First Report of Biochemical Mechanisms of Insecticide Resistance in the Field Population of Culex pipiens (Diptera: Culicidae) from Sari, Mazandaran, North of Iran.
Culex pipiens play an important role in transmission of infectious diseases. Vector control by chemical pesticides, leads inevitably to resistance development. Understanding the underlying resistance mechanisms can help improve the control programmes and insecticide resistance management. The total contents of cytochrome p450s and the activities of glutathione S-transferases, alpha- and beta-esterases and inhibition rates of acetylcholine esterase (by propoxur) were measured in the field population of Cx. pipiens collected from Sari County, North of Iran, in 2016 and the results were compared with those of the laboratory susceptible strain according to the biochemical assay methods of WHO for adult mosquitoes. Independent sample t-test was used to compare the mean values of enzyme activities/contents between filed and laboratory susceptible populations. The enzyme ratio of cytochrome p450s, alpha- and beta-esterases in the field population was 2.07, 3.72 and 1.36 respectively when compared with the results of the laboratory population. Although not statistically significant, the mean GSTs activities in the field population was marginally less than the laboratory population (ER=0.92). Acetylcholinesterase was insensitive to propoxur in 62.82% of the individuals of the tested field population. There was a significant difference (P< 0.05) between all values of the activities/contents of the enzyme in the field population except for GSTs compared with the laboratory susceptible strain. The highest enzyme activity was related to alpha esterase. The present study showed a range of metabolic mechanisms, comprising p450s and esterases combined with target site insensitivity of AChE, contributing to organophosphate, carbamate and pyrethroid resistance in the field population of Cx. pipiens.


Plasmodium falciparum: Multidrug resistance.
Malaria is the most lethal and debilitating disease caused by the protozoan parasite Plasmodium worldwide. The most severe forms of disease and the incidence rates of mortality are associated with P. falciparum infections. With the identification of disease source and symptoms, many chemical entities were developed naturally and synthetically for administration as a potential antimalarial drug. The major classes of approved antimalarial drugs that are governed as first-line treatment in tropical and subtropical areas include quinolines, naphthoquinones, antifolates, 8-aminoquinolines, and endoperoxides. However, the efficacy of antimalarial drugs has decreased due to ongoing multidrug resistance problem to current drugs. With increasing resistance to the current antimalarial artemisinin and its combination therapies, malaria prophylaxis has declined gradually. New-generation antimalarial and novel drug target are required to check the incidence of malaria resistance. This review summarizes the emergence of multidrug resistance to known antimalarial and the development of new antimalarial to resolve drug resistance condition. Few essential proteins are also discussed that can be considered as novel drug target against malaria in future.


Evolution of membrane oxygenator technology for utilization during pediatric cardiopulmonary bypass.
The development of the membrane oxygenator for pediatric cardiopulmonary bypass has been an incorporation of ideology and technological advancements with contributions by many investigators throughout the past two centuries. With the pursuit of this technological achievement, the ability to care for mankind in the areas of cardiac surgery has been made possible. Heart disease can affect anyone within the general population, but one such segment that it can affect from inception includes children. Currently, congenital heart defects are the most common birth defects nationally and worldwide. A large meta-analysis study from 1930 to 2010 was conducted in review of published medical literature totaling 114 papers with a study population of 24,091,867 live births, and divulged a staggering incidence of congenital heart disease involving 164,396 subjects with diverse cardiac illnesses. The prevalence of these diseases increased from 0.6 per 1,000 live births from 1930-1934 to 9.1 per 1,000 live births after 1995. These data reveal an emphasis on a growing public health issue regarding congenital heart disease. This discovery displays a need for heightened awareness in the scientific and medical industrial community to accelerate investigative research on emerging cardiovascular devices in an effort to confront congenital anomalies. One such device that has evolved over the past several decades is the pediatric membrane oxygenator. The pediatric membrane oxygenator, in conjunction with the heart lung machine, assists in the repair of most congenital cardiac defects. Numerous children born with congenital heart disease with or without congestive heart failure have experienced improved clinical outcomes in quality of life, survival, and mortality as a result of the inclusion of this technology during their cardiac surgical procedure. The purpose of this review is to report a summary of the published medical and scientific literature related to development of the pediatric membrane oxygenator from its conceptual evolutionary stages to artificially supporting whole body perfusion in the modern pediatric cardiac surgical setting.


Jun B expression is regulated differently by three mitogenic pathways in thyrocytes.
In dog thyrocytes in primary culture, thyrotropin (TSH), acting through cyclic AMP, induces proliferation and differentiation expression, while tetradecanoylphorbol acetate (TPA) or epidermal growth factor (EGF) induces proliferation and dedifferentiation. In this work, we have investigated the regulation of mRNA expression of the protooncogene jun B in these cells. TSH stimulated jun B expression very transiently with biphasic kinetics similar to those obtained for c-myc mRNA accumulation. Forskolin reproduced these effects, suggesting that they are, as other effects of TSH in this system, mediated by cyclic AMP. As shown by nuclear run-on experiments, jun B is regulated by the cAMP pathway at the transcriptional level. As in other cell types, EGF or TPA caused a more sustained increase in mRNA levels. In thyroid slices, in which DNA synthesis appears to be induced by the wounding process, jun B is also induced, suggesting a correlation with the proliferative status of the cell. Interestingly, two jun B mRNAs of 2.1 and 2.3 kb were induced by all the mitogenic pathways. The kinetics of their accumulation were different; i.e., TPA induced the smaller transcript with some delay after the longer one and cycloheximide induced the progressive shortening of the first appearing heavier mRNA. The 2.3-kb messenger has a longer poly(A) tail, and kinetics in the presence of actinomycin D suggested it could represent a precursor form of the 2.1-kb messenger. It is suggested that the specific kinetics of cyclic-AMP-induced accumulation of jun B mRNA could be related to the dual stimulation of differentiation and proliferation by TSH in dog thyrocytes.


Understandings and experiences of adherence to secondary prevention for patients with cardiovascular disease and comorbid depression or anxiety.
Over 20% of cardiovascular disease (CVD) patients have a comorbid mental health disorder, resulting in an increased risk of recurring major adverse cardiac events (MACE) and mortality. Despite the higher risk, patients with comorbid depression or anxiety disorders are twice as likely to be non-adherent to secondary prevention. Therefore, better understanding of the adherence experiences of this subgroup is needed to inform service delivery and enhance adherence for this higher risk group. This study aims to explore the perceptions, understandings, and experiences of adherence to secondary prevention amongst 33 cardiac patients with diagnosed depression and/or anxiety disorder. Participants were recruited as part of the Cardiovascular Health in Anxiety or Mood Problems Study. Semi-structured interviews were conducted and data were analysed via inductive thematic analysis. Patient understandings of adherence to secondary prevention were limited, with medication compliance considered the marker of adherence. Further, participants did not perceive unintentional non-adherence to constitute non-adherence, rather an intent to engage was viewed as defining adherence. Participants also reported that a lack of practitioner understanding and management around their mental health negatively impacted the practitioner-patient relationship and their engagement with secondary prevention. Results highlight that unique barriers, especially around management of comorbid mental health exist for this subgroup. Additionally, adherence to secondary prevention might be limited by patients' narrow understandings of adherence as the intent to engage and as medication compliance.


Block of the background K(+) channel TASK-1 contributes to arrhythmogenic effects of platelet-activating factor.
Platelet-activating factor (PAF), an inflammatory phospholipid, induces ventricular arrhythmia via an unknown ionic mechanism. We can now link PAF-mediated cardiac electrophysiological effects to inhibition of a two-pore domain K(+) channel [TWIK-related acid-sensitive K(+) background channel (TASK-1)]. Superfusion of carbamyl-PAF (C-PAF), a stable analog of PAF, over murine ventricular myocytes causes abnormal automaticity, plateau phase arrest of the action potential, and early afterdepolarizations in paced and quiescent cells from wild-type but not PAF receptor knockout mice. C-PAF-dependent currents are insensitive to Cs(+) and are outwardly rectifying with biophysical properties consistent with a K(+)-selective channel. The current is blocked by TASK-1 inhibitors, including protons, Ba(2+), Zn(2+), and methanandamide, a stable analog of the endogenous lipid ligand of cannabinoid receptors. In addition, when TASK-1 is expressed in CHO cells that express an endogenous PAF receptor, superfusion of C-PAF decreases the expressed current. Like C-PAF, methanandamide evoked spontaneous activity in quiescent myocytes. C-PAF- and methanandamide-sensitive currents are blocked by a specific protein kinase C (PKC) inhibitor, implying overlapping signaling pathways. In conclusion, C-PAF blocks TASK-1 or a closely related channel, the effect is PKC dependent, and the inhibition alters the electrical activity of myocytes in ways that would be arrhythmogenic in the intact heart.


Arsenic in US correctional facility drinking water, 2006-2011.
Little is known about the quality of drinking water in US correctional facilities (e.g. detention centers, prisons, jails, etc.). Our objective was to determine if incarcerated persons are at risk for chronic, elevated arsenic exposure relative to the non-incarcerated US population, particularly in the Southwestern US where public water and groundwater arsenic concentrations are high compared to the rest of the US. We analyzed 230,158 arsenic monitoring records from 37,086 community water systems (CWSs) from the Environmental Protection Agency's (EPA) Third Six Year Review of Contaminant Occurrence dataset (covering 2006-2011). We compared six-year average arsenic concentrations and the odds of exceeding the EPA's 10 μg/L maximum contaminant level (MCL) for CWSs exclusively serving correctional facilities versus all other CWSs in the Southwestern US, where groundwater arsenic concentrations are high. Average six-year water arsenic concentrations were higher for Southwestern correctional facility CWSs (6.41 μg/L, 95% CI 3.48, 9.34) compared to all other Southwestern CWSs (3.11 μg/L, 95% CI 2.97, 3.24) and to other CWSs across the rest of the US (1.39 μg/L, 95% CI 1.35, 1.42). In the Southwest, 26.1% (N = 6) of correctional facility CWSs versus 5.8% (509) of other CWSs reported six-year arsenic averages exceeding 10 μg/L, corresponding to an odds ratio of 5.70 (95% confidence interval 2.24, 14.52). Correctional facility CWSs in the Southwest were also more likely to report six-year averages exceeding 5 μg/L (the MCL for New Jersey and New Hampshire, N = 8, odds ratio 2.77, 95% CI 1.17, 6.54). Persons incarcerated in the Southwestern US were at disproportionate risk of elevated drinking water arsenic exposure and related disease from 2006 to 2011. Strict enforcement of EPA regulations and additional technical and financial support for CWSs serving correctional facilities in the Southwest is necessary to protect the health and human rights of incarcerated persons.


A composite bone graft substitute for anterior cervical fusion: assessment of osseointegration by quantitative computed tomography.
An ongoing prospective nonrandomized trial was conducted with 146 patients who underwent anterior cervical fusion with a bone graft substitute. The results for 78 patients followed up a minimum of 2 years are reported by an independent observer. To investigate the clinical efficacy and osseointegration of the implant. Most of the cervical cages provide an immediate load-bearing capacity, but not necessarily a solid fusion. The bony ingrowth can hardly be evaluated by flexion-extension plain radiographs, especially if a part of the implant is made of titanium. The composite bone graft substitute consists of a rectangular fenestrated titanium cage filled with a highly porous hydroxyapatite cylinder soaked with bone marrow aspirated from a vertebra. Altogether, 102 implants were inserted for anterior cervical fusion. The bone ingrowth was measured in 50 patients by quantitative computed tomography of the implant and the adjacent vertebrae after 1 week, then 6 and 12 months after surgery. The investigation was repeated in 24 randomly selected patients 24 months after surgery. The surgery benefited 42 of 48 patients with myelopathy, 27 of 28 patients with radiculopathy, and both patients with anterior horn cell syndrome. No graft substitute-related complications were observed, and no revision surgery was performed. Six months after surgery, quantitative computed tomography showed up to a 14% increase in the hydroxyapatite mass in the core of the implant, which was statistically significant (P < 0.001). At 1 year, the newly formed hydroxyapatite mass increased to 23%, and in 2 years reached 24%. The composite implant is effective as a bone graft substitute, providing immediate load-bearing capacity, an osteoconductive scaffold, and osteoinductive potency. After surgery, the increase in hydroxyapatite radiodensity was apparent in plain computed tomography scans and confirmed to be significant by quantitative computed tomography.


Effects of Neuromuscular Electrical Stimulation and Resistance Training on Knee Extensor/Flexor Muscles.
Neuromuscular electrical stimulation (NMES) has recently drawn a lot of attention as means for strengthening of voluntary muscle contraction both in sport and rehabilitation. NMES training increases maximal voluntary contraction (MVC) force output through neural adaptations. On the other hand, positive effects of resistance training (RT) on muscle strength are well known. The aim of this study was to investigate effects of a 5-week program of NMES compared to RT program of same duration. Sample of 15 students' of faculty of sport and physical education (age 22 ± 2) were randomized in two groups: NMES (N = 7) and RT (N = 8). NMES group performed NMES superimposed over voluntary muscle contraction, RT group performed resistance training with submaximal loads. Subjects were evaluated for knee isokinetic dynamometry on both sides (60° and 180° s). After intervention no significant difference between groups were observed in isokinetic dynamometry (p = 0.177). However, applying pair sample t test within each group revealed that peak torque increased in NMES-group (p = 0.002 for right knee extensors muscles, p = 0.003 for left, respectively, at 60° and p = 0.004 for left knee extensors muscles, at angular velocity 180°). In RT group (p = 0.033 for right knee extensors muscles, p = 0.029 for right knee flexor muscles, at angular velocity 60°). Our results indicate that NMES has equal potential if not in some way better than classical RT having in mind that overload on locomotor apparatus during NMES is minimal and force of muscle contraction is equal on both sides, for enhancement of knee muscles concentric peak torque.


ALPHA-Amylase activity in sweat and serum of patients with cystic fibrosis of the pancreas.
The hypothesis by Doggett and Harrison, according to which alpha-amylase is the pathogenic factor of the exocrinopathy in cystic fibrosis (C.F.), is investigated. No elevation of alpha-amylase in sweat and serum of C.F. patients, as compared with controls of similar age, is observed. It is concluded that the "C.F. factor" cannot be identified with alpha-amylase.


30 YEARS OF THE MINERALOCORTICOID RECEPTOR: Nongenomic effects via the mineralocorticoid receptor.
The mineralocorticoid receptor (MR) belongs to the steroid hormone receptor family and classically functions as a ligand-dependent transcription factor. It is involved in water-electrolyte homeostasis and blood pressure regulation but independent from these effects also furthers inflammation, fibrosis, hypertrophy and remodeling in cardiovascular tissues. Next to genomic effects, aldosterone elicits very rapid actions within minutes that do not require transcription or translation and that occur not only in classical MR epithelial target organs like kidney and colon but also in nonepithelial tissues like heart, vasculature and adipose tissue. Most of these effects can be mediated by classical MR and its crosstalk with different signaling cascades. Near the plasma membrane, the MR seems to be associated with caveolin and striatin as well as with receptor tyrosine kinases like EGFR, PDGFR and IGF1R and G protein-coupled receptors like AT1 and GPER1, which then mediate nongenomic aldosterone effects. GPER1 has also been named a putative novel MR. There is a close interaction and functional synergism between the genomic and the nongenomic signaling so that nongenomic signaling can lead to long-term effects and support genomic actions. Therefore, understanding nongenomic aldosterone/MR effects is of potential relevance for modulating genomic aldosterone effects and may provide additional targets for intervention.


Radioimmunoassay, acetylating radio-enzymatic assay, and microbioassay of gentamicin: a comparative study.
Gentamicin is an aminoglycoside antibiotic widely used to treat gram-negative bacillary infections. Because it has a low therapeutic index, monitoring of serum levels may help to insure adequacy of dosage and avoid toxicity. Microbiological assays are relatively slow and can be complicated by the presence of other antimicrobials. Radioimmunoassay (RIA) and acetylating radio-enzymatic assay (ARA) are new methods for gentamicin assay which offer the following advantages: rapidity (less than 3 hours); no interference by other antibiotics; RIA is extremely sensitive and ARA is versatile (being useful in the measurement of other aminoglycosides). Correlation coefficients determined by linear regression analysis of assays on 36 patient samples performed in duplicate on 2 different days demonstrated no significant difference in measurement of gentamicin by each of the methods. Factors such as numbers of specimens, cost, and time involved will affect the decision of the method to be applied in individual laboratories.


Study on nanocomposites based on carbonate@ceria.
Nanocomposites based on the ceria-carbonate composite have demonstrated as electrolytes in development of successful 300-600 degrees C fuel cell technology. In this paper, the nanocomposite electrolyte based on carbonate@SDC (SDC: samarium doped ceria) was directly synthesized from the co-precipitation method and characterized by XRD, SEM, TEM, BET, etc. It was proved that the carbonate@SDC was a two-phase material with average particle size about 14.5 nm (S(BET)) and crystalline size (D(XRD)) ranged from 12 to 14 nm. When the carbonate@SDC electrolyte was used to fabricate single SOFC, the cell shows remarkable performance with maximum power density 1000-1180 mw/cm2 at low temperature (300-550 degrees C).


An update on the pathophysiology and genetics of restless legs syndrome.
This update on restless legs syndrome (RLS) focuses on its pathophysiology, genetics, health impact, and treatment. Although symptoms are exquisitely responsive to dopaminergics, clear delineation of dopaminergic pathology has not emerged. Rather, heuristic models of alterations in spinal sensorimotor circuits and central nervous system iron deficiency are gaining more attention. Genome-wide association studies have recently identified polymorphisms in three genes with no obvious relationship to dopamine that account for 70% of the population risk for RLS. A single variant in the BTBD9 gene on chromosome 6 contributes to 50% of the population risk. Although the functions of BTBD9 remain uncertain, its biological plausibility is evidenced by its dose-dependent relationship to periodic limb movements of sleep, decrements in iron stores, and ethnic differences in RLS prevalence. RLS is also implicated in cardiovascular morbidity. Despite these advances in our knowledge, there remain unanswered questions about the pathophysiology of RLS as well as its genetics, health-related significance, and treatment.


Evaluating the impact of test-trace-isolate for COVID-19 management and alternative strategies.
There are many contrasting results concerning the effectiveness of Test-Trace-Isolate (TTI) strategies in mitigating SARS-CoV-2 spread. To shed light on this debate, we developed a novel static-temporal multiplex network characterizing both the regular (static) and random (temporal) contact patterns of individuals and a SARS-CoV-2 transmission model calibrated with historical COVID-19 epidemiological data. We estimated that the TTI strategy alone could not control the disease spread: assuming R0 = 2.5, the infection attack rate would be reduced by 24.5%. Increased test capacity and improved contact trace efficiency only slightly improved the effectiveness of the TTI. We thus investigated the effectiveness of the TTI strategy when coupled with reactive social distancing policies. Limiting contacts on the temporal contact layer would be insufficient to control an epidemic and contacts on both layers would need to be limited simultaneously. For example, the infection attack rate would be reduced by 68.1% when the reactive distancing policy disconnects 30% and 50% of contacts on static and temporal layers, respectively. Our findings highlight that, to reduce the overall transmission, it is important to limit contacts regardless of their types in addition to identifying infected individuals through contact tracing, given the substantial proportion of asymptomatic and pre-symptomatic SARS-CoV-2 transmission.


Identifying the genetic determinants of emotionality in humans; insights from rodents.
The identification of human quantitative trait genes underlying variation in emotionality has proved to be both difficult and expensive. Whilst several QTL have been consistently identified and independently replicated across both phenotypes and populations, little progression has been made towards the identification of underlying genes or even variants. In this review we discuss the potential for using cross-species QTL concordance as a tool for QTL dissection in behaviour genetics, using an affect-related locus mapped to human chromosome 1 as an example.


Clinical effects of glass ionomer restorations on residual carious dentin in primary molars.
To evaluate the mineralization and morphology of residual (infected and affected) carious dentin following the restoration of vital primary molars with viscous glass-ionomer cement (GIC). Encapsulated Fuji IX GP and Ketac-Molar Aplicap GICs were placed in cavities that were prepared using the atraumatic restorative treatment (ART) approach. Four suitable exfoliated teeth with intact restorations were sectioned and then examined by electron probe microanalysis (EPMA) and scanning electron microscopy (SEM). EPMA demonstrated the presence of fluorine and strontium that had penetrated into the underlying residual carious dentin from the adjacent GIC. The concentrations of these two elements, and those of calcium and phosphorous, varied with distance from the GIC/dentin interface. SEM showed varying degrees of dentin tubule destruction and intratubular (peritubular) dentin present immediately subjacent to the GIC/dentin interface. Incomplete removal of carious dentin was observed in all specimens, and GIC remained adherent to the tissue.


Microfluidic add-on for standard electrophysiology chambers.
We have developed a microfluidic brain slice device (microBSD) that marries an off-the shelf brain slice perfusion chamber with an array of microfluidic channels set into the bottom surface of the chamber substrate. As this device is created through rapid prototyping, once optimized, it is trivial to replicate and share the devices with other investigators. The device integrates seamlessly into standard physiology and imaging chambers and it is immediately available to the whole slice physiology community. With this technology we can address the flow of neurochemicals and any other soluble factors to precise locations in the brain slice with the temporal profile we choose. Dopamine (DA) was chosen as a model neurotransmitter and we have quantified delivery in brain tissue using cyclic voltammetry (CV) and fluorescence imaging.


Fluorescent in-situ hybridization on human embryos showing cleavage arrest after freezing and thawing.
Our current freezing-thawing policy is to transfer only embryos that cleave further in the 24 h following thawing. The purpose of our study was to investigate the incidence of numerical abnormalities for chromosomes X, Y and 1 in blastomeres of human preimplantation embryos that survived cryopreservation but did not cleave further after thawing. A total of 63 embryos surviving a freezing-thawing cycle but not cleaving further within 24 h after thawing were screened. Of the 63 screened embryos that showed cleavage arrest 24 h after thawing, 56 embryos (88.9%) remained arrested 48 h after thawing and slightly more than half of these (29/56; 51.8%) showed further deterioration in morphological quality. Seven embryos (11.1%) showed signs of further cleavage; five embryos showed additional cleavage of one blastomere and two developed a blastocoelic cavity. Fluorescent in-situ hybridization (FISH) with three specific probes for simultaneous detection of chromosomes X, Y and 1 was performed and was successful in 60 out of 63 embryos. Of these successfully labelled embryos, 26 (43.3%) were in the diploid range: 12 (20%) were uniformly diploid for the chromosomes X, Y and 1; three embryos showed aneuploidy in all their blastomeres (two were XXY-karyotype and one was monosomy 1) and in 11 embryos nondisjunction was detected. Thirteen embryos were categorized as being either haploid, triploid, tetraploid or hexaploid. Nine embryos were classified as mosaic and 12 as being highly abnormal or chaotic. These preliminary results suggest that a large proportion of embryos that do not cleave further after freezing and thawing carry chromosomal aberrations. This finding supports our policy of not transferring cryopreserved embryos which do not cleave further 24 h following thawing.


Muscle- and Region-Specific Associations Between Muscle Size and Muscular Strength During Hip Extension and Knee Flexion in the Hamstrings.
Strength deficits of the hamstrings following sports injuries decrease athletic performance and increase the risk of injury recurrence. Previous studies have shown a high correlation between the muscular strength during hip-extension and knee-flexion and total muscle size of the hamstrings. However, it remains unclear which region of the individual hamstring muscles is closely associated with muscular strength. To investigate the relationship between the size of each region of the individual hamstring muscles and muscular strength during hip extension and knee flexion. Within-subject repeated measures. University laboratory. Twenty healthy young male volunteers who regularly engaged in sports activities. Anatomical cross-sectional areas were acquired from the proximal, middle, and distal regions of the biceps femoris long head, biceps femoris short head, semitendinosus, and semimembranosus. Hip-extension and knee-flexion strength were measured during maximal voluntary isometric and concentric contractions (angular velocities of 60°/s and 180°/s). The anatomical cross-sectional area of the distal regions in biceps femoris long head (r = .525-.642) and semitendinosus (r = .567) were significantly correlated with hip-extension strength under all conditions and only at an angular velocity of 180°/s, respectively. Meanwhile, anatomical cross-sectional areas of the distal regions in biceps femoris short head (r = .587-.684) and semimembranosus (r = .569-.576) were closely associated with knee-flexion strength under all conditions. These results suggest that muscle size in the distal regions of biceps femoris long head and semitendinosus greatly contributes to the production of hip-extension strength, whereas that of biceps femoris short head and semimembranosus significantly contributes to the generation of knee-flexion strength. These findings could be useful for designing training and rehabilitation programs to efficiently improve strength deficits following sports injuries such as strain injury and anterior cruciate ligament tears.


Role of the glutamic and aspartic residues in Na+-ATPase function in the ZrENA1 gene of Zygosaccharomyces rouxii.
The effect of replacement of negatively charged amino acid residues on the function of Na+ transport proteins of the salt-tolerant yeast Zygosaccharomyces rouxii was examined by heterologous expression of mutant proteins in a strain of Saccharomyces cerevisiae, RH16.6, that lacks native Na+-ATPase activity due to null mutations of ENA1, ENA2, ENA3, and ENA4. Mutants of Na+/H+ antiporter gene (ZrSOD2) and Na+-ATPase gene (ZrENA1) of Z. rouxii were generated by site-directed mutagenesis. The significance of two aspartic residues arranged in tandem (D265 and D266) was demonstrated in Z. rouxii Na+/H+ antiporter. Some Z. rouxii Na+-ATPase mutant genes, namely E778A, D852A, and E981A present in the transmembrane domains (TMDs) and D736A, D743A, D748A, D749A, D759A, and D760A present in the cytoplasmic space were constructed. A lower level of salt tolerance was bestowed by the mutant genes D852A and E981A present in TMDs and D748A and D749A present in cytoplasmic space, compared with the wild-type gene (ZrENA1).


Azithromycin found to be comparable to levofloxacin for the treatment of US travelers with acute diarrhea acquired in Mexico.
Increased drug resistance among enteropathogens is an emergent problem in travelers' diarrhea. This randomized, double-blind trial was conducted in Guadalajara, Mexico, during the summers of 1999-2001 to compare azithromycin with levofloxacin for the treatment of travelers' diarrhea. A total of 217 US adults were randomized to receive a single oral dose of azithromycin (1000 mg; 108 persons) or levofloxacin (500 mg; 109 persons), with a follow-up period of 4 days. Three patients in each group dropped out of the study. The median time between initiation of therapy and passage of the last unformed stool (azithromycin group, 22.3 h; levofloxacin group, 21.5 h) and the number of unformed stools passed during the 4-day follow-up period (azithromycin group, 6.5; levofloxacin group, 5.5) were similar. Treatment failure occurred in 10 patients (9.5%) receiving azithromycin and 8 patients (7.5%) receiving levofloxacin. Possible minor, self-limiting adverse events occurred in 57 patients in each treatment group. Azithromycin was found to be a safe and effective alternative to levofloxacin for the treatment of acute travelers' diarrhea in US adult travelers to Mexico.


Use of lux genes in applied biochemistry.
Bioluminescence has emerged in the last decade as a major tool for the study of bacterial adaptation and survival. In addition to the advantages of sensitivity and the real-time, non-invasive nature of this reporter, the imaging potential of using low-light and photon-counting video cameras has been particularly influential in establishing its ascendancy-over more traditional reporter systems. This review provides a reflection of personal activity in this field through applications in Food Microbiology and collaboration with colleagues both in the UK and beyond.


Identification and characteristics of extracellular vesicles from bovine blastocysts produced in vitro.
Extracellular vesicles (EVs) have been identified within different body fluids and cell culture media. However, there is very little information on the secretion of these vesicles during early embryonic development. The aims of this work were first to demonstrate the secretion of extracellular vesicles by pre-implantation bovine embryos and second to identify and characterize the population of EVs secreted by bovine blastocysts during the period from day seven to nine of embryo culture and its correlation with further embryo development up to day 11. Bovine embryos were produced by in vitro fertilization (IVF) or parthenogenetic activation (PA) and cultured until blastocyst stage. Blastocyst selection was performed at day 7 post IVF/PA considering two variables: stage of development and quality of embryos. Selected blastocysts were cultured in vitro for 48 hours in groups (exp. 1) or individually (exp. 2) in SOF media depleted of exosomes. At day 9 post IVF/PA the media was collected and EVs isolated by ultracentrifugation. Transmission electron microscopy revealed the presence of heterogeneous vesicles of different sizes and population: microvesicles (MVs) and exosomes (EXs) of rounded shape, enclosed by a lipid bi-layer and ranging from 30 to 385 nm of diameter. Flow cytometry analysis allowed identifying CD63 and CD9 proteins as exosome markers. Nanoparticle tracking analysis generated a large number of variables, which required the use of multivariate statistics. The results indicated that the concentration of vesicles is higher in those blastocysts with arrested development from day 9 up to day 11 of in vitro development (6.7 x 108 particles/ml) derived from IVF (p <0.05), compared to PA blastocysts (4.7 x 108 particles/ml). Likewise, the profile (concentration and diameter) of particles secreted by embryos derived from IVF were different from those secreted by PA embryos. In conclusion, we demonstrated that bovine blastocysts secrete MVs/EXs to the culture media. Data suggest that characteristics of the population of EVs vary depending on embryo competence.


Discovery of GPCR ligands for probing signal transduction pathways.
G protein-coupled receptors (GPCRs) are seven integral transmembrane proteins that are the primary targets of almost 30% of approved drugs and continue to represent a major focus of pharmaceutical research. All of GPCR targeted medicines were discovered by classical medicinal chemistry approaches. After the first GPCR crystal structures were determined, the docking screens using these structures lead to discovery of more novel and potent ligands. There are over 360 pharmaceutically relevant GPCRs in the human genome and to date about only 30 of structures have been determined. For these reasons, computational techniques such as homology modeling and molecular dynamics simulations have proven their usefulness to explore the structure and function of GPCRs. Furthermore, structure-based drug design and in silico screening (High Throughput Docking) are still the most common computational procedures in GPCRs drug discovery. Moreover, ligand-based methods such as three-dimensional quantitative structure-selectivity relationships, are the ideal molecular modeling approaches to rationalize the activity of tested GPCR ligands and identify novel GPCR ligands. In this review, we discuss the most recent advances for the computational approaches to effectively guide selectivity and affinity of ligands. We also describe novel approaches in medicinal chemistry, such as the development of biased agonists, allosteric modulators, and bivalent ligands for class A GPCRs. Furthermore, we highlight some knockout mice models in discovering biased signaling selectivity.


C-terminal deletions of Merkel cell polyomavirus large T-antigen, a highly specific surrogate marker for virally induced malignancy.
In 67-100% of cutaneous Merkel cell carcinomas (MCC) the Merkel cell polyomavirus (MCPyV) integrates into the host genome. Mutations and deletions truncating the C-terminal helicase domain of the T-antigen (TAg) protein have been detected in these MCCs, but not in healthy skin specimens. C-terminal deletions of the TAg nucleic acid sequences are characteristic for about 38% of these cases. While the association of MCPyV with MCC has been proven, it is unknown whether MCPyV may play a similar role in other tumor entities. We describe in detail the development and validation of a novel Merkel cell polyomavirus TAg C-terminus deletion assay (MCPyV ΔC-TAg). The triplex real-time PCR quantifies the N- and C-terminal part of the MCPyV TAg gene and the cellular β-globin gene. By comparing the copy numbers of the N- and C-terminal part, deletions of the MCPyV TAg C-terminus are rapidly identified. MCPyV ΔC-TAg was used to assess the physical state of MCPyV TAg in a large series of 55 MCCs, 15 cutaneous lymphomas and 47 forehead smears of healthy individuals. Neither DNA positivity nor viral load was able to discriminate MCCs from the other different types of samples. However, deleted TAg C-terminus sequences were detected only in MCPyV positive MCCs (39%). Consequently, detection of deleted C-terminal TAg sequences appears to be a highly specific surrogate marker for virally induced malignancy and should be used to support novel assumed MCPyV-tumor associations. The study further supports the notion that MCPyV does not play a role in cutaneous lymphoma pathogenesis.


The Effects of Transcutaneous and Intraoral Low-Level Laser Therapy After Extraction of Lower Third Molars: A Randomized Single Blind, Placebo Controlled Dual-Center Study.
The surgical removal of impacted third molars is one of the most common procedures performed by oral and maxillofacial surgeons. The purpose of this study is to determine whether either transcutaneous or intraoral low-level laser therapy (LLLT) reduces postoperative pain and assists in the healing of mandibular third molar extraction. This randomized, placebo controlled, single-blind, split-mouth design study was conducted on 60 patients with full bony impacted similar position mandibular third molars bilaterally. The patients were divided into two groups of 30 each: transcutaneous LLLT and intraoral LLLT and the other side of each group treated with nonactive laser (60 teeth). The laser treatment consisted of administering laser energy immediately before and after the extraction procedure with gallium aluminum arsenide (GaAlAs) 830 nm diode lasers. Postoperative pain and healing of the sockets were compared in transcutaneous and intraoral group with placebo for 1 week following the extraction. Descriptive and bivariate statistics was computed, and the p-value was set at 0.05. Intraoral LLLT application resulted in a statistically significant reduction of postoperative pain in comparison with transcutaneous laser group and placebo. The unhealed socket numbers were compared in two groups at seventh day and no differences were observed. The results of this study suggest that single-session intraoral LLLT is more effective than extraoral application for reducing postoperative pain. It was postulated that the differences between skin and mucosa could have effect on the results. Although intraoral use would allow closer application to the surgical site, the size of some laser devices precludes their use intraorally.


Polyamines directly promote antizyme-mediated degradation of ornithine decarboxylase by the proteasome.
Ornithine decarboxylase (ODC), a ubiquitin-independent substrate of the proteasome, is a homodimeric protein with a rate-limiting function in polyamine biosynthesis. Polyamines regulate ODC levels by a feedback mechanism mediated by ODC antizyme (OAZ). Higher cellular polyamine levels trigger the synthesis of OAZ and also inhibit its ubiquitin-dependent proteasomal degradation. OAZ binds ODC monomers and targets them to the proteasome. Here, we report that polyamines, aside from their role in the control of OAZ synthesis and stability, directly enhance OAZ-mediated ODC degradation by the proteasome. Using a stable mutant of OAZ, we show that polyamines promote ODC degradation in Saccharomyces cerevisiae cells even when OAZ levels are not changed. Furthermore, polyamines stimulated the in vitro degradation of ODC by the proteasome in a reconstituted system using purified components. In these assays, spermine shows a greater effect than spermidine. By contrast, polyamines do not have any stimulatory effect on the degradation of ubiquitin-dependent substrates.


Molecular Simulations of the Hydration Behavior of a Zwitterion Brush Array and Its Antifouling Property in an Aqueous Environment.
We carried out umbrella sampling and molecular dynamics (MD) simulations to investigate molecular interactions between sulfobetaine zwitterions or between sulfobetaine brushes in different media. Simulation results show that it is more energetically favorable for the two sulfobetaine zwitterions or brushes to be fully hydrated in aqueous solutions than in vacuum where strong ion pairs are formed. Structural properties of the hydrated sulfobetaine brush array and its antifouling behavior against a foulant gel are subsequently studied through steered MD simulations. We find that sulfobetaine brush arrays with different grafting densities have different structures and antifouling mechanisms. At a comparably higher grafting density, the sulfobetaine brush array exhibits a more organized structure which can hold a tightly bound hydration water layer at the interface. Compression of this hydration layer results in a strong repulsive force. However, at a comparably lower grafting density, the brush array exhibits a randomly oriented structure in which the antifouling of the brush array is through the deformation of the sulfobetaine branches.


Classification and application of cementation alternatives.
More important than any resin cement is the marginal fit and retentive nature of the ceramic restoration to be bonded. Cementation should not be intended to compensate for poor dentistry, but to assist in producing superior aesthetic results. Responding to the demand for all-ceramic restorations, dental manufacturers continue to refine the resin cements used in luting for this modality. The ideal resin cement for ceramic restorations must demonstrate low viscosity; easy loading; a film thickness which facilitates complete seating high strength; and excellent wear resistance. This article reviews the available resin cements, with particular focus on a new type of initiator and stabilizer for light- and dual-cured resin cements.


Alterations of epidermal lectin binding sites in acute contact dermatitis.
We investigated alterations of epidermal lectin binding sites, as well as of pemphigus and bullous pemphigoid antigens, in 28 human patch test reactions, both allergic (nickel, formaldehyde, N,N'-1,3-dimethylbutyl-N'-phenylenediamine) and irritant (sodium lauryl sulfate). The epidermal reactivity to a panel of lectins and human antisera to pemphigus vulgaris and bullous pemphigoid antigens was compared with samples obtained from normal skin and from skin under tape occlusion. We observed selective perturbations of lectin and antibody binding in acute contact dermatitis, whether allergic or irritant. The main findings were a loss of terminal sialic acids and longer bi- and triantennary mannosyl residues as well as a loss of pemphigus vulgaris antigen. The only difference between allergic and irritant patch test reactions was in topography of loss of WGA binding sites: in the former, it was most pronounced in the lower and middle epidermis, whereas in the latter it was seen in the uppermost subcorneal layers. Our findings support a common pathway of cell membrane alterations of keratinocytes in acute contact dermatitis.


KCNQ1 Gene Variants in Large Asymptomatic Populations: Considerations for Genomic Screening of Military Cohorts.
The advances in genomic technology of large populations make the potential for genomic screening of military cohorts and recruits feasible, affording the potential to identify at-risk individuals before occurrence of potentially life-threatening events. Exploring sudden cardiac death, known to cause significant morbidity and mortality in young military service members, we focused on the most common gene associated with long QT syndrome (LQTS), KCNQ1. Using the publicly available database Exome Aggregation Consortium as a surrogate for a military population, variants in KCNQ1 were filtered on the basis of population prevalence, classification as a disease mutation in the Human Gene Mutation database, and classification as pathogenic or likely pathogenic in the ClinVar database. Variant prevalence and penetrance estimates were derived using reports from the medical literature. We showed that in a population of over 60,000 individuals, at least 97 (0.2%) individuals would harbor a potentially pathogenic mutation in KCNQ1, which is more prevalent than expected on the basis of current medical literature (p = 0.0004). KCNQ1 variant penetrance was estimated to be only 9% to 17%. Identifying the importance of large genomic studies, our study demonstrates that 46% of pathogenic mutations in KCNQ1 had a population frequency of less than 1:50,000. Screening a large database with genomic screening for a condition that is relevant to active duty service members results in the identification of many individuals with potentially pathogenic mutations in the KCNQ1 gene, which has profound implications for screening military or other adult cohorts in terms of over diagnosis, overtreatment, and increased medical resource usage. This study of KCNQ1 provides a platform for consideration of other genes that cause sudden cardiac death as well as other medically actionable hereditary disorders for which genomic screening is available. We review the potential benefits of genomic screening and also present the complex hurdles that will be encountered as such technologies unfold.


Safety and effectiveness of conventional systemic therapy and biological drugs in patients with moderate to severe psoriasis and HIV infection: a retrospective multicenter study.
Background: The management of HIV-positive patients with psoriasis is controversial and limited to individual cases or short series of patients. Objectives: To evaluate the safety and effectiveness of conventional and biologic immunosuppressive drugs in the treatment of patients with psoriasis and concomitant HIV infection. Methods: A retrospective multicenter study was conducted. The study included data from 2008 to 2016. Inclusion criteria were: HIV adult patients with moderate-to-severe psoriasis, HIV viral load determinations at baseline and at least after 6 months of treatment, and systemic immunosuppressive treatment for at least 6 months. A descriptive analysis was performed. Results: Twenty-three patients with plaque-type psoriasis and HIV infection (five with AIDS) were included. Median follow-up time was 3.2 years. The main drugs used were etanercept, methotrexate, and ustekinumab. In most cases, viral load and CD4 cell count not only remained stable but also improved throughout the follow-up. Six patients presented severe adverse events during the follow-up, four of them in the AIDS stage. At the end of the follow-up period, 76.5% of the patients had achieved a PASI 75. Conclusion: Biologic drugs, both anti-TNF alpha agents and ustekinumab, seem to have an acceptable safety profile and high effectiveness in HIV-positive patients.


Birth registration coverage according to the sex of the head of household: an analysis of national surveys from 93 low- and middle-income countries.
Within-country inequalities in birth registration coverage (BRC) have been documented according to wealth, place of residence and other household characteristics. We investigated whether sex of the head of household was associated with BRC. Using data from nationally-representative surveys (Demographic and Health Survey or Multiple Indicator Cluster Survey) from 93 low and middle-income countries (LMICs) carried out in 2010 or later, we developed a typology including three main types of households: male-headed (MHH) and female-led with or without an adult male resident. Using Poisson regression, we compared BRC for children aged less than 12 months living the three types of households within each country, and then pooled results for all countries. Analyses were also adjusted for household wealth quintiles, maternal education and urban-rural residence. BRC ranged from 2.2% Ethiopia to 100% in Thailand (median 79%) while the proportion of MHH ranged from 52.1% in Ukraine to 98.3% in Afghanistan (median 72.9%). In most countries the proportion of poor families was highest in FHH (no male) and lowest in FHH (any male), with MHH occupying an intermediate position. Of the 93 countries, in the adjusted analyses, FHH (no male) had significantly higher BRC than MHH in 13 countries, while in eight countries the opposite trend was observed. The pooled analyses showed t BRC ratios of 1.01 (95% CI: 1.00; 1.01) for FHH (any male) relative to MHH, and also 1.01 (95% CI: 1.00; 1.01) for FHH (no male) relative to MHH. These analyses also showed a high degree of heterogeneity among countries. Sex of the head of household was not consistently associated with BRC in the pooled analyses but noteworthy differences in different directions were found in specific countries. Formal and informal benefits to FHH (no male), as well as women's ability to allocate household resources to their children in FHH, may explain why this vulnerable group has managed to offset a potential disadvantage to their children.


The activity of the carbamoyl phosphate synthase 1 promoter in human liver-derived cells is dependent on hepatocyte nuclear factor 3-beta.
Carbamoyl phosphate synthase 1 (CPS1) is the rate-limiting enzyme in the first step of the urea cycle and an indispensable enzyme in the metabolism of human liver. However, CPS1 epigenetic regulation involves promoter analysis and the role of liver-enriched transcription factors (LETFs), which is not fully elucidated. In this work, the promoter region of hCPS1 gene was cloned, and its activity was investigated. An LETF, hepatocyte nuclear factor 3-beta (HNF3β), was found to promote the transcriptional expression of CPS1 in liver-derived cell lines. In addition, dual-luciferase reporter assay shows that the essential binding sites of the HNF3β may exist in the oligonucleotide -70 nt to +73 nt. Two putative binding sites are available for HNF3β. Mutation analysis results show that the binding site 2 of HNF3β was effective, and the transcriptional activity of CPS1 promoter significantly decreased after mutation. Electrophoretic mobile shift assay (EMSA) and ChIP assay confirmed that HNF3β can interact with the binding site in the CPS1 promoter region of -70 nt to +73 nt promoter region in vivo and in vitro to regulate the transcription of CPS1. Moreover, HNF3β overexpression enhanced the transcription of CPS1 and consequently improved the mRNA and protein levels of CPS1, whereas the knockdown of HNF3β showed the opposite effects. Finally, urea production in cells was measured, and ammonia detoxification improved significantly in cells after transfection with HNF3β. HNF3β plays a vital role in regulation of CPS1 gene and could promote the metabolism of ammonia by regulating CPS1 expression.


[Alpha-fetoprotein in the serum of puerperae].
Alpha-fetoprotein was determined in the sera of parturients with normal pregnancy and delivery, parturients with preterm delivery and with preeclampsia. Alpha-fetoprotein was determined by the plates of Behring Institute. The authors established that after delivery alpha-fetoprotein did not differ substantially in the separate groups. Furthermore till the eight day after delivery the reduction of alpha-fetoprotein was minimal with the exception in the parturients with preterm delivery.


Oncologic Outcome and Morbidity in the Elderly Rectal Cancer Patients After Preoperative Chemoradiotherapy and Total Mesorectal Excision: A Multi-institutional and Case-matched Control Study.
To determine the toxicity and oncologic outcome of neoadjuvant chemoradiotherapy (CRT) followed by curative total mesorectal excision (TME) in the elderly (≥70 yrs) and younger (<70 yrs) rectal cancer patients. Sufficient data for elderly rectal cancer patients who received definitive trimodality have not been accumulated yet. A total of 1232 rectal cancer patients who received neoadjuvant CRT and TME were enrolled in this study. After propensity-score matching, 310 younger patients and 310 elderly patients were matched with 1:1 manner. Treatment response, toxicity, surgical outcome, recurrence, and survival were assessed and compared between the 2 groups of patients. The median age was 58 years for the younger patient group and 74 years for the elderly group. Pathologic complete response rates were not significantly different between the 2 groups (younger and elderly: 17.1% vs 14.8%, P = 0.443). The 5-year recurrence-free survival (younger and elderly: 67.7% vs 65.5%, P = 0.483) and overall survival (younger and elderly: 82.9% vs. 79.5%, P = 0.271) rates were not significantly different between the 2 groups either. Adjuvant chemotherapy after surgery was less frequently delivered to the elderly than that to younger patients (83.9% vs 69.0%). Grade 3 or higher acute hematologic toxicity was observed more frequently in the elderly than that in the younger group (9.0% vs 16.1%, P = 0.008). Late complication rate was higher in the elderly group compared with that in the younger group without statistical significance (2.6% vs 4.5%, P = 0.193). Although acute hematologic toxicity was observed more frequently in the elderly patients than that in the younger patients, elderly rectal cancer patients with good performance status who received preoperative CRT and TME showed favorable tumor response and recurrence-free survival similar to younger patients.


Protective effect of c-Myc/Rab7a signal pathway in glioblastoma cells under hypoxia.
Glioblastoma multiforme (GBM) is the most common primary brain tumor, and is associated with a poor prognosis. Hypoxia is prevalent in the microenvironment of GBM, and promotes tumorigenesis and resistance to anticancer therapy. However, its mechanism remains incompletely understood. We used immunohistochemistry, quantitative real-time PCR, and Western blots to assess c-Myc and Rab7a expression levels in 12 GBM specimens from a single institution. A luciferase reporter assay was conducted to confirmed whether Rab7a is transcriptionally regulated by c-Myc. To clarify the precise role of c-Myc/Rab7a on GBM cell proliferation, we did in vitro and in vivo analyses with lentivirus vectors. Cell viability was assessed using a cell counting kit-8 assay in the context of hypoxia. Autophagy was measured using transmission electron microscopy and Western blot, and apoptosis was measured using flow cytometry and Western blot. Gene and protein expression of c-Myc and Rab7a were significantly upregulated in GBM specimens. Moreover, c-Myc regulated Rab7a by specifically interacting with the Rab7a promoter. Furthermore, hypoxia activated the c-Myc/Rab7a pathway, which protects GBM cells from damage caused by hypoxia. Importantly, c-Myc/Rab7a inhibited apoptosis and induced autophagy in vitro and in vivo. Collectively, our results suggest that the c-Myc/Rab7a pathway protects GBM cells from hypoxic injury via regulation of apoptosis and autophagy, contributing to the growth of GBM.


14-Benzoyl-mesaconine hydro-chloride methanol monosolvate.
The title compound, C(31)H(44)N(3)O(10) (+)·Cl(-)·CH(4)O, is the methanol solvate of 8-benzo-yloxy-,9,11,11a-tetra-hydroxy-6,10,13-trimeth-oxy-3-meth-oxy-methyl-1-methyl-tetra-deca-hydro-1H-3,6a,12-(epiethane-1,1,2-tri-yl)-7,9-methanona-phtho[2,3-b]azocin-1-ium chloride, the amine-protonated hydro-chloride of 14-benzoyl-mesaconine hydro-chloride. The cation has an aconitine carbon skeleton with four six-membered rings of which three display chair conformations and one a boat conformation, and two five-membered rings with envelope conformations. In the crystal, the components are connected into an infinite chain by inter- and intra-molecular O-H⋯O, N-H⋯O and O-H⋯Cl hydrogen bonds.


HYDROXYUREA: MECHANISM OF ACTION.
Acetohydroxamic acid has been identified, by paper chromatography, in the blood of three patients with chronic myelogenous leukemia on hydroxyurea therapy. This suggests that the drug is hydrolyzed yielding hyroxylamine, which then cleaves thioesters, in particular acetyl-coenzyme A.


The battered child syndrome.
This is a report of socio-medical aspects of 23 episodes of physical abuse among 21 children, and findings include one death, a predominant age below two years, no unusual race or sex distribution, a high incidence of head trauma, skin lesions and bone changes, with a common history of previous abuse in the patient and siblings. Intact families had few children in the house. Parents were usually young, had a high incidence of illegitimate births, mental disease and previous criminal records. Abuse was often inflicted by a close relative, usually the mother. Incidence and severity of physical abuse in California and participation in therapy are discussed.


Diffuse large B cell lymphoma within an anal fistula: a case report with an intriguing possible aetiology.
Described is a case of diffuse large B cell lymphoma that presented within a typical fistula tract, possibly secondary to oxidative stress within the fistula tract itself and consequent malignant change rather than a fistula as a consequence of necrosis in a lymphoma. If so it would be unique in the world literature. A 42-year-old fitness instructor presented with a typical appearing left lateral anal fistula. Biopsy of the fistulous tract revealed B cell lymphoma, graded 1E. Although chemotherapy cured the lymphoma, surgical treatment by ligation of the inter-sphincteric fistula tract was required to heal the fistula. At 3-year follow-up, there has been no recurrence of the lymphoma or the fistula. Neoplasia arising secondary to oxidative stress within an anal fistula is a well-established phenomenon. Early diagnosis of rare conditions associated with anal fistula can only be accomplished by routine biopsy of every fistula tract.


Analysis of the craniofacial skeleton in cleft children with otitis media with effusion.
Various aspects have been proposed as the cause of otitis media with effusion (OME) in cleft lip and/or palate (CL/P) populations (i.e. abnormal anatomic relation of Eustachian tube (ET) musculature and soft palate; the lack of intact palatal partition; reduction of nasal patency or chronic rhinitis; timing and type of surgery). However, the role of deviated craniofacial skeleton (CFS) in CL/P has been neglected, although the role of the craniofacial development on poor ET function has been advocated in non-cleft children. In this study, we evaluated clinical and cephalometric data of 37 Japanese children with unilateral complete cleft lip and palate (UCLP, 25) or isolated cleft palate (ICP, 12) and compared them to 40 non-cleft children, who were proportionally matched for age and sex. Data showed that OME was more often in children with unilateral cleft lip and palate-UCLP (76%) and those with isolated cleft palate-ICP (67%) than non-clefts (10.00%). In addition to a number of skeletal abnormalities (i.e. short dimensions related to the posterior cranial base and backward and upward position of the maxilla) detected in CL/P cases, mastoid depth and height were also shorter in cleft cases than normal subjects. On the other hand, a small tendency to recurrent upper airway infection (RUAI) was observed in cleft cases with OME. Further, it was found that the following differences in the mastoid-middle ear-Eustachian tube (M-ME-ET) system were associated with a tendency to OME in UCLP cases: more horizontal ET in relation to the posterior cranial base; short bony ET; short height and antero-posterior depth of the mastoid air cell system.


Epigenetic markers of renal function in african americans.
Chronic kidney disease (CKD) is an increasing concern in the United States due to its rapidly rising prevalence, particularly among African Americans. Epigenetic DNA methylation markers are becoming important biomarkers of chronic diseases such as CKD. To better understand how these methylation markers play a role in kidney function, we measured 26,428 DNA methylation sites in 972 African Americans from the Genetic Epidemiology Network of Arteriopathy (GENOA) study. We then evaluated (1) whether epigenetic markers are associated with estimated glomerular filtration rate (eGFR), (2) whether the significantly associated markers are also associated with traditional risk factors and/or novel biomarkers for eGFR, and (3) how much additional variation in eGFR is explained by epigenetic markers beyond established risk factors and biomarkers. The majority of methylation markers most significantly associated with eGFR (24 out of the top 30) appeared to function, at least in part, through pathways related to aging, inflammation, or cholesterol. However, six epigenetic markers were still able to significantly predict eGFR after adjustment for other risk factors. This work shows that epigenetic markers may offer valuable new insight into the complex pathophysiology of CKD in African Americans.


Structural basis for intramolecular interaction of post-translationally modified H-Ras•GTP prepared by protein ligation.
Ras undergoes post-translational modifications including farnesylation, proteolysis, and carboxymethylation at the C terminus, which are necessary for membrane recruitment and effector recognition. Full activation of c-Raf-1 requires cooperative interaction of the farnesylated C terminus and the activator region of Ras with its cysteine-rich domain (CRD). However, the molecular basis for this interaction remains unclear because of difficulties in preparing modified Ras in amounts sufficient for structural studies. Here, we use Sortase A-catalyzed protein ligation to prepare modified Ras in sufficient amounts for NMR and X-ray crystallographic analyses. The results show that the farnesylated C terminus establishes an intramolecular interaction with the catalytic domain and brings the farnesyl moiety to the proximity of the activator region, which may be responsible for their cooperative recognition of c-Raf-1-CRD.


The use of plant community attributes to detect habitat quality in coastal environments.
The monitoring of biodiversity has mainly focused on the species level. However, researchers and land managers are making increasing use of complementary assessment tools that address higher levels of biological organization, i.e. communities, habitats and ecosystems. Recently, a variety of frameworks have been proposed for assessing the conservation status of communities or ecosystems. Among the various criteria proposed, all the protocols suggest considering (i) spatial aspects (range and area), and (ii) qualitative aspects of specific structures and functions. However, changes to ecological function are difficult to quantify and many protocols end up by using qualitative criteria. The aim of this work was to test the efficacy of some plant community attributes for the detection of vegetation quality in sand dune plant communities. We chose plant community attributes that either help to distinguish a habitat from others (diagnostic components) or play a significant role in habitat function and persistence over time. We used a diachronic approach by contrasting up-to-date vegetation data with data from previous studies carried out within the same areas. Changes in species composition were detected through detrended correspondence analyses (detrended correspondence analyses), Multi-Response Permutation Procedures and Indicator Species Analysis, while structural changes were analyzed by comparing species richness, total species cover, ecological groups of species and growth forms through null models. Ecological groups such as native focal species and aliens, and growth forms proved their efficacy in discriminating between habitat types and in describing their changes over time. The approach used in this study may provide an instrument for the assessment of plant community quality that can be applied to other coastal ecosystems.


Increased metaboreflex activity is related to exercise intolerance in heart transplant patients.
Heart transplantation does not normalize exercise capacity or the ventilatory response to exercise. We hypothesized that excessive muscle reflex activity, as assessed by the muscle sympathetic nerve activity (MSNA) response to handgrip exercise, persists after cardiac transplantation and that this mechanism is related to exercise hyperpnea in heart transplant recipients (HTRs). We determined the MSNA, ventilatory, and cardiovascular responses to isometric and dynamic handgrips in 11 HTRs and 10 matched control subjects. Handgrips were followed by a post-handgrip ischemia to isolate the metaboreflex contribution to exercise responses. HTRs and control subjects also underwent recordings during isocapnic hypoxia and a maximal, symptom-limited, cycle ergometer exercise test. HTRs had higher resting MSNA (P < 0.01) and heart rate (P < 0.01) than the control subjects. Isometric handgrip increased MSNA in HTRs more than in the controls (P = 0.003). Dynamic handgrip increased MSNA only in HTRs. During post-handgrip ischemia, MSNA and ventilation remained more elevated in HTRs (P < 0.05). The MSNA and ventilatory responses to hypoxia were also higher in HTRs (both P < 0.04). In HTRs, metaboreflex overactivity was related to the ventilatory response to exercise, characterized by the regression slope relating ventilation to CO(2) output (r = +0.8; P < 0.05) and a lower peak ventilation (r = +0.81; P < 0.05) during cycle ergometer exercise tests. However, increased chemoreflex sensitivity (r = +0.91; P < 0.005), but not metaboreflex activity, accounted for the lower peak ventilation during exercise in a stepwise regression analysis. In conclusion, heart transplantation does not normalize muscle metaboreceptor activity; both increased metaboreflex and chemoreflex control are related to exercise intolerance in HTRs.


Developmental effects of intrauterine growth retardation on cerebral amino acid transport.
Early restriction of nutrients during the perinatal period of life can modify the development of the mammalian fetus and have marked repercussions on the ontogeny of the CNS. The brain is vulnerable to undernutrition, with delayed morphologic and biochemical maturation leading to impaired functions. The aim of the present investigation was to assess whether modified brain neurotransmitter and amino acid concentrations found in an animal model of intrauterine growth retardation were related to modified blood-brain amino acid transport properties. Four amino acids were tested: alanine and taurine, plus two neurotransmitter precursors, tryptophan and tyrosine. Intrauterine growth retardation was induced by restriction of maternal-fetal blood flow from the 17th d of gestation. Blood-brain transport of these amino acids was measured by i.v. injection of radiolabeled amino acids in 7-d-old, 21-d-old, and 60-d-old intrauterine growth-retarded or control rats. No major statistical differences were revealed either for brain regional transport or between intrauterine growth-retarded animals and controls at any age studied. Transfer coefficients and influxes remained statistically similar for almost all brain regions in both groups. A significant decrease and different time course for amino acid transport with age related to the blood-brain barrier maturation are confirmed in this model. Our results are related to a major role of the blood-brain barrier as a part of mechanisms leading to "brain growth sparing."


Single-base substitutions in the CHM promoter as a cause of choroideremia.
Although over 150 unique mutations affecting the coding sequence of CHM have been identified in patients with the X-linked chorioretinal disease choroideremia (CHM), no regulatory mutations have been reported, and indeed the promoter has not been defined. Here, we describe two independent families affected by CHM bearing a mutation outside the gene's coding region at position c.-98: C>A and C>T, which segregated with the disease. The male proband of family 1 was found to lack CHM mRNA and its gene product Rab escort protein 1, whereas whole-genome sequencing of an affected male in family 2 excluded the involvement of any other known retinal genes. Both mutations abrogated luciferase activity when inserted into a reporter construct, and by further employing the luciferase reporter system to assay sequences 5' to the gene, we identified the CHM promoter as the region encompassing nucleotides c.-119 to c.-76. These findings suggest that the CHM promoter region should be examined in patients with CHM who lack coding sequence mutations, and reveals, for the first time, features of the gene's regulation.


Two new skinks (Scincidae: Glaphyromorphus) from rainforest habitats in north-eastern Australia.
Tropical rainforest is largely restricted in Australia to the fairly continuous Wet Tropics region and disconnected patches to the north on Cape York. The Wet Tropics is relatively well explored and studied, whereas the rainforests of Cape York have received less attention due to their remoteness. Here we describe two new species of Glaphyromorphus skinks from rainforest areas on Cape York. The two new species are most similar to each other and to G. fuscicaudis and G. nigricaudis, but both are readily diagnosed on numerous traits. Glaphyromorphus othelarrni sp. nov. is diagnosed from all similar species by its supralabial count (typically 8 vs 7), high number of subdigital lamellae beneath the 4th finger (14-15 vs < 14), and its relatively longer limbs. Glaphyromorphus nyanchupinta sp. nov. is diagnosed from all similar species by its small body size (max SVL = ~ 54 mm vs > 85 mm) and slender body shape, low number of subdigital lamellae beneath the 4th toe (17-20 vs generally 20 or more), and head and body pattern. Both species also differ from each other and similar congeners in other aspects of body shape, scalation and colour pattern. Glaphyromorphus othelarrni sp. nov. is restricted to boulder-strewn rainforest of the Melville Range, whilst Glaphyromorphus nyanchupinta sp. nov. is known only from upland rainforest in the McIlwraith Range. We discuss patterns of rainforest vertebrate endemism on Cape York, and the importance of lithorefugia in generating these.


Increased Risk of Rheumatoid Arthritis in Patients With Asthma: A Genetic Association Study Using Two-Sample Mendelian Randomization Analysis.
Observational studies have explored the association between asthma and some types of arthritis, including rheumatoid arthritis and osteoarthritis, but the results are largely contradictory. We aimed to investigate the causal effects of asthma on arthritis, including osteoarthritis, rheumatoid arthritis, gout, and ankylosing spondylitis. Two-sample Mendelian randomization (MR) analysis was used to investigate the causal effects of asthma on each arthritis. The genetic instruments for asthma were obtained from a large genome-wide association study of asthma. The inverse-variance weighted (IVW) method was used as the main analysis of MR. Bonferroni-adjusted P value threshold was used to account for multiple comparisons. MR-IVW analysis suggested that adult-onset asthma (AOA) was associated with increased risk of rheumatoid arthritis. The odds ratio for rheumatoid arthritis associated with AOA and childhood-onset asthma (COA) were 1.018 (95% confidence interval [95% CI], 1.011-1.025; P < 0.001) and 1.006 (95% CI 1.001-1.012; P = 0.046), respectively. For osteoarthritis, gout, or ankylosing spondylitis, all the MR analyses showed no significant causal effects of AOA or COA on them. We also performed a reverse MR analysis to explore the causal effects of rheumatoid on all asthma, allergic asthma, or nonallergic asthma and found no significant causal effects on them. Genetically predicted AOA predisposes patients to an increased risk of rheumatoid arthritis but has no causal effects on osteoarthritis, gout, and ankylosing spondylitis. The result of COA on rheumatoid arthritis is suggestive of potential causal relationship but needs to be confirmed in further studies.


Evolutionary Phase Transitions in Random Environments.
We present analytical results for long-term growth rates of structured populations in randomly fluctuating environments, which we apply to predict how cellular response networks evolve. We show that networks which respond rapidly to a stimulus will evolve phenotypic memory exclusively under random (i.e., nonperiodic) environments. We identify the evolutionary phase diagram for simple response networks, which we show can exhibit both continuous and discontinuous transitions. Our approach enables exact analysis of diverse evolutionary systems, from viral epidemics to emergence of drug resistance.


Reciprocal regulation of β2-adrenoceptor-activated cAMP response-element binding protein signalling by arrestin2 and arrestin3.
Activation of Gs coupled receptors (e.g. β2-adrenoreceptor (β2AR)) expressed within the uterine muscle layer (myometrium), promotes intracellular cAMP generation, inducing muscle relaxation through short-term inhibition of contractile proteins, and longer-term modulation of cellular phenotype to promote quiescence. In the myometrium cAMP-driven modulation of cell phenotype is facilitated by CREB activity, however despite the importance of CREB signalling in the promotion of myometrial quiescence during pregnancy, little is currently known regarding the molecular mechanisms involved. Thus, we have characterised β-adrenoceptor-stimulated CREB signalling in the immortalised ULTR human myometrial cell line. The non-selective β-adrenoceptor agonist isoprenaline induced time- and concentration-dependent CREB phosphorylation, which was abolished by the β2AR selective antagonist ICI118,551. β2AR-stimulated CREB phosphorylation was mediated through a short-term PKA-dependent phase, and longer-term Src/p38 MAPK-dependent/PKA-independent phase. Since in model cells, arrestin2 can facilitate β2AR-mediated Src/p38 recruitment, we examined whether CREB signalling was activated through a similar process in myometrial cells. Depletion of arrestin2 attenuated p38 phosphorylation, whilst arrestin3 depletion enhanced and prolonged isoprenaline-stimulated p38 signals, which was reversed following inhibition of Src. Knockdown of arrestin2 led to enhanced short-term (up to 10min), and attenuated longer-term (>10min) isoprenaline-stimulated CREB phosphorylation. Contrastingly, removal of arrestin3 enhanced and prolonged isoprenaline-stimulated CREB phosphorylation, whilst depletion of both arrestins abolished CREB signals at time points >5min. In summary, we have delineated the molecular mechanisms coupling β2AR activity to CREB signalling in ULTR myometrial cells, revealing a biphasic activation process encompassing short-term PKA-dependent, and prolonged Src/arrestin2/p38-dependent components. Indeed, our data highlight a novel arrestin-mediated modulation of CREB signalling, suggesting a reciprocal relationship between arrestin2 and arrestin3, wherein recruitment of arrestin3 restricts the ability of β2AR to activate prolonged CREB phosphorylation by precluding recruitment of an arrestin2/Src/p38 complex.


Disentangling 1/f noise from confined ion dynamics.
Ion transport through biological and solid-state nanochannels is known to be a highly noisy process. The power spectrum of current fluctuations is empirically known to scale like the inverse of frequency, following the long-standing yet poorly understood Hooge's law. Here, we report measurements of current fluctuations across nanometer-scale two-dimensional channels with different surface properties. The structure of fluctuations is found to depend on the channel's material. While in pristine channels current fluctuations scale like 1/f1+a with a = 0-0.5, the noise power spectrum of activated graphite channels displays different regimes depending on frequency. Based on these observations, we develop a theoretical formalism directly linking ion dynamics and current fluctuations. We predict that the noise power spectrum takes the form 1/f × Schannel(f), where 1/f fluctuations emerge in fluidic reservoirs on both sides of the channel and Schannel describes fluctuations inside it. Deviations to Hooge's law thus allow direct access to the ion transport dynamics of the channel - explaining the entire phenomenology observed in experiments on 2D nanochannels. Our results demonstrate how current fluctuations can be used to characterize nanoscale ion dynamics.


Identification of novel inhibitors for TNFα, TNFR1 and TNFα-TNFR1 complex using pharmacophore-based approaches.
Tumor necrosis factor α (TNFα) is a multifunctional cytokine with a potent pro-inflammatory effect. It is a validated therapeutic target molecule for several disorders related to autoimmunity and inflammation. TNFα-TNF receptor-1 (TNFR1) signaling contributes to the pathological processes of these disorders. The current study is focused on finding novel small molecules that can directly bind to TNFα and/or TNFR1, preventing the interaction between TNFα or TNFR1, and regulating downstream signaling pathways. Cheminformatics pipeline (pharmacophore modeling, virtual screening, molecular docking and in silico ADMET analysis) was used to screen for novel TNFα and TNFR1 inhibitors in the Zinc database. The pharmacophore-based models were generated to screen for the best drug like compounds in the Zinc database. The 39, 37 and 45 best hit molecules were mapped with the core pharmacophore features of TNFα, TNFR1, and the TNFα-TNFR1 complex respectively. They were further evaluated by molecular docking, protein-ligand interactions and in silico ADMET studies. The molecular docking analysis revealed the binding energies of TNFα, TNFR1 and the TNFα-TNFR1 complex, the basis of which was used to select the top five best binding energy compounds. Furthermore, in silico ADMET studies clearly revealed that all 15 compounds (ZINC09609430, ZINC49467549, ZINC13113075, ZINC39907639, ZINC25251930, ZINC02968981, ZINC09544246, ZINC58047088, ZINC72021182, ZINC08704414, ZINC05462670, ZINC35681945, ZINC23553920, ZINC05328058, and ZINC17206695) satisfied the Lipinski rule of five and had no toxicity. The new selective TNFα, TNFR1 and TNFα-TNFR1 complex inhibitors can serve as anti-inflammatory agents and are promising candidates for further research.


Cyclooxygenase-2 differentially directs uterine angiogenesis during implantation in mice.
Increased vascular permeability and angiogenesis at the site of blastocyst apposition in the uterus are two hallmarks of the implantation process. The present investigation shows that although the proangiogenic vascular endothelial growth factor (VEGF) and its receptor, Flk-1, are primarily important for uterine vascular permeability and angiogenesis prior to and during the attachment phase of the implantation process, VEGF in complementation with the angiopoietins and their receptor, Tie-2, directs angiogenesis during decidualization following implantation. Mice with null mutation for the gene encoding cyclooxygenase-2 (COX-2), a rate-limiting enzyme in prostaglandin (PG) biosynthesis, show implantation and decidualization failure. Using reporter and mutant mice, we show here that COX-2-derived prostaglandins (PGs) are important for uterine vascular permeability and angiogenesis during implantation and decidualization, suggesting that one cause of the failure of these latter processes in Cox-2-/- mice is the deregulated vascular events in the absence of COX-2. The attenuation of uterine angiogenesis in these mice is primarily due to defective VEGF signaling and not due to the defective angiopoietin system.


A novel risk stratification model, involving preoperative lymphocyte-monocyte ratio and standard pathological factors, for overall survival in patients with bladder cancer undergoing radical cystectomy.
To evaluate the ability of preoperative lymphocyte-monocyte ratio to predict prognosis and determine post-operative risk stratification in patients with bladder cancer undergoing radical cystectomy. A retrospective review of the 210 patients who had undergone radical cystectomy for bladder cancer from 2006 to 2013 identified 181 patients with sufficient data to evaluate the prognostic significance of the lymphocyte-monocyte ratio. Overall survival was assessed by the Kaplan-Meier method. The association of clinicopathological findings with overall survival was evaluated by a multivariate Cox proportional model, and a novel risk stratification model to predict prognosis was established. Median follow-up after radical cystectomy was 6.0 years. The 5-year overall survival rate was significantly lower for patients with low than high lymphocyte-monocyte ratio (27.6 vs 80.7%, P < 0.001). Multivariable analyses showed that pT ≥2, pN ≥1, positive margins and low lymphocyte-monocyte ratio were independent predictors of overall survival. A post-operative risk stratification model using these factors showed significant differences among the three subgroups (low, intermediate and high risk) with a concordance index of 0.84. The 5-year overall survival rates in patients at low, intermediate and high risk were 85.4, 45.5 and 0%, respectively (P < 0.001). Preoperative lymphocyte-monocyte ratio, pathological tumor and lymph node stage and positive margins are significantly associated with overall survival in patients who have undergone radical cystectomy for bladder cancer.


Effects of ginseng ingestion on growth hormone, testosterone, cortisol, and insulin-like growth factor 1 responses to acute resistance exercise.
Ginseng, an herbal plant, has been ingested by many athletes in Oriental regions of the world in order to improve stamina and to facilitate rapid recovery from injuries. However, adequate investigation has not been conducted to examine the ergogenic effects of ginseng. To examine the effects of ginseng supplements on hormonal status following acute resistance exercise, eight male college students were randomly given water (control; CON) or 20 g of ginseng root extract (GIN) treatment immediately after a standardized exercise bout. Venous blood samples were drawn before and immediately after exercise and at 4 time points during a 2-hour recovery period. Human growth hormone, testosterone, cortisol, and insulin-like growth factor 1 (IGF-1) levels were determined by radioimmunoassay. The responses of plasma hormones following ginseng consumption were not significant between CON and GIN treatments during the 2-hour recovery period. These results do not support the use of ginseng to promote an anabolic hormonal status following resistance exercise.


Clostridium muellerianum sp. nov., a carbon monoxide-oxidizing acetogen isolated from old hay.
An acid/alcohol-producing, Gram-stain-positive, obligately anaerobic, rod-shaped, non-motile, non-spore forming acetogen, designated as strain P21T, was isolated from old hay after enrichment with CO as the substrate. Spores not observed even after prolonged incubation (30 days). Phylogenetic analysis of the 16S rRNA gene sequence of strain P21T showed it was closely related to Clostridium carboxidivorans DSM 15243T (97.9%), Clostridium scatologenes DSM 757T (97.7 %) and Clostridium drakei DSM 12750T (97.7 %). The genome is 5.6 Mb and the G+C content is 29.4 mol%. Average nucleotide identity between strain P21T, C. carboxidivorans, C. scatologenes and C. drakei was 87.1, 86.4, 86.4 %, respectively. Strain P21T grew on CO:CO2, H2:CO2, l-arabinose, ribose, xylose, fructose, galactose, glucose, lactose, mannose, cellobiose, sucrose, cellulose, starch, pyruvate, choline, glutamate, histidine, serine, threonine and casamino acids. End products of metabolism were acetate, butyrate, caproate, ethanol and hexanol. Dominant cellular fatty acids (>10 %) were C16 : 0 (41.5 %), C16 : 1 ω7c/C16 : 1 ω6c (10.0 %), and a summed feature containing cyclo C17 : 1/C18 : 0 (17.3 %). Based on the phenotypic, chemotaxonomic, phylogenetic and phylogenomic analyses, strain P21T represents a new species in the genus Clostridium, for which the name Clostridium muellerianum sp. nov. is proposed. The type strain is P21T (=DSM 111390T=NCIMB 15261T).


The C-terminal, third conserved motif of the protein activator PACT plays an essential role in the activation of double-stranded-RNA-dependent protein kinase (PKR).
One of the key mediators of the antiviral and antiproliferative actions of interferon is double-stranded-RNA-dependent protein kinase (PKR). PKR activity is also involved in the regulation of cell proliferation, apoptosis and signal transduction. We have recently identified PACT, a novel protein activator of PKR, as an important modulator of PKR activity in cells in the absence of viral infection. PACT heterodimerizes with PKR and activates it by direct protein-protein interactions. Endogenous PACT acts as an activator of PKR in response to diverse stress signals, such as serum starvation and peroxide or arsenite treatment, and is therefore a novel, stress-modulated physiological activator of PKR. In this study, we have characterized the functional domains of PACT that are required for PKR activation. Our results have shown that, unlike the N-terminal conserved domains 1 and 2, the third conserved domain of PACT is dispensable for its binding of double-stranded RNA and inter action with PKR. However, a deletion of domain 3 results in a loss of PKR activation ability, in spite of a normal interaction with PKR, thereby indicating that domain 3 plays an essential role in PKR activation. Purified recombinant domain 3 could also activate PKR efficiently in vitro. Our results indicate that, although dispensable for PACT's high-affinity interaction with PKR, the third motif is essential for PKR activation. In addition, domain 3 and eukaryotic initiation factor 2alpha both interact with PKR through the same region within PKR, which we have mapped to lie between amino acid residues 318 and 551.


ETV1 inhibition depressed M2 polarization of tumor-associated macrophage and cell process in gastrointestinal stromal tumor via down-regulating PDE3A.
M2-type polarization of tumor associated-macrophage (TAM) is involved in the malignancy of gastrointestinal stromal tumor (GIST) progression. ETS variant 1 (ETV1) has been previously validated to regulate GIST pathogenesis. Our study intended to explore the role and mechanism of ETV1 in mediating the M2-polarization of TAM in GIST progression. First, we analyzed the correlation between ETV1 expression and M2-polarization in GIST tissues. IL-4 was used to treat THP-1-derived TAM cells and IL-4-stimulated TAM were co-cultured with GIST-T1 cells to mimic the GIST microenvironment. A loss-of-function assay was performed to explore the role of ETV1. Results showed that ETV1 elevation was positively correlated with M2-polarization. IL-4-induced TAM promoted ETV1 expression, silencing ETV1 inhibited proliferation, invasion and KIT activation in IL-4-treated GIST cells, while cell apoptosis was enhanced. Besides, co-culture of ETV1-silenced GIST cells significantly depressed M2-polarization in TAM, presented as decreased levels of CD206, Agr-1 and cytokines, as well as the proportion of CD206-positive TAM. PDE3A was positively correlated with ETV1 and M2-polarization. Overexpressing PDE3A reversed the inhibitory effects of ETV1 silencing. Generally, ETV1 inhibition depressed M2-polarization of TAM in GIST and its promotion on pathological aggravation via down-regulating PDE3A. This evidence may provide a new target for GIST regulation.


[Fatal urinary tract malformations: their place and incidence in a series of 450 neonatal autopsies].
In our serie of 450 neonatal necropsies, 26 cases presenting with urinary tract anomalies have been observed (5,7% of the total and 15,5% in those with malformations). From an etiological point of view, two groups could be distinguished: one (17 cases) with severe oligoamnios sequence and the second (9 cases) with hereditary malformations. In this second heterogeneous group thourough pathological examination seems to be the essential for diagnosis and efficient genetic counselling.


Production of hepatitis B virus in vitro by transient expression of cloned HBV DNA in a hepatoma cell line.
Transfection of human hepatoma cell lines with cloned HBV DNA resulted in the secretion of large amounts of hepatitis B surface antigen (HBsAg) and core-related antigens (HBc/HBeAg) if well-differentiated cell lines were employed. Synthesis of both viral antigens was the highest in cell line HuH-7 and continued for approximately 25 days. Particles resembling hepatitis B virions (Dane particles) by morphology, density and by the presence of the preS1 surface antigen were released from the transfected HuH-7 cells into the culture medium. These particles produced in vitro were also indistinguishable from the naturally occurring hepatitis B virions in containing the virus-associated DNA polymerase and mature HBV genomes. Restriction analysis of these DNA molecules was compatible with the nucleotide sequence of the transfecting HBV DNA sequence. Viral surface antigens and core proteins present in the culture medium were fractionated and characterized by immunoprecipitation and SDS--PAGE after labeling with [35S]methionine. Antisera specific for X-gene products identified in cell extracts two hitherto unknown HBV gene products. This system thus provides a new approach to open questions regarding HBV-related gene function and HBV replication.


Lamivudine (3TC), a Nucleoside Reverse Transcriptase Inhibitor, Prevents the Neuropathological Alterations Present in Mutant Tau Transgenic Mice.
The dysregulation of transposable elements contributes to neurodegenerative disorders. Previous studies have reported an increase in retrotransposon transcription in Drosophila models as well as in human tauopathies. In this context, we tested the possible protective effects of a reverse transcriptase inhibitor, namely lamivudine (also known as 3TC), in P301S mice, an animal model of Alzheimer's disease based on FTDP-17-tau overexpression. Transgenic P301S mice administered lamivudine through drinking water showed a decrease in the following histopathological marks typical of tauopathies: tau phosphorylation; inflammation; neuronal death; and hippocampal atrophy. Lamivudine treatment attenuated motor deficits (Rotarod test) and improved short-term memory (Y-maze test). To evaluate the role of tau in retrotransposition, we cotransfected HeLa cells with a plasmid containing a complete LINE-1 sequence and a neomycin reporter cassette designed for retrotransposition assays, and a plasmid with the tau sequence. LINE-1 insertion increased considerably in the cotransfection compared to the transfection without tau. In addition, lamivudine inhibited the insertion of LINE-1. Our data suggest that the progression of the tauopathy can be attenuated by the administration of lamivudine upon the first symptoms of neuropathology.


A review of human milk banking and public health policy in Australia.
Breastmilk is the perfect food for human infants. It is markedly different from, and uniquely superior to, artificial baby milk. Human milk banks are services which collect, screen, process and distribute donated breastmilk. Recipients are generally ill and premature infants whose mothers are unable to breastfeed them. This review of human milk banking in Australian public health policy draws from local and international research. This history of human milk banking and contemporary Australian policies, pertaining to breastfeeding and milk banking, are examined. Human milk banking is noted to be largely invisible from national breastfeeding policies. The barriers to establishing human milk banks in the Australian context are explored. Strategies which have helped generate support for human milk banking are discussed. International research has demonstrated the cost-effectiveness of banked donor milk. It is time for human milk banking to become an integral component of Australian breastfeeding policies, viewed as one of several initiatives to protect and support breastfeeding.


Synthesis of Biaryls via Benzylic C-C Bond Cleavage of Styrenes and Benzyl Alcohols.
A metal-free oxidative coupling of styrenes and benzyl alcohols with arenes has been developed for the synthesis of biaryls. The reaction features a conspicuous benzylic C-C bond cleavage of styrenes and benzyl alcohols. The reaction with both substrates proceeds through a common aldehydic intermediate formed through oxidative C-C bond cleavage of alkene and oxidation of benzyl alcohols. The reaction proceeds efficiently over a broad range of substrates with excellent functional group tolerance.


Effects of dentin characteristics on interfacial nanoleakage.
Water emanating from dentinal tubules during air-drying and light-curing of adhesives leads to entrapment of droplets at the resin-dentin interface and contributes to nanoleakage. This study tested the null hypothesis that characteristics of substrate dentin and type of adhesive used for bonding would not affect the occurrence of nanoleakage. Three self-etch adhesives were used to bond to 4 types of dentin with different characteristics in 12 groups. After silver challenge, nanoleakage percentage was measured within the hybrid layer of each sample. The deep dentin cut perpendicular to tubules always showed a significantly higher nanoleakage percentage compared with that of the other 3 types of dentin. The percentages of nanoleakage within the hybrid layers were not statistically different among adhesives. However, when bonding to deep perpendicular dentin, both all-in-one adhesives revealed more distinct nanoleakage within the adhesive layer compared with that achieved with Clearfil SE Bond, a two-step self-etch adhesive. The results did not support the null hypothesis.


Identification of a new dysfunctional platelet P2Y12 receptor variant associated with bleeding diathesis.
Defects of the platelet P2Y12 receptor (P2Y12R) for adenosine diphosphate (ADP) are associated with increased bleeding risk. The study of molecular abnormalities associated with inherited qualitative defects of the P2Y12R protein is useful to unravel structure-function relationships of the receptor. We describe the case of 2 brothers, sons of first cousins, with lifelong history of abnormal bleeding, associated with dysfunctional P2Y12R and a previously undescribed missense mutation in the encoding gene. ADP (4-20 µM)-induced aggregation of patients' platelets was markedly reduced and rapidly reversible. Other agonists induced borderline-normal aggregation. Inhibition of vasodilator-stimulated phosphoprotein phosphorylation and prostaglandin E1-induced increase in cyclic adenosine monophosphate (cAMP) by ADP was impaired, whereas inhibition of cAMP increase by epinephrine was normal. [(3)H]PSB-0413, a selective P2Y12R antagonist, bound to a normal number of binding sites; however, its affinity, and that of the agonists ADP and 2-methylthio-adenosine-5'-diphosphate, was reduced. Patients' DNA showed a homozygous c.847T>A substitution that changed the codon for His-187 to Gln (p.His187Gln). Crystallographic data and molecular modeling studies indicated that His187 in transmembrane 5 is important for agonist and nucleotide antagonist binding and located in a region undergoing conformational changes. These studies delineate a region of P2Y12R required for normal function after ADP binding.


Glycerol-Assisted Construction of Long-Life Three-Dimensional Surface-Enhanced Raman Scattering Hot Spot Matrix.
An evaporative self-assembly strategy for constructing a long-life 3D surface-enhanced Raman scattering (SERS) hot spot matrix is proposed with the assistance of glycerol to improve the spectral sensitivity and reproducibility. Different from the traditional wetting-state or drying-state methods, silver nanoparticles (AgNPs) in the glycerol-stabilized 3D SERS hot spot matrix can be maintained in the translation state for more than 7 days with the maximal uniformity of the interparticle distance. Brownian dynamics simulations reveal that more hot spots emerge in the glycerol-stabilized AgNPs matrix, and the distances between the AgNPs are not fixed but balanced in a small range by the interplay of the van der Waals attraction and the electrostatic repulsion. A 2 orders of magnitude extra SERS enhancement, more stable peak frequencies, and a narrower peak full width at half-maximum (fwhm) can therefore be obtained, which ensure extremely stable and reproducible SERS signals. Single-molecule detection sensitivity utilizing the glycerol-stabilized 3D SERS hot spot matrix has been demonstrated by collecting the SERS spectra of dye molecules at a concentration of as low as 0.5 aM (5 × 10-19 M) with a good signal-to-noise ratio. A long lifetime, ultrahigh SERS enhancement, and extremely stable peak shape make the 3D SERS hot spot matrix a sensitive, practical, and reliable tool for the detection and analysis of analytes at ultralow concentration or even at the single-molecule level in complex matrixes.