 7, 8, 10. mpm is a malignancy arising from the pleural sierosa with an fdg - avidity 17, and this characteristic has been proposed to evaluate chemotherapeutic response 18 - 21. pleural inflammation, induced by tp and mpm, produces pleural thickening or nodularity detected by radiologic examination, and the resulting increase in suv uptake leads to biases in the post - chemotherapy evaluation. increased fdg uptake, with an associated increase in the thickness of non calcified pleural lesions, or new fdg - avid pleural lesions without associated calcification, are suspicious for malignancy, whereas fdg uptake in calcified pleural lesions suggests benign tp - induced changes. currently, little clinical data is available on which is the best metabolic index to reduce chemotherapy evaluation bias in mpm patients previously treated with tp. in our experience, considering that morphologic changes shown by ct - scan predict response to chemotherapy 26, we observed that the metabolic response measured by suvmean seems to be in better agreement with the radiologic evaluation compared to the suvmax, but without reaching statistical significance. this may probably be due to the relatively small number of cases in our study, and therefore a larger prospective study is required to confirm our hypothesis. | purpose : talc pleurodesis (tp) is employed worldwide for the management of persistent pneumothorax or pleural effusion, particularly of malignant origin. however, there are very little available data on 18f - fluorodeoxyglucose positron - emission tomography / computed tomography (18f fdg pet / ct) response evaluation in malignant pleural mesothelioma (mpm) patients treated with tp and chemotherapy.methods : patients with histologically confirmed mpm underwent tp and fdg pet / ct staging and restaging after 3 - 4 courses of chemotherapy. all patients fasted and received a dose of 5.18 mbq 18f - fdg per kilogram of body weight. whole - body emission scans were acquired with and without ordered subset expectation maximization (osem) iterative reconstruction algorithm.results : from january 2004 to march 2010, 8 patients with biopsy confirmed mpm (7 epithelial, 1 biphasic), with a median age of 65 years (range : 54 - 77), were evaluated. median follow - up was 31 months (range : 4 - 44). after tp treatment, there was a mean interval of 14 days (range : 9 - 22) and 125 days (range : 76 - 162) between fdg pet / ct staging and restaging. according to modified recist and eortc criteria, there was a concordance between the radiologic and metabolic suvmean and suvmax responses in 6 (75%) and 3 (37.5%) patients, respectively.conclusion : tp produces an increased fdg pet uptake which may interfere with the post - chemotherapy disease evaluation. in our case series, the metabolic response measured by suvmean seems to be in better agreement with the radiologic response compared to the suvmax. |
the united states department of defense and department of veteran affairs are dedicated to providing superior rehabilitation care for veterans and service members who have suffered combat - associated traumatic limb loss. current literature reports that 70% of amputees with multiple limb loss are bothered by sweating and skin irritation inside of the prosthesis socket. we present a review of the literature examining the effectiveness of botulinum toxin a (btx - a) and btx - b for the treatment of hyperhidrosis related to prosthesis use. while both btx - a and btx - b are approved by the food and drug administration (fda), indications for use are limited. hyperhidrosis continues to be a significant barrier to prosthesis comfort, and treatment with btx has the potential to address this concern. a review of the literature was conducted using the pubmed database, focusing on hyperhidrosis treatment after a traumatic limb amputation. we used the key search words : btx ; hyperhidrosis ; amputation ; trauma - related amputation ; prosthesis comfort. articles discussing hyperhidrosis treatment for axillary, palmar, plantar and amputations secondary to chronic medical conditions were excluded. botulinum toxin a is currently approved by the fda as botox (allergan, inc., irvine, california, usa) supplied in 50 and 100 unit vials, and dysport (ipsen, berkshire, united kingdom) supplied in 300 and 500 unit vials, to treat cervical dystonia, severe primary axillary hyperhidrosis, strabismus, blepharospasm, and for temporary improvement of moderate to severe glabellar lines. the benefits of using btx - a for these conditions is well - established and it has few adverse effects at their respective therapeutic doses, which range from 20 to 360 units. the most common adverse effects reported are muscle weakness, fatigue, flu - like symptoms, dry mouth, dizziness, discomfort at the injection site and skin rash. large volume injections of btx - a > 600 units have been associated with systemic weakness related to injection dose and frequency, which should be a consideration in the treatment of the residual limb hyperhidrosis. current literature emphasizes the positive effects of btx - a on pain perception, itch and inflammation while simultaneously producing an anhidrotic effect. a study conducted by charrow. found that injections of btx - a successfully reduced residual limb hyperhidrosis and improved prosthesis fit and function when evaluated three weeks after treatment. in this study, 300 - 500 units of btx - a at a dilution of 100 units in 1 ml of 0.9% isotonic saline were injected intradermally with 2 - 3 units at 1 cm intervals in a circumferential pattern on the skin covered by the prosthesis socket. this approach however, did not have any effect on the residual limb pain (rlp) or phantom limb pain (plp). however, jin. demonstrated in three cases that injection of btx - a at 200, 300, or 500 units using electromyographic guidance and targeting areas of the residual limb where strong fasciculations were present achieved marked improvement in rlp and plp lasting up to three months. three separate case studies utilized a starch - iodine test to identify hyperhidrotic areas on a residual limb, then injected btx - a at 100 u in 5 - 10 unit aliquots, or 300 units in 25 units aliquots. in all three reports, btx - a was diluted in preservative - free saline and produced anhidrotic effects lasting three months. because our facility is a government - funded hospital with extensive medical coverage for military personnel, poor prosthesis satisfaction due to hyperhidrosis is the only indication needed for treatment. while large volume injections can be costly, the literature has shown that improving prosthesis comfort is directly correlated with an improved quality of life. to inject btx - a at 1 - 2 cm intervals in a grid - like pattern over a surface area of 600 cm would require over 140 intradermal injections, typically achieved with a 30-gauge needle. the procedure lasts up to 30 min and is criticized as painful. a recent study by torrisi. investigated the use of pocketed microneedles (pmn) as an intradermal delivery system for btx - a. the width of the pmn used in this study is 340 m, which could serve as a less invasive administration method of btx - a in hyperhidrotic areas. botulinum toxin type b, supplied in 2500, 5000, and 10,000 units vials is approved by the fda as myobloc (solstice neurosciences, inc. the use of btx - b for the treatment of palmar hyperhidrosis was deemed effective by baumann. and further studied by kern. in the treatment of the residual limb hyperhidrosis for lower limb amputees. btx - b administered in a low dose is believed to have a higher affinity for sympathetic nerve endings and better diffusion than btx - a. furthermore, btx - b and was found to be effective in reducing residual limb sweating in nine lower limb amputees. treated nine lower limb amputees with 1750 units of btx - b injected intracutaneously using a 27-gauge needle (20 injection sites 2 - 4 cm apart). in addition to subjective identification of hyperhidrotic areas, a starch - iodine test was used on six participants, to help guide injection locations. study reported a significant reduction in residual limb sweating and a significant improvement in use of the prosthetic device, duration of use and quality of life evaluated 4 weeks and 3 months after treatment. btx - b also decreased rlp and plp as well as improved quality of life and prosthesis use. reports that btx - b injections in muscular trigger points decreased rlp and involuntary movements of the stump for 4 - 12 weeks, which was dependent on the btx - b dose and location of amputation. two of the four patients had a trauma - related amputation, and all found the injections very painful. current literature investigating the use of btx to treat hyperhidrosis in trauma - related amputations is limited. however, current case studies all report positive outcomes related to treating hyperhidrosis with either btx - a or btx - b. hyperhidrosis, discomfort, and skin irritation related to prosthesis use in trauma - related limb loss remains a topic deserving of more attention as a larger, double - blind, placebo - controlled study evaluating the efficacy of btx - a and btx - b in treatment. | hyperhidrosis - related to prosthesis use in patients who have suffered a traumatic limb amputation presents itself as a barrier to comfort, prosthesis use and overall quality of life. this review intends to encourage dermatologists to consider the use of botulinum toxin a or b for the treatment of hyperhidrosis in the residual limb and may serve as a stimulus for a modern, in - depth, and more comprehensive study. a review of the literature was conducted using the pubmed database, focusing on hyperhidrosis treatment after traumatic limb amputation. articles discussing hyperhidrosis treatment for amputations secondary to chronic medical conditions were excluded. seven case studies published over the last 12 years have demonstrated positive outcomes of this treatment strategy. overall, there is little data examining this topic and current publications focus primarily on small case series. a larger, double - blind, placebo - controlled study would likely benefit veterans, service members, and civilians. |
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