Omalizumab is an effective and rapidly acting therapy in difficult-to-treat chronic urticaria: a retrospective clinical analysis.
Omalizumab (anti-IgE) therapy is effective and safe in chronic urticaria (CU) in placebo-controlled clinical trials but real life clinical data are scarce. To better understand the effects of omalizumab in CU patients treated outside of clinical trials. In this retrospective clinical analysis, we assessed responder rates, optimal dosage, response to up-/downdosing, time to relief of symptoms, rates of return and time of relapse after omalizumab administration, and safety in 51 CU patients, 20 with chronic spontaneous urticaria (CSU) alone, 21 with different forms of chronic inducible urticaria (CindU) and 10 with both. Omalizumab treatment led to complete remission in 83% of CSU and 70% of CindU patients. When starting with 150mg omalizumab 4 weekly, only 2/15 CSU and 7/17 CindU patients required updosing to achieve complete remission. In CSU, 57% of complete responses occurred within week one, all on the first day. Relapses were 2-8 weeks in all but six patients, where they were <4 months. Omalizumab was safe. Efficacy was not correlated to baseline IgE levels. Clinical experience from more than 1250 injections in 51 patients over four years indicates that omalizumab is a rapidly acting, highly effective and safe drug in CSU and CindU patients. Our observations in a real life clinical setting support the recommendation of current EAACI/GA(2)LEN/EDF/WAO guideline for the management of urticaria to use omalizumab to treat urticaria patients.


Systemic Oxidative Stress Is Increased in Postmenopausal Women and Independently Associates with Homocysteine Levels.
Oxidative stress plays a pivotal role in the pathogenesis of cardiovascular diseases (CVD). Postmenopausal women have an increased risk of developing CVD due to decreased estrogen availability, which is accompanied by increased oxidative stress. Serum free thiols (R-SH) provide a robust and powerful read-out of systemic oxidative stress. In this study, we aimed to establish serum levels of free thiols and explore associations between free thiols and demographic, clinical, and biochemical parameters related to obesity and the risk for developing CVD in both pre- and postmenopausal women. Serum free thiols were measured in a cohort consisting of healthy pre- (n = 223) and postmenopausal (n = 118) Omani women. Postmenopausal women had significantly lower levels of serum free thiols as compared to premenopausal women (762.9 ± 85.3 vs. 780 ± 80.9 μM, age-adjusted p < 0.001). Women's age was positively associated with serum free thiol levels in premenopausal women (β = 0.36, p = 0.002), whereas an inverse association was observed in postmenopausal women (β = -0.29, p = 0.002). Homocysteine levels were significantly inversely associated with serum free thiol levels in both pre- (β = -0.19, p = 0.005) and postmenopausal (β = -0.20, p = 0.032) women, independent from known cardiovascular risk factors. In this study, we show that postmenopausal women are affected by increased systemic oxidative stress, which independently associates with homocysteine levels.


A humanized Smn gene containing the SMN2 nucleotide alteration in exon 7 mimics SMN2 splicing and the SMA disease phenotype.
Proximal spinal muscular atrophy (SMA) is a neurodegenerative disease caused by low levels of the survival motor neuron (SMN) protein. In humans, SMN1 and SMN2 encode the SMN protein. In SMA patients, the SMN1 gene is lost and the remaining SMN2 gene only partially compensates. Mediated by a C>T nucleotide transition in SMN2, the inefficient recognition of exon 7 by the splicing machinery results in low levels of SMN. Because the SMN2 gene is capable of expressing SMN protein, correction of SMN2 splicing is an attractive therapeutic option. Although current mouse models of SMA characterized by Smn knock-out alleles in combination with SMN2 transgenes adequately model the disease phenotype, their complex genetics and short lifespan have hindered the development and testing of therapies aimed at SMN2 splicing correction. Here we show that the mouse and human minigenes are regulated similarly by conserved elements within in exon 7 and its downstream intron. Importantly, the C>T mutation is sufficient to induce exon 7 skipping in the mouse minigene as in the human SMN2. When the mouse Smn gene was humanized to carry the C>T mutation, keeping it under the control of the endogenous promoter, and in the natural genomic context, the resulting mice exhibit exon 7 skipping and mild adult onset SMA characterized by muscle weakness, decreased activity and an alteration of the muscle fibers size. This Smn C>T mouse represents a new model for an adult onset form of SMA (type III/IV) also know as the Kugelberg-Welander disease.


Impact of a physical fitness program in a blue-collar workforce.
The purpose of this study was to examine the impact of participation in the Wellness--Paths to Health Physical Fitness Program on selected physiological and personal health variables including blood pressure, pulse, body weight, trunk flexibility, cardiovascular endurance, abdominal strength, percent body fat and a cumulative fitness score. The subjects (N = 129) of this study were divided into four levels of adherence to this exercise-based health program. Data analysis examined relationships between adherence levels and changes in the selected physiological and personal health variables across two measures. In addition, data from high-risk participants were analyzed. The results of the study found statistical significant changes in the selected variables between repeated assessment measures. In addition, statistically significant relationships were found between levels of adherence to the program and six of the dependent variables.


Mid-infrared supercontinuum generation in tapered chalcogenide fiber for producing octave-spanning frequency comb around 3 μm.
We demonstrate mid-infrared (mid-IR) supercontinuum generation (SCG) with instantaneous bandwidth from 2.2 to 5 μm at 40 dB below the peak, covering the wavelength range desirable for molecular spectroscopy and numerous other applications. The SCG occurs in a tapered As(2)S(3) fiber prepared by in-situ tapering and is pumped by femtosecond pulses from the subharmonic of a mode-locked Er-doped fiber laser. Interference with a narrow linewidth c.w. laser verifies that the coherence properties of the near-IR frequency comb have been preserved through these cascaded nonlinear processes. With this approach stable broad mid-IR frequency combs can be derived from commercially available near-IR frequency combs without an extra stabilization mechanism.


Metabolic cleavage and translocation efficiency of selected cell penetrating peptides: a comparative study with epithelial cell cultures.
We investigated the metabolic stability of four cell penetrating peptides (CPPs), namely SAP, hCT(9-32)-br, [Palpha] and [Pbeta], when in contact with either subconfluent HeLa, confluent MDCK or Calu-3 epithelial cell cultures. Additionally, through analysis of their cellular translocation efficiency, we evaluated possible relations between metabolic stability and translocation efficiency. Metabolic degradation kinetics and resulting metabolites were assessed using RP-HPLC and MALDI-TOF mass spectrometry. Translocation efficiencies were determined using fluorescence-activated cell sorting (FACS) and confocal laser scanning microscopy (CLSM). Between HeLa, MDCK and Calu-3 we found the levels of proteolytic activities to be highly variable. However, for each peptide, the individual degradation patterns were quite similar. The metabolic stability of the investigated CPPs was in the order of CF-SAP = CF-hCT(9-32)-br > [Pbeta]-IAF > [Palpha] and we identified specific cleavage sites for each of the four peptides. Throughout, we observed higher translocation efficiencies into HeLa cells as compared to MDCK and Calu-3, corresponding to the lower state of differentiation of HeLa cell cultures. No direct relation between metabolic stability and translocation efficiency was found, indicating that metabolic stability in general is not a main limiting factor for efficient cellular translocation. Nevertheless, translocation of individual CPPs may be improved by structural modifications aiming at increased metabolic stability.


The identification and determination of selected 1,4-benzodiazepines by an optimised capillary electrophoresis-electrospray mass spectrometric method.
An optimised capillary electrophoresis-electrospray mass spectrometric method is presented for the identification and determination of diazepam and its metabolites N'-desmethyldiazepam, oxazepam and temazepam. By investigating constituent parts of the capillary electrophoresis-electrospray mass spectrometric interface and optimising their function, a relatively fast and reproducible method is described for the identification and determination of selected 1,4-benzodiazepines. Optimisation of sheath and auxiliary gas flows, capillary tip tapering, capillary tip positioning, sheath liquid composition and flow rate and pressure application during the separation step have led to acceptable relative standard deviation (RSD) values for migration time and peak area, correlation coefficients and limits of detection. This has been achieved as a result of stabilising the electrospray current prior to analysis, a procedure that takes a matter of minutes when using the method described. Sequential product ion fragmentation (MS(n)) characterisation of 15 1,4-benzodiazepines is also presented and mechanisms for the observed fragmentation patterns proposed.


p21 is decreased in polycystic kidney disease and leads to increased epithelial cell cycle progression: roscovitine augments p21 levels.
Autosomal dominant polycystic kidney disease (ADPKD) is a common genetic disease with few treatment options other than renal replacement therapy. p21, a cyclin kinase inhibitor which has pleiotropic effects on the cell cycle, in many cases acts to suppress cell cycle progression and to prevent apoptosis. Because defects in cell cycle arrest and apoptosis of renal tubular epithelial cells occur in PKD, and in light of earlier reports that polycystin-1 upregulates p21 and that the cyclin-dependent kinase inhibitor roscovitine arrests progression in a mouse model, we asked whether (1) p21 deficiency might underlie ADPKD and (2) the mechanism of the salutary roscovitine effect on PKD involves p21. p21 levels in human and animal tissue samples as well as cell lines were examined by immunoblotting and/or immunohistochemisty. Apoptosis was assessed by PARP cleavage. p21 expression was attenuated in a renal tubular epithelial cell line by antisense methods, and proliferation in response to p21 attenuation and to roscovitine was assessed by the MTT assay. We show that p21 is decreased in human as well as a non-transgenic rat model of ADPKD. In addition, hepatocyte growth factor, which induces transition from a cystic to a tubular phenotype, increases p21 levels. Furthermore, attenuation of p21 results in augmentation of cell cycle transit in vitro. Thus, levels of p21 are inversely correlated with renal tubular epithelial cell proliferation. Roscovitine, which has been shown to arrest progression in a murine model of PKD, increases p21 levels and decreases renal tubular epithelial cell proliferation, with no affect on apoptosis. The novelty of our study is the demonstration in vivo in humans and rat models of a decrement of p21 in cystic kidneys as compared to non-cystic kidneys. Validation of a potential pathogenetic model of increased cyst formation due to enhanced epithelial proliferation and apoptosis mediated by p21 suggests a mechanism for the salutary effect of roscovitine in ADPKD and supports further investigation of p21 as a target for future therapy.


Assessment of transesophageal Doppler echography in dissecting aortic aneurysm.
To assess the clinical value of transesophageal Doppler echography in the diagnosis of dissecting aortic aneurysm, both transesophageal and conventional echograms were performed in 22 cases of dissecting aortic aneurysm. Of the 22 patients, 17 underwent angiography; 8, X-ray computed tomography; 4, both; and 12, surgery. The performance of each method was assessed in the following four segments: A, ascending aorta; B, aortic arch; C, thoracic descending aorta; and D, upper abdominal aorta. The results by angiography were presumed to be correct. In the group of 17 patients who underwent angiography, the rate of correct detection of an intimal flap using the transesophageal approach was 100% in all four segments, significantly better than detection by the conventional approach (segment A, 65%; segment B, 47%; segment C, 35%; segment D, 53%) (p less than 0.01), and the rate of correct detection of the entry sites using the transesophageal approach was 100%, significantly better than that by conventional approach (42%) (p less than 0.05). X-ray computed tomography was not capable of detecting the site of entry in all cases. The presence of thrombus, aortic regurgitation and pericardial hemorrhage were all revealed clearly by the transesophageal approach, and the results were partly proved by other methods. In conclusion, transesophageal Doppler echography provides a rapid and accurate method of diagnosing and evaluating dissecting aortic aneurysm and permits prompt initiation of appropriate treatment.


COPD classification methods and informativeness on mortality: contrasting evidences.
Understanding whether the ABCD GOLD classification method is informative with respect to the spirometric classification of severity in predicting mortality of patients with chronic obstructive pulmonary disease (COPD) is subject of debate. The results of a study performed on a sample of the Norwegian population (HUNT2) were recently published. Such data showed the inferiority of ABCD classification in predicting mortality compared to the spirometric classification, which was considered the gold standard up to the 2011 version of GOLD guidelines. This result is not in line with the results of other studies that have shown the equivalence of the two classifications. The new GOLD classification seemed to be a step forward for what concerns understanding patient's needs, but it seems clear that the insertion of a single clinical variable to the spirometric data may not be exhaustive in describing all the phenomena related to a heterogeneous disease such as COPD. The publication of the HUNT study provides an opportunity to analyze how the evidence has been produced, which scientific speculations it offers, what considerations could be drawn and what further research would be appropriate.


Is higher serum cholesterol associated with altered tendon structure or tendon pain? A systematic review.
Tendon pain occurs in individuals with extreme cholesterol levels (familial hypercholesterolaemia). It is unclear whether the association with tendon pain is strong with less extreme elevations of cholesterol. To determine whether lipid levels are associated with abnormal tendon structure or the presence of tendon pain. We conducted a systematic review and meta-analysis. Relevant articles were found through an electronic search of 6 medical databases-MEDLINE, Cochrane, AMED, EMBASE, Web of Science and Scopus. We included all case-control or cross-sectional studies with data describing (1) lipid levels or use of lipid-lowering drugs and (2) tendon structure or tendon pain. 17 studies (2612 participants) were eligible for inclusion in the review. People with altered tendon structure or tendon pain had significantly higher total cholesterol, low-density lipoprotein cholesterol and triglycerides, as well as lower high-density lipoprotein cholesterol; with mean difference values of 0.66, 1.00, 0.33, and -0.19 mmol/L, respectively. The results of this review indicate that a relationship exists between an individual's lipid profile and tendon health. However, further longitudinal studies are required to determine whether a cause and effect relationship exists between tendon structure and lipid levels. This could lead to advancement in the understanding of the pathoaetiology and thus treatment of tendinopathy.


Anexelekto/MER tyrosine kinase inhibitor ONO-7475 arrests growth and kills FMS-like tyrosine kinase 3-internal tandem duplication mutant acute myeloid leukemia cells by diverse mechanisms.
Nearly one-third of patients with acute myeloid leukemia have FMS-like tyrosine kinase 3 mutations and thus have poor survival prospects. Receptor tyrosine kinase anexelekto is critical for FMS-like tyrosine kinase 3 signaling and participates in FMS-like tyrosine kinase 3 inhibitor resistance mechanisms. Thus, strategies targeting anexelekto could prove useful for acute myeloid leukemia therapy. ONO-7475 is an inhibitor with high specificity for anexelekto and MER tyrosine kinase. Herein, we report that ONO-7475 potently arrested growth and induced apoptosis in acute myeloid leukemia with internal tandem duplication mutation of FMS-like tyrosine kinase 3. MER tyrosine kinase-lacking MOLM13 cells were sensitive to ONO-7475, while MER tyrosine kinase expressing OCI-AML3 cells were resistant, suggesting that the drug acts via anexelekto in acute myeloid leukemia cells. Reverse phase protein analysis of ONO-7475 treated cells revealed that cell cycle regulators like cyclin dependent kinase 1, cyclin B1, polo-like kinase 1, and retinoblastoma were suppressed. ONO-7475 suppressed cyclin dependent kinase 1, cyclin B1, polo-like kinase 1 gene expression suggesting that anexelekto may regulate the cell cycle, at least in part, via transcriptional mechanisms. Importantly, ONO-7475 was effective in a human FMS-like tyrosine kinase 3 with internal tandem duplication mutant murine xenograft model. Mice fed a diet containing ONO-7475 exhibited significantly longer survival and, interestingly, blocked leukemia cell infiltration in the liver. In summary, ONO-7475 effectively kills acute myeloid leukemia cells in vitro and in vivo by mechanisms that involve disruption of diverse survival and proliferation pathways.


On the issues related to compliance of LF pulsed exposures with safety standards and guidelines.
In this paper, procedures to determine compliance of low-frequency pulsed exposures are investigated. Current methods specified by international standards or guidelines (e.g., from the ICNIRP or IEEE) are recognized to be conservative in order to account for uncertainties coming from the assessment procedures. In this way, protection of workers and the general public should be guaranteed. However, overly conservative procedures could hinder the application of technologies employing complex, intermittent, or pulsed waveforms without improving safety. Besides over conservatism, variabilities among the results of several procedures are examined for the first time. These limits pose several concerns on the applicability of the existing compliance formulae. A more stable technique, which is still easy to implement, is therefore proposed.


Targeting AXL and mTOR Pathway Overcomes Primary and Acquired Resistance to WEE1 Inhibition in Small-Cell Lung Cancer.
Purpose: Drugs targeting DNA repair and cell-cycle checkpoints have emerged as promising therapies for small-cell lung cancer (SCLC). Among these, the WEE1 inhibitor AZD1775 has shown clinical activity in a subset of SCLC patients, but resistance is common. Understanding primary and acquired resistance mechanisms will be critical for developing effective WEE1 inhibitor combinations.Experimental Design: AZD1775 sensitivity in SCLC cell lines was correlated with baseline expression level of 200 total or phosphorylated proteins measured by reverse-phase protein array (RPPA) to identify predictive markers of primary resistance. We further established AZD1775 acquired resistance models to identify mechanism of acquired resistance. Combination regimens were tested to overcome primary and acquired resistance to AZD1775 in in vitro and in vivo SCLC models.Results: High-throughput proteomic profiling demonstrate that SCLC models with primary resistance to AZD1775 express high levels of AXL and phosphorylated S6 and that WEE1/AXL or WEE1/mTOR inhibitor combinations overcome resistance in vitro and in vivo Furthermore, AXL, independently and via mTOR, activates the ERK pathway, leading to recruitment and activation of another G2-checkpoint protein, CHK1. AZD1775 acquired resistance models demonstrated upregulation of AXL, pS6, and MET, and resistance was overcome with the addition of AXL (TP0903), dual-AXL/MET (cabozantinib), or mTOR (RAD001) inhibitors.Conclusions: AXL promotes resistance to WEE1 inhibition via downstream mTOR signaling and resulting activation of a parallel DNA damage repair pathway, CHK1. These findings suggest rational combinations to enhance the clinical efficacy of AZD1775, which is currently in clinical trials for SCLC and other malignancies. Clin Cancer Res; 23(20); 6239-53. ©2017 AACR.


The role of neurocritical care: a brief report on the survey results of neurosciences and critical care specialists.
Neurocritical care is a new subspecialty field in medicine that intersects with many of the neuroscience and critical care specialties, and continues to evolve in its scope of practice and practitioners. The objective of this study was to assess the perceived need for and roles of neurocritical care intensivists and neurointensive care units among physicians involved with intensive care and the neurosciences. An online survey of physicians practicing critical care medicine, and neurology was performed during the 2008 Leapfrog initiative to formally recognize neurocritical care training. The survey closed in July 2009 and achieved a 13% response rate (980/7524 physicians surveyed). Survey respondents (mostly from North America) included 362 (41.4%) neurologists, 164 (18.8%) internists, 104 (11.9%) pediatric intensivists, 82 (9.4%) anesthesiologists, and 162 (18.5%) from other specialties. Over 70% of respondents reported that the availability of neurocritical care units staffed with neurointensivists would improve the quality of care of critically ill neurological/neurosurgical patients. Neurologists were reported as the most appropriate specialty for training in neurointensive care by 53.3%, and 57% of respondents responded positively that neurology residency programs should offer a separate training track for those interested in neurocritical care. Broad level of support exists among the survey respondents (mostly neurologists and intensivists) for the establishment of neurological critical care units. Since neurology remains the predominant career path from which to draw neurointensivists, there may be a role for more comprehensive neurointensive care training within neurology residencies or an alternative training track for interested residents.


Risk of locoregional recurrence by receptor status in breast cancer patients receiving modern systemic therapy and post-mastectomy radiation.
We assessed differences in locoregional outcome based on receptor status combinations in a cohort of stage II-III breast cancer patients treated with modern trimodality therapy. Medical records of 582 consecutively treated patients receiving post-mastectomy radiation (PMRT) between 1/1999 and 12/2009 were reviewed. Rate of local regional recurrence (LRR) was estimated by the method of cumulative incidence allowing for competing risks. The effect of prognostic factors was examined by Gray's test and by Fine and Gray's modeling approach. Median follow-up was 44.7 months. Five-year progression-free survival (PFS) was 73.9% and overall survival (OS) was 84%. The cumulative 5-year incidence of LRR as first site of failure was 6.2% (95% CI 4.2-8.7). Five-year cumulative incidence of LRR was 8.6 versus 4.4% for estrogen receptor (ER) negative versus ER positive (P = 0.017), 8.5 versus 3.4% for progesterone receptor (PR) negative versus PR positive (P = 0.011), and 1.7 versus 7.5% for HER2 positive (86% received trastuzamab) versus HER2 negative (P = 0.032). Five-year cumulative incidence of LRR was 11.8% for the triple negative subtype and 3.9% for other receptor combinations (P < 0.001). Among patients whose disease is ER positive, 5-year LRR rate was 7.8 versus 3.4% for PR negative versus PR positive (P = 0.130). The prognostic value of the triple negative and HER2 negative subtypes was maintained on multivariate analysis. In the era of HER-2 targeted therapy, tumors that are HER-2 over expressing and are treated with trastuzumab have a very low rate of LRR. ER negative, PR negative, and triple negative status are associated with increased risk of LRR.


Wolbachia infection and Lepidoptera of conservation concern.
Conservation of at-risk species requires multi-faceted and carefully-considered management approaches to be successful. For arthropods, the presence of endosymbiotic bacteria, such as Wolbachia (Rickettsiales: Rickettsiaceae), may complicate management plans and exacerbate the challenges faced by conservation managers. Wolbachia poses a substantial and underappreciated threat to the conservation of arthropods because infection may induce a number of phenotypic effects, most of which are considered deleterious to the host population. In this study, the prevalence of Wolbachia infection in lepidopteran species of conservation concern was examined. Using standard molecular techniques, 22 species of Lepidoptera were screened, of which 19 were infected with Wolbachia. This rate is comparable to that observed in insects as a whole. However, this is likely an underestimate because geographic sampling was not extensive and may not have included infected segments of the species' ranges. Wolbachia infections may be particularly problematic for conservation management plans that incorporate captive propagation or translocation. Inadvertent introduction of Wolbachia into uninfected populations or introduction of a new strain may put these populations at greater risk for extinction. Further sampling to investigate the geographic extent of Wolbachia infections within species of conservation concern and experiments designed to determine the nature of the infection phenotype(s) are necessary to manage the potential threat of infection.


Spatial and Temporal Clustering of Chikungunya Virus Transmission in Dominica.
Using geo-referenced case data, we present spatial and spatio-temporal cluster analyses of the early spread of the 2013-2015 chikungunya virus (CHIKV) in Dominica, an island in the Caribbean. Spatial coordinates of the locations of the first 417 reported cases observed between December 15th, 2013 and March 11th, 2014, were captured using the Global Positioning System (GPS). We observed a preponderance of female cases, which has been reported for CHIKV outbreaks in other regions. We also noted statistically significant spatial and spatio-temporal clusters in highly populated areas and observed major clusters prior to implementation of intensive vector control programs suggesting early vector control measures, and education had an impact on the spread of the CHIKV epidemic in Dominica. A dynamical identification of clusters can lead to local assessment of risk and provide opportunities for targeted control efforts for nations experiencing CHIKV outbreaks.


Extinction of metastable stochastic populations.
We investigate the phenomenon of extinction of a long-lived self-regulating stochastic population, caused by intrinsic (demographic) noise. Extinction typically occurs via one of two scenarios depending on whether the absorbing state n=0 is a repelling (scenario A) or attracting (scenario B) point of the deterministic rate equation. In scenario A the metastable stochastic population resides in the vicinity of an attracting fixed point next to the repelling point n=0 . In scenario B there is an intermediate repelling point n=n1 between the attracting point n=0 and another attracting point n=n2 in the vicinity of which the metastable population resides. The crux of the theory is a dissipative variant of WKB (Wentzel-Kramers-Brillouin) approximation which assumes that the typical population size in the metastable state is large. Starting from the master equation, we calculate the quasistationary probability distribution of the population sizes and the (exponentially long) mean time to extinction for each of the two scenarios. When necessary, the WKB approximation is complemented (i) by a recursive solution of the quasistationary master equation at small n and (ii) by the van Kampen system-size expansion, valid near the fixed points of the deterministic rate equation. The theory yields both entropic barriers to extinction and pre-exponential factors, and holds for a general set of multistep processes when detailed balance is broken. The results simplify considerably for single-step processes and near the characteristic bifurcations of scenarios A and B.


Asymptomatic aortic stenosis in the elderly: a clinical review.
Aortic stenosis is the most common form of valvular heart disease. Progression of aortic stenosis is very slow and highly variable. Decisions about when to perform valve surgery are made by subjective assessment of patient symptoms and objective measures of the valve and ventricular function by transthoracic echocardiography. To review current concepts regarding the development, progression, and assessment of aortic stenosis; the appropriate monitoring intervals for transthoracic echocardiography; and the indications for valve procedures. Guidelines and literature search. Angina, exertional syncope, and heart failure are key symptoms indicating a need for intervention. The frequency of valvular monitoring by transthoracic echocardiography is guided by the disease severity. Despite evidence of severe disease, valve procedures can safely be deferred if patients experience no symptoms and have normal left ventricular ejection fraction. Asymptomatic patients with severe aortic stenosis may subconsciously curtail their activities to avoid symptoms. Apparently, asymptomatic patients can undergo a carefully monitored exercise stress test to confirm both their asymptomatic status and hemodynamic response to exercise. Bioprosthetic valves are a good replacement alternative for older patients who are good surgical candidates and who have no need for warfarin therapy. For patients who are at high or very high risk of cardiac surgery, transcutaneous aortic valve implantation is an increasing available and preferred over medical management. Asymptomatic patients with severe aortic stenosis require frequent monitoring of their subjective symptoms combined with objective measurement of aortic valve gradient and ventricular function by transthoracic echocardiography. Although conventional surgical replacement remains the mainstay of therapy for aortic stenosis, transcutaneous aortic valve implantation options are evolving.


The interception and retention of 238Pu deposition by orange trees.
Radioisotope thermoelectric generators (RTG) transform the heat produced during the alpha decay of 238Pu into electrical energy for use by deep-space probes, such as the Voyager spacecraft, which have returned images and other data from Jupiter, Saturn and Uranus. Future missions involving RTGs may be launched aboard the space shuttle, and there is a remote possibility that an explosion of liquid-hydrogen and liquid-oxygen fuel could rupture the RTGs and disperse 238Pu into the atmosphere over central Florida. Research was performed to determine the potential transport to man of atmospherically dispersed Pu via contaminated orange fruits. The results indicate that the major contamination of oranges would result from the interception and retention of 238Pu deposition by fruits. The resulting surface contamination could enter human food chains through transfer to internal tissues during peeling or in the reconstituted juices and flavorings made from orange skins. The interception of 238Pu deposition by fruits is especially important because the results indicate no measurable loss of Pu from fruit surfaces through time or with washing. Approximately 1% of the 238Pu deposited onto an orange grove would be harvested in the year following deposition.


Inhibition by anions of human red cell carbonic anhydrase B: physiological and biochemical implications.
The hydration rate of CO2 catalyzed by human red cell carbonic anhydrase B is 92 percent reduced by the normal concentrations of chloride and bicarbonate in red cells. This reflects a general sensitivity of this reaction to halides and other anions, up to 87 times greater than the effect on red cell carbonic anhydrase C. The catalytic hydration of CO2 is generally more (up to 24 times) sensitive to inhibition by anions and sulfonamides than the dehydration of HCO3-, probably reflecting different mechanisms. The sensitivity of enzyme B to anion inhibition also depends upon the substrate, being much greater for CO2 than for certain esters. On the basis of the very low catalytic activity of B for CO2 in the presence of physiological concentration of chloride, and the fact that carbonic anhydrase C is effective for CO2 hydration (in the presence of chloride) at a rate 340 times greater than that of CO2 output from tissues, it appears that the biological role of enzyme B is not that of a carbonic anhydrase.


A study of the connection between gambling and crime in Hungarian prisons.
In the following study, we examine the connection between crime and gambling addictions. In addition to examining the playing habits of 140 detainees prior to their imprisonment, we also study their demographic characteristics, and their relations.(2) We previously studied these correlations as a part of a data recording in 2009 (Tessényi-Kovács, 2011).(3) By the 10th of October 2012, the use of gaming machines was terminated in Hungary. Therefore, our survey has gained a repeated actuality, since a significant cause of addiction has disappeared from the market. We question whether this factor has had a measurable effect. Although the elapsed time is not necessarily enough to measure the presumed changes in crime due to gambling, we nevertheless make the comparison with the results from 2009. For the study of pathological problem gambling, we applied the original version of the SOGS-questionnaire, complemented with demographics and other relevant questions from the perspective of our survey.


Estimating a Preference-Based Single Index Measuring the Quality-of-Life Impact of Self-Management for Diabetes.
Self-management is becoming increasingly important in diabetes but is neglected in conventional preference-based measures. The objective of this paper was to generate health state utility values for a novel classification system measuring the quality-of-life impact of self-management for diabetes, which can be used to generate quality-adjusted life years (QALYs). A large online survey was conducted using a discrete choice experiment (DCE), with duration as an additional attribute, on members of the UK general population ( n = 1,493) to elicit values for health (social limitations, mood, vitality, hypoglycaemia) and non-health (stress, hassle, control, support) aspects of self-management in diabetes. The data were modelled using a conditional fixed-effects logit model and utility estimates were anchored on the one to zero (full health to dead) scale. The model produced significant and consistent coefficients, with one logical inconsistency and 3 insignificant coefficients for the milder levels of some attributes. The anchored utilities ranged from 1 for the best state to -0.029 for the worst state (meaning worse than dead) defined by the classification system. The results presented here can potentially be used to generate utility values capturing the day to day impact of interventions in diabetes on both health and self-management. These utility values can potentially be used to generate QALYs for economic models of the cost-effectiveness of interventions in diabetes.


Species distribution, antifungal susceptibility and clonal relatedness of Candida isolates from patients in neonatal and pediatric intensive care units at a medical center in Turkey.
The aim of this study was to assess species distribution, antifungal susceptibility and clonal relationships among Candida strains isolated from a group of pediatric/neonatal intensive care (PICU/NICU) patients that had a very high mortality rate (76%). The cases of 21 patients (19 with candidemia, 2 with Candida meningitides) treated over a 1-year period in a Turkish hospital PICU and NICU were retrospectively analyzed. Twenty-eight Candida isolates were detected from blood (20), cerebrospinal fluid (CSF) (2) and other specimens (6). Candida species were identified using the API ID 32C System. Susceptibility testing was done (all 28 isolates) for amphotericin B, fluconazole and itraconazole using the broth microdilution method. Arbitrarily primed polymerase chain reaction (AP-PCR) was used for molecular typing of the 3 most common ones; C. albicans (15), C. parapsilosis (6), and C. pelliculosa (4). Electrophoretic karyotyping (EK) was done to check clonal identity obtained by AP-PCR. Of the 20 blood isolates, 8 (40%) were C. albicans, 12 (60%) were non-albicans Candida, and one of the 2 CSF isolates was C. albicans. The overall species distribution was as follows: 15 C. albicans isolates, 6 C. parapsilosis isolates, 4 C. pelliculosa isolates, 2 C. famata isolates and 1 C tropicalis isolate. Amphotericin B had the best antifungal activity with a MIC90 of 0.125 microg/ml, and the rates of susceptibility to fluconazole and itraconazole were 93% and 82%, respectively. AP-PCR revealed 11 genotypes (4 were identical pairs, 7 were distinct) among the 15 C. albicans isolates, 2 genotypes (5 were classified in the same type) among the 6 C. parapsilosis isolates, and 4 separate genotypes for the 4 C. pelliculosa isolates. Karyotyping results correlated well with the AP-PCR findings. As indicated in the previous research, our results confirmed that non-albicans Candida species have become more frequently causative agents for invasive fungal infections in the ICU. Transmission of C. albicans and C. pelliculosa was relatively low, but transmission of C. parapsilosis was high, suggesting that more effective control and very strict treatment protocols are needed for patients having high mortality and invasive fungal infection in ICU.


Carabidae population dynamics and temporal partitioning: response to coupled neonicotinoid-transgenic technologies in maize.
Insecticidal Bt crops and seed treatments represent additional pest management tools for growers, prompting ecological studies comparing their impact on farm system inputs and effects to nontarget organisms compared with conventional practices. Using high taxonomic and temporal resolution, we contrast the dominance structure of carabids and dynamics of the most abundant species in maize (both sweet and field corn) agroecosystems using pest management tactics determined by the purchase of seed and application of pyrethroid insecticides. In the seed-based treatments, sweet corn contained Cry1Ab/c proteins, whereas field corn contained the coupled technology of Cry3Bb1 proteins for control of corn rootworm and neonicotinoid seed treatments aimed at secondary soil-borne pests. The insecticide treatments involved foliar pyrethroids in sweet corn and at-planting pyrethroids in field corn. The carabid community, comprised of 49 species, was dominated by four species, Scarites quadriceps Chaudoir, Poecilus chalcites Say, Pterostichus melanarius Illiger, and Harpalus pensylvanicus DeGeer, that each occupied a distinct temporal niche during the growing season. Two species, Pt. melanarius and H. pensylvanicus, exhibited differences between treatments over time. Only H. pensylvanicus had consistent results in both years, in which activity densities in field corn were significantly higher in the control in July and/or August. These results, along with laboratory bioassays, led us to hypothesize that lower adult captures resulted from decrease in prey availability or exposure of H. pensylvanicus larvae to soil-directed insecticides-either the neonicotinoid seed treatment in the transgenic field corn or an at-planting soil insecticide in the conventional field corn.


Brain lesions in birds: effects on discrimination acquisition and reversal.
Bobwhite quail (Colinus virginianus) with lesions of the cortex and the dorsal portion of the hyperstriatum (or " wulst") showed a considerable deficit in ability to reverse a learned discrimination between horizontal and vertical stripes. Two birds that had been run on 25 such reversals before ablation showed the same result. Lesioned birds did not, however, differ from controls in the Original acquisition of the discrimination. Analysis of performance indicated that the reversal deficit was not due to difficulties in discrimination, interference with motor ability, or lack of motivation. The deficit appeared to be based on perseveration of response to the previous positive stimulus during reversal training.


Proline-containing beta-turns. IV. Crystal and solution conformations of tert.-butyloxycarbonyl-L-prolyl-D-alanine and tert.-butyloxycarbonyl-L-prolyl-D-alanyl-L-alanine.
The conformations of the dipeptide t-Boc-Pro-DAla-OH and the tripeptide t-Boc-Pro-DAla-Ala-OH have been determined in the crystalline state by X-ray diffraction and in solution by CD, n.m.r. and i.r. techniques. The unit cell of the dipeptide crystal contains two independent molecules connected by intermolecular hydrogen bonds. The urethane-proline peptide bond is in the cis orientation in both the molecular forms while the peptide bond between Pro and DAla is in the trans orientation. The single dipeptide molecule exhibits a "bent" structure which approximates to a partial beta-turn. The tripeptide adopts the 4----1 hydrogen-bonded type II beta-turn with all trans peptide bonds. In solution, the CD and i.r. data on the dipeptide indicate an ordered conformation with an intramolecular hydrogen bond. N.m.r. data indicate a significant proportion of the conformer with a trans orientation at the urethane-proline peptide bond. The temperature coefficient of the amide proton of this conformer in DMSO-d6 points to a 3----1 intramolecular hydrogen bond. Taken together, the data on the dipeptide in solution indicate the presence (in addition to the cis conformer) of a C7 conformation which is absent in the crystalline state. The spectral data on the tripeptide indicate the presence of the type II beta-turn in solution in addition to the nonhydrogen-bonded conformer with the cis peptide bond between the urethane and proline residues. The relevance of these data to studies on the substrate specificity of collagen prolylhydroxylase is pointed out.


Prenatal screening for β-thalassemia major reveals new and rare mutations in the Pakistani population.
β-Thalassemia is the most common genetic disorder in Pakistan, where more than 6000 affected children are born annually, and the carrier population is around 10 million. The objective was to study β-globin gene mutations in chorionic villous biopsy samples. Prenatal screening of 383 pregnant women between 2003 and 2010 was carried out using a panel of 13 mutation primers and amplification refractory mutations system (ARMS)-PCR. In addition, DNA sequencing was used to confirm uncharacterized mutations and in some cases fetal disease status was confirmed by linkage analysis. Families enrolled in this study represented major ethnic groups in Pakistan. Of the 13 mutations tested, three mutations accounted 71% of the total, including IVS1-5(G-C)[HBB:c.92+5G>C], codon 8/9(+G) [HBB:c.27_28insG] and del 619[NG_000007.3:g71609-72227del619]. Mutations in four uncharacterized samples were later confirmed by DNA sequencing as -88(C-T)[HBB:c.-138C>G], -90(C-T)[HBB:c.-140C>T] and codon 59(+T)[HBB:c.178_179insT]. To our knowledge, this is the first report of these mutations in Pakistan. Moreover, 19.2% fetal samples were normal and 52.3% heterozygous, whereas 26.4% were affected with thalassemia major. IVS1-5:IVS1-5 was the most common genotype in fetal samples. Prenatal diagnosis of β-thalassemia using ARMS PCR is an efficient approach for reducing the burden of this disease in Pakistan. In addition, rare mutations reported in this study should be incorporated in the diagnostic strategy.


Flavobacterium frigidarium sp. nov., an aerobic, psychrophilic, xylanolytic and laminarinolytic bacterium from Antarctica.
A psychrophilic, aerobic bacterium designated A2iT was isolated from marine sediment recovered from shallow waters surrounding Adelaide Island, Antarctica (67 degrees 34' S, 68 degrees 07' W). The organism exhibited xylanolytic and laminarinolytic activity and was halotolerant. Basic characterization showed that it was gram-negative, non-motile, yellow-pigmented (beta,beta-carotene-3,3'-diol) and positive for oxidase and catalase synthesis. Analysis of the 16S rDNA sequence suggests that the organism belongs to the Flexibacter-Cytophaga-Bacteroides phylum. On the basis of its 16S rDNA sequence, the bacterium is 96.8% similar to Flavobacterium columnare ATCC 43622--its closest relation. The genomic DNA G+C content was 35 mol%. Growth on xylan occurs optimally at 15 degrees C, though growth also occurs at 0 degrees C, and the doubling times are 9.6 and 34.8 h, respectively. The maximum growth temperature on xylan is at 24 degrees C. The bacterium is a neutrophile, growing across the pH range 5.6-8.4 and having an optimum at pH 7.5. Analysis of the 16S rDNA sequence, together with phenotypic characterization, suggests that the organism is a member of the genus Flavobacterium. DNA-DNA hybridization experiments have shown that it is a novel species; it is proposed, therefore, that the organism be designated as the type strain of Flavobacterium frigidarium sp. nov. (= ATCC 700810T = NCIMB 13737T).


Light-Controlled Fermentations for Microbial Chemical and Protein Production.
Microbial cell factories offer a sustainable alternative for producing chemicals and recombinant proteins from renewable feedstocks. However, overburdening a microorganism with genetic modifications can reduce host fitness and productivity. This problem can be overcome by using dynamic control: inducible expression of enzymes and pathways, typically using chemical- or nutrient-based additives, to balance cellular growth and production. Optogenetics offers a non-invasive, highly tunable, and reversible method of dynamically regulating gene expression. Here, we describe how to set up light-controlled fermentations of engineered Escherichia coli and Saccharomyces cerevisiae for the production of chemicals or recombinant proteins. We discuss how to apply light at selected times and dosages to decouple microbial growth and production for improved fermentation control and productivity, as well as the key optimization considerations for best results. Additionally, we describe how to implement light controls for lab-scale bioreactor experiments. These protocols facilitate the adoption of optogenetic controls in engineered microorganisms for improved fermentation performance.


Microbial populations and enzyme activities in soil in situ under transgenic corn expressing cry proteins from Bacillus thuringiensis.
Transgenic Bt crops produce insecticidal Cry proteins that are released to soil in plant residues, root exudates, and pollen and that may affect soil microorganisms. As a continuation of studies in the laboratory and a plant-growth room, a field study was conducted at the Rosemount Experiment Station of the University of Minnesota. Three Bt corn varieties that express the Cry1Ab protein, which is toxic to the European corn borer (Ostrinia nubilalis Hübner), and one Bt corn variety that expresses the Cry3Bb1 protein, which is toxic to the corn rootworm complex (Diabrotica spp.), and their near-isogenic non-Bt varieties were evaluated for their effects on microbial diversity by classical dilution plating and molecular (polymerase chain reaction-denaturing gradient gel electrophoresis) techniques and for the activities of some enzymes (arylsulfatases, acid and alkaline phosphatases, dehydrogenases, and proteases) involved in the degradation of plant biomass. After 4 consecutive years of corn cultivation (2003-2006), there were, in general, no consistent statistically significant differences in the numbers of different groups of microorganisms, the activities of the enzymes, and the pH between soils planted with Bt and non-Bt corn. Numbers and types of microorganisms and enzyme activities differed with season and with the varieties of corn, but these differences were not related to the presence of the Cry proteins in soil. The Cry1Ab protein of Bt corn (events Bt11 and MON810) was detected in most soils during the 4 yr, whereas the Cry3Bb1 protein was not detected in soils of Bt corn (event MON863) expressing the cry3Bb1 gene.


Superresolution Imaging of Clinical Formalin Fixed Paraffin Embedded Breast Cancer with Single Molecule Localization Microscopy.
Millions of archived formalin-fixed, paraffin-embedded (FFPE) specimens contain valuable molecular insight into healthy and diseased states persevered in their native ultrastructure. To diagnose and treat diseases in tissue on the nanoscopic scale, pathology traditionally employs electron microscopy (EM), but this platform has significant limitations including cost and painstaking sample preparation. The invention of single molecule localization microscopy (SMLM) optically overcame the diffraction limit of light to resolve fluorescently labeled molecules on the nanoscale, leading to many exciting biological discoveries. However, applications of SMLM in preserved tissues has been limited. Through adaptation of the immunofluorescence workflow on FFPE sections milled at histological thickness, cellular architecture can now be visualized on the nanoscale using SMLM including individual mitochondria, undulations in the nuclear lamina, and the HER2 receptor on membrane protrusions in human breast cancer specimens. Using astigmatism imaging, these structures can also be resolved in three dimensions to a depth of ~800 nm. These results demonstrate the utility of SMLM in efficiently uncovering ultrastructural information of archived clinical samples, which may offer molecular insights into the physiopathology of tissues to assist in disease diagnosis and treatment using conventional sample preparation methods.


Subcutaneous tissue response to titanium, poly(ε-caprolactone), and carbonate-substituted hydroxyapatite-coated poly(ε-caprolactone) plates: a rabbit study.
The aim of this study was to evaluate the soft tissue response to poly(ε-caprolactone) (PCL) implants with and without carbonate-substituted hydroxyapatite (CHA) coating compared to the commonly used titanium alloy (Ti-6Al-4V)-machined surface. Experimental materials were implanted subcutaneously in New Zealand white rabbits for 5 weeks. The tissue attachment strength, as evaluated by a tissue peel test, histological and histomorphology analysis, as well as scanning electron microscopy were compared between groups. The peel test result revealed no statistically significant difference between groups. Histological analysis found fibrous capsule formation around all implant materials. The fibrous capsule around PCL implants with and without CHA coating was significantly thinner compared with the capsule thickness around the titanium implants. However, the inflammatory cells, as present at the fibrous capsule-implant interface, were found to be significantly lower in the Ti-group. In conclusion, the current data do not prove that PCL or PCL with a CHA coating results in a superior soft tissue response compared with a machined titanium implant.


FcγRIIb on myeloid cells and intrinsic renal cells rather than B cells protects from nephrotoxic nephritis.
FcγRIIb is the sole inhibitory FcR for IgG in humans and mice, where it is involved in the negative regulation of Ab production and cellular activation. FcγRIIb-deficient mice show exacerbated disease following the induction of nephrotoxic nephritis (NTN). In this study, we determined the cellular origin of the FcγRIIb-knockout phenotype by inducing NTN in mice with a deficiency of FcγRIIb on either B cells alone (FcγRIIB(fl/fl)/CD19Cre(+)) or myeloid cells (FcγRIIB(fl/fl)/CEBPαCre(+)). Deletion of FcγRIIb from B cells did not increase susceptibility to NTN, compared with wild-type (WT) mice, despite higher Ab titers in the FcγRIIB(fl/fl)/CD19Cre(+) mice compared with the WT littermate controls. In contrast, mice lacking FcγRIIb on myeloid cells had exacerbated disease as measured by increased glomerular thrombosis, glomerular crescents, albuminuria, serum urea, and glomerular neutrophil infiltration when compared with WT littermate controls. The role for FcγRIIb expression on radioresistant intrinsic renal cells in the protection from NTN was then investigated using bone marrow chimeric mice. FcγRIIb(-/-) mice transplanted with FcγRIIb(-/-) bone marrow were more susceptible to NTN than WT mice transplanted with FcγRIIb(-/-) bone marrow, indicating that the presence of WT intrinsic renal cells protects from NTN. These results demonstrate that FcγRIIb on myeloid cells plays a major role in protection from NTN, and therefore, augmentation of FcγRIIb on these cells could be a therapeutic target in human Ab-mediated glomerulonephritis. Where there was a lack of FcγRIIb on circulating myeloid cells, expression of FcγRIIb on intrinsic renal cells provided an additional level of protection from Ab-mediated glomerulonephritis.


Value of Routine Dengue Diagnostic Tests in Urine and Saliva Specimens.
Dengue laboratory diagnosis is essentially based on detection of the virus, its components or antibodies directed against the virus in blood samples. Blood, however, may be difficult to draw in some patients, especially in children, and sampling during outbreak investigations or epidemiological studies may face logistical challenges or limited compliance to invasive procedures from subjects. The aim of this study was to assess the possibility of using saliva and urine samples instead of blood for dengue diagnosis. Serial plasma, urine and saliva samples were collected at several time-points between the day of admission to hospital until three months after the onset of fever in children with confirmed dengue disease. Quantitative RT-PCR, NS1 antigen capture and ELISA serology for anti-DENV antibody (IgG, IgM and IgA) detection were performed in parallel on the three body fluids. RT-PCR and NS1 tests demonstrated an overall sensitivity of 85.4%/63.4%, 41.6%/14.5% and 39%/28.3%, in plasma, urine and saliva specimens, respectively. When urine and saliva samples were collected at the same time-points and tested concurrently, the diagnostic sensitivity of RNA and NS1 detection assays was 69.1% and 34.4%, respectively. IgG/IgA detection assays had an overall sensitivity of 54.4%/37.4%, 38.5%/26.8% and 52.9%/28.6% in plasma, urine and saliva specimens, respectively. IgM were detected in 38.1% and 36% of the plasma and saliva samples but never in urine. Although the performances of the different diagnostic methods were not as good in saliva and urine as in plasma specimens, the results obtained by qRT-PCR and by anti-DENV antibody ELISA could well justify the use of these two body fluids to detect dengue infection in situations when the collection of blood specimens is not possible.


Gap between flexion and extension ranges of motion: a novel indicator to predict the loss of cervical lordosis after laminoplasty in patients with cervical spondylotic myelopathy.
Kyphotic deformity resulting from the loss of cervical lordosis (CL) is a rare but serious complication after cervical laminoplasty (CLP), and it is essential to recognize the risk factors. Previous studies have demonstrated that a greater flexion range of motion (fROM) and smaller extension ROM (eROM) in the cervical spine are associated with the loss of CL after CLP. Considering these facts together, one can hypothesize that an indicator representing the gap between fROM and eROM (gROM) is highly useful in predicting postoperative CL loss. In the present study, the authors aimed to investigate the risk factors of marked CL loss after CLP for cervical spondylotic myelopathy (CSM), including the gROM as a potential predictor. Patients who had undergone CLP for CSM were divided into those with and those without a loss of more than 10° in the sagittal Cobb angle between C2 and C7 at the final follow-up period compared to preoperative measurements (CL loss [CLL] group and no CLL [NCLL] group, respectively). Demographic characteristics, surgical information, preoperative radiographic measurements, and posterior paraspinal muscle morphology evaluated with MRI were compared between the two groups. fROM and eROM were examined on neutral and flexion-extension views of lateral radiography, and gROM was calculated using the following formula: gROM (°) = fROM - eROM. The performance of variables in discriminating between the CLL and NCLL groups was assessed using the receiver operating characteristic (ROC) curve. This study included 111 patients (mean age at surgery 68.3 years, 61.3% male), with 10 and 101 patients in the CLL and NCLL groups, respectively. Univariate analyses showed that fROM and gROM were significantly greater in the CLL group than in the NCLL group (40.2° vs 26.6°, p < 0.001; 31.6° vs 14.3°, p < 0.001, respectively). ROC curve analyses revealed that both fROM and gROM had excellent discriminating capacities; gROM was likely to have a higher area under the ROC curve than fROM (0.906 vs 0.860, p = 0.094), with an optimal cutoff value of 27°. The gROM is a highly useful indicator for predicting a marked loss of CL after CLP. For CSM patients with a preoperative gROM exceeding 30°, CLP should be carefully considered, since kyphotic changes can develop postoperatively.


Identification of the Ferredoxin-Binding Site of a Ferredoxin-Dependent Cyanobacterial Nitrate Reductase.
An in silico model for the 1:1 ferredoxin (Fd)/nitrate reductase (NR) complex, using the known structure of Synechocystis sp. PCC 6803 Fd and the in silico model of Synechococcus sp. PCC 7942 NR, is used to map the interaction sites that define the interface between Fd and NR. To test the electrostatic interactions predicted by the model complex, five positively charged NR amino acids (Arg43, Arg46, Arg197, Lys201, and Lys614) and a negatively charged amino acid (Glu219) were altered using site-directed mutagenesis and characterized by activity measurements, metal analysis, and electron paramagnetic resonance (EPR) studies. All of the charge replacement variants retained wild-type levels of activity with reduced methyl viologen (MV), but a significant decrease in activity was observed for the R43Q, R46Q, K201Q, and K614Q variants when reduced Fd served as the electron donor. EPR analysis as well as the Fe and Mo analyses showed that loss of activity observed with these variants was not the consequence of perturbation of the Mo center or [4Fe-4S] cluster. Therefore, the loss of the Fd-linked specific activity observed with these variants can be explained only by invoking a role for Arg43, Arg46, Lys201, and Lys614 in Fd binding. The R43Q, R46Q, K201Q, and K614Q NR variants also showed a decreased binding affinity for Fd, compared to that of wild-type NR, supporting a key role of these four positively charged residues in the productive binding of Fd.


Allotype-specific analysis of anti-(Tyr,Glu)-Ala-Lys antibodies produced by Ir-1A high and low responder chimeric mice.
Katz et al. (1) have demonstrated a restriction in lymphoid cell interaction when the antigen used is under immune response (Ir) gene control. T cells from (low responder x high responder) F(1) mice primed to the terpolymer L-glutamic acid, L-lysine, L-tyrosine (GLT) can collaborate with 2,4-dinitrophenyl (DNP)-primed B cells from the Ir-GLT high responder but not low responder strain in response to DNP-GLT (1). In contrast are the studies of Bechtol et al. and Bechtol and McDevitt (2,3), who examined the antibody responses of tetraparental mice immunized with the synthetic polypeptide poly-L(Tyr,Glu)-poly D,L-Ala- poly-L-Lys ((T,G)-A-L), an antigen under Ir-1A genetic control. Several tetraparental mice produced anti(T-,G)-A-L antibody of low responder strain immunoglobulin (Ig) allotype (2,3). These results indicated that he Ir-1A gene was not expressed in B cells and implied that interactions among genetically dissimilar cell populations could occur when tolerance existed to H-2 antigenic differences. Recent studies with bone marrow cell chimeric mice have shown that chimeric T cells can interact with H-2 histoincompatible B cells in response to antigens not under Ir gene control (4-6). To clarify whether lymphoid cell chimerism, with presumed tolerance to H-2 incompatibility, would permit effective cell interactions in response to antigens under Ir gene control, bone marrow cell chimeric mice were prepared by using strains differing both for Ig allotype and for high versus low responsiveness to (T,G)-A-L. An antigen-specific and allotype- specific antibody assay was used to discriminate the responses produced by high and low responder strain B cells in these chimeras. The results suggest that lymphoid cell chimerism per se is not sufficient to obviate Ir gene-mediated restriction in cell interaction.


'They are called Imperfect men': Male Infertility and Sexual Health in Early Modern England.
Scholars of early modern gender and medicine have tended to focus on female infertility. Discussions that have included male reproductive failure have considered sexual ability and impotence, rather than infertility. Nonetheless, fathering children was important to male social standing and the fulfilment of their patriarchal roles. This article will demonstrate that male infertility was not absent from medical literature, but appeared in a variety of settings including tests for infertility, seventeenth-century handbills for treatments, and surgical treatises. It will show that medical and surgical writers accepted that men could be rendered infertile, but still sexually capable, in a variety of ways. Moreover, the article will show that seventeenth-century surgeons expected male readers to be concerned about their reproductive potential and constructed a framework of efficacy based upon their ability to secure on-going fertility.


Removal of hexavalent chromium from aqueous solution by calcined Zn/Al-LDHs.
In this study, Zn/Al-layered double hydroxides (Zn/Al-LDHs) were synthesized by a co-precipitation method and characterized with X-ray diffraction, Fourier transform infrared spectroscopy, and scanning electron microscopy. Then the hexavalent chromium Cr(VI) adsorption experiments on calcined Zn/Al-LDHs were carried out to analyze the effects of pH, temperature, adsorption time, initial Cr(VI) concentration and adsorbent dosage on the removal of Cr(VI) from aqueous solutions. The maximum adsorption capacity for Cr(VI) on calcined Zn/Al-LDHs under optimal conditions was found to be over 120 mg/g. The kinetic and isotherm of Cr(VI) adsorption on calcined Zn/Al-LDHs can be described with the pseudo-second-order kinetic model and Langmuir isotherm, respectively.


Experimental phasing for structure determination using membrane-protein crystals grown by the lipid cubic phase method.
Despite the marked increase in the number of membrane-protein structures solved using crystals grown by the lipid cubic phase or in meso method, only ten have been determined by SAD/MAD. This is likely to be a consequence of the technical difficulties associated with handling proteins and crystals in the sticky and viscous hosting mesophase that is usually incubated in glass sandwich plates for the purposes of crystallization. Here, a four-year campaign aimed at phasing the in meso structure of the integral membrane diacylglycerol kinase (DgkA) from Escherichia coli is reported. Heavy-atom labelling of this small hydrophobic enzyme was attempted by pre-labelling, co-crystallization, soaking, site-specific mercury binding to genetically engineered single-cysteine mutants and selenomethionine incorporation. Strategies and techniques for special handling are reported, as well as the typical results and the lessons learned for each of these approaches. In addition, an assay to assess the accessibility of cysteine residues in membrane proteins for mercury labelling is introduced. The various techniques and strategies described will provide a valuable reference for future experimental phasing of membrane proteins where crystals are grown by the lipid cubic phase method.


Should Graphic Warning Labels Proposed for Cigarette Packages Sold in the United States Mention the Food and Drug Administration?
Under the US Family Smoking Prevention and Tobacco Control Act, the US Food and Drug Administration (FDA) has the authority to implement graphic warning labels (GWLs) on cigarette packages. Neither the original labels proposed by the FDA nor the revised labels include a source to indicate sponsorship of the warnings. This study tests the potential impact of adding a sponsor to the content of GWLs. We recruited adult smokers (N = 245) and middle-school youth (N = 242) from low-income areas in the Northeastern US. We randomly assigned participants to view one of three versions of the original FDA-proposed warning labels in a between-subjects experiment: no sponsor, "US Food and Drug Administration," or "American Cancer Society" sponsor. We tested the effect of varying sponsorship on source attribution and source credibility. Compared to unsponsored labels, FDA sponsorship increased source attributions that the FDA sponsored the labels among both middle-school, largely nonsmoking youth and adult smokers. However, sponsorship had no effect on source credibility among either population. We found no evidence that adding FDA as the source is likely to boost source credibility judgments, at least in the short term; though doing so would not appear to have adverse effects on credibility judgments. As such, our data are largely consistent with the Tobacco Control Act's provisions that allow, but do not require, FDA sponsorship on the labels. This study addresses the FDA's regulatory efforts by informing the possible design and content of future cigarette warning labels. Our results do not offer compelling evidence that adding the FDA name on GWLs will directly increase source credibility. Future work may test more explicit FDA source labeling and continue to examine the credibility of tobacco message content among high-priority populations.


High-resolution genomic profiling to predict 10-year overall survival in node-negative breast cancer.
Women with clinically node-negative breast cancer have a better prognosis than do those with axillary lymph node metastasis. Nonetheless, approximately 20% of node-negative patients die within 15 years of diagnosis, and thus additional prognostic markers are greatly needed. To identify specific copy number alterations (CNAs) that differed in frequency between 10-year survivors and deceased patients with node-negative breast cancer, array comparative genomic hybridization (aCGH) was applied to 41 primary node-negative breast tumors. Fisher's exact test was used to identify significantly different CNAs between 10-year survivors and deceased patients. Losses at 8p21.2 approximately p21.3, 8p23.1 approximately p23.2, Xp21.3, and Xp22.31 approximately p22.33 were significantly more common in tumors from deceased patients, suggesting that these alterations may contribute to tumor aggressiveness. Gains at 1q25.2 approximately q25.3 and 1q31.3 approximately q41 were more prevalent in tumors from survivors; specific gains at these genomic regions may inhibit further tumor progression, resulting in a less aggressive form of node-negative breast cancer. Evaluation of the identified CNAs in an independent external data set verified the prognostic potential of the 1q31.3 approximately q41 region. Although further extensive validation is needed, the prognostic CNAs identified in this work may in time facilitate the clinical assessment of breast cancer.


Differential cytokine regulation by NF-kappaB and AP-1 in Jurkat T-cells.
Activator protein (AP)-1 and nuclear factor (NF)-kappaB largely control T-cell activation, following binding of foreign antigens to the T-cell receptor leading to cytokine secretion. Elevated levels of pro-inflammatory cytokines and chemokines such as TNF, IL-6 and CXCL8 are associated with several human diseases including cystic fibrosis, pulmonary fibrosis and AIDS. The aim of this study was to investigate the role of the transcription factors, AP-1 and NF-kappaB, in IL-6 and CXCL8 regulation in Jurkat T-cells. Phorbol myristate acetate (PMA) exposure resulted in an up-regulation of AP-1 and down-regulation of NF-kappaB activity, however, exposure to heat killed (HK) Escherichia. coli MG1655 resulted in a dose-dependent increase in NF-kappaB activity without affecting AP-1. The cytokine profile revealed an up-regulation of the chemokine CXCL8 and the pro-inflammatory cytokines TNF, IL-2 and IL-6 following treatment with both PMA and HK E. coli, while the levels of the anti-inflammatory cytokine IL-10 were not affected by PMA but were significantly down-regulated by HK E. coli. AP-1 activation was significantly increased 2 h after PMA exposure and continued to increase thereafter. In contrast, NF-kappaB responded to PMA exposure by a rapid up-regulation followed by a subsequent down-regulation. Increased intracellular Ca2+ concentrations countered the down-regulation of NF-kappaB by PMA, while similar treatment with calcium ionophore resulted in a reduced NF-kappaB activity following induction with HK E. coli. In order to further study NF-kappaB activation, we considered two up-stream signalling proteins, PKC and Bcl10. Phosphorylated-PKC levels increased in response to PMA and HK E. coli, while Bcl10 levels significantly decreased following PMA treatment. Using an NF-kappaB activation inhibitor, we observed complete inhibition of IL-6 expression while CXCL8 levels only decreased by 40% at the highest concentration. Treatment of Jurkat T-cells with PMA in the presence of JNK-inhibitor suppressed both CXCL8 and IL-6 while PKC-inhibitor primarily decreased CXCL8 expression. The present study shows that NF-kappaB regulated IL-6 but not CXCL8. This complex regulation of CXCL8 suggests that there is a need to further evaluate the signalling pathways in order to develop new treatment for diseases with elevated CXCL8 levels, such as AIDS and autoimmune diseases.


Reslizumab and Eosinophilic Asthma: One Step Closer to Precision Medicine?
Human eosinophils represent approximately 1% of peripheral blood leukocytes. However, these cells have the propensity to leave the blood stream and migrate into inflamed tissues. Eosinophilic inflammation is present in a significant proportion of patients with severe asthma. Asthma is a chronic inflammatory disorder that affects more than 315 million people worldwide, with 10% having severe uncontrolled disease. Although the majority of patients can be efficiently treated, severe asthmatics continue to be uncontrolled and are at risk of exacerbations and even death. Interleukin-5 (IL-5) plays a fundamental role in eosinophil differentiation, maturation, activation and inhibition of apoptosis. Therefore, targeting IL-5 is an appealing approach to the treatment of patients with severe eosinophilic asthma. Reslizumab, a humanized anti-IL-5 monoclonal antibody, binds with high affinity to amino acids 89-92 of IL-5 that are critical for binding to IL-5 receptor α. Two phase III studies have demonstrated that reslizumab administration in adult patients with severe asthma and eosinophilia (≥400 cells/μL) improved lung function, asthma control, and symptoms. Thus, the use of blood eosinophils as a baseline biomarker could help to select patients with severe uncontrolled asthma who are likely to achieve benefits in asthma control with reslizumab. In conclusion, targeted therapy with reslizumab represents one step closer to precision medicine in patients with severe eosinophilic asthma.


Foliar uptake of radiocaesium from irrigation water by paddy rice (Oryza sativa): an overlooked pathway in contaminated environments.
Flooded (paddy) rice (Oryza sativa) can take up ions from the irrigation water by foliar uptake via the exposed stem base. We hypothesised that the stem base uptake of radiocaesium (RCs) is a pathway for rice grown in RCs-contaminated environments. We developed a bi-compartmental device which discriminates the stem base from root RCs uptake from solutions, thereby using RCs isotopes (137 Cs and 134 Cs) with < 2% solution leak between the compartments. Radiocaesium uptake was linear over time (0-24 h). Radiocaesium uptake to the entire plant, expressed per dry weight of the exposed parts, was sixfold higher for the roots than for the exposed stem base. At equal RCs concentrations in both compartments, the exposed stem base and root uptake contributed almost equally to the total shoot RCs concentrations. Reducing potassium supply to the roots not only increased the root RCs uptake but also increased RCs uptake by the stem base. This study was the first to experimentally demonstrate active and internally regulated RCs uptake by the stem base of rice. Scenario calculations for the Fukushima-affected area predict that RCs in irrigation water could be an important source of RCs in rice as indirectly suggested from field data.


Effects of genetic merit for carcass weight, breed type and slaughter weight on performance and carcass traits of beef × dairy steers.
Crossbreeding of Holstein-Friesian dairy cows with both early maturing (e.g. Aberdeen Angus (AA)) and late maturing (e.g. Belgian Blue (BB)) beef breeds is commonly practised. In Ireland, genetic merit for growth rate of beef sires is expressed as expected progeny difference for carcass weight (EPD(CWT)). The objective of this study was to compare the progeny of Holstein-Friesian cows, sired by AA and BB bulls of low (L) and high (H) EPD(CWT) for performance and carcass traits. A total of 118 spring-born male progeny from 20 (9 AA and 11 BB) sires (8 L and 12 H) were managed together from shortly after birth to about 19 months of age. They were then assigned to one of two mean slaughter weights (560 kg (light) or 620 kg (heavy)). Following slaughter, carcasses were graded for conformation class and fat class, the 6th to 10th ribs joint was dissected as an indicator of carcass composition, and samples of subcutaneous fat and musculus longissimus were subjected to Hunterlab colour measurements. A sample of m. longissimus was also chemically analysed. Slaughter and carcass weights per day of age for AAL, AAH, BBL and BBH were 747, 789, 790 and 805 (s.e. 10.5) g, and 385, 411, 427 and 443 (s.e. 4.4) g, respectively. Corresponding carcass weight, kill-out proportion, carcass conformation class (scale 1 to 5) and carcass fat class (scale 1 to 5) values were 289, 312, 320 and 333 (s.e. 4.0) kg, 516, 522, 542 and 553 (s.e. 3.5) g/kg, 2.5, 2.4, 3.0 and 3.1 (s.e. 0.10), and 3.4, 3.5, 2.9 and 2.8 (s.e. 0.11). There were few breed type × genetic merit interactions. Delaying slaughter date increased slaughter weight, carcass weight and all measures of fatness. It also reduced the proportion of carcass weight in the hind quarter and the proportions of bone and muscle in the ribs joint. None of these effects accompanied the increase in carcass weight due to higher EPD(CWT). It is concluded that BB have superior production traits to AA. Selection of sires for higher EPD(CWT) increases growth rate, kill-out proportion and carcass weight of progeny with little effect on carcass or muscle traits. The extra carcass weight due to higher EPD(CWT) is more valuable commercially than a comparable carcass weight increment from a delay in slaughter date because it comprises a higher proportion of muscle.


Lafora disease.
Lafora disease (LD) is an autosomal recessive progressive myoclonus epilepsy due to mutations in the EPM2A (laforin) and EPM2B (malin) genes, with no substantial genotype-phenotype differences between the two. Founder effects and recurrent mutations are common, and mostly isolated to specific ethnic groups and/or geographical locations. Pathologically, LD is characterized by distinctive polyglucosans, which are formations of abnormal glycogen. Polyglucosans, or Lafora bodies (LB) are typically found in the brain, periportal hepatocytes of the liver, skeletal and cardiac myocytes, and in the eccrine duct and apocrine myoepithelial cells of sweat glands. Mouse models of the disease and other naturally occurring animal models have similar pathology and phenotype. Hypotheses of LB formation remain controversial, with compelling evidence and caveats for each hypothesis. However, it is clear that the laforin and malin functions regulating glycogen structure are key. With the exception of a few missense mutations LD is clinically homogeneous, with onset in adolescence. Symptoms begin with seizures, and neurological decline follows soon after. The disease course is progressive and fatal, with death occurring within 10 years of onset. Antiepileptic drugs are mostly non-effective, with none having a major influence on the progression of cognitive and behavioral symptoms. Diagnosis and genetic counseling are important aspects of LD, and social support is essential in disease management. Future therapeutics for LD will revolve around the pathogenesics of the disease. Currently, efforts at identifying compounds or approaches to reduce brain glycogen synthesis appear to be highly promising.


Risk of stroke after surgery in patients with and without chronic atrial fibrillation.
The extent to which chronic atrial fibrillation affects the risk of postoperative stroke is largely unknown. We sought to determine the 30-day rate of stroke among patients with and without chronic AF who underwent 10 different types of surgery. The crude incidence of stroke was retrospectively determined using a population-based linked administrative database of hospitalized patients who underwent specified operations between 1 January 1996 and 30 November 2005. The risk of stroke in patients with AF was adjusted for age, race, sex, presence of diabetes, heart failure, hypertension and prior stroke. The overall 30-day rate of stroke in 69 202 patients with chronic AF was 1.8% (95% CI, 1.7-1.9%) vs. 0.6% (CI, 0.58-0.62%) in 2 470 649 patients without AF. The risk-adjusted odds of a postoperative stroke in patients with chronic AF were 2.1 (CI, 2.0-2.3). The highest incremental difference in the crude rate of stroke was observed in patients undergoing neurologic or vascular surgery, with a difference of approximately 2%. Patients with chronic AF had twice the risk of postoperative stroke. Randomized trials are needed to determine if aggressive perioperative anticoagulation can reduce the incidence of postoperative stroke in patients with AF.


Identification of high-grade serous ovarian cancer miRNA species associated with survival and drug response in patients receiving neoadjuvant chemotherapy: a retrospective longitudinal analysis using matched tumor biopsies.
Neoadjuvant chemotherapy (NACT) has been recognized as a reliable therapeutic strategy in patients with unresectable advanced epithelial ovarian cancer (EOC). The molecular events leading to platinum (Pt) response in NACT settings have hitherto not been explored. In the present work, longitudinal changes of miRNA expression profile were investigated to identify miRNA families with prognostic role in high-grade serous EOC patients who received the NACT regimen. One hundred sixty-four matched tumor biopsies taken at initial laparoscopic evaluation and at interval-debulking surgery (IDS) after four courses of Pt-based therapy were selected from 82 stage IIIC-IV high-grade serous-EOC patients that were judged unsuitable for complete primary debulking and subjected the NACT protocol. miRNA profiling by microarray, real-time PCR and immuno-histochemical staining for Smad2 phosphorylation (P-Smad2) were used for data analysis. Analysis revealed that 369 miRNAs were differentially expressed in matched biopsies (referred to as DEMs). DEMs were not scattered across the genome, but clustered into families: miR-199, let-7, miR-30, miR-181 and miR-29. Multivariate analysis showed that miR-199a-3p, miR-199a-5p, miR-181a-5p and let-7g-5p associated with overall and progression-free survival (P < 0.05); miR-199a-3p, miR-199a-5p and miR-181a-5p associated with residual tumor volume and Pt-free interval (P < 0.05). Immuno-histochemical staining confirmed an enrichment of P-Smad2, a marker of transforming growth factor-β activation, in tumors from patients with shorter PFS and OS, and with high levels of expression of miR-181a-5p (P < 0.05). Kaplan-Meier curves plotting concomitant expression of P-Smad2 and miR-181a-5p show significant differences in PFS and OS compared with those depicting the expression of each biomarker alone (P < 0.001). This study describes several miRNA families with a prognostic role in the NACT setting. It also confirms that concomitant analysis of P-Smad2 and miR-181a-5p in surgical samples may be capable of identifying those ovarian cancer patients with poor outcome and little chance of response to Pt-based NACT.


Development of spontaneous leg movements in infants with and without periventricular leukomalacia.
The main question asked in the present study was whether support could be found for the notion that supraspinal influences on the generation of spontaneous kicking movements become increasingly apparent in the first half-year after birth. In comparing groups of infants with and without damage in tracts connected with the cortex surrounding the central sulcus, such support would consist of the finding that similar patterns of spontaneous kicking are observed early in development, whereas differences between groups should occur with increasing age. Using 3-D registrations, the spontaneous kicking movements of 19 infants with differing degrees of periventricular, lobar, and subcortical leukomalacia based on white matter (WM) abnormalities on ultrasound were compared to those of 10 healthy control infants at 6, 12, 18, and 26 weeks of corrected age. Magnetic resonance imaging recordings were used to identify the location and severity of the brain lesions. Infants with extensive lesions in the periventricular and lobar WM with or without diffuse lesions in the subcortical WM showed a decreased variability on some spatial and temporal parameters of kicks. More importantly, these infants showed a different developmental course for intralimb couplings when compared to the other infants; they were unable to dissociate tight intralimb couplings at 18 and 26 weeks. As all of these infants had substantial damage of the corticospinal tracts, these findings suggest that these tracts are involved in the regulation of intralimb joint dissociations between 4 and 6 months of age. However, caution is needed as areas outside those in which the corticospinal tracts are located could be damaged as well and most of the infants with moderate to severe lesions in the corticospinal tract had additional psychomotor problems. For interlimb couplings and most of the spatial and temporal parameters of kicks, no differences were found between groups. This strengthens the claim that inter- and intralimb couplings are organized in fundamentally different ways.


Reproductive health and the industrialized food system: a point of intervention for health policy.
What food is produced, and how, can have a critical impact on human nutrition and the environment, which in turn are key drivers of healthy human reproduction and development. The US food production system yields a large volume of food that is relatively low in cost for consumers but is often high in calories and low in nutritional value. In this article we examine the evidence that intensive use of pesticides, chemical fertilizers, hormones, antibiotics, and fossil fuel in food production, as well as chemicals in food packaging, are potentially harmful to human reproductive and developmental health. We conclude that policies to advance a healthy food system are necessary to prevent adverse reproductive health effects and avoid associated health costs among current and future generations. These policies include changes to the Farm Bill and the Toxic Substances Control Act, and greater involvement by the health care sector in supporting and sourcing food from urban agriculture programs, farmers' markets, and local food outlets, as well as increasing understanding by clinicians of the links between reproductive health and industrialized food production.


Genome-Wide Analysis of Corynespora cassiicola Leaf Fall Disease Putative Effectors.
Corynespora cassiicola is an Ascomycetes fungus with a broad host range and diverse life styles. Mostly known as a necrotrophic plant pathogen, it has also been associated with rare cases of human infection. In the rubber tree, this fungus causes the Corynespora leaf fall (CLF) disease, which increasingly affects natural rubber production in Asia and Africa. It has also been found as an endophyte in South American rubber plantations where no CLF outbreak has yet occurred. The C. cassiicola species is genetically highly diverse, but no clear relationship has been evidenced between phylogenetic lineage and pathogenicity. Cassiicolin, a small glycosylated secreted protein effector, is thought to be involved in the necrotrophic interaction with the rubber tree but some virulent C. cassiicola isolates do not have a cassiicolin gene. This study set out to identify other putative effectors involved in CLF. The genome of a highly virulent C. cassiicola isolate from the rubber tree (CCP) was sequenced and assembled. In silico prediction revealed 2870 putative effectors, comprising CAZymes, lipases, peptidases, secreted proteins and enzymes associated with secondary metabolism. Comparison with the genomes of 44 other fungal species, focusing on effector content, revealed a striking proximity with phylogenetically unrelated species (Colletotrichum acutatum, Colletotrichum gloesporioides, Fusarium oxysporum, nectria hematococca, and Botrosphaeria dothidea) sharing life style plasticity and broad host range. Candidate effectors involved in the compatible interaction with the rubber tree were identified by transcriptomic analysis. Differentially expressed genes included 92 putative effectors, among which cassiicolin and two other secreted singleton proteins. Finally, the genomes of 35 C. cassiicola isolates representing the genetic diversity of the species were sequenced and assembled, and putative effectors identified. At the intraspecific level, effector-based classification was found to be highly consistent with the phylogenomic trees. Identification of lineage-specific effectors is a key step toward understanding C. cassiicola virulence and host specialization mechanisms.


Impact of CFTR DeltaF508 mutation on prostaglandin E2 production and type IIA phospholipase A2 expression by pulmonary epithelial cells.
Cystic fibrosis (CF) is characterized by an exacerbated inflammatory pulmonary response with excessive production of inflammatory mediators. We investigated here the impact of cystic fibrosis transmembrane conductance regulator (CFTR) dysfunction on prostaglandin E2 (PGE2) production and type IIA secreted phospholipase A2 (sPLA2-IIA) expression. We show that both resting and LPS-stimulated human respiratory epithelial cell line bearing DeltaF508 mutation on CFTR (CF cells) released more PGE2 than control cell line. This was accompanied by enhanced expression and activity of cyclooxygenase-2 in CF cells. PGE2 release was attenuated after experimentally induced retrafficking of the DeltaF508-CFTR at the plasma membrane. sPLA2-IIA expression occurred at higher levels in CF cells than in control cells and was enhanced by LPS and PGE2. Suppression of PGE2 synthesis by aspirin led to an inhibition of LPS-induced sPLA2-IIA expression. Higher activation of NF-kappaB was observed in CF cells compared with control cells and was enhanced by LPS. However, addition of PGE2 or aspirin had no effect on NF-kappaB activation. LPS-induced sPLA2-IIA expression was reduced by an NF-kappaB inhibitor. We suggest that the lack of the CFTR in the plasma membrane results in a PGE2 overproduction and an enhanced sPLA2-IIA expression. This expression is upregulated by NF-kappaB and amplified by PGE2 via a unidentified signaling pathway.


Extended transcervical thymectomy in the treatment of myasthenia gravis.
The ideal operative technique for thymectomy in myasthenia gravis remains controversial. Most surgeons perform thymectomy via median sternotomy, some supplementing this with an even more extensive mediastinal and cervical dissection designed to remove all areas of possible ectopic thymic tissue. We and others have advocated a transcervical approach that is less morbid and costly than sternotomy approaches. The transcervical approach allows a complete extracapsular thymic resection, but it does not address all areas of potential ectopic thymic tissue. We have published our experience with 151 extended transcervical thymectomies (TCT). At mean follow-up of 53 months (complete follow-up in 97%), Kaplan-Meier estimates of complete stable remission were 33% and 35% at 3 and 6 years. If one includes patients who became asymptomatic but remained on low dose, single-drug immunosuppression as complete remissions (CRs), then the CR rates were 43% and 45% at 3 and 6 years. Longer term (mean 83 months) follow-up of the earliest 84 patients in the series showed preserved CR rates. On multivariate analysis, only preoperative Osserman Class (group mean 2.3) was significantly associated with improved CR rate. These results were obtained with a major operative complication rate of 0.7% and minor complication rate of 6.6%, and nearly every operation was performed without the need for overnight hospital admission. We believe that these response rates following TCT are sufficiently similar to those following transsternal techniques of thymectomy to allow us to recommend this less morbid and less costly operation as an eminently reasonable choice in the surgical treatment of myasthenia gravis.


Mapping the risk for transmission of urban schistosomiasis in the Brazilian Northeast.
This is an analysis of the risk of schistosomiasis transmission in the city of Recife in the Northeast of Brazil based on the number of schistosomiasis cases (Schistosoma mansoni) registered for the period 2007-2017 together with data resulting from active search of breeding sites of the Biomphalaria snail intermediate host. The analyses were performed using Kernel Density Estimation (KDE), SaTScan and Map Algebra methodology using human socio-demographic data and biotic and abiotic data from the snail breeding sites. Investigating 44 breeding sites resulted in a total of 3.800 snails, 31.8% of which were positive for S. mansoni DNA. These data were considered in relation to total of 652 schistosomiasis cases. The KDE showed two high-risk and two medium-risk clusters, while three significant clusters were identified by SaTScan. Combining these data with the Map Algebra methodology showed that all high-risk neighbourhoods had breeding sites with snails positive for S. mansoni. It was concluded that schistosomiasis transmission cannot be controlled without basic sanitation and sewage management in the presence of Biomphalaria snails. The technique of Map Algebra was found to be fundamental for the analysis and demonstration of areas with a high probability of schistosomiasis transmission.


Signal requirements for the generation of CD4+ and CD8+ T-cell responses to human allogeneic microvascular endothelium.
Microvascular endothelium has been implicated as a major target in the rejection of vascularized allografts. In an attempt to dissect the stepwise generation of the T-cell-mediated immune response to microvascular endothelial cells (ECs), we analyzed the requirements for the two major T-cell subsets, CD4+ and CD8+, in the triggering of proliferative and cytotoxic responses to allogeneic ECs in vitro. Results demonstrate that resting ECs are unable to stimulate a functional response by purified T, CD4+, and CD8+ cells in the absence of costimulatory signals. T cells and CD8+ cells develop both proliferative and cytotoxic anti-EC responses by the addition of as little as 2 units/ml interleukin-2, 10 units/ml interleukin-1, or irradiated monocytes autologous to the responder lymphocytes, whereas only autologous monocytes are capable of triggering CD4+ T-cell precursors to proliferate and become anti-EC-specific effector cytotoxic T lymphocytes. CD8+ cell-mediated anti-EC cytotoxicity is directed toward allogeneic major histocompatibility complex (MHC) class I determinants on ECs and involves recognition by the CD3/T-cell receptor complex and the CD8 molecule on the effector T cells. CD4+ cells can be driven to proliferate, produce interleukin-2, and become anti-EC-specific cytotoxic T lymphocytes despite the lack of detectable membrane MHC class II determinants on the target cells. Chloroquine inhibition experiments demonstrate that autologous monocytes/macrophages are required by CD4+ T-cell precursors for the processing of EC-derived alloantigens and their subsequent presentation in the context of self-MHC molecules. These results are in agreement with the adoptive transfer experiments in experimental allograft models and suggest that the coordinate engagement of cells of the CD4, CD8, and monocyte/macrophage series is more effective than the individual cell subsets in the


[Future research needs and strategies for orofacial pain and oral motor disorders].
Musculoskeletal disorders in orofacial region are known to induce mastication disability and decrease the patients' quality of life levels. Additionally, the prevalence of those disorders tends to increase in elderly. However, since the existing therapies are not fully effective and medical expenses for those disorders are expanding, a prompt solution is necessary especially in aging societies like Japan. In addition, investigations for those disorders are important, because the research findings would be useful for the investigation of similar disease in other organs. Therefore, it is possible to consider that they can directly associate with general health and the universality of prosthodontics. In order to solve the above problems, two different research strategies are desirable. First, to improve the current curative effect, the effectiveness of existing diagnostic system and treatment modalities should be evaluated using high quality clinical studies. Second, as the research strategies on a long-term basis, investigations, which target the pathology and the contributing factors in a biological point of view are essential. This article will discuss about the research strategy to elucidate the pathology and the factors, which play important roles in the disease mechanisms, based on the previous and ongoing physiological research findings. Furthermore, I would like to review the useful newly developed technologies for the research strategy and new research fields, to enhance the value of prosthodontics and discuss about our future research direction.


Lactobacillus casei rhamnosus Lcr35 in children with chronic constipation.
The purpose of the present paper was to evaluate the efficacy of probiotics (Lactobacillus casei rhamnosus, Lcr35) for treating children with chronic constipation and to compare its effect with magnesium oxide (MgO) and placebo. This double-blind placebo-controlled, randomized study enrolled 45 children under 10 years old with chronic constipation. They were randomly assigned to receive Lcr35 (8 x 10(8) c.f.u./day; n = 18), MgO (50 mg/kg/day; n = 18), or placebo (n = 9) orally twice daily for 4 weeks. Lactulose use (1 mL/kg per day) was allowed when no stool passage for 3 days was noted. Glycerin enema was used only when no defecation was noted for >5 days or abdominal pain was suffered due to stool impaction. Bacterial cultures of stool were performed before and after treatment to evaluate the change of intestinal flora. Comparisons of the frequency of defecation, consistency of stool and the use of lactulose or enema during the period of treatment were made among the three groups. The patients who received MgO or probiotics had a higher defecation frequency (P = 0.03), higher percentage of treatment success (P = 0.01), less use of glycerin enema (P = 0.04) and less hard stool (P = 0.01) than the placebo group. There was no significant difference between MgO and probiotic groups in the aforementioned comparisons. The first effect of MgO (second week) on constipation was slightly earlier than that of probiotic (second to third week). Abdominal pain occurred less frequently in the probiotic group than in both the MgO and the placebo groups (P = 0.03). There was no statistically significant difference among the three groups in the use of lactulose, episodes of fecal soiling, and change of appetite. No adverse effect was noted in probiotic and placebo groups. Only one patient in the MgO group suffered from mild diarrhea. Lcr35 was effective in treating children with chronic constipation. There is no statistically significant difference in efficacy between MgO and Lcr35, but less abdominal pain occurred when using Lcr35. Study with larger case number and longer follow up is needed in the future.


The organization of histone H3 modifications as revealed by a panel of specific monoclonal antibodies.
Histone modifications play critical roles in the epigenetic regulation of gene expression and in the maintenance of genome integrity. Acetylation and methylation of histone H3 are particularly important in gene activation and silencing. We generated and characterized a panel of mouse monoclonal antibodies that specifically recognize different modifications on K4, K9, and K27 residues on histone H3. By using these antibodies for chromatin immunoprecipitation and immunoblotting, we analyzed the relationship between different modifications in nearby nucleosomes in human cells. Within a few nucleosome neighbors, trimethyl-K4 was associated with acetyl-K27, rather than with dimethyl-K4 and acetyl-K9, consistent with their co-localization on active promoters. Furthermore, simultaneous immunofluorescence using directly-labeled antibodies revealed that di- and tri-methylation on K4 was diminished during replicative senescence. These highly-reliable and fully-characterized monoclonal antibodies may facilitate future epigenomic studies on healthy and diseased cells.