Pharmacodynamics of ginsenosides: antioxidant activities, activation of Nrf2, and potential synergistic effects of combinations.
Ginseng has long been used in Asian countries for more than 2000 years. Currently, in the "Western World or Western Medicines", many reports have indicated that they have used herbal medicines, and ginseng is one of the most popular herbs. Several recent reports have indicated that the antioxidant/antioxidative stress activities of ginseng play a role in the benefits of ginseng; however, the precise mechanism is lacking. The antioxidant response element (ARE) is a critical regulatory element for the expression of many antioxidant enzymes and phase II/III drug metabolizing/transporter genes, mediated by the transcription factor nuclear factor (erythroid-derived 2)-like 2 (Nrf2). The aim of this study was to examine the potential activation and synergism of Nrf2-ARE-mediated transcriptional activity between three common ginsenosides present in ginseng, ginsenoside Rb1 (Rb1), ginsenoside Rg1 (Rg1), and ginsenoside 20(S)-protopanaxatriol (20S). We tested whether these ginsenosides and their combinations could induce Nrf2-ARE activities in HepG2-C8 cells with stably transfected ARE luciferase reporter gene. Cell proliferation, antioxidant and ARE activities, Western blotting of Nrf2 protein, and qPCR of mRNA of Nrf2 were conducted for Rb1, Rg1, and 20S as well as the combinations of 20S with Rb1 or Rg1. To determine the combination effects, the combination index (CI) was calculated. Rb1 and Rg1 are relatively nontoxic to the cells, while 20S at 50 μM or above significantly inhibited the cell proliferation. Rb1, Rg1, or 20S induced total antioxidant activity and ARE activity in a concentration-dependent manner. Furthermore, combinations of 20S with either Rb1 or Rg1 induced total antioxidant and ARE activity synergistically. The induction of Nrf2 protein and mRNA was also found to be synergistic with the combination treatments. In summary, in this study, we show that ginsenosides Rb1, Rg1, and 20S possess antioxidant activity, transcriptionally activating ARE as well as the potential of synergistic activities. The Nrf2-ARE-mediated antioxidant pathway could play a role for the overall antioxidative stress activities, which could be important for ginseng's health beneficial effects such as cancer chemopreventive activities.


Automated classification of three-dimensional reconstructions of coral reefs using convolutional neural networks.
Coral reefs are biologically diverse and structurally complex ecosystems, which have been severally affected by human actions. Consequently, there is a need for rapid ecological assessment of coral reefs, but current approaches require time consuming manual analysis, either during a dive survey or on images collected during a survey. Reef structural complexity is essential for ecological function but is challenging to measure and often relegated to simple metrics such as rugosity. Recent advances in computer vision and machine learning offer the potential to alleviate some of these limitations. We developed an approach to automatically classify 3D reconstructions of reef sections and assessed the accuracy of this approach. 3D reconstructions of reef sections were generated using commercial Structure-from-Motion software with images extracted from video surveys. To generate a 3D classified map, locations on the 3D reconstruction were mapped back into the original images to extract multiple views of the location. Several approaches were tested to merge information from multiple views of a point into a single classification, all of which used convolutional neural networks to classify or extract features from the images, but differ in the strategy employed for merging information. Approaches to merging information entailed voting, probability averaging, and a learned neural-network layer. All approaches performed similarly achieving overall classification accuracies of ~96% and >90% accuracy on most classes. With this high classification accuracy, these approaches are suitable for many ecological applications.


Premastication: the second arm of infant and young child feeding for health and survival?
Premastication of foods for infants was a crucial behavioural adaptation to neoteny that ensured nutritional adequacy during the period of complementary feeding throughout the course of human evolution until recent times.While the paps and gruels of agricultural systems provided an alternative and modern food technology appears to make it unnecessary, we argue that, in addition to its role in nutrition, premastication also played a crucial role in supporting infant health. Its abandonment, particularly in poor communities, has placed children at increased risk of inadequate nutrition and decreased ability to confront infections associated with the introduction of complementary foods.We present two empirical studies. Section I is a cross-cultural study of the ethnographic literature in order to estimate prevalence in non-Western societies.One-third of ethnographies in the worldwide sample with data on infant feeding report premastication. Section II presents the results of a qualitative study in China, conducted in order to provide data on the likelihood that this percent is incorrect due to under-reporting.The finding that 63% of Chinese university students received premasticated food as infants, whereas none of eight ethnographic studies performed in Han China identified premastication in their reports, provides support for the conclusion that the cross-cultural study grossly underestimates its prevalence in non-Western societies. Section III is a discussion of potential benefits and risks of infant exposure to maternal saliva.We conclude with the argument for a concerted research effort to determine whether premastication can solve not only the 'weanling dilemma' in poor countries but also some of the health problems among the better-off.


Does familiality predispose to both emergence and persistence of psychosis? A follow-up study.
It as been suggested that in schizophrenia an association exists between family history of schizophrenia and poor outcome on the one hand, and family history of affective disorders and good outcome on the other. We tested for associations between four-year outcome and familial loading for psychotic disorders in a mixed sample of 150 consecutively admitted patients with functional psychosis (schizophrenia, psychotic affective disorders, other psychotic disorders) of recent onset. For each proband, a familial loading score for (i) broadly defined psychotic disorder, (ii) schizophrenia, and (iii) affective disorder was calculated using information on relatives obtained through the Family History Research Diagnostic Criteria method and direct interviews of relatives with the Schedule for Affective Disorders and Schizophrenia. In our sample of psychotic patients, familial loading for psychotic disorder predicted persistent negative symptoms over the follow-up period (OR 1.5; 95% CI 1-2.2), especially in schizophrenia, and was also associated with more time hospitalised (P < 0.05) [corrected], and more social disability at follow-up (P < 0.05). Greater familial loading for schizophrenia predicted a greater likelihood of non-recovery (OR 2.2; 95% CI 1.1-4.4) and a greater likelihood to have had persistent negative symptoms over the follow-up period (OR 1.7; 95% CI 0.9-3.1). No association was found between outcome and familial loading for affective disorder. We conclude that familial loading may be a continuous risk factor for some dimensions of clinical outcome in the functional psychoses. This suggests that there is a continuum of genetic liability not only to the emergence of psychotic illness, but also the subsequent chronicity of the disorder.


Location, location, location: how the tissue microenvironment affects inflammation in RA.
Current treatments for rheumatoid arthritis (RA) do not work well for a large proportion of patients, or at all in some individuals, and cannot cure or prevent this disease. One major obstacle to developing better drugs is a lack of complete understanding of how inflammatory joint disease arises and progresses. Emerging evidence indicates an important role for the tissue microenvironment in the pathogenesis of RA. Each tissue is made up of cells surrounded and supported by a unique extracellular matrix (ECM). These complex molecular networks define tissue architecture and provide environmental signals that programme site-specific cell behaviour. In the synovium, a main site of disease activity in RA, positional and disease stage-specific cellular diversity exist. Improved understanding of the architecture of the synovium from gross anatomy to the single-cell level, in parallel with evidence demonstrating how the synovial ECM is vital for synovial homeostasis and how dysregulated signals from the ECM promote chronic inflammation and tissue destruction in the RA joint, has opened up new ways of thinking about the pathogenesis of RA. These new ideas provide novel therapeutic approaches for patients with difficult-to-treat disease and could also be used in disease prevention.


Risk for suicide among medicaid beneficiaries.
Demographic, diagnostic, and service expenditure characteristics of Florida Medicaid enrollees who died by suicide were investigated. Among persons receiving Medicaid and Supplemental Security Income (SSI), findings indicate the most powerful predictors of suicide were involuntary psychiatric examination, mental health hospitalization, and high mental health service use. Among Medicaid enrollees not receiving SSI, strongest suicide predictors were mental health hospitalization, high expenditures for physical health medications, and involuntary psychiatric examination. Findings suggest reducing involuntary psychiatric examinations and mental health hospitalizations while improving physical health may reduce suicide in the Medicaid population. Comprehensive hospital discharge planning, adherence monitoring with follow-up care, training mental health providers in assessing suicide lethality, and providing adequate assessment time are all crucial to achieve these objectives.


Associations between Infant Feeding Practices and Length, Weight, and Disease in Developing Countries.
The health benefits of exclusive breastfeeding are well-known, but the relative detrimental impacts of other foods on infant health are unknown. Because infants in developing countries are fed a wide range of food, quantifying the burden of these diverse feeding practices on infant health is essential for public health policy. We used data from the Demographic Health Survey from 20 developing countries over multiple years to examine the independent association of six different types of food (exclusive breastfeeding, non-exclusive breastfeeding, infant formula, milk liquids, non-milk liquids, and solid foods) with five measures of infant health (length, weight, diarrhea, fever, and cough). We estimated associations with regression analysis, controlling for confounding factors with infant, mother, and household factors and community-year fixed effects. We used these estimates in a simulation model to quantify the burden of different combinations of food on infant health. We show that for an infant younger than 6 months old, following current guidelines and exclusively breastfeeding instead of giving the infant solid foods may increase length by 0.75 cm and weight by 0.25 kg and decrease diarrhea, fever, and cough prevalence by 8, 12, and 11%, respectively. We found that the burden on infant health of some feeding practices is less than others. Although all other feeding practices are associated with worse health outcomes than exclusive breastfeeding, breastfeeding supplemented with liquids has a lower burden on infant health than solid foods and infant formula has a lower burden than milk or non-milk liquids as measured by four of five health metrics. Providing specific quantified burden estimates of these practices can help inform public health policy related to infant feeding practices.


Optimization of a Microfluidic Mixing Process for Gene Expression-Based Bio-dosimetry.
In recent decades advances in radiation imaging and radiation therapy have led to a dramatic increase in the number of people exposed to radiation. Consequently, there is a clear need for personalized biodosimetry diagnostics in order to monitor the dose of radiation received and adapt it to each patient depending on their sensitivity to radiation exposure (Hall E.J. and Brenner D. J., 2008). Similarly, after a large-scale radiological event such as a dirty bomb attack, there will be a major need to assess, within a few days the radiation doses received by tens of thousands of individuals. Current high throughput devices can handle only a few hundred individuals per day. Hence there is a great need for a very fast self-contained non-invasive biodosimetric device based on a rapid blood test.This paper presents a case study where regression methods and designed experiments are used to arrive at the optimal settings for various factors that impact the kinetics in a biodosimetric device. We use ridge regression to initially identify a set of potentially important variables in the mixing process which is one of the critical sub systems of the device. This was followed by a series of designed experiments to arrive at the optimal setting of the significant microfluidic cartridge and piezoelectric disk (PZT) (D. Sadler, F. Zenhausern, U.S. Patent 6,986,601; Lee, S. Y., Ko, B., Yang, W., 2005) related factors. This statistical approach has been utilized to study the microfluidic mixing to mix water and dye mixtures of 70 μl volume. The outcome of the statistical design, experimentation and analysis was then exploited for optimizing the design, fabrication and assembly of the microfluidic devices. As a result of the experiments that were performed, the system was fine tuned and the mixing time was reduced from 5.5 minutes to 2 minutes.


Effects of serum on mammary epithelial proliferation in vitro during mammary gland development.
The proliferative response of mammary gland epithelium from nonpregnant, pregnant, and lactating mice to mammary serum factor and insulin was studied in vitro. Mammary gland epiithelium from nonpregnant and lactating animals has a delayed proliferative response to mammary serum factor and insulin when compared to the response of epithelium from pregnant animals. The results show that as the animals go through pregnancy into lactation the mammary gland epithelium becomes less responsive to mammary serum factor while it retains its responsiveness to insulin. The concentration of mammary serum factor in sera from animals at various physiological stages is constant. Sera from hypophysectomized rats, on the other hand, show a 50% drop in mammary serum factor activity. This loss of activity cannot be reversed by injecting prolactin, 17-beta-estradiol, or growth hormone into the hypophysectomized animals. A hypothesis that the mammary gland is composed of two proliferative epithelial populations is developed, and the possible role of prolactin in stimulating DNA synthesis is discussed.


A LIM-homeobox gene is required for differentiation of Wnt-expressing cells at the posterior end of the planarian body.
Planarians have high regenerative ability, which is dependent on pluripotent adult somatic stem cells called neoblasts. Recently, canonical Wnt/β-catenin signaling was shown to be required for posterior specification, and Hedgehog signaling was shown to control anterior-posterior polarity via activation of the Djwnt1/P-1 gene at the posterior end of planarians. Thus, various signaling molecules play an important role in planarian stem cell regulation. However, the molecular mechanisms directly involved in stem cell differentiation have remained unclear. Here, we demonstrate that one of the planarian LIM-homeobox genes, Djislet, is required for the differentiation of Djwnt1/P-1-expressing cells from stem cells at the posterior end. RNA interference (RNAi)-treated planarians of Djislet [Djislet(RNAi)] show a tail-less phenotype. Thus, we speculated that Djislet might be involved in activation of the Wnt signaling pathway in the posterior blastema. When we carefully examined the expression pattern of Djwnt1/P-1 by quantitative real-time PCR during posterior regeneration, we found two phases of Djwnt1/P-1 expression: the first phase was detected in the differentiated cells in the old tissue in the early stage of regeneration and then a second phase was observed in the cells derived from stem cells in the posterior blastema. Interestingly, Djislet is expressed in stem cell-derived DjPiwiA- and Djwnt1/P-1-expressing cells, and Djislet(RNAi) only perturbed the second phase. Thus, we propose that Djislet might act to trigger the differentiation of cells expressing Djwnt1/P-1 from stem cells.


Efficacy of rituximab in non-paraneoplastic autoimmune retinopathy.
Autoimmune retinopathy (AIR) is a rare but potentially blinding condition that is often underdiagnosed. Common features in AIR presentation include rapidly progressive vision loss with abnormal electrophysiological responses of the retina associated with positive anti-retinal antibodies. AIR is also challenging to treat, and thus, the introduction of new potential therapeutic agents is welcomed. The goal of this communication is to assess the effects of rituximab infusions on electroretinogram (ERG) responses and visual function outcomes in patients with non-paraneoplastic autoimmune retinopathy (npAIR). Following infusion(s), three out of five patients showed no evidence of disease progression or improved, while two patients continued to progress on ERG. One patient demonstrated improvement in visual acuity (2 lines) in both eyes. ERG responses provided objective monitoring of patients' visual function and response to immunosuppression over time. These findings suggest that patients with npAIR unresponsive to other immunosuppression therapies may benefit from rituximab infusion, although stabilization rather than improvement was more frequently the outcome in our case series. Furthermore, regularly scheduled ERG follow-up examinations are recommended for monitoring patients' progression during treatment.


Clinical criteria for obtaining maxillofacial computed tomographic scans in trauma patients.
Over 150,000 patients present with maxillofacial trauma annually to emergency rooms in the United States. Although maxillofacial computed tomography is a sensitive screening tool for identifying facial fractures, indiscriminate use leads to unnecessary radiation exposure and substantial costs. A decision instrument is needed to ensure computed tomographic evaluation of patients at high risk for facial fracture and limit computed tomography use in low-risk patients. A retrospective review was conducted of all patients evaluated at a Level I trauma center over a 3-year period. Inclusion criteria were maxillofacial examination on presentation, maxillofacial computed tomography, and head computed tomography. A total of 525 patients met the enrollment criteria. Injury to the maxillofacial skeleton occurred in 332 patients (63.2 percent). The presence of any of the following five physical examination criteria identified patients at high risk for facial fracture: bony stepoff or instability, periorbital swelling or contusion, Glasgow Coma Scale score less than 14, malocclusion, or tooth absence. These criteria identified all but six of the 332 patients with a facial fracture (sensitivity, 98.2 percent; 95 percent confidence interval, 96.5 to 99.1 percent). The negative predictive value was 87.8 percent (95 percent confidence interval, 76.3 to 94.2 percent). No patient determined by these criteria to be at low risk for a facial fracture required surgical treatment. If these criteria had been applied to the study population, radiographic imaging could have been avoided in 9.3 percent of patients. A decision instrument based on clinical criteria can ensure appropriate screening of patients at high risk for facial fracture. Application of this instrument may reduce unnecessary maxillofacial imaging.


X-ray structure of the mouse serotonin 5-HT3 receptor.
Neurotransmitter-gated ion channels of the Cys-loop receptor family mediate fast neurotransmission throughout the nervous system. The molecular processes of neurotransmitter binding, subsequent opening of the ion channel and ion permeation remain poorly understood. Here we present the X-ray structure of a mammalian Cys-loop receptor, the mouse serotonin 5-HT3 receptor, at 3.5 Å resolution. The structure of the proteolysed receptor, made up of two fragments and comprising part of the intracellular domain, was determined in complex with stabilizing nanobodies. The extracellular domain reveals the detailed anatomy of the neurotransmitter binding site capped by a nanobody. The membrane domain delimits an aqueous pore with a 4.6 Å constriction. In the intracellular domain, a bundle of five intracellular helices creates a closed vestibule where lateral portals are obstructed by loops. This 5-HT3 receptor structure, revealing part of the intracellular domain, expands the structural basis for understanding the operating mechanism of mammalian Cys-loop receptors.


Physiological and psychological stress responses in adults with attention-deficit/hyperactivity disorder (ADHD).
According to self-report and unsystematic observational data adult patients with attention-deficit/hyperactivity disorder suffer from increased vulnerability to daily life stressors. The present study examined psychological and physiological stress responses in adult ADHD subjects in comparison to healthy controls under laboratory conditions. Thirty-six subjects (18 patients with DSM-IV ADHD diagnosis, 18 sex- and age-matched healthy controls) underwent the Trier Social Stress Test (TSST; Kirschbaum, C., Pirke, K.-M., Hellhammer, D.H., 1993. The "Trier Social Stress Test"--a tool for investigating psychobiological stress responses in a laboratory setting. Neuropsychobiology 28, 76-81), a standardized psychosocial stress protocol which contains a stress anticipation phase and a stress phase with a free speech assignment and subsequent performance of a mental arithmetic. Physiological stress measures were salivary cortisol as an indicator of the HPA axis, heart rate (HR), and time- and frequency-domain heart rate variability (HRV) parameters. Subjective stress experience was measured via self-report repeatedly throughout the experimental session. In line with previous theoretical and empirical work in the field of childhood ADHD, it was hypothesized that the ADHD and control group would exhibit comparable baseline levels in all dependent variables. For ADHD subjects, we expected attenuated responses of the physiological parameters during anticipation and presence of the standardized stressor, but elevated subjective stress ratings. Hypotheses were confirmed for the baseline condition. Consistent with our assumptions in regard to the psychological stress response, the ADHD group experienced significantly greater subjective stress. The results for the physiological variables were mixed. While ADHD subjects revealed an attenuated HR during the stress phase, no significant group differences were found for the other parameters, although a trend was observed for both the low frequency/high frequency (LF/HF) ratio of the HRV power spectral analysis and salivary cortisol (the latter possibly indicating generally lower cortisol levels in ADHD subjects). In summary, the present findings are the first to demonstrate a significant alteration of a specific physiological stress measure (HR) and, more clearly, of psychological aspects of the stress response in adults suffering from ADHD. In regard to the physiological stress response, it is recommended that future studies employ larger sample sizes and a more comprehensive range of physiological stress parameters. Additionally, the issue of transferability of laboratory results to real life stressors needs to be addressed.


Evaluation of the cardiac actin gene in Doberman Pinschers with dilated cardiomyopathy.
To evaluate the coding region of the cardiac actin gene in Doberman Pinschers with dilated cardiomyopathy (DCM) for mutations that could be responsible for the development of the condition 28 dogs (16 Doberman Pinschers with DCM and 12 mixed-breed control dogs). Ten milliliters of blood was collected from each dog for DNA extraction. Polymerase chain reaction (PCR) primers were designed to amplify canine exonic regions, using the sequences of exons 2 to 6 of the cardiac actin gene. Single-stranded conformational polymorphism analysis was performed for each exon with all samples. Autoradiographs were analyzed for banding patterns specific to affected dogs. The DNA sequencing was performed on a selected group of affected and control dogs. Molecular analysis of exons 2 to 6 of the cardiac actin gene did not reveal any differences in base pairs between affected dogs and control dogs selected for DNA evaluation. Mutations in exons 5 and 6 of the cardiac actin gene that have been reported in humans with familial DCM do not appear to be the cause of familial DCM in Doberman Pinschers. Additionally, evaluation of exons 2 to 6 for causative mutations did not reveal a cause for inherited DCM in these Doberman Pinschers. Although there is evidence that DCM in Doberman Pinschers is an inherited problem, a molecular basis for this condition remains unresolved. Evaluation of other genes coding for cytoskeletal proteins is warranted.


Suicidal ideation, plans, and attempts in chronic pain patients: factors associated with increased risk.
This study describes suicidal behavior in a cross-sectional sample of chronic pain patients and evaluates factors associated with increased risk for suicidal ideation. One hundred-fifty-three adults with nonmalignant pain (42% back pain) who were consecutively referred to a tertiary care pain center completed a Structured Clinical Interview for Suicide History, the McGill Pain Questionnaire, and the Beck Depression Inventory. Nineteen-percent reported current passive suicidal ideation (PSI), 13% had active thoughts of committing suicide (ASI), 5% had a current suicide plan, and 5% reported a previous suicide attempt. Drug overdose was the most commonly reported plan and method of attempt (75%). Thirteen-percent reported a family history of suicide attempt/completion. Pain-specific and traditional suicide risk factors were evaluated as predictors of current PSI and ASI. Logistic regression analyses revealed that a family history of suicide attempts/completions was associated with a 7.5 fold increase in risk of PSI (P=0.001) and a 6.6 fold increase in ASI (P=0.003), after adjusting for significant covariates. Having abdominal pain was associated with an adjusted 5.5 fold increase in PSI (P=0.05) and a 4.2 fold increase in ASI (P=0.10). Neuropathic pain significantly reduced risk for both PSI (P=0.002) and ASI (P=0.01). Demographics, pain severity, and depression severity were not associated with suicidal ideation in multivariate analyses. These findings highlight the need for routine evaluation and monitoring of suicidal behavior in chronic pain, especially for patients with family histories of suicide, those taking potentially lethal medications, and patients with abdominal pain.


A pilot study to determine the feasibility of enhancing cognitive abilities in children with sensory processing dysfunction.
Children with Sensory Processing Dysfunction (SPD) experience incoming information in atypical, distracting ways. Qualitative challenges with attention have been reported in these children, but such difficulties have not been quantified using either behavioral or functional neuroimaging methods. Furthermore, the efficacy of evidence-based cognitive control interventions aimed at enhancing attention in this group has not been tested. Here we present work aimed at characterizing and enhancing attentional abilities for children with SPD. A sample of 38 SPD and 25 typically developing children were tested on behavioral, neural, and parental measures of attention before and after a 4-week iPad-based at-home cognitive remediation program. At baseline, 54% of children with SPD met or exceeded criteria on a parent report measure for inattention/hyperactivity. Significant deficits involving sustained attention, selective attention and goal management were observed only in the subset of SPD children with parent-reported inattention. This subset of children also showed reduced midline frontal theta activity, an electroencephalographic measure of attention. Following the cognitive intervention, only the SPD children with inattention/hyperactivity showed both improvements in midline frontal theta activity and on a parental report of inattention. Notably, 33% of these individuals no longer met the clinical cut-off for inattention, with the parent-reported improvements persisting for 9 months. These findings support the benefit of a targeted attention intervention for a subset of children with SPD, while simultaneously highlighting the importance of having a multifaceted assessment for individuals with neurodevelopmental conditions to optimally personalize treatment.


Optical receiver design for high optical return loss.
A new fiber pigtailed optical receiver design using a single gradient-index rod lens with a beveled exit face achieved a broadband optical return loss of > 65 dB, which was limited by the diffuse reflection from the photodetector front surface. By contrast the optical return loss for a receiver with an unbeveled lens exit face and an on-axis optical path was limited to a smaller value by the specular reflection from the lens exit face.


Interactive Effects of Corrosion, Copper, and Chloramines on Legionella and Mycobacteria in Hot Water Plumbing.
Complexities associated with drinking water plumbing systems can result in undesirable interactions among plumbing components that undermine engineering controls for opportunistic pathogens (OPs). In this study, we examine the effects of plumbing system materials and two commonly applied disinfectants, copper and chloramines, on water chemistry and the growth of Legionella and mycobacteria across a transect of bench- and pilot-scale hot water experiments carried out with the same municipal water supply. We discovered that copper released from corrosion of plumbing materials can initiate evolution of >1100 times more hydrogen (H2) from water heater sacrificial anode rods than does presence of copper dosed as soluble cupric ions. H2 is a favorable electron donor for autotrophs and causes fixation of organic carbon that could serve as a nutrient for OPs. Dosed cupric ions acted as a disinfectant in stratified stagnant pipes, inhibiting culturable Legionella and biofilm formation, but promoted Legionella growth in pipes subject to convective mixing. This difference was presumably due to continuous delivery of nutrients to biofilm on the pipes under convective mixing conditions. Chloramines eliminated culturable Legionella and prevented L. pneumophila from recolonizing biofilms, but M. avium gene numbers increased by 0.14-0.76 logs in the bulk water and were unaffected in the biofilm. This study provides practical confirmation of past discrepancies in the literature regarding the variable effects of copper on Legionella growth, and confirms prior reports of trade-offs between Legionella and mycobacteria if chloramines are applied as secondary disinfectant residual.


Chikungunya virus-associated long-term arthralgia: a 36-month prospective longitudinal study.
Arthritogenic alphaviruses, including Chikungunya virus (CHIKV), are responsible for acute fever and arthralgia, but can also lead to chronic symptoms. In 2006, a Chikungunya outbreak occurred in La Réunion Island, during which we constituted a prospective cohort of viremic patients (n = 180) and defined the clinical and biological features of acute infection. Individuals were followed as part of a longitudinal study to investigate in details the long-term outcome of Chikungunya. Patients were submitted to clinical investigations 4, 6, 14 and 36 months after presentation with acute CHIKV infection. At 36 months, 22 patients with arthralgia and 20 patients without arthralgia were randomly selected from the cohort and consented for blood sampling. During the 3 years following acute infection, 60% of patients had experienced symptoms of arthralgia, with most reporting episodic relapse and recovery periods. Long-term arthralgias were typically polyarthralgia (70%), that were usually symmetrical (90%) and highly incapacitating (77%). They were often associated with local swelling (63%), asthenia (77%) or depression (56%). The age over 35 years and the presence of arthralgia 4 months after the disease onset are risk factors of long-term arthralgia. Patients with long-term arthralgia did not display biological markers typically found in autoimmune or rheumatoid diseases. These data helped define the features of CHIKV-associated chronic arthralgia and permitted an estimation of the economic burden associated with arthralgia. This study demonstrates that chronic arthralgia is a frequent complication of acute Chikungunya disease and suggests that it results from a local rather than systemic inflammation.


Visualizing the global secondary structure of a viral RNA genome with cryo-electron microscopy.
The lifecycle, and therefore the virulence, of single-stranded (ss)-RNA viruses is regulated not only by their particular protein gene products, but also by the secondary and tertiary structure of their genomes. The secondary structure of the entire genomic RNA of satellite tobacco mosaic virus (STMV) was recently determined by selective 2'-hydroxyl acylation analyzed by primer extension (SHAPE). The SHAPE analysis suggested a single highly extended secondary structure with much less branching than occurs in the ensemble of structures predicted by purely thermodynamic algorithms. Here we examine the solution-equilibrated STMV genome by direct visualization with cryo-electron microscopy (cryo-EM), using an RNA of similar length transcribed from the yeast genome as a control. The cryo-EM data reveal an ensemble of branching patterns that are collectively consistent with the SHAPE-derived secondary structure model. Thus, our results both elucidate the statistical nature of the secondary structure of large ss-RNAs and give visual support for modern RNA structure determination methods. Additionally, this work introduces cryo-EM as a means to distinguish between competing secondary structure models if the models differ significantly in terms of the number and/or length of branches. Furthermore, with the latest advances in cryo-EM technology, we suggest the possibility of developing methods that incorporate restraints from cryo-EM into the next generation of algorithms for the determination of RNA secondary and tertiary structures.


Reward sensitivity and action in Parkinson's disease patients with and without apathy.
Clinical apathy results in dysfunction of goal directed behaviour, a key component of which is the initiation of action. Previous work has suggested that blunting of reward sensitivity is an important mechanism underlying apathy. However, an additional component might be impoverished initiation of action itself. This study aims to investigate the link between motivation and motor output and its association with apathy and dopamine. An oculomotor task that measures pupillary and saccadic response to monetary incentives was used to assess reward sensitivity, first in 23 young and 18 elderly controls, and then in 22 patients with Parkinson's disease tested ON and OFF dopaminergic medication. To distinguish between pupillary responses to anticipated reward alone versus responses associated with motor preparation, a saccadic 'go/no-go' task was performed. Half of the trials required a saccade to be initiated to receive a reward and in the remaining trials no action was required but reward was still obtained. No significant difference in pupil response was demonstrated between the two conditions in all groups tested, suggesting pupillary responses to rewards are not contingent upon motor preparation in Parkinson's disease. Being ON or OFF dopamine did not influence this response either. Previous work demonstrated associations between apathy and pupillary reward insensitivity in Parkinson's disease. Here we observed this effect only when an action was required to receive a reward, and only in the ON state. These findings suggest that apathy in Parkinson's disease is linked to reduced reward sensitivity and that this is most prominently observed when actions have to be initiated to rewarding goals, with the effect modulated by being ON dopaminergic medication. OFF medication, there was no such strong relationship, and similarly in the 'no-go' conditions, either ON or OFF dopaminergic drugs. The results provide preliminary data which suggest that apathy in Parkinson's disease is associated with a reduction in reward sensitivity and this is most evident when associated with initiation of goal directed actions in the presence of adequate dopamine.


Coronary MR angiography: experimental results with a monomer-stabilized blood pool contrast medium.
To evaluate the signal-enhancing characteristics of monomer-coated very small superparamagnetic iron oxide (SPIO) particles used as a blood pool contrast medium for magnetic resonance (MR) angiography in the coronary arteries. The particles used in this study were coated with citrate as the monomer (VSOP-C91). The particles have a total diameter of 7 nm and show the following relaxivities at 0.47 T: T1, 19 L/mmol. sec(-1); T2, 29 L/mmol. sec(-1). Fifteen cardiac MR examinations were performed at 1.5 T in five pigs. Images were acquired from immediately to 35 minutes (equilibrium phase) after intravenous injection of gadopentetate dimeglumine, gadobenate dimeglumine, and the very small SPIO particles (n = 5 for each substance). Immediately after administration of gadopentetate dimeglumine, gadobenate dimeglumine, and the very small SPIO particles, respectively, increases in the signal-to-noise ratio in blood were 94%, 103%, and 102% and in myocardium were 83%, 83%, and 29% (P <.05, very small SPIO particles versus the low-molecular-weight gadolinium-based compounds). Differences in the blood-to-myocardium contrast-to-noise ratio and visualization of the coronary arteries and their branches were also significant. VSOP-C91 significantly improves visualization of the coronary arteries at MR angiography from immediately to 35 minutes after injection.


Integrating NTD mapping protocols: Can surveys for trachoma and urinary schistosomiasis be done simultaneously?
We determined whether the school-based "disease mapping" methodology used to assess urinary schistosomiasis (SCH) is useful for determining trachoma interventions and whether the district-based approach recommended for trachoma is useful for SCH control programs. We conducted two separate integrated surveys in eight districts of central Nigeria: school based and district based. A total of 17,189 children were examined for trachoma and 16,238 children were examined for hematuria from 363 schools and 2,149 households. School surveys identified 67 communities warranting praziquantel drug treatment of SCH and 142 trachoma-endemic communities warranting trachoma control activities. In district-level estimates, we identified 24 communities for praziquantel treatment and 0 for trachoma intervention. Integrating trachoma into SCH school-based surveys, and SCH into trachoma surveys, was quick and easy, but in this setting, school-based surveys were more useful for identifying communities where intervention is warranted.


Expanding the taxonomy of the diagnostic criteria for temporomandibular disorders.
There is a need to expand the current temporomandibular disorders' (TMDs) classification to include less common but clinically important disorders. The immediate aim was to develop a consensus-based classification system and associated diagnostic criteria that have clinical and research utility for less common TMDs. The long-term aim was to establish a foundation, vis-à-vis this classification system, that will stimulate data collection, validity testing and further criteria refinement. A working group [members of the International RDC/TMD Consortium Network of the International Association for Dental Research (IADR), members of the Orofacial Pain Special Interest Group (SIG) of the International Association for the Study of Pain (IASP), and members from other professional societies] reviewed disorders for inclusion based on clinical significance, the availability of plausible diagnostic criteria and the ability to operationalise and study the criteria. The disorders were derived from the literature when possible and based on expert opinion as necessary. The expanded TMDs taxonomy was presented for feedback at international meetings. Of 56 disorders considered, 37 were included in the expanded taxonomy and were placed into the following four categories: temporomandibular joint disorders, masticatory muscle disorders, headache disorders and disorders affecting associated structures. Those excluded were extremely uncommon, lacking operationalised diagnostic criteria, not clearly related to TMDs, or not sufficiently distinct from disorders already included within the taxonomy. The expanded TMDs taxonomy offers an integrated approach to clinical diagnosis and provides a framework for further research to operationalise and test the proposed taxonomy and diagnostic criteria.


Modeling short-term dynamics and variability for realistic interactive facial animation.
Modern modeling and rendering techniques have produced nearly photorealistic face models, but truly expressive digital faces also require natural-looking movements. Virtual characters in today's applications often display unrealistic facial expressions. Indeed, facial animation with traditional schemes such as keyframing and motion capture demands expertise. Moreover, the traditional schemes aren't adapted to interactive applications that require the real-time generation of context-dependent movements. A new animation system produces realistic expressive facial motion at interactive speed. The system relies on a set of motion models controlling facial-expression dynamics. The models are fitted on captured motion data and therefore retain the dynamic signature of human facial expressions. They also contain a nondeterministic component that ensures the variety of the long-term visual behavior. This system can efficiently animate any synthetic face. The video illustrates interactive use of a system that generates facial-animation sequences.


Diffuse alveolar damage in a patient treated with gemcitabine.
We present a case of diffuse alveolar damage (DAD) that occurred in a male aged 68 yrs treated with gemcitabine, a novel antineoplastic agent, that was given for hepatic relapse of a previously resected non-small cell lung cancer. The patient developed acute respiratory failure after the sixth drug dose, and died 4 days after admission. Autopsy revealed a pattern of DAD. No evidence of infection or other specific aetiologies could be found. To our knowledge, only three cases of pulmonary toxicity resulting from treatment with gemcitabine have been published; two of them were fatal and postmortem examination revealed a pattern consistent with acute respiratory distress syndrome. A careful survey may determine the incidence of pulmonary toxicity of this new drug in the future.


Sarilumab use in severe SARS-CoV-2 pneumonia.
Interleukin-6 signal blockade showed preliminary beneficial effects in treating inflammatory response against SARS-CoV-2 leading to severe respiratory distress. Herein we describe the outcomes of off-label intravenous use of Sarilumab in severe SARS-CoV-2-related pneumonia. 53 patients with SARS-CoV-2 severe pneumonia received intravenous Sarilumab; pulmonary function improvement or Intensive Care Unit (ICU) admission rate in medical wards, live discharge rate in ICU treated patients and safety profile were recorded. Sarilumab 400 mg was administered intravenously on day 1, with eventual additional infusion based on clinical judgement, and patients were followed for at least 14 days, unless previously discharged or dead. Of the 53 SARS-CoV-2pos patients receiving Sarilumab, 39(73·6%) were treated in medical wards [66·7% with a single infusion; median PaO2/FiO2:146(IQR:120-212)] while 14(26·4%) in ICU [92·6% with a second infusion; median PaO2/FiO2: 112(IQR:100-141.5)].Within the medical wards, 7(17·9%) required ICU admission, 4 of whom were re-admitted to the ward within 5-8 days. At 19 days median follow-up, 89·7% of medical inpatients significantly improved (46·1% after 24 h, 61·5% after 3 days), 70·6% were discharged from the hospital and 85·7% no longer needed oxygen therapy. Within patients receiving Sarilumab in ICU, 64·2% were discharged from ICU to the ward and 35·8% were still alive at the last follow-up. Overall mortality rate was 5·7%. IL-6R inhibition appears to be a potential treatment strategy for severe SARS-CoV-2 pneumonia and intravenous Sarilumab seems a promising treatment approach showing, in the short term, an important clinical outcome and good safety.


The overlapping burden of the three leading causes of disability and death in sub-Saharan African children.
Despite substantial declines since 2000, lower respiratory infections (LRIs), diarrhoeal diseases, and malaria remain among the leading causes of nonfatal and fatal disease burden for children under 5 years of age (under 5), primarily in sub-Saharan Africa (SSA). The spatial burden of each of these diseases has been estimated subnationally across SSA, yet no prior analyses have examined the pattern of their combined burden. Here we synthesise subnational estimates of the burden of LRIs, diarrhoea, and malaria in children under-5 from 2000 to 2017 for 43 sub-Saharan countries. Some units faced a relatively equal burden from each of the three diseases, while others had one or two dominant sources of unit-level burden, with no consistent pattern geographically across the entire subcontinent. Using a subnational counterfactual analysis, we show that nearly 300 million DALYs could have been averted since 2000 by raising all units to their national average. Our findings are directly relevant for decision-makers in determining which and targeting where the most appropriate interventions are for increasing child survival.


Defective bone formation and anabolic response to exogenous estrogen in mice with targeted disruption of endothelial nitric oxide synthase.
Nitric oxide (NO) is a pleiotropic signaling molecule that is produced by bone cells constitutively and in response to diverse stimuli such as proinflammatory cytokines, mechanical strain, and sex hormones. Endothelial nitric oxide synthase (eNOS) is the predominant NOS isoform expressed in bone, but its physiological role in regulating bone metabolism remains unclear. Here we studied various aspects of bone metabolism in female mice with targeted disruption of the eNOS gene. Mice with eNOS deficiency (eNOS KO) had reduced bone mineral density, and cortical thinning when compared with WT controls and histomorphometric analysis of bone revealed profound abnormalities of bone formation, with reduced osteoblast numbers, surfaces and mineral apposition rate. Studies in vitro showed that osteoblasts derived from eNOS KO mice had reduced rates of growth when compared with WT and were less well differentiated as reflected by lower levels of alkaline phosphatase activity. Mice with eNOS deficiency lost bone normally following ovariectomy but exhibited a significantly blunted anabolic response to high dose exogenous estrogen. We conclude that the eNOS pathway plays an essential role in regulating bone mass and bone turnover by modulating osteoblast function.


Hospital Factors Associated With Interhospital Transfer Destination for Stroke in the Northeast United States.
Background We aimed to determine if there is an association between hospital quality and the likelihood of a given hospital being a preferred transfer destination for stroke patients. Methods and Results Data from Medicare claims identified acute ischemic stroke transferred between 394 northeast US hospitals from 2007 to 2011. Hospitals were categorized as transferring (n=136), retaining (n=241), or receiving (n=17) hospitals based on the proportion of acute ischemic stroke encounters transferred or received. We identified all 6409 potential dyads of sending and receiving hospitals, and categorized dyads as connected if ≥5 patients were transferred between the hospitals annually (n=82). We used logistic regression to identify hospital characteristics associated with establishing a connected dyad, exploring the effect of adjusting for different quality measures and outcomes. We also adjusted for driving distance between hospitals, receiving hospital stroke volume, and the number of hospitals in the receiving hospital referral region. The odds of establishing a transfer connection increased when rate of alteplase administration increased at the receiving hospital or decreased at the sending hospital, however this finding did not hold after applying a potential strategy to adjust for clustering. Receiving hospital performance on 90-day home time was not associated with likelihood of transfer connection. Conclusions Among northeast US hospitals, we found that differences in hospital quality, specifically higher levels of alteplase administration, may be associated with increased likelihood of being a transfer destination. Further research is needed to better understand acute ischemic stroke transfer patterns to optimize stroke transfer systems.


Antiplatelet drugs and platelet reactivity: is it time to halt clinical research on tailored strategies?
Personalized medicine of antiplatelet drugs in cardiovascular patients has led to a significant enthusiasm. Indeed, numerous longitudinal studies showed an association between high platelet reactivity and the recurrence of ischemic events. The first small randomized trials of P2Y12 blockers tailored to each patient's platelet reactivity yielded encouraging reductions of coronary stent thrombosis in high-risk populations. The discovery of genetic variants contributing to the pharmacodynamic effect of clopidogrel has then paved the way toward a personalized antiplatelet therapy based on reliable and stable genetic tests. This enthusiasm was soon tempered by large interventional trials demonstrating that a platelet function testing-based strategy did not improve clinical outcome and that genetic variants discovered up to now only explained a small part of the pharmacodynamic effect of clopidogrel, thus limiting its clinical use. Looking back to the most recent trials, their target populations and the type of clinical setting, it seems that the one-size-fits all policy regarding antiplatelet drugs may be well acceptable for low-risk patients. On the contrary, integration of the clinical setting as well as other risk factors may help to identify subgroups of patients who could derive a benefit from a truly personalized management of antiplatelet therapy.


Computational and analytical challenges in single-cell transcriptomics.
The development of high-throughput RNA sequencing (RNA-seq) at the single-cell level has already led to profound new discoveries in biology, ranging from the identification of novel cell types to the study of global patterns of stochastic gene expression. Alongside the technological breakthroughs that have facilitated the large-scale generation of single-cell transcriptomic data, it is important to consider the specific computational and analytical challenges that still have to be overcome. Although some tools for analysing RNA-seq data from bulk cell populations can be readily applied to single-cell RNA-seq data, many new computational strategies are required to fully exploit this data type and to enable a comprehensive yet detailed study of gene expression at the single-cell level.


Evidence for karyogamy and exchange of genetic material in the binucleate intestinal parasite Giardia intestinalis.
The diplomonad parasite Giardia intestinalis contains two functionally equivalent nuclei that are inherited independently during mitosis. Although presumed to be asexual, Giardia has low levels of allelic heterozygosity, indicating that the two nuclear genomes may exchange genetic material. Fluorescence in situ hybridization performed with probes to an episomal plasmid suggests that plasmids are transferred between nuclei in the cyst, and transmission electron micrographs demonstrate fusion between cyst nuclei. Green fluorescent protein fusions of giardial homologs of meiosis-specific genes localized to the nuclei of cysts, but not the vegetative trophozoite. These data suggest that the fusion of nuclei, or karyogamy, and subsequently somatic homologous recombination facilitated by the meiosis gene homologs, occur in the giardial cyst.


Visual acuity as an outcome measure in clinical trials of retinal diseases.
Visual acuity (VA) is the primary outcome measure in many studies involving eye diseases. A standard statistical approach for comparing a continuous measurement such as a VA letter score between 2 treatment groups is to perform a t test comparing the means. However, frequently a binary variable is created from the continuous VA letter score based on whether or not there has been a worsening (or gain) of > or =15 letters (equivalent to > or =3 lines), and a chi square or similar statistical test is performed to compare the proportions of success (or failure) between groups. The purpose of this article is to contrast these 2 approaches. Clinical trial reports of retinal disorders were used to compare results using mean change in the VA letter score versus binary proportions created from the VA letter score. Additionally, analyses were performed using generated data to gain a perspective on the magnitude of differences that might be expected between the 2 methods. Studies from the literature showed that differences of 6% to 15% in > or =15-letter worsening corresponded to mean differences in letter scores between groups of 3.0 to 7.0 (approximately 0.6 to 1.4 lines). Analyses using generated data demonstrated that a mean improvement in the VA letter score of 5 corresponded to a doubling of the proportion of eyes with > or =15-letter improvement and a 28% relative reduction in the proportion of eyes with > or =15-letter worsening. How VA data should be analyzed in a clinical trial depends to large extent on the research question. The frequently used outcome of > or =15-letter change has several drawbacks, including loss of efficiency (need for a larger sample), misclassification of the outcome, and potential for a ceiling or floor effect. Therefore, for most clinical trials we believe that the primary outcome analysis should be a comparison of changes in the VA letter score, and created binary variables should be reported as secondary outcomes. This approach maximizes the information gained from the data and accommodates both improvement and worsening of acuity.


How I treat resistant cytomegalovirus infection in hematopoietic cell transplantation recipients.
Cytomegalovirus (CMV) infection is a significant complication in hematopoietic cell transplantation (HCT) recipients. Four antiviral drugs are used for preventing or treating CMV: ganciclovir, valganciclovir, foscarnet, and cidofovir. With prolonged and repeated use of these drugs, CMV can become resistant to standard therapy, resulting in increased morbidity and mortality, especially in HCT recipients. Antiviral drug resistance should be suspected when CMV viremia (DNAemia or antigenemia) fails to improve or continue to increase after 2 weeks of appropriately dosed and delivered antiviral therapy. CMV resistance is diagnosed by detecting specific genetic mutations. UL97 mutations confer resistance to ganciclovir and valganciclovir, and a UL54 mutation confers multidrug resistance. Risk factors for resistance include prolonged or previous anti-CMV drug exposure or inadequate dosing, absorption, or bioavailability. Host risk factors include type of HCT and degree of immunosuppression. Depending on the genotyping results, multiple strategies can be adopted to treat resistant CMV infections, albeit no randomized clinical trials exist so far, after reducing immunosuppression (if possible): ganciclovir dose escalation, ganciclovir and foscarnet combination, and adjunct therapy such as CMV-specific cytotoxic T-lymphocyte infusions. Novel therapies such as maribavir, brincidofovir, and letermovir should be further studied for treatment of resistant CMV.


Does adolescent depression predict obesity in black and white young adult women?
To examine whether adolescent depressive symptoms predict young adult body mass index (BMI) and obesity in black and white women. Participants included 1554 black and white adolescent girls from the National Heart, Lung, and Blood Institute Growth and Health Study (NGHS) who completed the Center for Epidemiological Studies--Depression Scale (CES-D) at ages 16 and 18 years. Regression analyses showed that depressive symptoms at both ages 16 and 18 were associated with increased risk of obesity (BMI > or = 30) and elevated BMI in young adulthood (age 21) in both black and white girls. Black girls exhibited a significantly greater likelihood of obesity and higher BMI (i.e. a main effect of race), but the race x CES-D interaction was not significant in any analysis. Depressive symptoms in adolescence appear to be predictive of obesity and elevated BMI in early adulthood for both black and white girls, even when taking prior BMI into account, indicating that depressive symptoms confer risk for obesity above and beyond the known tracking of body weight. Obesity prevention studies might consider assessing depressive symptoms in adolescence in order to more fully capture important risk variables.


Enrichment of individual KIR2DL4 sequences from genomic DNA using long-template PCR and allele-specific hybridization to magnetic bead-bound oligonucleotide probes.
DNA enrichment by allele-specific hybridization (DEASH) was used as a means to isolate individual alleles of the killer cell immunoglobulin-like receptor (KIR2DL4) gene from heterozygous genomic DNA. Using long-template polymerase chain reaction (LT-PCR), the complete KIR2DL4 gene was amplified from a cell line that had previously been characterized for its KIR gene content by PCR using sequence-specific primers (PCR-SSP). The whole gene amplicons were sequenced and we identified two heterozygous positions in accordance with the predictions of the PCR-SSP. The amplicons were then hybridized to allele-specific, biotinylated oligonucleotide probes and through binding to streptavidin-coated beads, the targeted alleles were enriched. A second PCR amplified only the exonic regions of the enriched allele, and these were then sequenced in full. We show DEASH to be capable of enriching single alleles from a heterozygous PCR product, and through sequencing the enriched DNA, we are able to produce complete coding sequences of the KIR2DL4 alleles in accordance with the typing predicted by PCR-SSP.


Sequence of a novel cytochrome CYP2B cDNA coding for a protein which is expressed in a sebaceous gland, but not in the liver.
The major phenobarbital-inducible rat hepatic cytochromes P-450, CYP2B1 and CYP2B2, are the paradigmatic members of a cytochrome P-450 gene subfamily that contains at least seven additional members. Specific oligonucleotide probes for these genomic members of the CYP2B subfamily were used to assess their tissue-specific expression. In Northern-blot analysis a probe specific to gene 4 (which is designated now as CYP2B12) hybridized to a single mRNA present in the preputial gland, an organ which is used as a model for sebaceous glands, but did not hybridize to mRNA isolated from the liver or from five other tissues of untreated or Aroclor 1254-treated rats. The cDNA sequence for the CYP2B12 RNA was determined from overlapping cDNA clones and contained a long open reading frame of 1476 bp. The nucleotide sequence of the CYP2B12 cDNA was 85% similar to the sequence of the CYP2B1 cDNA in its coding region and was different from any CYP2B cDNA characterized until now. The cDNA-derived primary structure of the CYP2B12 protein contains a signal sequence for its insertion into the endoplasmic reticulum and the putative haem-binding site characteristic of cytochromes P-450. A part of the potential haem pocket of CYP2B12 was identical with a similar structure in a bacterial protocatechuate dioxygenase. In immunoblot analysis of preputial-gland microsomes, antibodies against CYP2B1 recognized a single abundant protein with a lower apparent molecular mass than that of CYP2B1. Our results demonstrate that the CYP2B12 protein has the potential to be enzymically active and are the first demonstration that a member of the CYP2B subfamily is expressed exclusively and at high levels in an extrahepatic organ.


Iso-alpha acids from hops (Humulus lupulus) inhibit hepatic steatosis, inflammation, and fibrosis.
Non-alcoholic fatty liver disease (NAFLD) is considered to be the hepatic manifestation of the metabolic syndrome. Iso-alpha acids (IAAs), hop-derived bitter compounds in beer, have been shown to beneficially affect different components of the metabolic syndrome such as insulin resistance and dyslipidemia. However, IAAs have not yet been studied in the context of chronic liver disease. Here we analyzed the effect of IAA on the pathogenesis of NAFLD. Once, we applied IAA to mice in combination with a NAFLD-inducing Western-type diet (WTD), and observed that IAA significantly inhibited WTD-induced body weight gain, glucose intolerance, and hepatic steatosis. Fitting to this, IAA dose-dependently inhibited cellular lipid accumulation in primary human hepatocytes (PHH) in vitro. Reduced expression of PPAR-gamma and key enzymes of lipid synthesis as well as increased expression of PPAR-alpha, indicative for increased lipid combustion, were identified as underlying mechanisms of reduced hepatocellular steatosis in vitro and in vivo. Analysis of hepatic HMOX1 expression indicated reduced oxidative stress in IAA-treated mice, which was paralleled by reduced activation of the JNK pathway and pro-inflammatory gene expression and immune cell infiltration. Furthermore, IAA reduced hepatic stellate cell (HSC) activation and pro-fibrogenic gene expression. Similarly, IAA also dose-dependently reduced oxidative stress and JNK activation in steatotic PHH, inhibited HSC activation, and reduced proliferation and pro-fibrogenic gene expression in already activated HSC in vitro. In conclusion, IAAs inhibit different pathophysiological steps of disease progression in NAFLD. Together with previous studies, which demonstrated the safety of even long-term application of IAA in humans, our data suggest IAA as promising therapeutic agent for the prevention and treatment of (non)alcoholic (fatty) liver disease.


Pilot Study to Establish a Novel Five-Gene Biomarker Panel for Predicting Lymph Node Metastasis in Patients With Early Stage Endometrial Cancer.
Introduction: In the United States and Europe, endometrial endometrioid carcinoma (EEC) is the most prevalent gynecologic malignancy. Lymph node metastasis (LNM) is the key determinant of the prognosis and treatment of EEC. A biomarker that predicts LNM in patients with EEC would be beneficial, enabling individualized treatment. Current preoperative assessment of LNM in EEC is not sufficiently accurate to predict LNM and prevent overtreatment. This pilot study established a biomarker signature for the prediction of LNM in early stage EEC. Methods: We performed RNA sequencing in 24 clinically early stage (T1) EEC tumors (lymph nodes positive and negative in 6 and 18, respectively) from Cathay General Hospital and analyzed the RNA sequencing data of 289 patients with EEC from The Cancer Genome Atlas (lymph node positive and negative in 33 and 256, respectively). We analyzed clinical data including tumor grade, depth of tumor invasion, and age to construct a sequencing-based prediction model using machine learning. For validation, we used another independent cohort of early stage EEC samples (n = 72) and performed quantitative real-time polymerase chain reaction (qRT-PCR). Finally, a PCR-based prediction model and risk score formula were established. Results: Eight genes (ASRGL1, ESR1, EYA2, MSX1, RHEX, SCGB2A1, SOX17, and STX18) plus one clinical parameter (depth of myometrial invasion) were identified for use in a sequencing-based prediction model. After qRT-PCR validation, five genes (ASRGL1, RHEX, SCGB2A1, SOX17, and STX18) were identified as predictive biomarkers. Receiver operating characteristic curve analysis revealed that these five genes can predict LNM. Combined use of these five genes resulted in higher diagnostic accuracy than use of any single gene, with an area under the curve of 0.898, sensitivity of 88.9%, and specificity of 84.1%. The accuracy, negative, and positive predictive values were 84.7, 98.1, and 44.4%, respectively. Conclusion: We developed a five-gene biomarker panel associated with LNM in early stage EEC. These five genes may represent novel targets for further mechanistic study. Our results, after corroboration by a prospective study, may have useful clinical implications and prevent unnecessary elective lymph node dissection while not adversely affecting the outcome of treatment for early stage EEC.


Transapical aortic valve implantation - an Australian experience.
The aim of this study was to report our initial experience with the transapical approach to transcatheter aortic valve implantation (TAVI) at an Australian institution. All patients with severe, symptomatic aortic stenosis were assessed by our multidisciplinary team. A total of 32 patients received a transapical TAVI using an Edwards SAPIEN prosthesis. Data were prospectively collected and analysed according to the Valve Academic Research Consortium version 2 guidelines. Intraoperative outcomes included: 100% device success with no conversion to surgical valve replacement, extracorporeal membrane oxygenation was used electively in 15.6% and emergently in 6.3%, and no valve migration or malpositioning requiring prosthesis retrieval and re-implantation. Outcomes at 30 days post-TAVI included: No mortality, 3.1% myocardial infarction, no disabling stroke, 3.1% non-disabling stroke, no transient ischaemic attacks, 6.3% life-threatening bleeding, 15.6% major bleeding, 3.1% major vascular complications, and 12.5% postoperative acute kidney injury requiring renal replacement therapy. Mild paravalvular regurgitation was present in 29%, and there was no moderate or severe regurgitation. Mean follow-up time was 28.8±12.9 months. Cumulative results included: 9.4% mortality, 6.3% stroke, 6.3% myocardial infarction, and no repeat procedures. At one year postoperation, echocardiography demonstrated that the mean pressure across the prosthesis was 10.1±1.7mmHg, and the mean aortic valve area was 1.4±0.2cm(2). Good short-term outcomes and low or zero mortality are achievable with transapical TAVI at an Australian institution.


White matter abnormalities and impaired attention abilities in children born very preterm.
While attention impairments are commonly observed in very preterm (<32weeks' gestational age) children, neuroanatomical correlates of these difficulties are unclear. We aimed to determine whether the microstructural organization of key white matter tracts thought to be involved in attention (cingulum bundle, superior longitudinal fasciculi, reticular activating system, and corpus callosum) were altered in very preterm children compared with term-born controls. We also aimed to determine whether alterations in microstructural organization of these tracts were associated with attention functioning in very preterm children. One hundred and forty-nine very preterm children and 36 term-born controls underwent neuroimaging and assessment of their attention abilities at 7years. Constrained spherical deconvolution and probabilistic tractography was used to identify the key white matter tracts. Altered microstructural organization and reduced tract volume within reticular activating system and corpus callosum were found in the very preterm group compared with the control group. Diffusion and volume changes in the cingulum bundle, superior longitudinal fasciculi, reticular activating system, and corpus callosum were related to variations in attention functioning in the very preterm children. These findings emphasize that white matter tract integrity is associated with later attentional abilities in very preterm children.


Routine spinal immobilization in trauma patients: what are the advantages and disadvantages?
Routine spinal immobilization for trauma patients has become established in developed countries throughout the world. Cervical spinal injury is, however, relatively rare in trauma patients, and immobilization practice was developed largely without firm supporting evidence. In recent years, published evidence has suggested that spinal immobilization may in some cases be harmful. The purpose of this article is to critically review the evidence and the implications for trauma patient management and outcomes. We searched MEDLINE, the Cochrane Database, Index Medicus and article references with a broad search strategy. Relevant results were analysed and critically reviewed in the context of trauma patient management. Our findings present a growing body of evidence documenting the risks and complications of routine spinal immobilization. There is a possibility that immobilization could be contributing to mortality and morbidity in some patients and this warrants further investigation.


Haemoglobin recovery among HIV-1 infected patients on zidovudine-based antiretroviral therapy and other regimens in north-central Nigeria.
We conducted a study to assess trends in haemoglobin recovery among HIV-infected patients initiated on zidovudine-based combination antiretroviral therapy (cART) stratified by baseline haemoglobin level. Haemoglobin data from non-pregnant adult patients initiating cART in rural north-central Nigeria between June 2009 and May 2011 were analysed using a linear mixed effects model to assess the interaction between time, zidovudine-containing regimen and baseline haemoglobin level on the outcome of subsequent haemoglobin level. Best-fit curves were created for baseline haemoglobin in the 10th, 25th, 75th and 90th percentiles. We included 313 patients with 736 measures of haemoglobin in the analysis (239 on zidovudine and 74 on non-zidovudine-containing regimens). Median haemoglobin increased over time in both groups, with differences in haemoglobin response over time related to baseline haemoglobin levels and zidovudine use (p = 0.003). The groups of patients on zidovudine at the 10th and 90th percentiles had downward sloping curves while all other groups had upward trending haemoglobin levels. Although haemoglobin levels increased overall for patients on zidovudine-containing regimens, for those in the 10th and 90th percentiles haemoglobin levels trended downward over time. These results have implications for decisions regarding when to initiate, switch from or avoid the use of zidovudine.


Identification of differentially expressed genes in clinically distinct groups of serous ovarian carcinomas using cDNA microarray.
To identify changes in gene expression in serous epithelial ovarian cancers (SEOC), we utilized cDNA microarrays consisting of 2382 genes with cancer related properties to analyze tumors from 20 patients with defined clinical out-comes. The significance analysis of microarrays method was used to determine differentially expressed genes, leading to the identification of 134 up-regulated and 231 down-regulated genes overall. By increasing the stringency of the statistical selection criteria, 41 over-expressed and 51 under-expressed genes were identified. The median duration of follow-up of the 20 patients was 16.8 months with a median progression free survival of 7.0 months. We found 11 genes that were differentially over-expressed in patients with recurrent disease, and 3 genes (homo sapiens mRNA for Ins P3 5-phophatase, lipoma HMGIC fusion partner-like 2 and CD63 melanoma 1 antigen) in patients who were dead of disease. Subsequently, we examined the distribution of the differentially expressed genes in the cDNA library database from adult human tumor and normal tissues using the DigiNorthern method to identify a subset of genes with relatively restricted tissue distribution. Finally, protein expression of 5 selected genes were further examined using immunohistochemistry applied on a tissue microarray prepared from an independent panel of 93 SEOC tissues. The results provided validation for 2 under-expressed genes (E2F transcription factor 5 and CK14) and 3 over-expressed genes (Bcl2-like 1, COX-2, CD63). Our study demonstrates differential gene expression in clinically distinct groups of SEOC using cDNA microarray. These genes may potentially be useful as biomarkers and/or targets for therapeutic intervention.


ADAR2-dependent GluA2 editing regulates cocaine seeking.
Activation of AMPA receptors (AMPARs) in the nucleus accumbens is necessary for the reinstatement of cocaine-seeking behavior, an animal model of drug craving and relapse. AMPARs are tetrameric protein complexes that consist of GluA1-4 subunits, of which GluA2 imparts calcium permeability. Adenosine deaminase acting on RNA 2 (ADAR2) is a nuclear enzyme that is essential for editing GluA2 pre-mRNA at Q/R site 607. Unedited GluA2(Q) subunits form calcium-permeable AMPARs (CP-AMPARs), whereas edited GluA2(R) subunits form calcium-impermeable channels (CI-AMPARs). Emerging evidence suggests that the reinstatement of cocaine seeking is associated with increased synaptic expression of CP-AMPARs in the nucleus accumbens. However, the role of GluA2 Q/R site editing and ADAR2 in cocaine seeking is unclear. In the present study, we investigated the effects of forced cocaine abstinence on GluA2 Q/R site editing and ADAR2 expression in the nucleus accumbens. Our results demonstrate that 7 days of cocaine abstinence is associated with decreased GluA2 Q/R site editing and reduced ADAR2 expression in the accumbens shell, but not core, of cocaine-experienced rats compared with yoked saline controls. To examine the functional significance of ADAR2 and GluA2 Q/R site editing in cocaine seeking, we used viral-mediated gene delivery to overexpress ADAR2b in the accumbens shell. Increased ADAR2b expression in the shell attenuated cocaine priming-induced reinstatement of drug seeking and was associated with increased GluA2 Q/R site editing and surface expression of GluA2-containing AMPARs. Taken together, these findings support the novel hypothesis that an increased contribution of accumbens shell CP-AMPARs containing unedited GluA2(Q) promotes cocaine seeking. Therefore, CP-AMPARs containing unedited GluA2(Q) represent a novel target for cocaine addiction pharmacotherapies.


Neutralization of SARS-CoV-2 by Destruction of the Prefusion Spike.
There are as yet no licensed therapeutics for the COVID-19 pandemic. The causal coronavirus (SARS-CoV-2) binds host cells via a trimeric spike whose receptor binding domain (RBD) recognizes angiotensin-converting enzyme 2, initiating conformational changes that drive membrane fusion. We find that the monoclonal antibody CR3022 binds the RBD tightly, neutralizing SARS-CoV-2, and report the crystal structure at 2.4 Å of the Fab/RBD complex. Some crystals are suitable for screening for entry-blocking inhibitors. The highly conserved, structure-stabilizing CR3022 epitope is inaccessible in the prefusion spike, suggesting that CR3022 binding facilitates conversion to the fusion-incompetent post-fusion state. Cryogenic electron microscopy (cryo-EM) analysis confirms that incubation of spike with CR3022 Fab leads to destruction of the prefusion trimer. Presentation of this cryptic epitope in an RBD-based vaccine might advantageously focus immune responses. Binders at this epitope could be useful therapeutically, possibly in synergy with an antibody that blocks receptor attachment.


A 4D hyperspherical interpretation of q-space.
3D q-space can be viewed as the surface of a 4D hypersphere. In this paper, we seek to develop a 4D hyperspherical interpretation of q-space by projecting it onto a hypersphere and subsequently modeling the q-space signal via 4D hyperspherical harmonics (HSH). Using this orthonormal basis, we derive several well-established q-space indices and numerically estimate the diffusion orientation distribution function (dODF). We also derive the integral transform describing the relationship between the diffusion signal and propagator on a hypersphere. Most importantly, we will demonstrate that for hybrid diffusion imaging (HYDI) acquisitions low order linear expansion of the HSH basis is sufficient to characterize diffusion in neural tissue. In fact, the HSH basis achieves comparable signal and better dODF reconstructions than other well-established methods, such as Bessel Fourier orientation reconstruction (BFOR), using fewer fitting parameters. All in all, this work provides a new way of looking at q-space.


Cholera outbreak caused by drinking contaminated water from a lakeshore water-collection site, Kasese District, south-western Uganda, June-July 2015.
On 20 June 2015, a cholera outbreak affecting more than 30 people was reported in a fishing village, Katwe, in Kasese District, south-western Uganda. We investigated this outbreak to identify the mode of transmission and to recommend control measures. We defined a suspected case as onset of acute watery diarrhoea between 1 June and 15 July 2015 in a resident of Katwe village; a confirmed case was a suspected case with Vibrio cholerae cultured from stool. For case finding, we reviewed medical records and actively searched for cases in the community. In a case-control investigation we compared exposure histories of 32 suspected case-persons and 128 age-matched controls. We also conducted an environmental assessment on how the exposures had occurred. We found 61 suspected cases (attack rate = 4.9/1000) during this outbreak, of which eight were confirmed. The primary case-person had onset on 16 June; afterwards cases sharply increased, peaked on 19 June, and rapidly declined afterwards. After 22 June, eight scattered cases occurred. The case-control investigation showed that 97% (31/32) of cases and 62% (79/128) of controls usually collected water from inside a water-collection site "X" (ORM-H = 16; 95% CI = 2.4-107). The primary case-person who developed symptoms while fishing, reportedly came ashore in the early morning hours on 17 June, and defecated "near" water-collection site X. We concluded that this cholera outbreak was caused by drinking lake water collected from inside the lakeshore water-collection site X. At our recommendations, the village administration provided water chlorination tablets to the villagers, issued water boiling advisory to the villagers, rigorously disinfected all patients' faeces and, three weeks later, fixed the tap-water system.


Trends in utilization of FDA expedited drug development and approval programs, 1987-2014: cohort study.
To evaluate the use of special expedited development and review pathways at the US Food and Drug Administration over the past two decades. Cohort study. FDA approved novel therapeutics between 1987 and 2014. Publicly available sources provided each drug's year of approval, their innovativeness (first in class versus not first in class), World Health Organization Anatomic Therapeutic Classification, and which (if any) of the FDA's four primary expedited development and review programs or designations were associated with each drug: orphan drug, fast track, accelerated approval, and priority review. Logistic regression models evaluated trends in the proportion of drugs associated with each of the four expedited development and review programs. To evaluate the number of programs associated with each approved drug over time, Poisson models were employed, with the number of programs as the dependent variable and a linear term for year of approval. The difference in trends was compared between drugs that were first in class and those that were not. The FDA approved 774 drugs during the study period, with one third representing first in class agents. Priority review (43%) was the most prevalent of the four programs, with accelerated approval (9%) the least common. There was a significant increase of 2.6% per year in the number of expedited review and approval programs granted to each newly approved agent (incidence rate ratio 1.026, 95% confidence interval 1.017 to 1.035, P<0.001), and a 2.4% increase in the proportion of drugs associated with at least one such program (odds ratio 1.024, 95% confidence interval 1.006 to 1.043, P=0.009). Driving this trend was an increase in the proportion of approved, non-first in class drugs associated with at least one program for drugs (P=0.03 for interaction). In the past two decades, drugs newly approved by the FDA have been associated with an increasing number of expedited development or review programs. Though expedited programs should be strictly limited to drugs providing noticeable clinical advances, this trend is being driven by drugs that are not first in class and thus potentially less innovative.


The interleukin-1 receptor/Toll-like receptor superfamily: signal transduction during inflammation and host defense.
The signal transduction pathways activated by the proinflammatory cytokine interleukin-1 (IL-1) have been the focus of much attention because of the important role that IL-1 plays in inflammatory diseases. A number of proteins have been described that participate in the post-receptor activation of the transcription factor nuclear factor kappaB (NF-kappaB), and stress-activated protein kinases such as p38 mitogen-activated protein kinase (MAPK). It has also emerged that the type I IL-1 receptor (IL-1RI) is a member of an expanding receptor superfamily. These related receptors all have sequence similarity in their cytosolic regions. The family includes the Drosophila melanogaster protein Toll, the IL-18 receptor (IL-18R), and 10 Toll-like receptors (TLRs), TLR-1 to TLR-10, which bind to microbial products, activating host defense responses. Because of the similarity of IL-1RI to Toll, the conserved sequence in the cytosolic region of these proteins has been termed the Toll-IL-1 receptor (TIR) domain. The same proteins activated during signaling by IL-1RI also participate in signaling by other receptors with TIR domains. The receptor superfamily is evolutionarily conserved; members also occur in plants and insects, where they also function in host defense. The signaling proteins that are activated are also conserved across species. Differences are, however, starting to emerge in signaling pathways activated by different receptors. This receptor superfamily, therefore, represents an ancient signaling system that is a critical determinant of the innate immune and inflammatory responses.


The structure of P-TEFb (CDK9/cyclin T1), its complex with flavopiridol and regulation by phosphorylation.
The positive transcription elongation factor b (P-TEFb) (CDK9/cyclin T (CycT)) promotes mRNA transcriptional elongation through phosphorylation of elongation repressors and RNA polymerase II. To understand the regulation of a transcriptional CDK by its cognate cyclin, we have determined the structures of the CDK9/CycT1 and free cyclin T2. There are distinct differences between CDK9/CycT1 and the cell cycle CDK CDK2/CycA manifested by a relative rotation of 26 degrees of CycT1 with respect to the CDK, showing for the first time plasticity in CDK cyclin interactions. The CDK9/CycT1 interface is relatively sparse but retains some core CDK-cyclin interactions. The CycT1 C-terminal helix shows flexibility that may be important for the interaction of this region with HIV TAT and HEXIM. Flavopiridol, an anticancer drug in phase II clinical trials, binds to the ATP site of CDK9 inducing unanticipated structural changes that bury the inhibitor. CDK9 activity and recognition of regulatory proteins are governed by autophosphorylation. We show that CDK9/CycT1 autophosphorylates on Thr186 in the activation segment and three C-terminal phosphorylation sites. Autophosphorylation on all sites occurs in cis.


The elastin fiber system between and adjacent to collector channels in the human juxtacanalicular tissue.
To determine the composition and investigate the elastin fiber system in the juxtacanalicular tissue adjacent to and between collector channel orifices in normal human eyes. Normal human eyes (71.0 ± 8.6 years; mean ± SD; n = 4) were perfusion fixed at low (10 mm Hg) and high pressure (20 mm Hg) with 3% paraformaldehyde/0.1 M phosphate buffer. Frontal serial sections were cut from paraffin blocks, and regions with and without collector channels were selected. Sections were stained using Weigert's resorcin-fuchsin stain with oxidation. Immunohistochemistry was performed using antibodies against elastin, fibrillin-1, and microfibrillar-associated protein-1/2. Elastin, elaunin, and oxytalan fibers were identified within the juxtacanalicular tissue of the inner and outer walls in low- and high-pressure eyes. These fibers were found at collector channel orifices, between collector channels, and within collector channel walls. Fibrillin-1 was located at the base and lateral edges of Schlemm's canal endothelial cells. Microfibrillar-associated protein-1/2 was found with elastin-like fibers at the base of Schlemm's canal endothelium cells, in the juxtacanalicular tissue, and in the uveal region. Elastin, elaunin, oxytalan, and elastin-associated proteins fibrillin-1 and microfibrillar-associated protein-1/2 were identified within the juxtacanalicular tissue of the inner and outer walls and within collector channel walls of human eyes perfused at low and high pressure. No differences in labeling patterns for elastin, elaunin, and oxytalan were found in the juxtacanalicular tissue adjacent to or between collector channel orifices. The elastin fiber system appears to have a significant role in the support and distensibility of the juxtacanalicular region under collector channels.


Quantitative power Doppler ultrasonography is a sensitive measure of metacarpophalangeal joint synovial vascularity in rheumatoid arthritis and declines significantly following a 2-week course of oral low-dose corticosteroids.
To investigate the stability over 2 weeks of ultrasonographic assessments of synovial thickness and vascularity in all 10 metacarpophalangeal joints of subjects with rheumatoid arthritis (RA) with a range of disease activities as measured by the validated Disease Activity Score-28 joint score (DAS28-ESR). And in subjects with severe disease activity, to compare the sensitivity of these measurements, acute-phase markers, and vascular endothelial growth factor to change in response to 2 weeks of oral prednisolone (7.5 mg daily). A group of 38 subjects with RA were enrolled, 13 (mean DAS28 2.1), 14 (mean DAS28 5.2), and 11 (mean DAS28 5.7) meriting oral corticosteroid treatment. Synovial thickness and vascularity were assessed by ultrasonography at 3 timepoints. Images were ranked by semiquantitative scale. Vascularity was also measured by quantitative determination of the power Doppler area (PDA). In the whole RA cohort, baseline indices of synovial thickness and vascularity correlated with DAS28, as did PDA (r = 0.42, p < 0.05). In the RA groups on stable therapy, synovial thickness and vascularity showed little variation over 2 weeks. In the corticosteroid group, PDA had fold changes of -1.9-fold (p < 0.05) after 1 week and -2.2-fold (p < 0.05) after 2 weeks. These were the largest fold changes of all measured variables. Ultrasonographic measures can differentiate disease severity in RA correlating closely with DAS28. Quantitative power Doppler signal was significantly reduced within 1 week of oral prednisolone, a rapid kinetic suggesting that PDA may have value as a sensitive early marker of therapeutic response.


Trace Elements in Human Nutrition (II) - An Update.
The dietary requirement for an essential trace element is an intake level which meets a specified criterion for adequacy and thereby minimizes risk of nutrient deficiency or excess. Disturbances in trace element homeostasis may result in the development of pathologic states and diseases. This article is an update of a review article "Trace Elements in Human Nutrition-A Review" previously published in 2013. The previous review was updated to emphasis in detail the importance of known trace elements so far in humans' physiology and nutrition and also to implement the detailed information for practical and effective management of trace elements' status in clinical diagnosis and health care situations. Although various classifications for trace elements have been proposed and may be controversial, this review will use World Health Organization( WHO) classification as previously done. For this review a traditional integrated review format was chosen and many recent medical and scientific literatures for the new findings on bioavailability, functions, and state of excess/deficiency of trace elements were assessed. The results indicated that for the known essential elements, essentiality and toxicity are unrelated and toxicity is a matter of dose or exposure. Little is known about the essentiality of some of the probably essential elements. In regard to toxic heavy metals, a toxic element may nevertheless be essential. In addition, the early pathological manifestations of trace elements deficiency or excess are difficult to detect until more specific pathologically relevant indicators become available. Discoveries and many refinements in the development of new techniques and continual improvement in laboratory methods have enabled researchers to detect the early pathological consequences of deficiency or excess of trace elements. They all are promises to fulfill the gaps in the present and future research and clinical diagnosis of trace elements deficiencies or intoxications. However, further investigations are needed to complete the important gaps in our knowledge on trace elements, especially probably essential trace elements' role in health and disease status.


Replication of Epstein-Barr virus oriLyt: lack of a dedicated virally encoded origin-binding protein and dependence on Zta in cotransfection assays.
Using a transient replication assay in which cosmid DNAs were cotransfected into Vero cells, we had previously demonstrated that oriLyt replication required six Epstein-Barr virus (EBV)-encoded replication genes. No oriLyt origin-binding protein was identified in this study, but oriLyt replication in the cotransfection assay was also dependent on the three lytic cycle transactivators Zta, Rta, and Mta and an activity encoded by the EBV Sal/I F fragment. We have now used expression plasmids for the six known replication proteins to further examine the question of the requirement for an oriLyt origin-binding protein. The activity in Sal/I-F was shown to be encoded by BKRF3. The predicted product of this open reading frame is an enzyme, uracyl DNA glycosylase, not an origin-binding protein, and is dispensable for replication in assays using expression plasmids. BBLF2, which is positionally related to the gene for the herpes simplex virus (HSV) UL9 origin-binding protein, was confirmed to be expressed as a spliced transcript with BBLF3 and not as an independent product. Examination of the requirement for the EBV transactivators revealed that Rta, while contributing to replication efficiency, was dispensable. Mta could be substituted by HSV IE63, and in complementation experiments with HSV replication genes, Mta was no longer required for replication of EBV oriLyt, suggesting that the contribution of Mta to replication may be indirect. Zta continued to be required for detectable oriLyt replication both with the EBV replication proteins and in the complementation assays with HSV replication proteins. We conclude that EBV does not encode an equivalent of HSV UL9 and that Zta is the sole virally encoded protein serving an essential origin-binding function.


Retinoic acid-induced modulation of rat liver transglutaminase and total polyamines in vivo.
The effect of a single intraperitoneal injection of retinoic acid on liver transglutaminase (EC 184.108.40.206) activity and total putrescine, spermidine and spermine was studied. The results demonstrate that: (1) transglutaminase activity is increased over control values as early as 4-6 h after treatment, reaching a maximum (2-fold increase) at 12 h and returning to control values at 36 h; (2) the retinoic acid-induced form of enzyme is the soluble tissue transglutaminase; (3) actinomycin D treatment does not completely inhibit the early (6 h) increase of activity, while suppressing that at 12 h; (4) the immunoassay of the soluble transglutaminase shows that, 6 h after treatment, there is no increase in the protein, whereas at 12 and 24 h a significant increase is observed; (5) putrescine, but not spermidine and spermine, increases (5-7-fold) 6 and 18 h after the retinoic acid treatment. The possibility also that the expression of soluble transglutaminase is modulated in vivo by retinoic acid and the relationship to polyamine levels are discussed.


Comprehensive analysis of published phase I/II clinical trials between 1990-2010 in osteosarcoma and Ewing sarcoma confirms limited outcomes and need for translational investment.
High grade primary bone sarcomas are rare cancers that affect mostly children and young adults. Osteosarcoma and Ewing sarcoma are the most common histological subtypes in this age group, with current multimodality treatment strategies achieving 55-70% overall survival. As there remains an urgent need to develop new therapeutic interventions, we have reviewed published phase I/II trials that have been reported for osteosarcoma and Ewing sarcoma in the last twenty years. We conducted a literature search for clinical trials between 1990 and 2010, either for trials enrolling bone sarcoma patients as part of a general sarcoma indication or trials specifically in osteosarcoma and Ewing sarcoma. We identified 42 clinical trials that fulfilled our search criteria for general sarcoma that enrolled these patient groups, and eight and twenty specific trials for Ewing and osteosarcoma patients, respectively. For the phase I trials which enrolled different tumour types our results were incomplete, because the sarcoma patients were not mentioned in the PubMed abstract. A total of 3,736 sarcoma patients were included in these trials over this period, 1,114 for osteosarcoma and 1,263 for Ewing sarcoma. As a proportion of the worldwide disease burden over this period, these numbers reflect a very small percentage of the potential patient recruitment, approximately 0.6% for Ewing sarcoma and 0.2% for osteosarcoma. However, these data show an increase in recent activity overall and suggest there is still much room for improvement in the current trial development structures. Lack of resources and commercial investment will inevitably limit opportunity to develop sufficiently rapid improvements in clinical outcomes. International collaboration exists in many well founded co-operative groups for phase III trials, but progress may be more effective if there were also more investment of molecular and translational research into disease focused phase I/II clinical trials. Examples of new models for early translational and early phase trial collaboration include the European based EuroBoNeT network, the Sarcoma Alliance for Research through Collaboration network (SARC) and the new European collaborative translational trial network, EuroSarc.


Perioperative Implications of Vaping.
Over the past 10 - 15 years, there has been a significant increase in the use of electronic cigarettes. These devices are generally used to deliver nicotine through inhalation by aerosolization. While the long-term risk of lung cancer is yet to be known, the chemicals and impurities in the solutions may have other acute and chronic effects on the respiratory system including respiratory failure from adult respiratory distress syndrome. Recent concerns have been raised regarding the potential for significant acute and chronic health care risks of these devices including pneumonitis, airway reactivity and respiratory failure. Given that many of the acute effects are related to the respiratory system, anesthetic care may be required during diagnostic procedures including bronchoscopy to investigate the etiology of acute respiratory symptomatology. We present an adolescent who presented to the operating room for bronchoscopy and bronchoalveolar lavage to investigate the etiology of respiratory involvement following an episode of vaping. The healthcare and end-organ effects of nicotine, tobacco smoke and vaping are discussed, and potential anesthetic implications are presented.


Histopathology of the synovial tissue: perspectives for biomarker development in chronic inflammatory arthritides.
The histopathological and molecular analysis of the synovial tissue has contributed to fundamental advances in our comprehension of arthritis pathogenesis and of the mechanisms of action of currently available treatments. On the other hand, its exploitation in clinical practice for diagnostic or prognostic purposes as well as for the prediction of treatment response to specific disease-modifying anti-rheumatic drugs is still limited. In this review, we present an overview of recent advances in the field of synovial tissue research with specific reference to the methods for synovial tissue collection, approaches to synovial tissue analysis and current perspectives for the exploitation of synovial tissue-derived biomarkers in chronic inflammatory arthritides.