Epidemiology of West Nile Virus in the Eastern Mediterranean region: A systematic review.
West Nile Virus (WNV), a member of the genus Flavivirus, is one of the most widely distributed arboviruses in the world. Despite some evidence for circulation of WNV in countries summarized by the World Health Organization as the Eastern Mediterrian Regional Office (EMRO), comprehensive knowledge about its epidemiology remains largely unknown. This study aims to provide a concise review of the published literature on WNV infections in the Eastern Mediterranean Regional Office of WHO (EMRO). A systematic review of WNV prevalence studies on humans, animals and vectors in the EMRO region was performed by searching: Web of Science, Science Direct, Scopus, PubMed, Embase and Google Scholar. Finally, 77 citations were included, comprising 35 seroprevalence studies on general population (24460 individuals), 15 prevalence studies among patients (3439 individuals), 22 seroprevalence studies among animals (10309 animals), and 9 studies on vectors (184242 vector species). Of the 22 countries in this region, five had no data on WNV infection among different populations. These countries include Kuwait, Bahrain, Oman, Syria and Somalia. On the other hand, among countries with available data, WNV-specific antibodies were detected in the general population of all investigated countries including Djibouti (0.3-60%), Egypt (1-61%), Iran (0-30%), Iraq (11.6-15.1%), Jordan (8%), Lebanon (0.5-1%), Libya (2.3%), Morocco (0-18.8%), Pakistan (0.6-65.0%), Sudan (2.2-47%), and Tunisia (4.3-31.1%). WNV RNA were also detected in patient populations of Iran (1.2%), Pakistan (33.3%), and Tunisia (5.3% -15.9%). WNV-specific antibodies were also detected in a wide range of animal species. The highest seropositivity rate was observed among equids (100% in Morocco) and dogs (96% in Morocco). The highest seroprevalence among birds was seen in Tunisia (23%). In addition, WNV infection was detected in mosquitoes (Culex, and Aedes) and ticks (Argas reflexus hermanni). The primary vector of WNV (Culex pipiens s.l.) was detected in Djibouti, Egypt, Iran and Tunisia, and in mosquitoes of all these countries, WNV was demonstrated. This first systematic regional assessment of WNV prevalence provides evidence to support the circulation of WNV in the EMRO region as nearly all studies showed evidence of WNV infection in human as well as animal/vector populations. These findings highlight the need for continued prevention and control strategies and the collection of epidemiologic data for WNV epidemic status, especially in countries that lack reliable surveillance systems.


Biomedical database inter-connectivity: an experiment linking MIM, GENBANK, and META-1 via MEDLINE.
The linkage of disparate biomedical databases is an important goal of the Unified Medical Language (UMLS) Project. We conducted an experiment to investigate the feasibility of using UMLS resources to link databases in clinical genetics and molecular biology. References from MIM ("Mendelian Inheritance in Man") were lexically mapped to the equivalent citations in MEDLINE. The MeSH major subject headings by which the citations in a particular MIM entry had been indexed were used to develop a "genetic-disorder-centered view of the world" in Meta-1 (the first official version of the UMLS Metathesaurus). Our hypothesis was that these MeSH subject headings could provide access to a "semantic neighborhood" in Meta-1 that would be relevant to a particular genetic disorder. By browsing in this "semantic neighborhood," a user could select various combinations of terms with which to search MEDLINE through an interface between Meta-1 and Grateful Med. Such searches might retrieve citations that were more recent than those in MIM or that provided useful supplementary information. Since some MEDLINE records contain pointers to entries in GENBANK, information about genetic sequences related to a particular clinical genetic disorder could also be retrieved. This scenario was implemented for a small number of MIM entries, providing a concrete demonstration that linking disparate electronic databases in an important subdomain of biomedicine is relatively straightforward.


Amplitude variability over trials in hemodynamic responses in adolescents with ADHD: The role of the anterior default mode network and the non-specific role of the striatum.
It has been suggested that intra-individual variability (IIV) in performance on attention and other cognitive tasks might be a cognitive endophenotype in individuals with ADHD. Despite robust IIV findings in behavioral data, only sparse data exist on how what type of brain dysfunction underlies variable response times. In this study, we asked whether ADHD IIV in reaction time on a commonly-used test of attention might be related to variation in hemodynamic responses (HRs) observed trial-to-trial. Based on previous studies linking IIV to regions within the "default mode" network (DMN), we predicted that adolescents with ADHD would have higher HR variability in the DMN compared with controls, and this in turn would be related to behavioral IIV. We also explored the influence of social anxiety on HR variability in ADHD as means to test whether higher arousal associated with high trait anxiety would affect the neural abnormalities. We assessed single-trial variability of HRs, estimated from fMRI event-related responses elicited during an auditory oddball paradigm in adolescents with ADHD and healthy controls (11-18 years old; N = 46). Adolescents with ADHD had higher HR variability compared with controls in anterior regions of the DMN. This effect was specific to ADHD and not associated with traits of age, IQ and anxiety. However, an ADHD effect of higher HR variability also appeared in a basal ganglia network, but for these brain regions the relationships of HR variability and social anxiety levels were more complex. Performance IIV correlated significantly with variability of HRs in both networks. These results suggest that assessment of trial-to-trial HR variability in ADHD provides information beyond that detectable through analysis of behavioral data and average brain activation levels, revealing specific neural correlates of a possible ADHD IIV endophenotype.


Beta-adrenergic blockade after stroke. A preliminary closed cohort study.
To collect background data for a prospective clinical trial of beta-blocking agents in the prevention of deaths after stroke, the long-term prognosis in 60 patients discharged from a stroke unit on beta-blocker therapy was compared with the outcome in 60 matched patients with stroke but without beta-blockers. Matching included sex, age, type of stroke, and presence or absence of hypertension and cardiac failure. Thirteen patients (22%) in the beta-blocker group died during a median followup of 41 months. Of the 60 patients not on beta-blockade at discharge, 21 (35%) died during a median followup of 36 months. By life-table technique and log-rank test, the relative risk for death was 0.60:1.00 (p = 0.14). During followup, 12 recurrent strokes were observed in patients on beta-blockers and 19 in patients without beta-blockers (relative risk 0.57:1.00; p = 0.12). It appeared that the reduction in mortality could only marginally be ascribed to fewer deaths from myocardial infarction; other causes of death were also less frequent in beta-blocker-treated patients. The results emphasize that supplementary information on the effect of beta-blocking agents on mortality after stroke is needed before a larger trial of beta-blocker therapy in patients with manifest cerebrovascular disease can be initiated.


Persistence of the efficacy of doramectin against Ostertagia ostertagi and Cooperia oncophora in cattle.
The persistence of the efficacy of doramectin injectable against Ostertagia ostertagi and Cooperia oncophora was evaluated in two studies in calves. In both, the calves were allocated to six groups of six. Calves in the first control group (C1) and first treated group (T1) received a daily infection of 200 L3 of O ostertagi and 200 L3 of C oncophora; the calves in groups C2 and T2 received a daily infection of 1000 L3 of each species, and groups C3 and T3 received 10,000 L3 of each species per day. The calves in the three treated groups each received 0.2 mg/kg doramectin injectable on day 0. In the first study, the calves were infected for 21 days with Cooperia and for 28 days with Ostertagia, and they were slaughtered on day 33. In the second study, the calves were infected for 21 days with both species, the infections with Cooperia and Ostertagia starting from eight and 15 days, respectively, after the treatment, and the animals were slaughtered on day 40. The calculation of the persistence of the activity of doramectin was based on its efficacy against the different developmental and adult stages of the two parasites. The data from both studies indicated that the efficacy of doramectin against Ostertagia persisted for at least five weeks, but no conclusions could be drawn about the effect of the size of the infective doses on the persistence of the activity. In contrast, the Cooperia worm counts from the second study suggested that the efficacy of doramectin against Cooperia persisted for at least four weeks when the calves were exposed to a low or moderate infection level, whereas at the highest infection level it persisted for between three and four weeks.


Removal of the blue component of light significantly decreases retinal damage after high intensity exposure.
Light causes damage to the retina (phototoxicity) and decreases photoreceptor responses to light. The most harmful component of visible light is the blue wavelength (400-500 nm). Different filters have been tested, but so far all of them allow passing a lot of this wavelength (70%). The aim of this work has been to prove that a filter that removes 94% of the blue component may protect the function and morphology of the retina significantly. Three experimental groups were designed. The first group was unexposed to light, the second one was exposed and the third one was exposed and protected by a blue-blocking filter. Light damage was induced in young albino mice (p30) by exposing them to white light of high intensity (5,000 lux) continuously for 7 days. Short wavelength light filters were used for light protection. The blue component was removed (94%) from the light source by our filter. Electroretinographical recordings were performed before and after light damage. Changes in retinal structure were studied using immunohistochemistry, and TUNEL labeling. Also, cells in the outer nuclear layer were counted and compared among the three different groups. Functional visual responses were significantly more conserved in protected animals (with the blue-blocking filter) than in unprotected animals. Also, retinal structure was better kept and photoreceptor survival was greater in protected animals, these differences were significant in central areas of the retina. Still, functional and morphological responses were significantly lower in protected than in unexposed groups. In conclusion, this blue-blocking filter decreases significantly photoreceptor damage after exposure to high intensity light. Actually, our eyes are exposed for a very long time to high levels of blue light (screens, artificial light LED, neons…). The potential damage caused by blue light can be palliated.


Tobacco and cancer in Turkey.
Although the smoking epidemic is decreasing steadily in other parts of the world, it continues to spread at an accelerated rate in underdeveloped and developing countries. Turkey, among other developing countries, faces the increasing threat of tobacco-related cancers, particularly lung cancer, which is the leading cause of cancer death in both sexes. We investigated the relationship between cigarette consumption and the relative mortality rates due to lung cancer in men and women between 1965 and 1992. We found a parallelism between the increasing total and per capita cigarette consumption and the rising relative mortality from lung cancer in both sexes. Total per capita cigarette consumption rose from 1230 cigarettes per year in 1985 to 1495 in 1991, and the per capita yearly cigarette consumption over the age of 15 increased from 1850 in 1965 to 2600 in 1992. During the same period, the relative mortality from lung cancer increased from 25 to 40% in men and from 11 to 16% in women. The tar, nicotine, and carbon monoxide determinations of locally produced and imported cigarettes suggested that the high tar and carbon monoxide content of most locally produced cigarettes smoked over many years could also be a contributory factor to the increased mortality rates due to lung cancer. Only two brands of locally produced cigarettes contained lower than 12 mg of tar per cigarette as allowed in European community states, whereas half of the imported brands of cigarettes met this standard. Four of the six imported brands of cigarettes contained higher tar and carbon monoxide compared with the same brands sold in England. These findings indicate that urgent measures are necessary not only to ban all activities promoting the sale of cigarettes but also to establish standards for both national and foreign brands of cigarettes while making a greater effort to reduce active and passive smoking in the Turkish population.


MSnbase, Efficient and Elegant R-Based Processing and Visualization of Raw Mass Spectrometry Data.
We present version 2 of the MSnbase R/Bioconductor package. MSnbase provides infrastructure for the manipulation, processing, and visualization of mass spectrometry data. We focus on the new on-disk infrastructure, that allows the handling of large raw mass spectrometry experiments on commodity hardware and illustrate how the package is used for elegant data processing, method development, and visualization.


Mesangial immune injury, hypertension, and progressive glomerular damage in Dahl rats.
Hypertension frequently accompanies chronic glomerulonephritis. Mesangial injury and glomerulosclerosis are common in glomerulonephritis and are often harbingers of progressive glomerular destruction. Thus, in a model of mesangial immune injury we studied the relationship between hypertension, mesangial injury, and glomerulosclerosis. We induced mesangial ferritin-antiferritin immune complex disease (FIC) in Dahl salt-sensitive (S) and salt-resistant (R) rats. S and R rats with FIC were fed chow containing 0.3% NaCl until 14 weeks of age and then switched to 8.0% NaCl chow until 28 weeks of age. Groups of control S and R rats (no FIC) were either fed 0.3% NaCl for 28 weeks or switched to 8.0% NaCl chow at 14 weeks of age. Blood pressure, serum creatinine, urinary protein, and glomerular injury (assessed by semiquantitative morphometric analysis) were determined at 14 and 28 weeks of age. R rats with or without FIC did not develop hypertension; mesangial injury was minimal. At 14 weeks of age, only S FIC rats developed hypertension, proteinuria, significant mesangial expansion and early glomerulosclerosis. At 28 weeks of age, proteinuria, mesangial expansion, and glomerulosclerosis were significantly more severe in hypertensive S rats with FIC than in those without FIC. These studies show that despite a normal salt intake, mesangial injury hastened the onset of hypertension, but only in rats genetically predisposed to hypertension (S FIC at 14 weeks). High dietary salt further aggravated hypertension, which, in turn, magnified both mesangial injury and glomerulosclerosis. Clinically, the different rates of progression of human glomerulonephritis associated with hypertension may be in part dependent on similar mechanisms.


Modeling facial movement: I. A dynamic analysis of differences based on skeletal characteristics.
To introduce a novel approach to analyze and model facial movements; and to quantify variations in facial movement caused by the extent of skeletal differences between the maxilla and mandible and the middle to lower facial heights. The hypothesis was that there are differences in facial movement related to the underlying facial skeleton which may be explained by the shape of the face rather than the pattern of movement. The study sample consisted of 43 subjects (23 men, 20 women) with a mean age of 18.5 years (SD = 11.90). Measures of the facial skeletal differences were made from lateral cephalometric radiographs, and subjects were classified as Class I and Class II, and normal to decreased lower anterior face height, respectively. Facial movements were recorded by a video-based tracking system. Descriptive and inferential statistics were performed on principal component scores generated from the movement data. A linear mixed-effects model was used to test for significant differences in movement among the different skeletal types. A dynamic modeling of facial movements was described that has numerous potential clinical applications. Also, differences in movement were found during the lip purse movement. Specifically, skeletal Class I individuals showed greater forward and upward movement during lip purse compared with individuals with severe skeletal Class II who moved their lips straight forward with less magnitude of movement. For most of the movements, apart from the lip purse, differences in motion were explained by static facial shape.


Using Emotional Intelligence as a Framework for Students' Professional Identity Formation.
Emotional intelligence and professional identity formation (PIF) intersect at various levels. Professional identity formation requires acute observation of others in the profession and the ability to decipher intentionality in behaviors. The developing pharmacist must make a deliberate effort to emulate positive norms and values that coincide with those associated with the profession while deliberately ignoring those that are incongruent. To learn from others in the profession, social skills are required, so one can ask questions, determine the best course of action, set goals, grow, and maintain relationships, and ask for help. The ability to manage one's emotions regardless of external circumstances can be valuable for any profession. Self-regulation and self-assessment of one's emotions and motivations can be useful for reevaluating our perspectives and priorities as pharmacists. Emotional intelligence is a critical component of building, demonstrating, and improving PIF. This commentary will provide strategies to facilitate and solidify the connection between the two.


Analysis of CC chemokine receptor 5 and 2 polymorphisms and renal transplant survival.
Chronic rejection is an immune process leading to graft failure. By regulating the trafficking of leukocytes, chemokines and chemokine receptors are thought to be one of the reasons causing acute renal rejection (ARE), which increases the possibility of chronic rejection and organ destruction. This study was designed to investigate, in the Turkish population, an association of chemokine receptor genetic variants, CCR2V641, CCR5-59029-A/G, CCR5-Delta32 and acute renal rejection after renal transplant surgery. We carried out our study in 85 Turkish renal transplant patients (45 men, 40 women; mean age 39 +/- 2 years) by polymerase chain reaction (PCR) and restriction fragment length polymorphism (RFLP) techniques. We found no significant difference in the incidence of rejection among patients possessing or lacking CCR5-Delta32. For the groups with and without acute renal rejection, we found a significant difference between the groups in A and G allele distribution in both CCR2V641and CCR559029 gene variants (p = 0.003 and p = 0.003, respectively). According to our findings, the risk of acute rejection in renal transplantation may be associated with genetic variation in the chemokine receptor genes CCR5-59029 and CCR2V641 in Turkey, and studies on these gene polymorphisms could be an ideal target for future interventions intended to prevent renal transplant loss.


Bryodin, a ribosome-inactivating protein from the roots of Bryonia dioica L. (white bryony).
Bryodin is a strongly basic (pI greater than or equal to 9.5) glycoprotein (neutral sugar content 6.3%) with Mr 30,000, purified from the roots of Bryonia dioica (white bryony). This protein inhibits protein synthesis by a rabbit reticulocyte lysate with and ID50 (concentration causing 50% inhibition) of 0.12 nM (3.6 ng/ml) and has much less effect on protein synthesis by whole cells, with ID50 values ranging from 46 nM to 2.27 microM (1.4-67 micrograms/ml). Bryodin acts by inactivating ribosomes, with a less-than-equimolar ratio, which suggests a catalytic action. Bryodin decreases the number of local lesions induced by tobacco mosaic virus in the leaves of Nicotiana glutinosa. From all its properties, bryodin can be considered to be a ribosome-inactivating protein, similar to those already known [reviews: Barbieri & Stirpe (1982) Cancer Surveys 1, 489-520; Stirpe & Barbieri (1986) FEBS Lett. 195, 1-8].


A Lipid-Inspired Highly Adhesive Interface for Durable Superhydrophobicity in Wet Environments and Stable Jumping Droplet Condensation.
Creating thin (<100 nm) hydrophobic coatings that are durable in wet conditions remains challenging. Although the dropwise condensation of steam on thin hydrophobic coatings can enhance condensation heat transfer by 1000%, these coatings easily delaminate. Designing interfaces with high adhesion while maintaining a nanoscale coating thickness is key to overcoming this challenge. In nature, cell membranes face this same challenge where nanometer-thick lipid bilayers achieve high adhesion in wet environments to maintain integrity. Nature ensures this adhesion by forming a lipid interface having two nonpolar surfaces, demonstrating high physicochemical resistance to biofluids attempting to open the interface. Here, developing an artificial lipid-like interface that utilizes fluorine-carbon molecular chains can achieve durable nanometric hydrophobic coatings. The application of our approach to create a superhydrophobic material shows high stability during jumping-droplet-enhanced condensation as quantified from a continual one-year steam condensation experiment. The jumping-droplet condensation enhanced condensation heat transfer coefficient up to 400% on tube samples when compared to filmwise condensation on bare copper. Our bioinspired materials design principle can be followed to develop many durable hydrophobic surfaces using alternate substrate-coating pairs, providing stable hydrophobicity or superhydrophobicity to a plethora of applications.


The case for the rope-pump in Africa: a comparative performance analysis.
The conventional handpump is the most popular technology choice for improved potable water supplies in rural sub-Saharan Africa. To date, however, it has failed to deliver satisfactory levels of sustainability, largely due to inadequate maintenance capacity. An alternative option to standardised imported handpumps is the locally manufactured rope-pump, which is considerably cheaper and easier to maintain but has been rejected in the past due to fears of impaired water quality. This paper presents the key aspects of a study in northern Ghana which compared the performance of rope-pumps with that of conventional handpumps, to determine whether or not the rope-pump provides a viable alternative for community water supplies across the subcontinent. User interviews, sanitary surveys, water quality analyses and technical performance measurements were used to develop a comparative performance analysis for the two pump types. The findings of the study indicated that the rope-pump out-performed the conventional handpump on the majority of counts and that, contrary to widespread perceptions, there was no significant difference between pump types with respect to the impact on microbiological water quality. Consequently, the rope-pump provides a significant technological opportunity to improve water supply sustainability in Africa.


Monoclonal antibody (mAb)-induced down-regulation of RON receptor tyrosine kinase diminishes tumorigenic activities of colon cancer cells.
Overexpression of the RON receptor tyrosine kinase contributes to pathogenesis of epithelial cancers and disruption of RON signals has potential for therapeutic intervention. Here, we report the inhibitory effects of monoclonal antibodies (Zt/g4, Zt/f2 and Zt/c9) on RON expression and tumorigenic activities in colon cancer cells. Persistent treatment of colon SW620 or other cells with Zt/g4 dramatically down-regulated RON expression as evident by Western blot and cell surface fluorescent analyses. The effect was both concentration and time-dependent and specific to RON but not to structure-related MET or -unrelated EGFR. The cause of reduction was antibody-induced receptor internalization followed by protein degradation through lysosome and proteasome-mediated pathways. Down-regulation of RON impaired intracellular signaling events. Phosphorylation of Erk1/2 and AKT was dramatically reduced after Zt/g4 treatment. Zt/g4 treatment also affects activities of DVL and GSK-3beta, which results in diminished beta-catenin nuclear translocation. Functional studies revealed that Zt/g4 treatment changes cellular morphology and affects colony formation in soft agar. It also increased the sensitivity of SW620 cells in response to gemcitabine-induced cytotoxicity. In this case, the death of SW620 cells was significantly increased when Zt/g4 was used in combination with gemcitabine. We conclude that persistent treatment of cancer cells with antibodies specific to RON extracellular domains results in down-regulation of RON expression. The reduced RON expression is accompanied with impaired signaling events, diminished tumorigenic activities and enhanced sensitivity towards cytotoxic drugs. Thus, Zt/g4-directed targeting could have therapeutic implication for controlling tumorigenic phenotypes of cancer cells.


Improvement in functional recovery with administration of Cerebrolysin after experimental closed head injury.
Cerebrolysin is a unique peptide preparation that mimics the action of neurotrophic factors. This study was designed to investigate the effects of acute treatment of experimental closed head injury (CHI) in rats with Cerebrolysin on neurological function. Adult male Wistar rats (n = 60) were subjected to impact acceleration-induced CHI. Closed head injured rats received intraperitoneal injection of saline (n = 30) or Cerebrolysin (2.5 ml/kg, n = 30) starting 1 hour postinjury and administered once daily until they were killed (2 or 14 days after CHI). To evaluate functional outcome, the modified neurological severity score (mNSS), foot fault, adhesive removal, and Morris water maze (MWM) tests were performed. Animals were killed on Day 14 (n = 20) after injury, and their brains were removed and processed for measurement of neuronal cells, axonal damage, apoptosis, and neuroblasts. The remaining rats (n = 40) were killed 2 days postinjury to evaluate cerebral microvascular patency by fluorescein isothiocyanate (FITC)-dextran perfusion (n = 16) and to measure the expression of vascular endothelial growth factor (VEGF) and matrix metalloproteinase-9 (MMP-9) by using real-time reverse transcriptase-polymerase chain reaction (RT-PCR, n = 8) and by immunohistochemical analysis (n = 16). At 14 days post-CHI, the Cerebrolysin treatment group exhibited significant improvements in functional outcomes (the adhesive removal, mNSS, foot-fault, and MWM tests), and significantly more neurons and neuroblasts were present in the dentate gyrus (DG) (p < 0.05) compared with the saline-treated group (p < 0.05). At 2 days post-CHI, the Cerebrolysin group exhibited a significantly higher percentage of phosphorylated neurofilament H (pNF-H)-positive staining area in the striatum (p < 0.05), a significant increase in the percentage of FITC-dextran perfused vessels in the brain cortex (p < 0.05), a significant increase in the number of VEGF-positive cells (p < 0.05), and a significant reduction in the MMP-9 staining area (p < 0.05) compared with the saline-treated group. There was no significant difference in mRNA levels of MMP-9 and VEGF in the hippocampus and cortex 48 hours postinjury between Cerebrolysin- and saline-treated rats that sustained CHI. Acute Cerebrolysin treatment improves functional recovery in rats after CHI. Cerebrolysin is neuroprotective for CHI (increased neurons in the dentate gyrus and the CA3 regions of the hippocampus and increased neuroblasts in the dentate gyrus) and may preserve axonal integrity in the striatum (significantly increased percentage of pNF-H-positive tissue in the striatum). Reduction of MMP-9 and elevation of VEGF likely contribute to enhancement of vascular patency and integrity as well as neuronal survival induced by Cerebrolysin. These promising results suggest that Cerebrolysin may be a useful treatment in improving the recovery of patients with CHI.


Importance of atrial surface area and refractory period in sustaining atrial fibrillation: testing the critical mass hypothesis.
The critical mass hypothesis for atrial fibrillation (AF) was proposed in 1914; however, there have been few studies defining the relationship between atrial surface area and AF. This study evaluated the effect of tissue area and effective refractory period (ERP) on the probability of sustaining AF in an in vivo model. Domestic pigs (n = 9) underwent median sternotomy. Epicardial activation maps were constructed from bipolar electrograms recorded from form-fitting electrode templates placed on the atria. Baseline ERPs were determined. ERP was lowered with a continuous infusion of acetylcholine (0.005-0.04 mg/Kg/min) until AF could be sustained after burst pacing. The atria were sequentially partitioned using bipolar radiofrequency ablation. ERPs were lowered using acetylcholine until AF could be sustained in each subdivision of atrial tissue. Each subdivision was further divided until AF was no longer inducible. At study completion, the heart was excised and the surface area of each section was measured. Over a range of ERPs from 75 to 250 ms, the probability of AF was correlated with increasing tissue area (range, 19.5-105 cm(2)) and decreasing ERP. Logistic regression analysis identified shorter ERP (P < .001) and larger area (P = .006) as factors predictive of an increased probability of sustained AF (area under the curve of the receiver-operator characteristic = 0.878). The probability of sustained AF was significantly associated with increasing tissue area and decreasing ERP. These data may lead to a greater understanding of the mechanism of AF and help to design better interventional procedures.


International Workshops on Genotoxicity Testing (IWGT): Origins, achievements and ambitions.
Over the past thirty years, the International Workshops on Genotoxicity Testing (IWGT) became one of the leading groups in the field of regulatory genotoxicology, not only due to the diversity of participants with respect to geography and professional affiliation, but also due to the unique setup of recurring IWGT meetings every four years. The hallmarks of the IWGT process have been diligent initial planning approaches of the working groups, collection of data so as to stimulate data-driven discussions and debate, and striving to reach consensus recommendations. The scientific quality of the Working Groups (WGs) has been exceptional due to the selection of highly regarded experts on each topic. As a result, the IWGT working group reports have become important documents. The deliberations and publications have provided guidance on test systems and testing protocols that have influenced the development or revision of test guidelines of the Organisation for Economic Co-operation and Development (OECD), guidance by the International Council for Harmonisation (ICH), and strategic testing or data analysis approaches in general. This article summarizes the history of the IWGT, identifies some of its major achievements, and provides an outlook for the future.


Randomized, open-label, 5-way crossover study to evaluate the pharmacokinetic/pharmacodynamic interaction between furosemide and the non-steroidal anti-inflammatory drugs diclofenac and ibuprofen in healthy volunteers.
Nonsteroidal anti-inflammatory drugs (NSAIDs) can induce renal complications in patients taking loop diuretics. This study investigated the pharmacokinetic/pharmacodynamic effects and safety profile of orally administered diclofenac sodium, ibuprofen and diclofenac epolamine topical patch (DETP) on furosemide in healthy adult subjects. This open-label, randomized, 5-way crossover study was conducted in 40 subjects (aged 19 - 45 y). Diclofenac (75 mg taken orally twice daily), DETP (1.3% applied topically twice daily), or ibuprofen (800 mg taken orally thrice daily) was administered for 3 consecutive days, followed by co-administration with furosemide (given intravenously as 20 mg/2 min). Plasma furosemide and NSAID concentrations, urine furosemide, sodium and potassium concentrations and urine output were determined throughout the 24 h period following furosemide administration. Orally administered ibuprofen significantly increased furosemide AUC(0-t) (37%) and AUC(0-inf) (36%) and decreased total body CL (27%), R(max) (19%) and CLR (23%) geometric mean ratios compared with furosemide control. Oral and topical diclofenac had no pharmacokinetic effects on furosemide. Ibuprofen increased sodium excretion (Ae(0-24), 16%) and decreased sodium R(max) (15%), and oral diclofenac decreased urine output (Vu(0-24), 15%). DETP had no effect on furosemide pharmacodynamics; total systemic exposure to diclofenac during DETP treatment was < 1% that of oral diclofenac. Treatments were generally safe, with 25 subjects reporting a total of 112 adverse events. Pharmacodynamic effects were seen with oral diclofenac (urine output) and ibuprofen (urine sodium excretion). Furosemide also affected plasma and urine pharmacokinetic profiles. Pharmacologic effects of DETP on furosemide were not observed under these conditions. Additional research is warranted to delineate the potential interactions of other NSAIDs with furosemide and other loop diuretics.


Gastrodin inhibits high glucose‑induced human retinal endothelial cell apoptosis by regulating the SIRT1/TLR4/NF‑κBp65 signaling pathway.
Diabetic retinopathy (DR), one of the most common complications of late‑phase diabetes, is associated with the ectopic apoptosis of microvascular cells. Gastrodin, a phenolic glucoside derived from Gastrodia elata Blume, has been reported to have antioxidant and anti‑inflammation activities. The aim of the present study was to investigate the effects of gastrodin on high glucose (HG)‑induced human retinal endothelial cell (HREC) injury and its underlying mechanism. The results demonstrated that HG induced cell apoptosis in HRECs, which was accompanied by increased levels of reactive oxygen species production. Gastrodin treatment significantly alleviated HG‑induced apoptosis and oxidative stress. Furthermore, HG stimulation decreased the levels of SIRT1, which was accompanied by an increase in Toll‑like receptor 4 (TLR4) expression and the levels of phosphorylated nuclear factor (NF)‑κBp65. However, the administration of gastrodin significantly inhibited the activation of the sirtuin 1 (SIRT1)/TLR4/NF‑κBp65 signaling pathway in HRECs exposed to HG. Collectively, the present study demonstrated that gastrodin may be effective against HG‑induced apoptosis and its action may be exerted through the regulation of the SIRT1/TLR4/NF‑κBp65 signaling pathway.


Fabrication of selective interface of ZnO/CdS heterostructures for more efficient photocatalytic hydrogen evolution.
In previous studies, photocatalytic heterostructures between two components have usually been distributed randomly at the material's surface. It is significant and important to fabricate a selective heterostructure interface with more efficient charge separation and transfer. In this study, active pores were first investigated through competitive adsorption, degradation efficiency, and selective corrosion. It was found that selective adsorption-induced photosensitization along with active facets led to selective photocorrosion around the pores in ZnO nanosheets. Then, comparison between the properties of selectively and randomly distributed ZnO/CdS heterostructures is presented, namely, phase composition, morphology, pore size, absorbance, electronic band structure, photocurrent density, electrochemical impedance, and hydrogen evolution. Due to the Z-scheme, ZnO/CdS heterostructure selectively bound at the active pores, due to which more efficient charge separation and higher hydrogen evolution were achieved for ZnO/CdS-S. Thus, fabrication of selective heterostructure interface endows ZnO/CdS with more efficient hydrogen evolution.


Tropomyosin isoform switching in tumorigenic human fibroblasts.
We identified six tropomyosin (Tm) isoforms in diploid human fibroblasts. We used computerized microdensitometry of 2-dimensional protein profiles to measure the relative rates of synthesis and abundance of the individual Tm isoforms and actin, the two major structural constituents of microfilaments. In carcinogen-transformed human fibroblasts (HuT-14), the rates of synthesis of three Tm isoforms (Tm1, Tm2, and Tm6) were greatly decreased relative to normal diploid parental fibroblasts and to actin. In contrast, related nontumorigenic HuT fibroblasts which are "immortalized" and anchorage independent exhibited both slight down-regulation of Tm1 and Tm6 and 3.5-fold up-regulation of Tm3. Thus, Tm isoform switching from the predominance of the larger more avid Tm isoforms (Tm1, Tm2, Tm3, and Tm6) to the smaller, less avid Tm isoforms (Tm4 and Tm5) in microfilaments was a transformation-induced change correlated with tumorigenicity in human fibroblasts.


Cost-effectiveness of etanercept treatment in early active rheumatoid arthritis followed by dose adjustment.
To explore the cost-effectiveness of early biologic treatment, followed by dose-reduction in the case of remission, of active rheumatoid arthritis (RA), compared with standard treatment with methotrexate (MTX) in Sweden. Effectiveness (function, disease activity, erosions) in early RA for both alternatives was taken from a clinical trial comparing etanercept (ETA) combined with MTX to MTX alone. Patients discontinuing treatment can switch to another or their first biologic treatment. For patients in remission (Disease Activity Score [DAS28] < 2.6), ETA is reduced to half the dose. Return to full dose occurs when DAS28 reaches ≥ 3.2 again. Costs and utilities by level of functional capacity from an observational study are used. The model is analyzed as a micro-simulation and results are presented from the societal perspective for Sweden, for 10 years; costs (€2008) and effects are discounted at 3 percent. Sensitivity analysis was performed for the perspective, the time horizon, switching, and dose-reduction. The main analysis conservatively assumes 50 percent switching at discontinuation. The cost per quality-adjusted life-year (QALY) gained with early ETA/MTX treatment is €13,500 (societal perspective, incremental cost of €15,500 and incremental QALYs of 1.15). With 75 percent switching, the cost per QALY gained was €10,400. Over 20 years, the cost per QALY gained was €8,200. Results were further sensitive to the time patients remained on half dose and the perspective. CONCLUSIONS AND POLICY IMPLICATIONS: This study combines clinical trial and clinical practice data to explore cost-effective treatment scenarios in early RA, including the use of biologics. Our results indicate that a situation where a considerable proportion of patients achieve remission, dose-adjustments will increase the cost-effectiveness of treatment.


The Transcriptional Repressor ZNF503/Zeppo2 Promotes Mammary Epithelial Cell Proliferation and Enhances Cell Invasion.
The NET (nocA, Nlz, elB, TLP-1) subfamily of zinc finger proteins is an important mediator during developmental processes. The evolutionary conserved zinc finger protein ZNF503/Zeppo2 (zinc finger elbow-related proline domain protein 2, Zpo2) plays critical roles during embryogenesis. We found that Zpo2 is expressed in adult tissue and examined its function. We found that ZPO2 is a nuclearly targeted transcriptional repressor that is expressed in mammary epithelial cells. Elevated Zpo2 levels increase mammary epithelial cell proliferation. Zpo2 promotes cellular invasion through down-regulation of E-cadherin and regulates the invasive phenotype in a RAC1-dependent manner. We detect elevated Zpo2 expression during breast cancer progression in a MMTV-PyMT transgenic mouse model. Tumor transplant experiments indicated that overexpression of Zpo2 in MMTV-PyMT mammary tumor cell lines enhances lung metastasis. Our findings suggest that Zpo2 plays a significant role in mammary gland homeostasis and that deregulation of Zpo2 may promote breast cancer development.


[The no-care of patients with spinal cord injuries in primary care: bioethical challenges for health policies].
It was aimed to investigate what actions taken in Basic Health Units of the Family-(BHUF) to meet the demands of adults with spinal cord injury (SCI) post-rehabilitation, list facilities/difficulties to meet the demands of users. Descriptive study conducted in twenty BHUF. Twenty doctors and twenty nurses, randomly selected, participated in the study. To collect and analyze data, it was used semi-structured interviews and analysis of content. The categories were based on the integrality, universality, charity and justice principles. It was found: professional disqualification to assist people with SCI, fragmented care, difficult to schedule consultations and examinations, difficulties in the reference and counter reference system, and lack of ongoing actions to take care of individuals with SCI. The BHUF not ensure the protection of all citizens, there are challenges to the construction practices of health solidarity, warm and consequently more effective and decisive for individuals with SCI.


Measurement of serum bile acids concentrations for diagnosis of hepatobiliary disease in cats.
Serum bile acid concentrations were measured after food had been withheld for 12 hours (fasting serum bile acid [FSBA] concentration) and 2 hours after a meal (post-prandial serum bile acid [PSBA] concentration) using a direct enzymatic procedure in 108 cats clinically suspected of having hepatobiliary disease. In all cats, liver tissue was examined histologically to confirm the diagnosis. Twenty-six cats did not have histologic evidence of hepatobiliary disease and served as controls. The remaining 82 cats had hepatobiliary disease including hepatic lipidosis (n = 20), portosystemic vascular anomaly (n = 24), hepatic necrosis (n =13), hepatic neoplasia (n = 8), or cholestatic hepatic disease(n = 17). Sensitivity and specificity of measuring FSBA and PSBA concentrations were calculated for each test alone and when results were interpreted in combination (ie, in series and in parallel), and were compared with sensitivity and specificity of routinely used serum biochemical tests, including measuring serum activities of alanine aminotransferase, aspartate aminotransferase, alkaline phosphatase, and gamma-glutamyltransferase, and measuring serum concentrations of cholesterol, BUN, and total bilirubin. When tests were considered individually, determination of FSBA and PSBA concentrations had higher specificity than did the other tests (using a cutoff of 15 mumol/L for FSBA concentration and of 20 mumol/L for PSBA concentration). Determination of PSBA concentration had the highest sensitivity of all single tests in cats with hepatic lipidosis, portosystemic vascular anomaly, or cholestasis; determination of alanine aminotransferase activity or PSBA concentration had the highest sensitivity for cats with hepatic necrosis.(ABSTRACT TRUNCATED AT 250 WORDS)


Identification of arsenic resistant endophytic bacteria from Pteris vittata roots and characterization for arsenic remediation application.
Mitigation of arsenic (As) pollution is a topical environmental issue of high R&D priority. The present investigation was carried out to isolate As resistant endophytes from the roots of Indian ecotype Pteris vittata and characterize their As transformation and tolerance ability, plant growth promoting characteristics and their role to facilitate As uptake by the plant. A total of 8 root endophytes were isolated from plants grown in As amended soil (25 mg As kg(-1)). These isolates were studied for minimum inhibitory concentration (MIC), arsenite As(III) - arsenate As(V) transformation ability, plant growth promoting (PGP) characteristics through siderophore, indole acetic acid (IAA) production, phosphatase, ACC deaminase activity, and presence of arsenite oxidase (aox) and arsenite transporter (arsB) genes. On the basis of 16S rDNA sequence analysis, these isolates belong to Proteobacteria, Firmicutes and Bacteroidetes families under the genera Bacillus, Enterobacter, Stenotrophomonas and Rhizobium. All isolates were found As tolerant, of which one isolates showed highest tolerance up to 1000 mg L(-1) concentration in SLP medium. Five isolates were IAA positive with highest IAA production up to 60 mg/L and two isolates exhibited siderophore activity. Phosphatase activity was shown by only one isolate while ACC deaminase activity was absent in all the isolates. The As transformation study by silver nitrate test showed that only two strains had dual characteristics of As(III) oxidation and As (V) reduction, four strains exhibited either of the characteristics while other two didn't confirmed any of the two characteristics. Presence of aox gene was detected in two strains and arsB gene in six isolates. The strain with highest As tolerance also showed highest IAA production and occurrence of arsB gene. Present investigation may open up further scope of utilizing these endophytes for up gradation of phytoextraction process.


Repair of pretibial atonic wound with long cross-leg flap.
Soft-tissue defects of the pretibial region constitute a major reconstructive challenge because of the insufficiency of the local tissues. In such cases, a long cross-leg flap may be an alternative option. This study presents a case with an atonic wound in the pretibial region caused by a previous traffic accident, which had received cross-leg flap repair following the excision.


Efficacy of side airbags in reducing driver deaths in driver-side car and SUV collisions.
To estimate the efficacy of side airbags in preventing driver deaths in passenger vehicles struck on the driver side. Risk ratios for driver deaths per driver-side collision were computed for side airbag-equipped cars and SUVs, relative to vehicles without side airbags. Driver fatality ratios also were calculated for the same vehicles in front and rear impacts, and these were used to adjust the side crash risk ratios for differences in fatality risk unrelated to side airbags. Risk ratios were calculated separately for side airbags providing torso-only protection and side airbags with head protection; almost all head protecting airbags also had airbags protecting the torso. Car driver death risk in driver-side crashes was reduced by 37 percent for head protecting airbags and 26 percent for torso-only side airbags. Car driver death risk was reduced for older and younger drivers, males and females, and drivers of small and midsize cars, and when the striking vehicle was an SUV/pickup or a car/minivan. Death risk for drivers of SUVs was reduced by 52 percent with head protecting side airbags and by 30 percent with torso-only airbags. The effectiveness of side airbags could not be assessed for pickups and minivans due to the small number of these vehicles with airbags involved in crashes. Side airbags substantially reduce the risk of car and SUV driver death in driver-side collisions. Making side airbags with head protection available to drivers and right front passengers in all passenger vehicles could reduce the number of fatalities in motor vehicle crashes in the United States by about 2,000 each year.


Harnessing Induced Essentiality: Targeting Carbonic Anhydrase IX and Angiogenesis Reduces Lung Metastasis of Triple Negative Breast Cancer Xenografts.
Triple Negative Breast Cancer (TNBC) is aggressive, metastatic and drug-resistant, limiting the spectrum of effective therapeutic options for breast cancer patients. To date, anti-angiogenic agents have had limited success in the treatment of systemic breast cancer, possibly due to the exacerbation of tumor hypoxia and increased metastasis. Hypoxia drives increased expression of downstream effectors, including Carbonic Anhydrase IX (CAIX), a critical functional component of the pro-survival machinery required by hypoxic tumor cells. Here, we used the highly metastatic, CAIX-positive MDA-MB-231 LM2-4 orthotopic model of TNBC to investigate whether combinatorial targeting of CAIX and angiogenesis impacts tumor growth and metastasis in vivo to improve efficacy. The administration of a small molecule inhibitor of CAIX, SLC-0111, significantly reduced overall metastatic burden, whereas exposure to sunitinib increased hypoxia and CAIX expression in primary tumors, and failed to inhibit metastasis. The administration of SLC-0111 significantly decreased primary tumor vascular density and permeability, and reduced metastasis to the lung and liver. Furthermore, combining sunitinib and SLC-0111 significantly reduced both primary tumor growth and sunitinib-induced metastasis to the lung. Our findings suggest that targeting angiogenesis and hypoxia effectors in combination holds promise as a novel rational strategy for the effective treatment of patients with TNBC.


Effect of halothane on cardiovascular and plasma catecholamine responses to tracheal intubation.
The catecholamine and cardiovascular responses to intubation were investigated during halothane anaesthesia. Thirty patients were allocated randomly to two groups. Following induction of anaesthesia and muscle relaxation, group 1 was ventilated with 70% nitrous oxide in oxygen before intubation; group 2 received 1% halothane in addition. After intubation, both groups received 0.5% halothane. Arterial pressure and heart rate, and plasma noradrenaline and adrenaline concentrations were measured throughout the induction sequence. In group 1 intubation was associated with increases (from pre-induction values) in systolic arterial pressure of 13% and diastolic arterial pressure of 35%, although the plasma concentrations of noradrenaline did not alter significantly. In group 2, although there was a pressor response to intubation, no overall change in systolic arterial pressure and only a 13% increase in diastolic arterial pressure occurred when compared with pre-induction values. This response was associated with a 78% increase in the noradrenaline concentration; the adrenaline concentration did not alter significantly.


Effect of the Dutch school-based education programme 'Taste Lessons' on behavioural determinants of taste acceptance and healthy eating: a quasi-experimental study.
To assess the effect of the Dutch school-based education programme 'Taste Lessons' on children's behavioural determinants towards tasting unfamiliar foods and eating healthy and a variety of foods. In a quasi-experimental study design, data on behavioural determinants were collected at baseline, four weeks and six months after the intervention in both the intervention and control group. Children completed consecutively three questionnaires in which knowledge, awareness, skills, attitude, emotion, subjective norm and intention towards the two target behaviours were assessed. Teachers implemented on average a third of the programme activities. Multilevel regression analyses were conducted to compare individual changes in the determinants in the intervention group with those in the control group, corrected for children's gender and age. Effect sizes were expressed as Cohen's d. Dutch elementary schools. Forty-nine classes (1183 children, 9-12 years old) in grades 5-8 of twenty-one elementary schools. The intervention group showed a higher increase in knowledge (d=0·26, P<0·01), which persisted after six months (d=0·23, P<0·05). After four weeks, the intervention group showed a higher increase in number of foods known (d=0·22, P<0·05) and tasted (d=0·21, P<0·05), subjective norm of the teacher (d=0·17, P<0·05) and intention (d=0·16, P<0·05) towards the target behaviours. Partial implementation of Taste Lessons during one school year showed small short-term effects on increasing behavioural determinants in relation to tasting unfamiliar foods and eating healthy and a variety of foods. Full and repeated implementation of Taste Lessons in subsequent years might result in larger effects.


Validation of a New Methodology to Determine 3-Dimensional Endograft Apposition, Position, and Expansion in the Aortic Neck After Endovascular Aneurysm Repair.
To validate a novel methodology employing regular postoperative computed tomography angiography (CTA) scans to assess essential factors contributing to durable endovascular aneurysm repair (EVAR), including endograft deployment accuracy, neck adaptation to radial forces, and effective apposition of the fabric within the aortic neck. Semiautomatic calculation of the apposition surface between the endograft and the infrarenal aortic neck was validated in vitro by comparing the calculated surfaces over a cylindrical silicon model with known dimensions on CTA reconstructions with various slice thicknesses. Interobserver variabilities were assessed for calculating endograft position, apposition, and expansion in a retrospective series of 24 elective EVAR patients using the repeatability coefficient (RC) and the intraclass correlation coefficient (ICC). The variability of these calculations was compared with variability of neck length and diameter measurements on centerline reconstructions of the preoperative and first postoperative CTA scans. In vitro validation showed accurate calculation of apposition, with deviation of 2.8% from the true surface for scans with 1-mm slice thickness. Excellent agreement was achieved for calculation of the endograft dimensions (ICC 0.909 to 0.996). Variability was low for calculation of endograft diameter (RC 2.3 mm), fabric distances (RC 5.2 to 5.7 mm), and shortest apposition length (RC 4.1 mm), which was the same as variability of regular neck diameter (RC 0.9 to 1.1 mm) and length (RC 4.0 to 8.0 mm) measurements. This retrospective validation study showed that apposition surfaces between an endograft and the infrarenal neck can be calculated accurately and with low variability. Determination of the (ap)position of the endograft in the aortic neck and detection of subtle changes during follow-up are crucial to determining eventual failure after EVAR.


[Sexuality and schizophrenia].
The sex behaviour of patients suffering from schizophrenia has been largely overlooked. This study is aimed at describing the pattern of sexual responses and conducts in 113 inpatients with schizophrenia (DSM-III-R). A high rate of sexual dysfunction was found in both males (62.9%) and females (50%). These rates are higher than found in other previous studies. The possible cause factors of sexual dysfunctions in this group of patients and the methodological problems related to this type of study are reviewed.


Relationship between fluctuations of pH and pressure in the human stomach and duodenum.
The normal patterns of pressure activity in the antrum, pylorus and duodenum and associated pH changes were monitored in 9 healthy volunteers under fasted conditions and after ingestion of 300 ml of chocolate milk. The manometric assembly incorporated three side holes in the antrum, four side holes in the duodenum, a sleeve sensor positioned across the pylorus, and pH electrodes situated in the terminal antrum and proximal duodenum. The most common motor pattern recorded under fasting conditions consisted of regular co-ordinated contractions, most of which involved the antrum and duodenum, showed evidence of propagation and was associated with transient decreases in duodenal pH and transient increases in antral pH. Ingestion of milk changes the motor pattern to one that was composed of pressure waves which were confined to the pylorus with few or no pressure waves in the terminal antrum or proximal duodenum. Isolated pyloric pressure waves were gradually replaced by propagated antropyloroduodenal contractions which were associated with transient decreases in duodenal pH. Isolated pyloric pressure waves were not associated with episodes of duodenal acidification under fed conditions.


Mesenteric lymph nodes as alternative site for pancreatic islet transplantation in a diabetic rat model.
Islet transplantation has progressively become a safe alternative to pancreas transplantation for the treatment of type 1 diabetes. However, the long-term results of islet transplantation could be significantly increased by improving the quality of the islet isolation technique even exploring alternative islet transplantation sites to reduce the number of islets required to mitigate hyperglycemia. The goal of the study was to test the lymph node as a suitable anatomical location for islet engraftment in a rodent model. Forty Lewis rats, 6-8 weeks old, body weight 250-300 g, have been used as islet donors and recipients in syngeneic islet transplantation experiments. Ten rats were rendered diabetic by one injection of 65 mg/Kg of streptozotocin. After pancreas retrieval from non diabetic donors, islet were isolated and transplanted in the mesenteric lymph nodes of 7 diabetic rats. Rats were followed for 30 days after islet transplantation. A total of 7 islet transplantations in mesenteric lymph nodes have been performed. Two rats died 24 and 36 h after transplantation due to complications. No transplanted rat acquired normal glucose blood levels and insulin independence after the transplantation. However, the mean blood levels of glycemia were significantly lower in transplanted rats compared with diabetic rats (470.4 mg/dl vs 605 mg/dl, p 0.04). Interestingly, transplanted rats have a significant weight increase after transplantation compared to diabetic rats (mean value 295 g in transplanted rats vs 245 g in diabetic rats, p < 0.05), with an overall improvement of social activities and health. Immunohistochemical analysis of the 5 mesenteric lymph nodes of transplanted rats demonstrated the presence of living islets in one lymph node. Although islet engraftment in lymph nodes is possible, islet transplantation in lymph nodes in rats resulted in few improvements of glucose parameters.


Highlights from ASCO 2020: updates on the treatment of chronic lymphocytic leukemia.
Because of the global coronavirus pandemic, the 2020 annual scientific meeting of the American Society of Clinical Oncology took place virtually, 29-30 May. At the meeting, results from key studies about the treatment of chronic lymphocytic leukemia (cll) were disseminated. Studies examined the efficacy and safety of ibrutinib, acalabrutinib, zanubrutinib, and venetoclax as monotherapy or in combination with novel agents for patients with treatment-naïve and relapsed or refractory cll. Our meeting report describes the foregoing studies and presents interviews with investigators and commentaries by Canadian hematologists about potential effects on Canadian practice.


Construction of Novel Yeast Strains from Candida tropicalis KBKTI 10.5.1 and Saccharomyces cerevisiae DBY1 to Improve the Performance of Ethanol Production Using Lignocellulosic Hydrolysate.
Increased consumption of xylose-glucose and yeast tolerance to lignocellulosic hydrolysate are the keys to the success of second-generation bioethanol production. Candida tropicalis KBKTI 10.5.1 is a new isolated strain that has the ability to ferment xylose. In contrast to Saccharomyces cerevisiae DBY1 which only can produce ethanol from glucose fermentation. The research objective is the application of the genome shuffling method to increase the performance of ethanol production using lignocellulosic hydrolysate. Mutants were selected on xylose and glucose substrates separately and using random amplified polymorphic DNA (RAPD) analysis. The ethanol production using lignocellulosic hydrolysate by parents and mutants was evaluated using a batch fermentation system. Concentrations of ethanol, residual sugars, and by-products such as glycerol, lactate and acetate were measured using HPLC machine equipped with Hiplex H for carbohydrate column and a refraction index detector (RID). Ethanol produced by Fcs1 and Fcs4 mutants on acid hydrolysate increased by 26.58% and 24.17% from parent DBY1, by 14.94% and 21.84% from parent KBKTI 10.5.1. In contrast to the increase in ethanol production on alkaline hydrolysate, Fcs1 and Fcs4 mutants only experienced an increase in ethanol production by 1.35% from the parent KBKTI 10.5.1. Ethanol productivity by Fcs1 and Fcs4 mutants on acid hydrolysate reached 0.042 g/L/h and 0.044 g/L/h. The recombination of the genomes of different yeast species resulted in novel yeast strains that improved resistance performance and ethanol production on lignocellulosic hydrolysates.


Screening for asthma: results from a rural cohort.
Asthma, the leading cause of chronic illness in children, must be managed in both the home and school environments. Identification of children who have risk factors associated with asthma is the first step toward achieving one of the Healthy People 2010 (2000) objectives, which identifies that 25 states will establish a system of surveillance to track asthma mortality, morbidity, access to care, and asthma management. The purposes of this research were to: a) identify rural children who are at risk for asthma through written screening; b) assess parameters of respiratory health status of rural school-aged children as indicated by forced expiratory volume at l second (FEV1), forced vital capacity (FVC), peak expiratory flow (PEF), mean mid-expiratory flow (FEF(25-75)); and c) identify the number of rural school-aged children who sought and obtained follow-up from their primary health care provider and were given a definitive diagnosis of asthma. The Vulnerable Populations Framework (Flaskerud and Winslow, 1998) was used to organize this study. A prospective descriptive design was utilized for this research. Approximately 12% of the children screened were referred to their primary care provider (PCP) for follow-up care. Of these approximately half of the children were seen by their PCP. Barriers to seeking follow-up care were: a) the child was not symptomatic all the time, b) reluctance to be diagnosed with asthma, and c) others, such as cost and time. Children who were not well controlled identified that they ran out of medicine and their parents did not refill their prescription. Results from this descriptive study indicate that screening for asthma in school may be a way to identify those children who are at risk for asthma, and who are not diagnosed as well as those who are diagnosed with asthma but are not optimally managed. While many parents wanted their children to be screened, follow-up care was not critical to them. Nurses working in a school setting are in a prime position to help identify those children with signs and symptoms of asthma. In addition, use of written screenings with or without spirometry may be helpful in identifying children at risk for asthma. Further studies need to be undertaken to determine if written screening is as efficacious as spirometry for school and other ambulatory care settings.


U7 snRNP-specific Lsm11 protein: dual binding contacts with the 100 kDa zinc finger processing factor (ZFP100) and a ZFP100-independent function in histone RNA 3' end processing.
The 3' cleavage generating non-polyadenylated animal histone mRNAs depends on the base pairing between U7 snRNA and a conserved histone pre-mRNA downstream element. This interaction is enhanced by a 100 kDa zinc finger protein (ZFP100) that forms a bridge between an RNA hairpin element upstream of the processing site and the U7 small nuclear ribonucleoprotein (snRNP). The N-terminus of Lsm11, a U7-specific Sm-like protein, was shown to be crucial for histone RNA processing and to bind ZFP100. By further analysing these two functions of Lsm11, we find that Lsm11 and ZFP100 can undergo two interactions, i.e. between the Lsm11 N-terminus and the zinc finger repeats of ZFP100, and between the N-terminus of ZFP100 and the Sm domain of Lsm11, respectively. Both interactions are not specific for the two proteins in vitro, but the second interaction is sufficient for a specific recognition of the U7 snRNP by ZFP100 in cell extracts. Furthermore, clustered point mutations in three phylogenetically conserved regions of the Lsm11 N-terminus impair or abolish histone RNA processing. As these mutations have no effect on the two interactions with ZFP100, these protein regions must play other roles in histone RNA processing, e.g. by contacting the pre-mRNA or additional processing factors.


Clinical significance of morphologic subdivision in diffuse large cell lymphoma.
Although diffuse large cell lymphomas can be morphologically divided into large cell (DLC) and immunoblastic (IBL) subtypes, the clinical significance of this subdivision remains controversial. The initial diagnostic materials from 85 patients with recorded diagnoses of diffuse large cell lymphoma who were treated at Stanford between 1975 and 1986 with cyclophosphamide, doxorubicin (Adriamycin; Adria Laboratories, Columbus, OH), vincristine, and prednisone (CHOP); methotrexate, bleomycin, doxorubicin, cyclophosphamide, vincristine, and dexamethasone (M-BACOD); or methotrexate, doxorubicin, cyclophosphamide, vincristine, prednisone, and bleomycin (MACOP-B) chemotherapy were retrospectively reviewed by a panel of hematopathologists and classified according to morphologic criteria of the Working Formulation. Based on the criterion of agreement of two of three observers, 60 patients were classified as having DLC, 19 as having IBL, and the lymphomas in 6 patients could not be additionally classified. No significant differences in complete response (CR) rate, freedom from disease progression (FFP), or overall survival were found between the DLC and IBL groups. There was also no significant difference in prognosis between DLC cases additionally subclassified as large cleaved cell (16 patients) and those with large non-cleaved cell (36 patients). Although IBL is considered to be a high-grade lymphoma, the authors concluded that it does not differ significantly in prognosis from DLC lymphoma and, therefore, does not justify a modified treatment selection based on IBL morphologic type alone. Definitive evaluation of the prognostic significance of morphologic subdivision may require a larger cohort of uniformly treated patients.


Closing the "know-do" gap: training public health professionals in eating disorders prevention via case-method teaching.
Expansion of our societies' capacity to prevent eating disorders will require strategic integration of the topic into the curricula of professional training programs. An ideal way to integrate new content into educational programs is through the case-method approach, a teaching method that is more effective than traditional teaching techniques. The Strategic Training Initiative for the Prevention of Eating Disorders has begun developing cases designed to be used in classroom settings to engage students in topical, high-impact issues in public health approaches to eating disorders prevention and screening. Dissemination of these cases will provide an opportunity for students in public health training programs to learn material in a meaningful context by actively applying skills as they are learning them, helping to bridge the "know-do" gap. The new curriculum is an important step toward realizing the goal that public health practitioners be fully equipped to address the challenge of eating disorders prevention.


Effect of nonsteroidal anti-inflammatory drugs on the inflammatory response of bovine endometrial epithelial cells in vitro.
Chronic postpartum uterine infection detrimentally affects subsequent fertility. Nonsteroidal anti-inflammatory drugs (NSAID) are used to alleviate pain and treat inflammatory conditions in transition dairy cows with varying success. To screen the efficacy of NSAID in the absence of animal experiments, we have established an in vitro model to study uterine inflammation. Inflammation was induced in cultured bovine endometrial epithelial cells by challenging cells with an inflammation cocktail: lipopolysaccharide and proinflammatory cytokines, interleukin-1β (IL1β) and tumor necrosis factor α (TNFα). Release of the inflammation markers, serum amyloid A (SAA) and α-1-acid glycoprotein (αAGP), was measured by ELISA. Concentration of these markers was used to indicate the effectiveness in dampening inflammation of 5 NSAID: meloxicam, flunixin meglumine, aspirin, ketoprofen, and tolfenamic acid. Three NSAID, meloxicam, flunixin meglumine, and tolfenamic acid, were successful at dampening the release of SAA and αAGP into cell-culture supernatant, and the corresponding treated cells were selected for down-stream mRNA expression analysis. Expression of 192 genes involved in regulation of inflammatory pathways were investigated using Nanostring. Of the genes investigated, 81 were above the mRNA expression-analysis threshold criteria and were included in expression analysis. All SAA genes investigated (SAA2, SAA3, M-SAA3.2) were upregulated in response to the inflammation cocktail, relative to mRNA expression in control cells; however, AGP mRNA expression was below the expression analysis threshold and was, therefore, excluded from analysis. Treatment with NSAID downregulated genes involved in regulating chemokine signaling (e.g., CXCL2, CXCR4, CXCL5, and CXCL16) and genes that regulate the eicosanoid pathway (e.g., LTA4H, PTGS2, PLA2G4A, and PTGDS). Of the 5 NSAID investigated, meloxicam, flunixin meglumine, and tolfenamic acid are recommended for further investigation into treatment of postpartum uterine inflammation. The results from this study confirm the immunomodulatory properties of the endometrial epithelium in response to inflammatory stimuli and suggest that NSAID may be beneficial in alleviating uterine inflammation.


The N-terminal cysteine-rich domain of tobacco class I chitinase is essential for chitin binding but not for catalytic or antifungal activity.
The vacuolar chitinases of class I possess an N-terminal cysteine-rich domain homologous to hevein and chitin-binding lectins such as wheat germ agglutinin and Urtica dioica lectin. To investigate the significance of this domain for the biochemical and functional characteristics of chitinase, chimeric genes encoding the basic chitinase A of tobacco (Nicotiana tabacum) with and without this domain were constructed and constitutively expressed in transgenic Nicotiana sylvestris. The chitinases were subsequently isolated and purified to homogeneity from the transgenic plants. Chromatography on colloidal chitin revealed that only the form with the N-terminal domain, and not the one without it, had chitin-binding properties, demonstrating directly that the domain is a chitin-binding domain (CBD). Under standard assay conditions with radioactive colloidal chitin, both forms of chitinase had approximately the same catalytic activity. However, kinetic analysis demonstrated that the enzyme without CBD had a considerably lower apparent affinity for its substrate. The pH and temperature optima of the two chitinases were similar, but the form with the CBD had an approximately 3-fold higher activation energy and retained a higher activity at low pH values. Both chitinases were capable of inhibiting growth of Trichoderma viride, although the form with the CBD was about three times more effective than the one without it. Thus, the CBD is not necessary for catalytic or antifungal activity of chitinase.


Long-Term Risks of Parkinson's Disease, Surgery, and Colorectal Cancer in Patients With Slow-Transit Constipation.
Long-term outcomes of constipation have not been evaluated fully. We investigated the incidence of Parkinson's disease, constipation-related surgery, and colorectal cancer (CRC) in patients with constipation and slow-transit constipation (STC), followed up for up to 20 years. We collected data from 2165 patients (33.1% men; median patient age, 54 y; median symptom duration, 5.0 y) with a diagnosis of constipation (based on Rome II criteria) who underwent an anorectal function test and a colonic transit time study, from 2000 through 2010, at a tertiary university hospital in Seoul, South Korea. The presence of STC was determined from colonic transit time. We used the Kaplan-Meier method to analyze and compare cumulative probabilities of a new diagnosis of Parkinson's disease or CRC according to the presence of STC. The patients were followed up until the end of 2019. During a median follow-up period of 4.7 years (interquartile range, 0.7-8.3 y), 10 patients underwent constipation-related surgery. The cumulative probabilities of constipation-related surgery were 0.7% at 5 years and 0.8% at 10 years after a diagnosis of constipation. Twenty-nine patients (1.3%) developed Parkinson's disease; the cumulative probabilities were 0.4% at 1 year, 1.0% at 5 years, and 2.6% at 10 years after a diagnosis of constipation. At 10 years, 1.3% of patients with STC required constipation-related surgery and 3.5% of patients with STC developed Parkinson's disease; in contrast, none of the patients without STC required constipation-related surgery (P = .003), and 1.5% developed Parkinson's disease (P = .019). In multivariate analysis, patient age of 65 years or older at the diagnosis of constipation (hazard ratio, 4.834; 95% CI, 2.088-11.190) and the presence of STC (hazard ratio, 2.477; 95% CI, 1.046-5.866) were associated independently with the development of Parkinson's disease. Only 5 patients had a new diagnosis of CRC during the follow-up period. The risk of CRC did not differ significantly between patients with vs without STC (P = .575). In a long-term follow-up study of patients with constipation in Korea, most patients had no severe complications. However, patients older than age 65 years with a new diagnosis of STC might be considered for Parkinson's disease screening.


Impact of the Dimethyl Sulfoxide Reductase Superfamily on the Evolution of Biogeochemical Cycles.
The dimethyl sulfoxide reductase (or MopB) family is a diverse assemblage of enzymes found throughout Bacteria and Archaea. Many of these enzymes are believed to have been present in the last universal common ancestor (LUCA) of all cellular lineages. However, gaps in knowledge remain about how MopB enzymes evolved and how this diversification of functions impacted global biogeochemical cycles through geologic time. In this study, we perform maximum likelihood phylogenetic analyses on manually curated comparative genomic and metagenomic data sets containing over 47,000 distinct MopB homologs. We demonstrate that these enzymes constitute a catalytically and mechanistically diverse superfamily defined not by the molybdopterin- or tungstopterin-containing [molybdopterin or tungstopterin bis(pyranopterin guanine dinucleotide) (Mo/W-bisPGD)] cofactor but rather by the structural fold that binds it in the protein. Our results suggest that major metabolic innovations were the result of the loss of the metal cofactor or the gain or loss of protein domains. Phylogenetic analyses also demonstrated that formate oxidation and CO2 reduction were the ancestral functions of the superfamily, traits that have been vertically inherited from the LUCA. Nearly all of the other families, which drive all other biogeochemical cycles mediated by this superfamily, originated in the bacterial domain. Thus, organisms from Bacteria have been the key drivers of catalytic and biogeochemical innovations within the superfamily. The relative ordination of MopB families and their associated catalytic activities emphasize fundamental mechanisms of evolution in this superfamily. Furthermore, it underscores the importance of prokaryotic adaptability in response to the transition from an anoxic to an oxidized atmosphere. IMPORTANCE The MopB superfamily constitutes a repertoire of metalloenzymes that are central to enduring mysteries in microbiology, from the origin of life and how microorganisms and biogeochemical cycles have coevolved over deep time to how anaerobic life adapted to increasing concentrations of O2 during the transition from an anoxic to an oxic world. Our work emphasizes that phylogenetic analyses can reveal how domain gain or loss events, the acquisition of novel partner subunits, and the loss of metal cofactors can stimulate novel radiations of enzymes that dramatically increase the catalytic versatility of superfamilies. We also contend that the superfamily concept in protein evolution can uncover surprising kinships between enzymes that have remarkably different catalytic and physiological functions.


Classification and Segmentation of Longitudinal Road Marking using Convolutional Neural Networks for Dynamic Retroreflection Estimation.
Road markings constitute one of the most important elements of the road. Moreover, they are managed according to specific standards, including a criterion for a luminous contrast, which can be referred to as retroreflection. Retroreflection can be used to measure the reflection properties of road markings or other road facilities. It is essential to manage retroreflection in order to improve road safety and sustainability. In this study, we propose a dynamic retroreflection estimation method for longitudinal road markings, which employs a luminance camera and convolutional neural networks (CNNs). The images that were captured by a luminance camera were input into a classification and regression CNN model in order to determine whether the longitudinal road marking was accurately acquired. A segmentation model was also developed and implemented in order to accurately present the longitudinal road marking and reference plate if a longitudinal road marking was determined to exist in the captured image. The retroreflection was dynamically measured as a driver drove along an actual road; consequently, the effectiveness of the proposed method was demonstrated.


Diagnostic challenges in low-grade central osteosarcoma.
Low-grade central osteosarcoma (LGCOS), a rare type of osteosarcoma, often has misleading radiological and pathological features that overlap with those of other bone tumours, thereby complicating diagnosis and treatment. We aimed to analyze the clinical, radiological, and pathological features of patients with LGCOS, with a focus on diagnosis, treatment, and outcomes. We retrospectively analyzed the medical records of 49 patients with LGCOS (Broder's grade 1 to 2) treated between January 1985 and December 2017 in a single institute. We examined the presence of malignant features on imaging (periosteal reaction, cortical destruction, soft-tissue invasion), the diagnostic accuracy of biopsy, surgical treatment, and oncological outcome. Based on imaging, 35 of 49 patients (71.4%) exhibited malignant features. Overall, 40 of 49 patients (81.6%) had undergone a biopsy before en-bloc resection: 27 of 40 patients (67.5%) were diagnosed on the first biopsy, which was more accurate when carried out by open rather than needle biopsy (91.3% vs 35.3% diagnostic accuracy, respectively; p < 0.001). Of the 40 patients treated by en-bloc resection, surgical margins were wide in 38 (95.0%) and marginal in two (5.0%). Furthermore, nine of 49 patients (18.4%) underwent curettage (intralesional margin) without previous biopsy. All patients with a positive margin developed local recurrence. Distant metastases occurred in five of 49 patients (10.2%). The mean five-year overall survival (OS) and distant relapse-free survival (D-RFS) were 89.3% (SD 5.1%) and 85.7% (SD 5.5%), respectively. Univariate analysis showed that the occurrence of distant metastasis was a poor prognostic factor for OS (hazard ratio 11.54, 95% confidence interval (CI) 1.92 to 69.17; p < 0.001). Local recurrence was a poor prognostic factor for D-RFS (HR 8.72, 95% CI 1.69 to 45.0; p = 0.002). The diagnosis of LGCOS can be challenging because it may present with non-malignant features and has a low diagnostic accuracy on biopsy. If precisely diagnosed, LGCOS can be successfully treated by surgical excision with wide margins.


Transcriptional dynamics of LTR retrotransposons in early generation and ancient sunflower hybrids.
Hybridization and abiotic stress are natural agents hypothesized to influence activation and proliferation of transposable elements in wild populations. In this report, we examine the effects of these agents on expression dynamics of both quiescent and transcriptionally active sublineages of long terminal repeat (LTR) retrotransposons in wild sunflower species with a notable history of transposable element proliferation. For annual sunflower species Helianthus annuus and H. petiolaris, neither early generation hybridization nor abiotic stress, alone or in combination, induced transcriptional activation of quiescent sublineages of LTR retrotransposons. These treatments also failed to further induce expression of sublineages that are transcriptionally active; instead, expression of active sublineages in F1 and backcross hybrids was nondistinguishable from, or intermediate relative to, parental lines, and abiotic stress generally decreased normalized expression relative to controls. In contrast to findings for early generation hybridization between H. annuus and H. petiolaris, ancient sunflower hybrid species derived from these same two species and which have undergone massive proliferation events of LTR retrotransposons display 2× to 6× higher expression levels of transcriptionally active sublineages relative to parental sunflower species H. annuus and H. petiolaris. Implications and possible explanations for these findings are discussed.


Purification, crystallization and preliminary X-ray analysis of Methanococcus jannaschii TATA box-binding protein (TBP).
TATA box-binding protein (TBP) from Methanococcus jannaschii has been crystallized by the hanging-drop vapour-diffusion method using PEG MME 2000 as a precipitant. The crystal belongs to space group P21, with unit-cell parameters a = 53.2, b = 55.5, c = 123.4 A,a = 90.0, fi = 91.0, y = 90.0 degrees, and contains four molecules in the asymmetric unit. A data set was collected to 1.9 A resolution using synchrotron radiation. A molecular-replacement solution was found using the structure of TBP from Sulfolobus acidocaldarius as a model. Crystallographic refinement is in progress.


Neonatal respiration, feeding and neurobehavioural state. Effects of intrapartum bupivacaine, pethidine and pethidine reversed by naloxone.
The effects on mature newborn have been compared at 0.5, 4, 8 12 24 and 48 hr after birth, of maternally administered epidural bupivacaine (11 babies) or pethidine (18 babies) or pethidine reversed by naloxone administered intramuscularly to the newborn (15 babies). Bupivacaine (mean dose 130 mg) had less effect that pethidine (mean dose 183.3 mg) on alveolar carbon dioxide tension (PACO2) at 0.5 hr after birth, but had a similar effect to pethidine on feeding, elicited reflexes and produced more depression of muscle tone up to 48 hr. Bupivacaine had more effect on PACO2 feeding measures, elicited reflexes and muscle tone at almost all examination periods than pethidine (mean dose 157.0 mg) reversed by naloxone (200 micrograms intramuscularly). Except at delivery, the effects of bupivacaine or pethidine on respiration and feeding up to 48 hr after birth were similar. There were more signs of depression with both drugs than when pethidine had been reversed by naloxone.


Protein phosphatase 2A (PP2A) has a potential role in CAPE-induced apoptosis of CCRF-CEM cells via effecting human telomerase reverse transcriptase activity.
Caffeic acid phenethyl ester (CAPE) is one of the most effective components of propolis which is collected by honey bees. The aim of this study was to investigate the cytotoxic and apoptotic effects of CAPE in the CCRF-CEM cell line and to clarify the role of serine/threonine protein phosphatase 2A (PP2A) and human telomerase reverse transcriptase (hTERT) activity as an underlining mechanism of CAPE-induced apoptosis. Trypan blue dye exclusion test and XTT methods were used to evaluate the cytotoxicity and ELISA based oligonucleotide detection, which can be seen during apoptosis, was used to determine apoptosis. Acridine orange/ethidium bromide dye technique was also used to evaluate apoptosis. The cytotoxic effect of CAPE was detected in a dose and time dependent manner with the IC(50) of 1 muM. ELISA and acridine orange/ethidium bromide methods have shown remarkable apoptosis at 48th hour in CAPE treated cells. To investigate the role of PP2A in CAPE-induced apoptosis of CCRF-CEM cells, we performed combination studies with CAPE and, Calyculin A and Okadaic acid, which are very well known inhibitors of PP2A, in IC(20) of inhibitors and IC(50) of CAPE. Combination studies revealed synergistic effect of both drugs by concomitant use. Western blot analyses of PP2A catalytic and regulatory subunits showed down-regulation of expression of PP2A catalytic subunit in CAPE treated cells at 48th hour. Since, PP2A is important in hTERT (telomerase catalytic subunit) activation and deactivation, we also performed hTERT activity in CAPE treated cells simultaneously. Treating cells with IC(50) of CAPE for 96 h with the intervals of 24 h showed marked reduction of hTERT activity. The reduction of hTERT activity in CAPE treated CCRF-CEM cells was more prominent in the initial 48 h. The variation of hTERT activity in CAPE treated CCRF-CEM cells may be the reason for the protein phosphatase interaction that occurred after treatment with CAPE.


Nanosized (Ni1-x Zn x )Fe2O4 for water oxidation.
Performing water splitting for H2 production is an interesting method to store different energies. For water splitting, an efficient and stable water-oxidizing catalyst is important. Ni-Fe (hydr)oxides are among the best catalysts for water oxidation in alkaline electrolytes. An Fe amount higher than 50% in Ni-Fe (hydr)oxides increases the overpotential for water oxidation. Thus, Ni-Fe (hydr)oxides with a high ratio of Fe to Ni have rarely been focused on for water oxidation. Herein, we report water oxidation using nanosized (Ni1-x Zn x )Fe2O4. The catalyst was characterized via some methods and tested at pH values of 3, 7 and 11 in phosphate buffer. Nanosized (Ni1-x Zn x )Fe2O4 is a good catalyst for water oxidation only under alkaline conditions. In the next step, amperometry studies showed current densities of 3.50 mA cm-2 and 11.50 mA cm-2 at 1.25 V in 0.10 M and 1.0 M KOH solution, respectively. The amperometric measurements indicated high catalyst stability in both 0.10 M and 1.0 M KOH. Tafel plots were obtained in KOH solution at concentrations of both 0.10 M and 1.0 M. At pH = 13 in KOH solution (0.10 M), linearity of lg(j) vs. potential was shown, with two slopes relating to both relatively low (170.9 mV per decade) and high overpotentials (484.2 mV per decade). In 1.0 M KOH solution, the Tafel plot showed linearity of lg(j) vs. potential, with two slopes relating to both relatively low (192.5 mV per decade) and high overpotentials (545.7 mV per decade). After water oxidation, no significant change was observed in the catalyst.


Stabilization of heterochromatin by CLOCK promotes stem cell rejuvenation and cartilage regeneration.
Accumulating evidence indicates an association between the circadian clock and the aging process. However, it remains elusive whether the deregulation of circadian clock proteins underlies stem cell aging and whether they are targetable for the alleviation of aging-associated syndromes. Here, we identified a transcription factor-independent role of CLOCK, a core component of the molecular circadian clock machinery, in counteracting human mesenchymal stem cell (hMSC) decay. CLOCK expression was decreased during hMSC aging. In addition, CLOCK deficiency accelerated hMSC senescence, whereas the overexpression of CLOCK, even as a transcriptionally inactive form, rejuvenated physiologically and pathologically aged hMSCs. Mechanistic studies revealed that CLOCK formed complexes with nuclear lamina proteins and KAP1, thus maintaining heterochromatin architecture and stabilizing repetitive genomic sequences. Finally, gene therapy with lentiviral vectors encoding CLOCK promoted cartilage regeneration and attenuated age-related articular degeneration in mice. These findings demonstrate a noncanonical role of CLOCK in stabilizing heterochromatin, promoting tissue regeneration, and mitigating aging-associated chronic diseases.


Topography and chemical reactivity of the active-inactive transition-sensitive SH-group in the mitochondrial NADH:ubiquinone oxidoreductase (Complex I).
The spatial arrangement and chemical reactivity of the activation-dependent thiol in the mitochondrial Complex I was studied using the membrane penetrating N-ethylmaleimide (NEM) and non-penetrating anionic 5,5'-dithiobis-(2-nitrobenzoate) (DTNB) as the specific inhibitors of the enzyme in mitochondria and inside-out submitochondrial particles (SMP). Both NEM and DTNB rapidly inhibited the de-activated Complex I in SMP. In mitochondria NEM caused rapid inhibition of Complex I, whereas the enzyme activity was insensitive to DTNB. In the presence of the channel-forming antibiotic alamethicin, mitochondrial Complex I became sensitive to DTNB. Neither active nor de-activated Complex I in SMP was inhibited by oxidized glutathione (10 mM, pH 8.0, 75 min). The data suggest that the active/de-active transition sulfhydryl group of Complex I which is sensitive to inhibition by NEM is located at the inner membrane-matrix interface. These data include the sidedness dependency of inhibition, effect of pH, ionic strength, and membrane bilayer modification on enzyme reactivity towards DTNB and its neutral analogue.


Isolation of mitochondria from Saccharomyces cerevisiae using magnetic bead affinity purification.
Isolated mitochondria are widely used to study the function of the organelle. Typically, mitochondria are prepared using differential centrifugation alone or in conjunction with density gradient ultracentrifugation. However, mitochondria isolated using differential centrifugation contain membrane or organelle contaminants, and further purification of crude mitochondria by density gradient ultracentrifugation requires large amounts of starting material, and is time-consuming. Mitochondria have also been isolated by irreversible binding to antibody-coated magnetic beads. We developed a method to prepare mitochondria from budding yeast that overcomes many of the limitations of other methods. Mitochondria are tagged by insertion of 6 histidines (6xHis) into the TOM70 (Translocase of outer membrane 70) gene at its chromosomal locus, isolated using Ni-NTA (nickel (II) nitrilotriacetic acid) paramagnetic beads and released from the magnetic beads by washing with imidazole. Mitochondria prepared using this method contain fewer contaminants, and are similar in ultrastructure as well as protein import and cytochrome c oxidase complex activity compared to mitochondria isolated by differential centrifugation. Moreover, this isolation method is amenable to small samples, faster than purification by differential and density gradient centrifugation, and more cost-effective than purification using antibody-coated magnetic beads. Importantly, this method can be applied to any cell type where the genetic modification can be introduced by CRISPR or other methods.


Clinical outcome of living donor kidney transplantation across simultaneous ABO and HLA incompatibility: Single center experience of first ten cases.
Preconditioning using different protocols has been tested to prevent antibody mediated rejection (ABMR) individually for ABO and HLA incompatibility. However, simultaneous presence of both barriers is still less explored. The aim of this study was to report outcomes of institutional desensitization protocol in renal transplant recipients with simultaneous ABO and HLA incompatibility. This was a retrospective study conducted from October 2015 to December 2018. All patients with a clinical diagnosis of dialysis dependent chronic kidney disease (CKD), who were prospective coexistent HLA and ABO incompatible renal transplant recipients were included in the study. Patients were followed up and graft function and patient survival was assessed at 1 y from the date of transplant. Median and mode baseline anti-A titers were 64, while median and mode baseline anti-B titers were 256. All recipients were discharged by tenth postoperative day. None of the patients had any bleeding complications. Post transplant infection rate was found to be 20 %. A total of 54 therapeutic plasma exchange (TPE) procedures were performed before transplant and 8 were performed after transplant. Graft survival and patient survival was 100 % at 3, 6, 9, and 12 months. Range and mean follow-up period was 15-42 months and 23 months respectively. Mean glomerular filtration rate (GFR) at 1 y using the CKD-EPI equation was 85.25 ± 13.76 mL/min. Biopsy proven ABMR was observed in one case only which was managed with TPE and immunosuppression. Simultaneous ABO and HLA incompatibility in renal transplant recipients can be managed successfully with adequate preconditioning and careful monitoring.


Metabolic modelling of polyhydroxyalkanoate copolymers production by mixed microbial cultures.
This paper presents a metabolic model describing the production of polyhydroxyalkanoate (PHA) copolymers in mixed microbial cultures, using mixtures of acetic and propionic acid as carbon source material. Material and energetic balances were established on the basis of previously elucidated metabolic pathways. Equations were derived for the theoretical yields for cell growth and PHA production on mixtures of acetic and propionic acid as functions of the oxidative phosphorylation efficiency, P/O ratio. The oxidative phosphorylation efficiency was estimated from rate measurements, which in turn allowed the estimation of the theoretical yield coefficients. The model was validated with experimental data collected in a sequencing batch reactor (SBR) operated under varying feeding conditions: feeding of acetic and propionic acid separately (control experiments), and the feeding of acetic and propionic acid simultaneously. Two different feast and famine culture enrichment strategies were studied: (i) either with acetate or (ii) with propionate as carbon source material. Metabolic flux analysis (MFA) was performed for the different feeding conditions and culture enrichment strategies. Flux balance analysis (FBA) was used to calculate optimal feeding scenarios for high quality PHA polymers production, where it was found that a suitable polymer would be obtained when acetate is fed in excess and the feeding rate of propionate is limited to approximately 0.17 C-mol/(C-mol.h). The results were compared with published pure culture metabolic studies. Acetate was more conducive toward the enrichment of a microbial culture with higher PHA storage fluxes and yields as compared to propionate. The P/O ratio was not only influenced by the selected microbial culture, but also by the carbon substrate fed to each culture, where higher P/O ratio values were consistently observed for acetate than propionate. MFA studies suggest that when mixtures of acetate and propionate are fed to the cultures, the catabolic activity is primarily guaranteed through acetate uptake, and the characteristic P/O ratio of acetate prevails over that of propionate. This study suggests that the PHA production process by mixed microbial cultures has the potential to be comparable or even more favourable than pure cultures.


Randomized double-blind placebo-controlled trial of homoeopathic 'proving' for Belladonna C30.
Homoeopathic drug pictures are developed by recording the symptomatic effects of homoeopathic remedies given to healthy volunteers (a 'proving'). In a double-blind randomized controlled trial we tested the hypothesis that individuals using an infinitesimal dilution of Belladonna (thirtieth potency, C30) would record more true symptoms, on a questionnaire that contained both true and false Belladonna proving symptoms, than those receiving placebo. 60 volunteers entered the study and 47 completed data collection. We were unable to distinguish between Belladonna C30 and placebo using our primary outcome measure. For the secondary outcome measure we analysed the number of individuals who proved to the remedy according to our predefined criteria: 4 out of 19 proved in the Belladonna C30 group and 1 out of 27 in the placebo group (difference not statistically significant). This pilot study does not demonstrate a clear proving reaction for Belladonna C30 versus placebo, but indicates how the question might be further investigated.


Effect of both local and systemically administered dexamethasone on long-term hearing and tissue response in a Guinea pig model of cochlear implantation.
Dexamethasone administered prior to cochlear implantation has been shown to reduce the loss of residual hearing in experimental settings. However, its effect on the tissue response around the implant has not been extensively studied. In this study dexamethasone sodium phosphate was administered to guinea pigs via local delivery to the round window (2% dexamethasone for 120 min prior to surgery, 'local 2/120', or 20% dexamethasone for 30 min prior to surgery) or intravenously (2 mg/kg dexamethasone for 60 min) prior to implantation. Auditory brainstem responses (ABR) were monitored for 3 months, after which the cochleae were embedded in Spurr's resin and sectioned. The extent of the tissue response and the survival of the neurosensory structures were analysed. Both local 2/120 and systemically delivered dexamethasone improved ABR thresholds when compared with control animals. Systemic dexamethasone also reduced the tissue response around the electrode. This suggests that whilst both locally and systemically administered dexamethasone can protect residual hearing after cochlear implantation, their effects upon the tissue response to implantation may differ.