Computation of DNP coupling factors of a nitroxide radical in toluene: seamless combination of MD simulations and analytical calculations.
Dynamic nuclear polarization (DNP) employs paramagnetic species to increase the NMR signal of nuclear spins. In liquids, the efficiency of the effect depends on the strength of the interaction between the electron and nuclear spins and the time scales on which this interaction is modulated by the physical motion of the spin-bearing molecules. An approach to quantitatively predict the contribution of molecular motions to the DNP enhancement using molecular dynamics (MD) simulations is developed and illustrated for the nitroxide radical TEMPOL in liquid toluene. A multi-resolution strategy that combines explicit treatment of the solvent at short distances from the free radical with implicit description at large intermolecular distances is adopted. Novel analytical expressions are obtained to correct for the finite spatial extent of the MD simulations. The atomistic and analytical descriptions are sewn seamlessly together by ensuring that for molecular trajectories that start in the near (explicit) region and end in the distant (implicit) region the analytical dipolar spectral densities reproduce the MD estimates. The spectral densities obtained from the developed approach are used to calculate DNP coupling factors separately for the ring and methyl protons of toluene. The agreement with previously reported experimental DNP data at a magnetic field of 3.4 T is noteworthy and encouraging. Maximum obtainable DNP enhancements at other magnetic fields are predicted.


[Manifestations of the "pretherapeutic phase" of malignant melanoma of the skin in Switzerland 1996-1998].
62 patients with a histologically confirmed diagnosis of cutaneous malignant melanoma were interviewed from 1996 to 1998. Lesions were found mostly on patients backs (30.6%) and they were noted most frequently because of a change in colour (darker) (50%) and increase in size (43.5%). The median time between first noticing the melanoma and visiting a physician was one month, but for 28% of the patients it was more than three months. The median time between a medical examination and surgical removal was shorter than one month, but for 22% of the patients it was more than three months. Patients with bleeding lesions sought medical help later (14 months). Misdiagnosis led to a six months delay in treatment and if no action was taken there was a 13 months delay.


Monitoring for overreaching in rugby league players.
The aim of this study was to identify indicators of non-functional overreaching (NFOR) in team sport athletes undertaking intensive training loads. Eighteen semi-professional rugby league players were randomly assigned into two pair matched groups. One group completed 6 weeks of normal training (NT) whilst the other group was deliberately overreached through intensified training (IT). Both groups then completed the same 7-day stepwise training load reduction taper. Multistage fitness test (MSFT) performance, VO2 (max), peak aerobic running velocity (V (max)), maximal heart rate, vertical jump, 10-s cycle sprint performance and body mass were measured pre- and post-training period and following the taper. Hormonal, haematological and immunological parameters were also measured pre-training and following weeks 2, 4 and 6 of training and post-taper. MANOVA for repeated measures with contrast analysis indicated that MSFT performance and VO2 (max) were significantly reduced in the IT group over time and condition, indicating that a state of overreaching was attained. However, the only biochemical measure that was significantly different between the IT and NT group was the glutamine to glutamate (Gln/Glu) ratio even though testosterone, testosterone to cortisol (T/C) ratio, plasma glutamate, and CK activity were significantly changed after training in both groups. Positive endurance and power performance changes were observed post-taper in the IT group confirming NFOR. These changes were associated with increases in the T/C ratio and the Gln/Glu ratio and decreases in plasma glutamate and CK activity. These results indicate that although there was no single reliable biochemical marker of NFOR in these athletes, the Gln/Glu ratio and MSFT test may be useful measures for monitoring responses to IT in team sport athletes.


Serological cross-reactivity between a human Ro/SS-A autoantigen (calreticulin) and the lambda Ral-1 antigen of Onchocerca volvulus.
We have cloned and sequenced a 46-kD Ro/SS-A autoantigen gene that is the human homologue of the calcium-binding protein, calreticulin. The sequence of this 46-kD Ro/SS-A protein (calreticulin) has significant homology to lambda Ral-1, a recombinant cDNA clone corresponding to a major antigen of the nematode, Onchocerca volvulus, the infectious agent of onchocerciasis. We therefore sought to determine whether antibodies produced by onchocerciasis patients might crossreact with the human 46-kD Ro/SS-A autoantigen (calreticulin). 20 of 22 sera from Liberian onchocerciasis patients who had no known evidence of autoimmune disease were found to contain antibodies that reacted with the 46-kD Ro/SS-A (calreticulin) by immunoblot analysis. Characteristic of sera reactive with Ro/SS-A antigens, some onchocerciasis sera also immunoprecipitated the Ro/SS-A-associated hY RNAs. In addition, a monoclonal antibody raised against O. volvulus organisms reacted to purified human WiL-2 cell 46 kD Ro/SS-A antigen (calreticulin) by ELISA. These results strongly suggest that onchocerciasis patients produce antibodies that crossreact with the 46-kD human Ro/SS-A autoantigen (calreticulin) and raise the possibility that infectious organisms such as O. volvulus might play a triggering or exacerbating role in the human Ro/SS-A autoimmune response.


MATLAB Simulation of UPQC for Power Quality Mitigation Using an Ant Colony Based Fuzzy Control Technique.
This paper proposes a fuzzy logic based new control scheme for the Unified Power Quality Conditioner (UPQC) for minimizing the voltage sag and total harmonic distortion in the distribution system consequently to improve the power quality. UPQC is a recent power electronic module which guarantees better power quality mitigation as it has both series-active and shunt-active power filters (APFs). The fuzzy logic controller has recently attracted a great deal of attention and possesses conceptually the quality of the simplicity by tackling complex systems with vagueness and ambiguity. In this research, the fuzzy logic controller is utilized for the generation of reference signal controlling the UPQC. To enable this, a systematic approach for creating the fuzzy membership functions is carried out by using an ant colony optimization technique for optimal fuzzy logic control. An exhaustive simulation study using the MATLAB/Simulink is carried out to investigate and demonstrate the performance of the proposed fuzzy logic controller and the simulation results are compared with the PI controller in terms of its performance in improving the power quality by minimizing the voltage sag and total harmonic distortion.


Respiratory Protection Considerations for Healthcare Workers During the COVID-19 Pandemic.
The COVID-19 pandemic has resulted in a surge of patients that exceeds available human and physical resources in many settings, triggering the implementation of crisis standards of care. High-quality respiratory protection is essential to reduce exposure among healthcare workers, yet dire shortages of personal protective equipment in the United States threaten the health and safety of this essential workforce. In the context of rapidly changing conditions and incomplete data, this article outlines 3 important strategies to improve healthcare workers' respiratory protection. At a minimum, healthcare workers delivering care to patients with confirmed or suspected COVID-19 should wear N95 respirators and full-face shields. Several mechanisms exist to boost and protect the supply of N95 respirators, including rigorous decontamination protocols, invoking the Defense Production Act, expanded use of reusable elastomeric respirators, and repurposing industrial N95 respirators. Finally, homemade facial coverings do not protect healthcare workers and should be avoided. These strategies, coupled with longer-term strategies of investments in protective equipment research, infrastructure, and data systems, provide a framework to protect healthcare workers immediately and enhance preparedness efforts for future pandemics.


Bioprospection of marine microorganisms: potential and challenges for Argentina.
The marine environments of Argentina have a remarkable extension, as well as high biological productivity and biodiversity of both macro- and microorganisms. Despite having a great potential for biotechnological applications, the microorganisms inhabiting these ecosystems remain mostly unexplored and unexploited. In this review, we study the research topics and the interactions among Argentinean laboratories, by analyzing current articles published on biotechnology-related marine microbiology by researchers of this country. In addition, we identify the challenges and opportunities for Argentina to take advantage of the genetic potential of its marine microorganisms. Finally, we suggest possible actions that could improve the development of this research field, as well as the utilization of this knowledge to solve societal needs.


Dual-energy micro-CT of the rodent lung.
The purpose of this work is to investigate the use of dual-energy micro-computed tomography (CT) for the estimation of vascular, tissue, and air fractions in rodent lungs using a postreconstruction three material decomposition method. Using simulations, we have estimated the accuracy limits of the decomposition for realistic micro-CT noise levels. Next, we performed experiments involving ex vivo lung imaging in which intact rat lungs were carefully removed from the thorax, injected with an iodine-based contrast agent, and then inflated with different volumes of air (n = 2). Finally, we performed in vivo imaging studies in C57BL/6 mice (n = 5) using fast prospective respiratory gating in end inspiration and end expiration for three different levels of positive end expiratory pressure (PEEP). Before imaging, mice were injected with a liposomal blood pool contrast agent. The three-dimensional air, tissue, and blood fraction maps were computed and analyzed. The results indicate that separation and volume estimation of the three material components of the lungs are possible. The mean accuracy values for air, blood, and tissue were 93, 93, and 90%, respectively. The absolute accuracy in determining all fraction materials was 91.6%. The coefficient of variation was small (2.5%) indicating good repeatability. The minimum difference that we could detect in material fractions was 15%. As expected, an increase in PEEP levels for the living mouse resulted in statistically significant increases in air fractions at end expiration but no significant changes at end inspiration. Our method has applicability in preclinical pulmonary studies where changes in lung structure and gas volume as a result of lung injury, environmental exposures, or drug bioactivity would have important physiological implications.


[Molecularly imprinted polymers in electro analysis of proteins].
In the review the main approaches to creation of recognition materials capable of competing with biological specific receptors, (polymeric analogs of antibodies or molecularly imprinted polymers, MIP) for the electro analysis of functionally significant proteins such as a myoglobin, troponin T, albumin, human ferritin, calmodulin are considered. The main types of monomers for MIP fabrication, and methods for MIP/protein interactions, such as a surface plasmon resonance (SPR), nanogravimetry with use of the quartz crystal resonator (QCM), spectral and electrochemical methods are discussed. Experimental data on electrochemical registration of a myoglobin using MIP/electrode are presented. For a development of electrochemical sensor systems based on MIPs, o-phenylenediamine (1,2-diaminobenzene was used as a monomer. It was shown that the imprinting factor Imax(MIP)/Imax(NIP), calculated as a myoglobin signal ratio when embedding in MIP to a myoglobin signal when embedding in the polymer received without molecular template (NIP) corresponds 2-4.


Single-cell genomics reveal low recombination frequencies in freshwater bacteria of the SAR11 clade.
The SAR11 group of Alphaproteobacteria is highly abundant in the oceans. It contains a recently diverged freshwater clade, which offers the opportunity to compare adaptations to salt- and freshwaters in a monophyletic bacterial group. However, there are no cultivated members of the freshwater SAR11 group and no genomes have been sequenced yet. We isolated ten single SAR11 cells from three freshwater lakes and sequenced and assembled their genomes. A phylogeny based on 57 proteins indicates that the cells are organized into distinct microclusters. We show that the freshwater genomes have evolved primarily by the accumulation of nucleotide substitutions and that they have among the lowest ratio of recombination to mutation estimated for bacteria. In contrast, members of the marine SAR11 clade have one of the highest ratios. Additional metagenome reads from six lakes confirm low recombination frequencies for the genome overall and reveal lake-specific variations in microcluster abundances. We identify hypervariable regions with gene contents broadly similar to those in the hypervariable regions of the marine isolates, containing genes putatively coding for cell surface molecules. We conclude that recombination rates differ dramatically in phylogenetic sister groups of the SAR11 clade adapted to freshwater and marine ecosystems. The results suggest that the transition from marine to freshwater systems has purged diversity and resulted in reduced opportunities for recombination with divergent members of the clade. The low recombination frequencies of the LD12 clade resemble the low genetic divergence of host-restricted pathogens that have recently shifted to a new host.


An ancient group I intron shared by eubacteria and chloroplasts.
Introns have been found in the genomes of all major groups of organisms except eubacteria. The presence of introns in chloroplasts and mitochondria, both of which are of eubacterial origin, has been interpreted as evidence either for the recent acquisition of introns by organelles or for the loss of introns from their eubacterial progenitors. The gene for the leucine transfer RNA with a UAA anticodon [tRNALeu (UAA)] from five diverse cyanobacteria and several major groups of chloroplasts contains a single group I intron. The intron is conserved in secondary structure and primary sequence, and occupies the same position, within the UAA anticodon. The homology of the intron across chloroplasts and cyanobacteria implies that it was present in their common ancestor and that it has been maintained in their genomes for at least 1 billion years.


The impact of the gut microbiome on memory and sleep in Drosophila.
The gut microbiome has been proposed to influence diverse behavioral traits of animals, although the experimental evidence is limited and often contradictory. Here, we made use of the tractability of Drosophila melanogaster for both behavioral analyses and microbiome studies to test how elimination of microorganisms affects a number of behavioral traits. Relative to conventional flies (i.e. with unaltered microbiome), microbiologically sterile (axenic) flies displayed a moderate reduction in memory performance in olfactory appetitive conditioning and courtship assays. The microbiological status of the flies had a small or no effect on anxiety-like behavior (centrophobism) or circadian rhythmicity of locomotor activity, but axenic flies tended to sleep for longer and displayed reduced sleep rebound after sleep deprivation. These last two effects were robust for most tests conducted on both wild-type Canton S and w1118 strains, as well for tests using an isogenized panel of flies with mutations in the period gene, which causes altered circadian rhythmicity. Interestingly, the effect of absence of microbiota on a few behavioral features, most notably instantaneous locomotor activity speed, varied among wild-type strains. Taken together, our findings demonstrate that the microbiome can have subtle but significant effects on specific aspects of Drosophila behavior, some of which are dependent on genetic background.


Inhibition of ethanol-inducible cytochrome P450IIE1 by 3-amino-1,2,4-triazole.
The p-nitrophenol hydroxylase activity of hepatic microsomes from acetone-treated rabbits was inhibited by 3-amino-1,2,4-triazole in a time- and NADPH-dependent manner. The loss of p-nitrophenol hydroxylation, an activity catalyzed predominately by P450IIE1, displayed a number of characteristics consistent with suicide inhibition of the enzyme. These include irreversibility, saturability, similarity of the effect of pH on the rate constant for inactivation and catalysis by the isozyme, protection by substrate, and the lack of an effect of exogenous nucleophiles on the inactivation. At pH 6.8, the KI for 3-amino-1,2,4-triazole for inactivation was 57 mM and the maximal rate of inactivation was 0.43 min-1. The inactivation of hepatic microsomes resulted in a loss of spectrally detectable P450 which was correlated with the concentration of P450IIE1 in various microsomal preparations. Purified P450IIE1 was rapidly autoinactivated (kinact of about 0.1 min-1) in the presence of NADPH and P450 reductase. However, the autoinactivation was completely prevented by the addition of catalase. In the presence of catalase, purified P450IIE1 was inactivated in a time- and concentration-dependent manner by 3-amino-1,2,4-triazole (KI was 10 mM and the maximal rate of inactivation was 0.44 min-1). The inactivation resulted in the loss of spectrally detectable P450 but did not cause the formation of P420 or a loss of heme as determined by the reduced pyridine hemochrome. The spectrum of the inactivated enzyme exhibited a decreased extinction in the Soret region with a broad maximum at 378 nm and a shoulder around 428 nm. Inactivated P450IIE1 did not show a characteristic low-spin spectrum in the presence of 4-methylpyrazole. When 3-amino-1,2,4-[5-14C]triazole was used in the inactivation reaction, there was no significant incorporation of radioactivity into the protein or heme; these results suggest that the inactivation may be due to covalent binding of the heme to the protein or the modification of residues near the heme, which prevent substrate interaction. The effective inhibition of P450IIE1 by 3-amino-1,2,4-triazole suggests that the compound may be useful for the identification of P450IIE1-dependent microsomal catalysis.


Association between asbestos exposure and pericardial and tunica vaginalis testis malignant mesothelioma: a case-control study and epidemiological remarks.
Objectives The purposes of this study are to describe the epidemiology of pericardial and tunica vaginalis testis mesothelioma and assess the role of asbestos exposure for these rare diseases. Methods Based on incident pericardial and tunica vaginalis testis mesothelioma cases collected from the Italian national mesothelioma registry (ReNaM) in the period 1993-2015, incidence rates, survival median period and prognostic factors have been evaluated. A case-control study has been performed to analyze the association with asbestos exposure (occupational and non-occupational) for these diseases. Results Between 1993 and 2015, 58 pericardial (20 women and 38 men) and 80 tunica vaginalis testis mesothelioma cases have been registered with a mean annual standardized (world standard population as reference) incidence rates of 0.049 (per million) in men and 0.023 in women for the pericardial site, and 0.095 for tunica vaginalis testis mesothelioma. Occupational exposure to asbestos was significantly associated with the risk of the diseases [odds ratio (OR) 3.68, 95% confidence interval (CI) 1.85-7.31 and OR 3.42, 95% CI 1.93-6.04 in pericardial and tunica vaginalis testis mesothelioma, respectively]. The median survival was 2.5 months for pericardial and 33.0 months for tunica vaginalis testis mesotheliomas. Age was the main predictive factor for survival for both anatomical sites. Conclusions For the first time in an analytical study, asbestos exposure was associated with pericardial and tunica vaginalis testis mesothelioma risk, supporting the causal role of asbestos for all anatomical sites. The extreme rarity of the diseases, the poor survival and the prognostic role of age have been confirmed based on population and nationwide mesothelioma registry data.


After-hyperpolarization currents and acetylcholine control sigmoid transfer functions in a spiking cortical model.
Recurrent networks are ubiquitous in the brain, where they enable a diverse set of transformations during perception, cognition, emotion, and action. It has been known since the 1970's how, in rate-based recurrent on-center off-surround networks, the choice of feedback signal function can control the transformation of input patterns into activity patterns that are stored in short term memory. A sigmoid signal function may, in particular, control a quenching threshold below which inputs are suppressed as noise and above which they may be contrast enhanced before the resulting activity pattern is stored. The threshold and slope of the sigmoid signal function determine the degree of noise suppression and of contrast enhancement. This article analyses how sigmoid signal functions and their shape may be determined in biophysically realistic spiking neurons. Combinations of fast, medium, and slow after-hyperpolarization (AHP) currents, and their modulation by acetylcholine (ACh), can control sigmoid signal threshold and slope. Instead of a simple gain in excitability that was previously attributed to ACh, cholinergic modulation may cause translation of the sigmoid threshold. This property clarifies how activation of ACh by basal forebrain circuits, notably the nucleus basalis of Meynert, may alter the vigilance of category learning circuits, and thus their sensitivity to predictive mismatches, thereby controlling whether learned categories code concrete or abstract information, as predicted by Adaptive Resonance Theory.


Resilience to bacterial infection: difference between species could be due to proteins in serum.
Vertebrates vary in resistance and resilience to infectious diseases, and the mechanisms that regulate the trade-off between these often opposing protective processes are not well understood. Variability in the sensitivity of species to the induction of damaging inflammation in response to equivalent pathogen loads (resilience) complicates the use of animal models that reflect human disease. We found that induction of proinflammatory cytokines from macrophages in response to inflammatory stimuli in vitro is regulated by proteins in the sera of species in inverse proportion to their in vivo resilience to lethal doses of bacterial lipopolysaccharide over a range of 10,000-fold. This finding suggests that proteins in serum rather than intrinsic cellular differences may play a role in regulating variations in resilience to microbe-associated molecular patterns between species. The involvement of circulating proteins as key molecules raises hope that the process might be manipulated to create better animal models and potentially new drug targets.


Estimation of connectional brain templates using selective multi-view network normalization.
The brain connectome encodes different facets of the brain construct such as function and structure in a network. Noting that a brain network captures the individual signature of a particular subject, it remains a formidable challenge to extract a shared and representative brain signature across a population of brain networks, let alone multi-view brain networks. In this paper, we propose netNorm, a method that can meet this challenge by normalizing a population of multi-view brain networks, where each brain network represents a particular view of the brain, acquired using a neuroimaging technique. While conventional methods integrate the network views equally at a global scale, we propose a selective technique which unfolds the fusion process at a local scale by first selecting for each local pairwise connectivity between two anatomical regions of interest the most representative cross-view feature vector in the population. By combining the selected cross-view feature vectors, we then estimate a population representative tensor. Such multi-view representation captures the most shared traits across all subjects and thereby occupies a centered location compared to all views. In the final step, netNorm non-linearly fuses the frontal views of the estimated representative population tensor into a single network depicting the final brain connectional template. We demonstrate the broad applicability of our method on four connectomic datasets and we show that netNorm (i) produces the most centered and representative connectional brain template (CBT) that consistently captures the unique and distinctive traits of a population of multi-view brain networks, and (ii) identifies disordered brain connections by comparing templates estimated using disordered and healthy brains, respectively, demonstrating the discriminative power of the estimated CBTs. This allows to rapidly and efficiently spot atypical deviations from the normal brain connectome for comparative studies, circumventing the need to use machine learning techniques for discriminative feature identification.


Description and implementation of an On-farm Wildlife Risk Mitigation Protocol at the wildlife-livestock interface: Tuberculosis in Mediterranean environments.
Animal tuberculosis (TB), which is caused by the Mycobacterium tuberculosis complex (MTC), is a zoonotic disease of global concern, and has a wide variety of wild and domestic reservoirs that can establish complex epidemiological systems. Of all the strategies employed to control TB, reducing the risks of interaction at the wildlife-livestock interface is a cornerstone. However, detailed protocols with which to assess and implement farm-specific preventive actions that can be employed against interactions with wildlife are lacking for extensive production systems. We describe an On-farm Wildlife Risk Mitigation Protocol that is applicable to beef cattle farming in Mediterranean environments in order to control the wildlife-livestock interaction and MTC transmission through the use of Farm-specific Action Plans (FsAP). We assessed the implementation and verification of FsAP in terms of its practical feasibility and acceptability by farmers (n=55 farms). Of the potential risk points, waterers (41.3 %) and waterholes (24.4 %) were the most common. Waterholes and springs were identified as the points with the greatest risks. Actions related to water management were essential on most farms (99 % of the high-risk points), as were those regarding wildlife management (36.4 % of the farms provided wild boar or cervids with supplementary food for hunting purposes). Overall, 75 % of the farmers adopted the plans to some extent, with an average of 31.8 % of actions implemented, but with high variability depending on the type of actions proposed. Farmers prioritised low-cost measures. Our results, in their entirety, indicate that the adoption of this On-farm Wildlife Risk Mitigation Protocol is practical and feasible in Mediterranean ecosystems, and can be easily transferred to professionals and adapted to other bioregions or epidemiological systems. The subsequent evaluation of FsAPs in terms of efficacy and cost-effectiveness, along with increasing their acceptance by farmers, are necessary steps for the further development of TB Risk Mitigation Programmes at a nationwide level.


Data-driven neuropathological staging and subtyping of TDP-43 proteinopathies.
TAR DNA-binding protein-43 (TDP-43) accumulation is the primary pathology underlying several neurodegenerative diseases. Charting the progression and heterogeneity of TDP-43 accumulation is necessary to better characterize TDP-43 proteinopathies, but current TDP-43 staging systems are heuristic and assume each syndrome is homogeneous. Here, we use data-driven disease progression modelling to derive a fine-grained empirical staging system for the classification and differentiation of frontotemporal lobar degeneration due to TDP-43 (FTLD-TDP, n = 126), amyotrophic lateral sclerosis (ALS, n = 141) and limbic-predominant age-related TDP-43 encephalopathy neuropathologic change (LATE-NC) with and without Alzheimer's disease (n = 304). The data-driven staging of ALS and FTLD-TDP complement and extend previously described human-defined staging schema for ALS and behavioural variant frontotemporal dementia. In LATE-NC individuals, progression along data-driven stages was positively associated with age, but negatively associated with age in individuals with FTLD-TDP. Using only regional TDP-43 severity, our data driven model distinguished individuals diagnosed with ALS, FTLD-TDP or LATE-NC with a cross-validated accuracy of 85.9%, with misclassifications associated with mixed pathological diagnosis, age and genetic mutations. Adding age and SuStaIn stage to this model increased accuracy to 92.3%. Our model differentiates LATE-NC from FTLD-TDP, though some overlap was observed between late-stage LATE-NC and early-stage FTLD-TDP. We further tested for the presence of subtypes with distinct regional TDP-43 progression patterns within each diagnostic group, identifying two distinct cortical-predominant and brainstem-predominant subtypes within FTLD-TDP and a further two subcortical-predominant and corticolimbic-predominant subtypes within ALS. The FTLD-TDP subtypes exhibited differing proportions of TDP-43 type, while there was a trend for age differing between ALS subtypes. Interestingly, a negative relationship between age and SuStaIn stage was seen in the brainstem/subcortical-predominant subtype of each proteinopathy. No subtypes were observed for the LATE-NC group, despite aggregating individuals with and without Alzheimer's disease and a larger sample size for this group. Overall, we provide an empirical pathological TDP-43 staging system for ALS, FTLD-TDP and LATE-NC, which yielded accurate classification. We further demonstrate that there is substantial heterogeneity amongst ALS and FTLD-TDP progression patterns that warrants further investigation in larger cross-cohort studies.


zA map for sequence analysis of the Arabidopsis thaliana genome.
Arabidopsis thaliana has emerged as a model system for studies of plant genetics and development, and its genome has been targeted for sequencing by an international consortium (the Arabidopsis Genome Initiative; http://genome-www. stanford.edu/Arabidopsis/agi.html). To support the genome-sequencing effort, we fingerprinted more than 20,000 BACs (ref. 2) from two high-quality publicly available libraries, generating an estimated 17-fold redundant coverage of the genome, and used the fingerprints to nucleate assembly of the data by computer. Subsequent manual revision of the assemblies resulted in the incorporation of 19,661 fingerprinted BACs into 169 ordered sets of overlapping clones ('contigs'), each containing at least 3 clones. These contigs are ideal for parallel selection of BACs for large-scale sequencing and have supported the generation of more than 5.8 Mb of finished genome sequence submitted to GenBank; analysis of the sequence has confirmed the integrity of contigs constructed using this fingerprint data. Placement of contigs onto chromosomes can now be performed, and is being pursued by groups involved in both sequencing and positional cloning studies. To our knowledge, these data provide the first example of whole-genome random BAC fingerprint analysis of a eucaryote, and have provided a model essential to efforts aimed at generating similar databases of fingerprint contigs to support sequencing of other complex genomes, including that of human.


A New biological proxy for deep-sea paleo-oxygen: Pores of epifaunal benthic foraminifera.
The negative consequences of fossil fuel burning for the oceans will likely include warming, acidification and deoxygenation, yet predicting future deoxygenation is difficult. Sensitive proxies for oxygen concentrations in ancient deep-ocean bottom-waters are needed to learn from patterns of marine deoxygenation during global warming conditions in the geological past. Understanding of past oxygenation effects related to climate change will better inform us about future patterns of deoxygenation. Here we describe a new, quantitative biological proxy for determining ocean paleo-oxygen concentrations: the surface area of pores (used for gas exchange) in the tests of deep-sea benthic foraminifera collected alive from 22 locations (water depths: 400 to 4100 m) at oxygen levels ranging from ~ 2 to ~ 277 μmol/l. This new proxy is based on species that are widely distributed geographically, bathymetrically and chronologically, and therefore should have broad applications. Our calibration demonstrates a strong, negative logarithmic correlation between bottom-water oxygen concentrations and pore surface area, indicating that pore surface area of fossil epifaunal benthic foraminifera can be used to reconstruct past changes in deep ocean oxygen and redox levels.


The effects of aerobic exercise training on mental health and self-esteem of type 2 diabetes mellitus patients.
Physical and mental health are very closely tied and either of them can exert a significant effect on the other. The aim of the present study was to examine the effects of aerobic exercise training on mental health and self-esteem of type 2 diabetes mellitus patients. 60 participants, aged 40 to 55 years, with blood sugar ranging between 150 to 250 mg/dL participated in the study. They were randomly divided into two groups of 30 patients. The inter-ventional group exercised for twelve weeks, three sessions per week, lasting from 45 to 60 minutes, followed by endurance training on treadmill. Training's intensity was considered equal to 60-70% of maximum oxygen consumption. According to the results of the study, 12-week aerobic exercise training had significant effects on self-esteem (P=0.001), and mental health (P=0.020), sub-scales of physical symptoms (P=0.001), and anxiety and insomnia (P=0.044). But it had no significant effects on the sub-scales of depression (P=0.078) and social functioning (P=0.207). Regular aerobic exercise training as an effective strategy plays an important role in improving self-esteem and mental health and also promoting life quality among diabetic patients.


Dry powder PA-824 aerosols for treatment of tuberculosis in guinea pigs.
Novel treatments for multidrug-resistant tuberculosis (MDR-TB), extensively drug-resistant tuberculosis (XDR-TB), or latent TB are needed urgently. Recently, we reported the formulation and characterization of the nitroimidazo-oxazine PA-824 for efficient aerosol delivery as dry powder porous particles and the subsequent disposition in guinea pigs after pulmonary administration. The objective of the present study was to evaluate the effects of these PA-824 therapeutic aerosols on the extent of TB infection in the low-inoculum aerosol infection guinea pig model. Four weeks after infection by the pulmonary route, animals received daily treatment for 4 weeks of either a high or a low dose of PA-824 dry powder aerosol. Animals received PA-824 cyclodextrin/lecithin suspensions orally as positive controls, and those receiving placebo particles or no treatment were negative controls. The lungs and spleens of animals receiving the high dose of inhaled PA-824 particles exhibited a lower degree of inflammation (indicated by wet tissue weights), bacterial burden, and tissue damage (indicated by histopathology) than those of untreated or placebo animals. Treatment with oral PA-824 cyclodextrin/lecithin suspension resulted in a more significant reduction in the bacterial burden of lungs and spleen, consistent with a dose that was larger than inhaled doses (eight times the inhaled low dose and four times the inhaled high dose). However, histopathological analysis revealed that the extent of tissue damage was comparable in groups receiving the oral or either inhaled dose. The present studies indicate the potential use of PA-824 dry powder aerosols in the treatment of TB.


The response of cancers to BRAF inhibition underscores the importance of cancer systems biology.
The BRAF inhibitor vemurafenib has become an important treatment option for melanoma patients, the majority of whom have a BRAF(V600E) mutation driving their malignancy. However, this same agent does not generally benefit colon cancer patients who have the BRAF(V600E) mutation. Recent work suggests that BRAF(V600E) inhibition by vemurafenib results in decreased negative feedback to the epidermal growth factor receptor (EGFR) pathway and that the different clinical responses are due to differences in the amount of EGFR present in these two cancers. The experimental work that identified the feedback signaling was an elegant mix of functional genomic approaches and focused, hypothesis-driven cellular and molecular biology. The results of these studies suggest that combined treatment of BRAF(V600E)-driven colon cancers with both vemurafenib and EGFR inhibitors is worth clinical evaluation.


Kinesin-8 motors act cooperatively to mediate length-dependent microtubule depolymerization.
Motor proteins in the kinesin-8 family depolymerize microtubules in a length-dependent manner that may be crucial for controlling the length of organelles such as the mitotic spindle. We used single-molecule microscopy to understand the mechanism of length-dependent depolymerization by the budding yeast kinesin-8, Kip3p. We found that after binding at a random position on a microtubule and walking to the plus end, an individual Kip3p molecule pauses there until an incoming Kip3p molecule bumps it off. Kip3p dissociation is accompanied by removal of just one or two tubulin dimers (on average). Such a cooperative mechanism leads to a depolymerization rate that is proportional to the flux of motors to the microtubule end and accounts for the length dependence of depolymerization. This type of feedback between length and disassembly may serve as a model for understanding how an ensemble of molecules can measure and control polymer length.


A double-blind, placebo-controlled study of fluvoxamine in adults with autistic disorder.
Autistic disorder is characterized by a fundamental disturbance in social interaction, impairments in communication, and a markedly restricted repertoire of activities and interests. Abnormalities in the serotonin neurotransmitter system have been identified in some persons with autism. No consistently effective and safe drugs have been developed for treating the symptoms of autism. Thirty adults with autistic disorder completed a 12-week double-blind, placebo-controlled trial of the potent and selective serotonin uptake inhibitor fluvoxamine maleate. Behavioral ratings were obtained at baseline and after 4, 8, and 12 weeks of treatment. Eight (53%) of 15 patients in the fluvoxamine-treated group were categorized as responders compared with none of 15 in the placebo group (P = .001). Fluvoxamine was superior to placebo in reducing repetitive thoughts and behavior (P < .001), maladaptive behavior (P < .001), and aggression (P < .03), and in improving some aspects of social relatedness (P < .04), especially language usage (P < .008). Treatment response was not correlated with age level of autistic behavior, or full-scale IQ. Other than mild sedation and nausea in a few patients, fluvoxamine was well tolerated. No dyskinesias, adverse cardiovascular events, or seizures occurred. Fluvoxamine is more effective than placebo in the short-term treatment of the symptoms of autistic disorder in adults. Controlled studies of fluvoxamine and other potent and selective serotonin uptake inhibitors seem warranted in children and adolescents with autism.


Metastasis-associated gene 1 (MTA1) enhances cisplatin resistance of malignant pleural mesothelioma by ATR-Chk1-mediated DNA repair.
Malignant pleural mesothelioma (MPM) chemoresistance remains a challenge to oncologists. In our previous study, we demonstrated that the aberrant expression of metastasis-associated gene 1 (MTA1) is associated with carcinogenesis and metastasis in MPM. The aim of the present study was to investigate the mechanism of MTA1 and chemo-resistance in MPM. Western blotting and real-time polymerase chain reaction were used to analyze the protein and mRNA levels. A stable clone with a knockdown of MTA1 was generated with shRNA via lentivirus technology in MPM cell lines. Cell Counting Kit-8 assay and crystal violet assay were used to measure cell viability. Immunochemical staining was employed to detect MTA1 expression in MPM tissues. The cell cycle of MPM cells was determined by phosphohistone H3 staining and flow cytometric analysis. The MTA1 protein was upregulated and enhanced cisplatin resistance in MPM. Cisplatin stabilized the expression of the MTA1 protein by inhibiting its ubiquitination, and MTA1 enhanced G2/M cell cycle delay and regulated and protected the tumor genome from chemotherapeutic drugs via participating in the phosphorylation of the ataxia telangiectasia mutated and rad3 related-checkpoint kinase 1 (ATR-Chk1) pathway. These data suggest that MTA1 enhances cisplatin resistance by ATR-Chk1-mediated DNA damage repairment and cisplatin stabilizes MTA1 expression via affecting on the ubiquitination pathway of MTA1 in MPM. Our findings indicate that MTA1 could serve as a novel therapeutic target to overcome chemoresistance in MPM.


University-based peer health education in China: the Shantou experience.
University-based peer health education is a recent development in China. The authors evaluated a newly implemented program in the Guangdong province. In September 2006, the authors conducted a cross-sectional study using self-administered questionnaires on 30 peer educators and 247 students. All peer educators and the majority of student respondents positively evaluated the program. Although students preferred to seek health information online, approximately one-quarter of the student respondents would contact peer educators. Third-year students were more than twice as likely (29.1%) to contact peer educators than were fourth-year students (13.1%). The peer educators perceived diet, physical activity, safer sex, and mental health as the most relevant student health topics. Peer educators cited acquiring factual information and medical skills, rather than personal development, as the most important things learned from the program. Despite some promising results, Western-based peer education models may require cultural adaptation for greater effectiveness in China.


Mitochondrial modulation of calcium signaling at the initiation of development.
Fertilization triggers cytosolic Ca(2+) oscillations that activate mammalian eggs and initiate development. Extensive evidence demonstrates that Ca(2+) is released from endoplasmic reticulum stores; however, less is known about how the increased Ca(2+) is restored to its resting level, forming the Ca(2+) oscillations. We investigated whether mitochondria also play a role in activation-associated Ca(2+) signaling. Mitochondrial dysfunction induced by the mitochondrial uncoupler FCCP or antimycin A disrupted cytosolic Ca(2+) oscillations, resulting in sustained increase in cytosolic Ca(2+), followed by apoptotic cell death. This suggests that functional mitochondria may participate in sequestering the released Ca(2+), contributing to cytosolic Ca(2+) oscillations and preventing cell death. By centrifugation, mouse eggs were stratified and separated into fractions containing both endoplasmic reticulum and mitochondria and fractions containing endoplasmic reticulum with no mitochondria. The former showed Ca(2+) oscillations by activation, whereas the latter exhibited sustained elevation in cytosolic Ca(2+) but no Ca(2+) oscillations, suggesting that mitochondria take up released cytosolic Ca(2+). Further, using Rhod-2 for detection of mitochondrial Ca(2+), we found that mitochondria exhibited Ca(2+) oscillations, the frequency of which was not different from that of cytosolic Ca(2+) oscillations, indicating that mitochondria are involved in Ca(2+) signaling during egg activation. Therefore, we propose that mitochondria play a crucial role in Ca(2+) signaling that mediates egg activation and development, and apoptotic cell death.


Clinical evaluation of a method for detecting superficial surgical transitional cell carcinoma of the bladder by light-induced fluorescence of protoporphyrin IX following the topical application of 5-aminolevulinic acid: preliminary results.
In bladder cancer, conventional white light endoscopic examination of the bladder does not provide adequate information about the presence of "flat" urothelial lesions such as carcinoma in situ. In the present investigation, we examine a new technique for the photodetection of such lesions by the imaging of protoporphyrin IX (PpIX) fluorescence following topical application of 5-aminolevulinic acid (ALA). Several hours after bladder instillation of an aqueous solution of ALA in 34 patients, a Krypton ion laser or a filtered Xenon arc-lamp was used to excite PpIX fluorescence. Tissue samples for histological analysis were taken while observing the bladder wall either by means of a video camera, or by direct endoscopic observation. A good correlation was found between the PpIX fluorescence and the histopathological diagnosis. On a total of 215 biopsies, 143 in fluorescent and 72 in nonfluorescent areas, all visible tumors on white light cytoscopy appeared in a bright red fluorescence with the photodetection technique. In addition, this method permitted to discover 47 unsuspected carcinomatous lesions on white light observation, among which 40% were carcinoma in situ. PpIX fluorescence induced by instillation into the bladder of 5-ALA is an efficient method of mapping the mucosa in bladder carcinoma.


Nature and type of patient-reported safety incidents in primary care: cross-sectional survey of patients from Australia and England.
Patient engagement in safety has shown positive effects in preventing or reducing adverse events and potential safety risks. Capturing and utilising patient-reported safety incident data can be used for service learning and improvement. The aim of this study was to characterise the nature of patient-reported safety incidents in primary care. Secondary analysis of two cross sectional studies. Adult patients from Australian and English primary care settings. Patients' self-reported experiences of safety incidents were captured using the validated Primary Care Patient Measure of Safety questionnaire. Qualitative responses to survey items were analysed and categorised using the Primary Care Patient Safety Classification System. The frequency and type of safety incidents, contributory factors, and patient and system level outcomes are presented. A total of 1329 patients (n=490, England; n=839, Australia) completed the questionnaire. Overall, 5.3% (n=69) of patients reported a safety incident over the preceding 12 months. The most common incident types were administration incidents (n=27, 31%) (mainly delays in accessing a physician) and incidents involving diagnosis and assessment (n=16, 18.4%). Organisation of care accounted for 27.6% (n=29) of the contributory factors identified in the safety incidents. Staff factors (n=13, 12.4%) was the second most commonly reported contributory factor. Where an outcome could be determined, patient inconvenience (n=24, 28.6%) and clinical harm (n=21, 25%) (psychological distress and unpleasant experience) were the most frequent. The nature and outcomes of patient-reported incidents differ markedly from those identified in studies of staff-reported incidents. The findings from this study emphasise the importance of capturing patient-reported safety incidents in the primary care setting. The patient perspective can complement existing sources of safety intelligence with the potential for service improvement.


Vibrational structure of dihydrofolate bound to R67 dihydrofolate reductase.
R67 is a Type II dihydrofolate reductase (DHFR) that catalyzes the reduction of dihydrofolate (DHF) to tetrahydrofolate by facilitating the addition of a proton to N5 of DHF and the transfer of a hydride ion from NADPH to C6. Because this enzyme is a plasmid-encoded DHFR from trimethoprim-resistant bacteria, extensive studies on R67 with various methods have been performed to elucidate its reaction mechanism. Here, Raman difference measurements, conducted on the ternary complex of R67.NADP(+).DHF believed to be an accurate mimic of the productive DHFR.NADPH.DHF complex, show that the pK(a) of N5 in the complex is less than 4. This is in clear contrast to the behavior observed in Escherichia coli DHFR, a substantially more efficient enzyme, where the pK(a) of bound DHF at N5 is increased to 6.5 compared with its solution value of 2.6. A comparison of the ternary complexes in R67 and E. coli DHFRs suggests that enzymic raising of the pK(a) at N5 can significantly increase the catalytic efficiency of the hydride transfer step. However, R67 shows that even without such a strategy an effective DHFR can still be designed.


Views of US physicians about controlling health care costs.
Physicians' views about health care costs are germane to pending policy reforms. To assess physicians' attitudes toward and perceived role in addressing health care costs. A cross-sectional survey mailed in 2012 to 3897 US physicians randomly selected from the AMA Masterfile. Enthusiasm for 17 cost-containment strategies and agreement with an 11-measure cost-consciousness scale. A total of 2556 physicians responded (response rate = 65%). Most believed that trial lawyers (60%), health insurance companies (59%), hospitals and health systems (56%), pharmaceutical and device manufacturers (56%), and patients (52%) have a "major responsibility" for reducing health care costs, whereas only 36% reported that practicing physicians have "major responsibility." Most were "very enthusiastic" for "promoting continuity of care" (75%), "expanding access to quality and safety data" (51%), and "limiting access to expensive treatments with little net benefit" (51%) as a means of reducing health care costs. Few expressed enthusiasm for "eliminating fee-for-service payment models" (7%). Most physicians reported being "aware of the costs of the tests/treatments [they] recommend" (76%), agreed they should adhere to clinical guidelines that discourage the use of marginally beneficial care (79%), and agreed that they "should be solely devoted to individual patients' best interests, even if that is expensive" (78%) and that "doctors need to take a more prominent role in limiting use of unnecessary tests" (89%). Most (85%) disagreed that they "should sometimes deny beneficial but costly services to certain patients because resources should go to other patients that need them more." In multivariable logistic regression models testing associations with enthusiasm for key cost-containment strategies, having a salary plus bonus or salary-only compensation type was independently associated with enthusiasm for "eliminating fee for service" (salary plus bonus: odds ratio [OR], 3.3, 99% CI, 1.8-6.1; salary only: OR, 4.3, 99% CI, 2.2-8.5). In multivariable linear regression models, group or government practice setting (β = 0.87, 95% CI, 0.29 to 1.45, P = .004; and β = 0.99, 95% CI, 0.20 to 1.79, P = .01, respectively) and having a salary plus bonus compensation type (β = 0.82; 95% CI, 0.32 to 1.33; P = .002) were positively associated with cost-consciousness. Finding the "uncertainty involved in patient care disconcerting" was negatively associated with cost-consciousness (β = -1.95; 95% CI, -2.71 to -1.18; P < .001). In this survey about health care cost containment, US physicians reported having some responsibility to address health care costs in their practice and expressed general agreement about several quality initiatives to reduce cost but reported less enthusiasm for cost containment involving changes in payment models.


c-erbB-2 oncoprotein expression in mammary and extramammary Paget's disease: an immunohistochemical study.
Over-expression of the c-erbB-2 oncogene occurs in a proportion of human adenocarcinomas and in breast carcinoma is associated with poorer prognosis. Sections of formalin-fixed, paraffin-embedded tumour tissue from 22 patients with mammary and extramammary Paget's disease have been stained immunohistochemically using a monoclonal antibody (NCL-CB11) raised against a synthetic peptide from the C-terminal end of the predicted sequence of the c-erbB-2 protein product. All 12 cases of mammary Paget's disease showed membrane staining of intra-epidermal cells, indicating c-erbB-2 over-expression. Sections of underlying ductal breast carcinoma were available in nine cases; all nine tumours were c-erbB-2 positive and in eight the in situ component was of comedo or solid type. There was membrane staining of tumour cells in four of the 10 cases of extramammary Paget's disease; staining intensity was generally weaker than that observed in the cases of mammary disease. The possible implications of these findings for the histogenesis of both mammary and extramammary Paget's disease are discussed.


Optimization of Peptide Vaccines to Induce Robust Antitumor CD4 T-cell Responses.
Substantial evidence indicates that immunotherapy is a feasible and effective approach for the treatment of numerous types of cancer. Among various immunotherapy options, peptide vaccines to generate antitumor T cells appear as promising candidates, because of their cost effectiveness and ease of implementation. Nevertheless, most peptide vaccines are notorious for being weekly immunogenic and, thus, optimization of the vaccination strategy is essential to achieve therapeutic effectiveness. In addition, effective peptide vaccines must stimulate both CD8 cytotoxic and CD4 helper T lymphocytes. Our group has been successful in designing effective peptide vaccination strategies for inducing CD8 T-cell responses in mouse tumor models. Here, we describe a somewhat similar, but distinct, peptide vaccination strategy capable of generating vast CD4 T-cell responses by combining synthetic peptides with toll-like receptor (TLR) agonists and OX40/CD40 costimulation. This vaccination strategy was efficient in overcoming immune tolerance to a self-tumor-associated antigen and generated significant antitumor effects in a mouse model of malignant melanoma. The optimized peptide vaccine also allowed the expansion of adoptively transferred CD4 T cells without the need for lymphodepletion and IL2 administration, generating effective antimelanoma responses through the enhancement of proliferative and antiapoptotic activities of CD4 T cells. These results have practical implications in the design of more effective T-cell-based immunotherapies. Cancer Immunol Res; 5(1); 72-83. ©2016 AACR.


Production of amyloid beta protein precursor as a proteinase inhibitor by human astrocytic tumors.
The production of amyloid beta protein precursor (APP), which is a potent inhibitor of matrix metalloproteinases and serine proteinases, in human astrocytic tumors (n = 17) and normal brain tissues (n = 3) was investigated. We found proteinase inhibitory activity at around 120 kD by trypsin reverse zymography in the culture media of explant cultures of anaplastic astrocytomas and glioblastomas, but not in those of astrocytomas and normal brain tissues. Immunohistochemistry using a monoclonal antibody against human APP demonstrated that APP was detectable mainly in tumor and endothelial cells. Semiquantative analysis of western blotting revealed that immunoreactivity for APP in the culture media of tumor explant cultures appeared to be increased associated with the malignancy of astrocytic tumors. These findings suggest that APP production may be related to the malignant progression of human astrocytic tumors.


Predictive value of early postoperative IOP and bleb morphology in Mitomycin-C augmented trabeculectomy.
Background: To determine the predictive value of postoperative bleb morphological features and intraocular pressure (IOP) on the success rate of trabeculectomy. Methods: In this prospective interventional case series, we analyzed for one year 80 consecutive primary open angle glaucoma patients who underwent mitomycin-augmented trabeculectomy. Bleb morphology was scored using the Indiana bleb appearance grading scale (IBAGS). Success was defined as IOP ≤15 mmHg at 12 months. We applied a multivariable regression analysis and determined the area under the receiver operating characteristic curve (AUC). Results: The mean age of participants was 62±12.3 years in the success and 63.2±16.3 years in the failure group (P= 0.430) with equal gender distribution (P=0.911). IOPs on day 1, 7 and 30 were similar in both (P= 0.193, 0.639, and 0.238, respectively.) The AUC of IOP at day 1, day 7 and 30 for predicting a successful outcome was 0.355, 0.452, and 0.80, respectively. The AUC for bleb morphology parameters of bleb height, extension, and vascularization, on day 14 were 0.368, 0.408, and 0.549, respectively. Values for day 30 were 0.428, 0.563, and 0.654. IOP change from day 1 to day 30 was a good predictor of failure (AUC=0.838, 95% CI: 0.704 to 0.971) with a change of more than 3 mmHg predicting failure with a sensitivity of 82.5% (95% CI: 68 to 91%) and a specificity of 87.5% (95% CI: 53 to 98%). Conclusions: IOP on day 30 had a fair to good accuracy while bleb features failed to predict success except bleb vascularity that had a poor to fair accuracy. An IOP increase more than 3 mmHg during the first 30 days was a good predictor of failure.


Sugar-sweetened beverages and genetic risk of obesity.
Temporal increases in the consumption of sugar-sweetened beverages have paralleled the rise in obesity prevalence, but whether the intake of such beverages interacts with the genetic predisposition to adiposity is unknown. We analyzed the interaction between genetic predisposition and the intake of sugar-sweetened beverages in relation to body-mass index (BMI; the weight in kilograms divided by the square of the height in meters) and obesity risk in 6934 women from the Nurses' Health Study (NHS) and in 4423 men from the Health Professionals Follow-up Study (HPFS) and also in a replication cohort of 21,740 women from the Women's Genome Health Study (WGHS). The genetic-predisposition score was calculated on the basis of 32 BMI-associated loci. The intake of sugar-sweetened beverages was examined prospectively in relation to BMI. In the NHS and HPFS cohorts, the genetic association with BMI was stronger among participants with higher intake of sugar-sweetened beverages than among those with lower intake. In the combined cohorts, the increases in BMI per increment of 10 risk alleles were 1.00 for an intake of less than one serving per month, 1.12 for one to four servings per month, 1.38 for two to six servings per week, and 1.78 for one or more servings per day (P<0.001 for interaction). For the same categories of intake, the relative risks of incident obesity per increment of 10 risk alleles were 1.19 (95% confidence interval [CI], 0.90 to 1.59), 1.67 (95% CI, 1.28 to 2.16), 1.58 (95% CI, 1.01 to 2.47), and 5.06 (95% CI, 1.66 to 15.5) (P=0.02 for interaction). In the WGHS cohort, the increases in BMI per increment of 10 risk alleles were 1.39, 1.64, 1.90, and 2.53 across the four categories of intake (P=0.001 for interaction); the relative risks for incident obesity were 1.40 (95% CI, 1.19 to 1.64), 1.50 (95% CI, 1.16 to 1.93), 1.54 (95% CI, 1.21 to 1.94), and 3.16 (95% CI, 2.03 to 4.92), respectively (P=0.007 for interaction). The genetic association with adiposity appeared to be more pronounced with greater intake of sugar-sweetened beverages. (Funded by the National Institutes of Health and others.).


What are the specific disability and limitation types underlying responses to the BRFSS disability questions?
Researchers rely on resources such as BRFSS data to understand the health status of people with disability. However, the survey data rely on a limited definition of disability resulting in imprecise inferences about the nature of disability. Understanding how health varies among people with different types of disability is vital to tailoring interventions for improving health and eliminating disparities. The purpose of this study was to utilize state added follow-up questions in the 2011 South Carolina BRFSS to describe the specific health conditions and limitations attributed to their disability and to compare health status across different types of disability. Participants reporting a disability were asked to name health condition(s) causing disability and describe their disability-related limitations. Descriptive statistics were calculated using weighted proportions. Logistic regression was used to model the associations of specific health conditions and limitations with the outcomes of self-rated general health and mental health status, controlling for demographic factors. The 5 most commonly reported health condition categories were (weighted percentage): musculoskeletal (68.56%); pulmonary (10.41%); neurologic (8.48%); heart disease (8%) and mental health (7.31%). The 5 most commonly reported limitation categories were: mobility/balance limitations (46.29%); pain (23.22%); breathing problems (12.36%); general weakness/fatigue (9.57%) and limited lifting (8.24%). There was substantial variation in the degree of association between categories of conditions and limitations and the outcomes of self-rated physical and mental health. Researchers and practitioners should consider variability in the nature of disability when designing interventions to improve the health of people with a disability.


Recurrence and chronic pain after mesh fixation with skin staples versus sutures in Lichtenstein's inguinal hernioplasty: a retrospective cohort study.
Reduction in operating time has been reported with skin staples instead of original technique of mesh fixation with sutures in Lichtenstein's hernioplasty. Few studies have been conducted, however, with inadequate follow-up and variable results. We have undertaken this study to compare the duration of surgery and incidence of recurrence (beyond 1 year) and chronic pain after mesh fixation with staples versus sutures. In this retrospective cohort study, adult males with primary unilateral inguinal hernia who underwent open Lichtenstein's hernioplasty from January 2009 till October 2010 were included. All patients with recurrent hernia, concomitant surgery, follow-up less than 1 year and missing data were excluded. Data collection was done via questionnaire and telephonic interviews. Outcomes are recurrence, chronic pain and duration of surgery. We reviewed 70 patients in each group. At median follow-up of 21 months (range 12-34), there was 0 % recurrence in both the groups and chronic pain was 4.4 % higher in suture versus staple group (11.4 vs. 7 %, p value = 0.7). Median duration of surgery was 30 min higher in suture as compared to staple group (90 vs. 60 min, p value = 0.004). Our study confirms that staples are superior to the sutures due to shorter operative time and do not cause any additive risk of recurrence or chronic pain. Prospective trial with long-term follow-up for each patient is required to validate these findings in order to generate definite guidelines.


Clinical characteristics, treatment, and survival outcome in pediatric patients with atypical teratoid/rhabdoid tumors: a retrospective study by the Japan Children's Cancer Group.
The prognosis of atypical teratoid/rhabdoid tumors (ATRTs) has improved in recent years with the use of multimodal therapy, mainly in cases not involving metastatic disease. The authors wanted to obtain historical control data and evaluate the suitable treatments in Japanese children with ATRTs that were proven negative for INI-1 immunostaining. The authors retrospectively collected clinical information on 38 pediatric patients with ATRTs treated from 2005 to 2016 and analyzed the data for this series. The median age of the patient population was 1.3 years, and the male/female ratio was approximately 2:1. Twenty-three patients (60.5%) had metastases. The effects of treatment on prognosis were analyzed for 34 patients after exclusion of 4 patients who could not receive curative treatment. At a median follow-up of 40.9 months, the mean (± SD) progression-free survival (PFS) and overall survival (OS) were 66.6% ± 8.3% and 45.9% ± 8.7% at 2 years and 44.2% ± 9.9% and 34.2% ± 8.9% at 5 years, respectively. The metastasis stage at diagnosis (M0-1 vs M2-4) (HR 2.68, 95% CI 1.08-6.65; p = 0.0338) and gross tumor resection (yes vs no) (HR 3.49, 95% CI 1.01-12.1; p = 0.0481) were prognostic factors for PFS but not for OS. Postoperative chemotherapy was performed in all 34 cases. High-dose chemotherapy was performed in 19 (55.8%) of 34 patients and showed a positive impact on OS (HR 0.31, 95% CI 0.11-0.86; p = 0.0254); the most commonly used regimen was a double-conditioning regimen of thiotepa plus melphalan. Local radiotherapy had a positive impact on both PFS and OS; however, craniospinal irradiation (CSI) performed in 12 patients as the primary therapy was associated with a poor outcome. Disseminated recurrence within 12 months from diagnosis was the most common pattern of treatment failure regardless of CSI. There has been an improvement in outcomes for pediatric ATRT patients since the introduction of multimodal therapy in Japan, mainly in patients without metastases. Even if selection bias is taken into consideration, CSI did not contribute to an improved prognosis. Novel treatment approaches are required for pediatric ATRT patients with metastases.


Outer membrane differences between pathogenic and environmental Yersinia enterocolitica biogroups probed with hydrophobic permeants and polycationic peptides.
Sensitivities to polycationic peptides and EDTA were compared in Yersinia enterocolitica pathogenic and environmental biogroups. As shown by changes in permeability to the fluorescent hydrophobic probe N-phenylnaphthylamine (NPN), the outer membranes (OMs) of pathogenic and environmental strains grown at 26 degrees C in standard broth were more resistant to poly-L-lysine, poly-L-ornithine, melittin, cecropin P1, polymyxin B, and EDTA than Escherichia coli OMs. At 37 degrees C, OMs of pathogenic biogroups were resistant to EDTA and polycations and OMs of environmental strains were resistant to EDTA whereas E. coli OMs were sensitive to both EDTA and polycations. Similar results were found when testing deoxycholate sensitivity after polycation exposure or when isogenic pairs with or without virulence plasmid pYV were compared. With bacteria grown without Ca++ available, OM permeability to NPN was drastically increased in pathogenic but not in environmental strains or E. coli. Under these conditions, OMs of pYV+ and pYV- cells showed small differences in NPN permeability but differences in polycation sensitivity could not be detected by fluorimetry. O:1,6 (environmental type) lipopolysaccharide (LPS), but not O:3 or O:8 LPS, was markedly rough at 37 degrees C, and this could explain the differences in polycation sensitivity. LPSs from serotypes O:3 and O:8 grown at 37 degrees C were more permeable to NPN than O:1,6 LPS, and O:8 LPS was resistant to polycation-induced permeabilization. These data suggest that LPSs relate to some but not all the OM differences described. It is hypothesized that the different OM properties of environmental and pathogenic biogroups reflect the adaptation of the latter biogroups to pathogenicity.


Adaptive Hybrid Surgery: Paradigm Shift for Patient-centered Neurosurgery.
The surgical management of cerebral and skull base lesions has evolved greatly in the last few decades. Still, a complete resection of lesions abutting critical neurovascular structures carries significant morbidity. Stereotactic radiosurgery (SRS) has emerged as an increasingly accepted treatment option. Minimally invasive, SRS results in excellent tumor control and low complication rates in patients with moderate-size tumors. The management of large cerebral and skull base tumors remains a formidable challenge. In such large tumors, radical surgical extirpation offers a significantly higher risk of neurological deficit, and SRS alone cannot be used because of the elevated incidence of radiation-induced complications known to be associated with large-volume tumors. With increasing treatment volumes, SRS-associated tumor control rates decrease and complication rates increase. Planned subtotal resection (STR) with adjuvant SRS (adaptive hybrid surgery [AHS]) has gained increasing interest in recent years as a multimodal approach. In AHS, a planned STR (aimed at decreasing surgical morbidity) followed by SRS to a preplanned residual tumor aids in harnessing advantages offered by both approaches. Although intuitive and reasonable, this paradigm shift from maximal resection at all cost has not been adopted widely. Combining open microsurgery with SRS requires a good understanding of both surgical and SRS modalities and their respective safety-efficacy features. We present a review and discussion on AHS as a modern, multidisciplinary treatment approach. Available data and views are discussed for vestibular schwannoma (VS) as a sample tumor. Other indications for AHS are mentioned in brief.


Crystal structures of Toxoplasma gondii adenosine kinase reveal a novel catalytic mechanism and prodrug binding.
Adenosine kinase (AK) is a key purine metabolic enzyme from the opportunistic parasitic protozoan Toxoplasma gondii and belongs to the family of carbohydrate kinases that includes ribokinase. To understand the catalytic mechanism of AK, we determined the structures of the T. gondii apo AK, AK:adenosine complex and the AK:adenosine:AMP-PCP complex to 2.55 A, 2.50 A and 1.71 A resolution, respectively. These structures reveal a novel catalytic mechanism that involves an adenosine-induced domain rotation of 30 degrees and a newly described anion hole (DTXGAGD), requiring a helix-to-coil conformational change that is induced by ATP binding. Nucleotide binding also evokes a coil-to-helix transition that completes the formation of the ATP binding pocket. A conserved dipeptide, Gly68-Gly69, which is located at the bottom of the adenosine-binding site, functions as the switch for domain rotation. The synergistic structural changes that occur upon substrate binding sequester the adenosine and the ATP gi phosphate from solvent and optimally position the substrates for catalysis. Finally, the 1.84 A resolution structure of an AK:7-iodotubercidin:AMP-PCP complex reveals the basis for the higher affinity binding of this prodrug over adenosine and thus provides a scaffold for the design of new inhibitors and subversive substrates that target the T. gondii AK.


Topoisomerase II cleavage in chromatin.
We have examined the effect of the anti-tumor drug VM-26 on purified Drosophila topoisomerase II, and used this drug to map (putative) topoisomerase II cleavage sites in chromatin. These studies indicate that VM-26 interferes with the strand breakage-rejoining catalytic cycle. VM-26 appears to stabilize the topoisomerase-II-cleavable complex and markedly enhances the formation of double-strand breaks in naked DNA. VM-26 also stimulates the formation of double-strand breaks in isolated Drosophila nuclei. Analysis of the parameters of the VM-26-stimulated cleavage reaction in nuclei strongly suggests that the double-strand scissions are generated by endogenous topoisomerase II. Finally, we have examined the distribution of (putative) cleavage sites for endogenous topoisomerase II in the chromatin of the 87A7 heat shock locus and the histone repeat unit. We have found that there are prominent VM-26-induced cleavage products from the 5' ends of the 87A7, the two heat shock protein 70 genes, and in the intergenic spacer separating these genes. Moreover, the pattern of VM-26-induced cleavage products is altered in nuclei prepared from heat-shocked cells. In the case of the histone repeat unit, only minor VM-26-induced cleavage products are observed in nuclei (in spite of the fact that experiments on naked DNA indicate that the histone repeat contains many major cleavage sites for purified topoisomerase II). These findings suggest that the nucleoprotein organization of different DNA segments may be important in determining whether specific sites are accessible to endogenous topoisomerase II in nuclei.


Occurrence and Characterization of mcr-1-Positive Escherichia coli Isolated From Food-Producing Animals in Poland, 2011-2016.
The emergence of plasmid-mediated colistin resistance (mcr genes) threatens the effectiveness of polymyxins, which are last-resort drugs to treat infections by multidrug- and carbapenem-resistant Gram-negative bacteria. Based on the occurrence of colistin resistance the aims of the study were to determine possible resistance mechanisms and then characterize the mcr-positive Escherichia coli. The research used material from the Polish national and EU harmonized antimicrobial resistance (AMR) monitoring programs. A total of 5,878 commensal E. coli from fecal samples of turkeys, chickens, pigs, and cattle collected in 2011-2016 were screened by minimum inhibitory concentration (MIC) determination for the presence of resistance to colistin (R) defined as R > 2 mg/L. Strains with MIC = 2 mg/L isolated in 2014-2016 were also included. A total of 128 isolates were obtained, and most (66.3%) had colistin MIC of 2 mg/L. PCR revealed mcr-1 in 80 (62.5%) isolates recovered from 61 turkeys, 11 broilers, 2 laying hens, 1 pig, and 1 bovine. No other mcr-type genes (including mcr-2 to -5) were detected. Whole-genome sequencing (WGS) of the mcr-1-positive isolates showed high diversity in the multi-locus sequence types (MLST) of E. coli, plasmid replicons, and AMR and virulence genes. Generally mcr-1.1 was detected on the same contig as the IncX4 (76.3%) and IncHI2 (6.3%) replicons. One isolate harbored mcr-1.1 on the chromosome. Various extended-spectrum beta-lactamase (bla SHV-12, bla CTX-M-1, bla CTX-M-15, bla TEM-30, bla TEM-52, and bla TEM-135) and quinolone resistance genes (qnrS1, qnrB19, and chromosomal gyrA, parC, and parE mutations) were present in the mcr-1.1-positive E. coli. A total of 49 sequence types (ST) were identified, ST354, ST359, ST48, and ST617 predominating. One isolate, identified as ST189, belonged to atypical enteropathogenic E. coli. Our findings show that mcr-1.1 has spread widely among production animals in Poland, particularly in turkeys and appears to be transferable mainly by IncX4 and IncHI2 plasmids spread across diverse E. coli lineages. Interestingly, most of these mcr-1-positive E. coli would remain undetected using phenotypic methods with the current epidemiological cut-off value (ECOFF). The appearance and spread of mcr-1 among various animals, but notably in turkeys, might be considered a food chain, and public health hazard.


Cancer stem cells, pluripotency, and cellular heterogeneity: a WNTer perspective.
Cancer stem cells (CSCs) are thought to represent the "beating heart" of malignant growth as they continuously fuel tumors through their ability to self-renew and differentiate. Moreover, they are also believed to underlie malignant behavior, local invasion, and metastasis in distal organ sites upon reversible epithelial-to-mesenchymal transitions (EMTs). Nevertheless, the CSC concept has been the object of controversy, mainly due to the absence of robust operational definitions and to the lack of consistency in the use of the often incorrect nomenclature employed to refer to these cells. Notwithstanding the controversies, it is now generally accepted that primary cancers are organized in hierarchical fashion with neoplastic stem-like cells able to give rise to new CSCs and to more committed malignant cells. Notably, these hierarchical structures are not unidirectional, but are rather characterized by a more dynamic equilibrium where stem-like and more committed cancer cells transit from one meta-state to the other partly because of cues from the microenvironment (niche), but also because of intrinsic and yet incompletely understood characteristics in the activation/silencing of specific signal transduction pathways. Here, we will focus on the Wnt/β-catenin signaling pathway as one of the major regulator of stemness in homeostasis and cancer, and on germ cell tumors as the type of malignancy that most closely mimics normal embryonic development and as such serve as a unique model to study the role of stem cells in neoplasia.


Localization of platelet-derived growth factor beta receptor expression in the periepithelial stroma of human breast carcinoma.
Platelet-derived growth factor (PDGF) BB is secreted by most human breast carcinoma cells; however, only recently have PDGF beta receptors been demonstrated in malignant breast tissue. In the present study, the tissue localization of PDGF beta receptor expression was studied in human breast carcinoma and nonmalignant breast tissues stained using both immunofluorescence and immunoperoxidase techniques. We examined a total of 29 cases of breast carcinomas, which showed both in situ and invasive components. PDGF beta receptor staining was localized in the periepithelial stroma and was particularly intense in regions immediately adjacent to carcinoma in situ components in all tumors examined. A diffuse low level of PDGF beta receptor staining was seen throughout the stroma of eight nonmalignant breast tissues as well as of nonmalignant regions of tumor tissues. Image analysis was used to assess the coincidence of staining of PDGF beta receptor with epithelial or stromal cells in 13 of the 29 tumor tissues studied. Less than 5% of malignant ductal epithelium or myoepithelium showed PDGF beta receptor staining. Analysis with stromal cell type-specific markers indicated significant localization of PDGF beta receptor primarily within alpha smooth muscle actin-staining cells (32%) and vascular endothelial cells (41%) in the periepithelial stroma. PDGF beta receptor positivity was strongly associated with periepithelial stromal cells adjacent to the basement membrane surrounding regions of carcinoma in situ but was less intense in regions of invasive carcinoma where basement membrane was degraded. The absence of PDGF beta receptors on carcinoma cells and their presence in the surrounding stroma suggest a paracrine stimulation of adjacent stromal tissue by malignant epithelial cells in human breast tumors.


Genetic and Phenotypic Characteristics of Staphylococcus aureus Isolates from Cystic Fibrosis Patients in Austria.
Cystic fibrosis (CF) is the most common life-limiting inherited disease in Caucasian populations. While pathological changes can be seen in various organs, morbidity and mortality are mainly related to the respiratory tract, with patients suffering from chronic bronchopulmonary infections with characteristic pathogens including Staphylococcus aureus. To date, there is only very limited data on the genetic and phenotypic characteristics of S. aureus in CF patients. Therefore, in our study, we characterized 58 S. aureus isolates collected from CF patients in Austria by spa typing, DNA microarray profiling, as well as antimicrobial susceptibility testing in order to determine common genomic and antimicrobial resistance features. The tested strain collection exhibited high genomic diversity. The 58 isolates were assigned to 16 clonal complexes and 48 spa types and differed greatly regarding their virulence and resistance gene profiles. The predominant clonal complexes were MLST CC30 (22%), CC15 (16%), CC45 (14%), and CC5 (12%), complexes that are highly prevalent worldwide among S. aureus strains isolated from humans colonized or infected with S. aureus. DNA microarray profiles showed a wide variety of genes encoding antimicrobial resistance and virulence factors such as various leukocidins, haemolysins, enterotoxins, exfoliative toxins, toxic shock syndrome toxin, as well as genes involved in adhesion and immune evasion. While a large number of strains exhibited resistance to one or several antimicrobial agents, methicillin-resistant S. aureus was found at a low prevalence of 3% (n = 2) only. The two methicillin-resistant S. aureus isolates were assigned to CC152/t355 (SCCmecV) and CC5/t001 (SCCmecI). This is the first study to genetically characterize S. aureus isolates in CF patients in Austria.


Coffee and tea drinking in relation to the risk of differentiated thyroid carcinoma: results from the European Prospective Investigation into Cancer and Nutrition (EPIC) study.
Coffee and tea constituents have shown several anti-carcinogenic activities in cellular and animal studies, including against thyroid cancer (TC). However, epidemiological evidence is still limited and inconsistent. Therefore, we aimed to investigate this association in a large prospective study. The study was conducted in the EPIC (European Prospective Investigation into Cancer and Nutrition) cohort, which included 476,108 adult men and women. Coffee and tea intakes were assessed through validated country-specific dietary questionnaires. During a mean follow-up of 14 years, 748 first incident differentiated TC cases (including 601 papillary and 109 follicular TC) were identified. Coffee consumption (per 100 mL/day) was not associated either with total differentiated TC risk (HRcalibrated 1.00, 95% CI 0.97-1.04) or with the risk of TC subtypes. Tea consumption (per 100 mL/day) was not associated with the risk of total differentiated TC (HRcalibrated 0.98, 95% CI 0.95-1.02) and papillary tumor (HRcalibrated 0.99, 95% CI 0.95-1.03), whereas an inverse association was found with follicular tumor risk (HRcalibrated 0.90, 95% CI 0.81-0.99), but this association was based on a sub-analysis with a small number of cancer cases. In this large prospective study, coffee and tea consumptions were not associated with TC risk.


Synthesis of Fibrils in Vitro by a Solubilized Cellulose Synthase from Acetobacter xylinum.
A digitonin-solubilized cellulose synthase was prepared from Acetobacter xylinum. When this enzyme was incubated under conditions known to lead to active synthesis of 1,4-beta-D-glucan polymer (cellulose), electron microscopy revealed that clusters of fibrils were assembled within minutes. Individual fibrils are 17 +/- 2 angstroms in diameter. Evidence that the fibrils were freshly synthesized and cellulosic in nature was their incorporation of the tritium from UDP-[(3)H]glucose (UDP, uridine 5'-diphosphate), their binding of gold-labeled cellobiohydrolase, and an electron diffraction pattern with 004, 200, and 012 reflections (characteristic of cellulose synthesized in vivo) but missing 110 and 110 reflections. The small size of the fibrils is atypical of native A. xylinum cellulose microfibrils. The fibrils synthesized in vitro resemble, in morphology and size, the fibrillar cellulose produced when A. xylinum is cultured in the presence of agents that interfere with the normal process of crystallization of the microfibrils. The solubilized enzyme unit may therefore be producing a basic fibrillar structure that, in vivo, interacts laterally with other fibrils to produce native cellulose microfibrils.


Voxel-based analysis of confounding effects of age and dementia severity on cerebral metabolism in Alzheimer's disease.
Alzheimer's disease is characterized by early hippocampal lesions, but neuropathological and functional imaging studies have also demonstrated involvement of associative cortices in patients suffering from this illness. New image-processing technologies have led to demonstration of predominant posteromedial cortical metabolic impairment in the disease. Confounding effects of both age and dementia severity on brain metabolism were assessed using categorical and correlational analyses performed with Statistical Parametric Mapping. Posterior cingulate and precuneus metabolism, assessed by positron emission tomography, was significantly correlated with age in a population of 46 patients with probable Alzheimer's disease. Metabolism in posterior cingulate and precuneus was higher in elderly than in younger patients with a diagnosis of Alzheimer's disease, even when dementia severity was taken as a confounding covariate. The data suggest that the sensitivity of positron emission tomography for the diagnosis of Alzheimer's disease is reduced in elderly cases, where less severe pathology is sufficient to induce clinical symptoms of dementia. Conversely, higher posteromedial metabolic impairment in early onset cases may reflect greater density of regional cerebral lesions or major decrease of functional afferences in a richly connected multimodal associative area. Posterior cingulate metabolism was also correlated to dementia severity, even when age was taken as a confounding covariate, whereas metabolism in the hippocampal formation was not shown to correlate with global cognitive deficit. Functional correlation was maintained between posterior cingulate and middle frontal cortex in demented patients as in elderly controls. The key role of posteromedial cortex in cognitive dysfunction assessed in Alzheimer's disease is probably related to its highly integrated position within attentional, visuospatial and memory neuronal networks.


Stenting in Non-Small Cell Lung Cancer: How Does It Affect the Outcomes?
Approximately 30% of lung cancer patients develop central airway obstruction (CAO) that remarkably shortens survival. There is little data about the benefits of stenting within this heterogeneous patient group. Our objective was to review their overall survival (OS) and their risk of hospitalization versus patients who did not have lesions requiring stent placement. We retrospectively reviewed charts of 171 non-small cell lung cancer (NSCLC) patients who underwent bronchoscopy in the University of Cincinnati Cancer Center from the year 2011 to 2013. Twenty-five patients with advanced lung cancer were evaluated by interventional pulmonology service for endobronchial stent placement for CAO. Eight patients did not require placement of a stent and 17 had obstructive lesions that required stenting by interventional pulmonology. Demographical parameters such as age and gender did not have a significant impact on the risk of hospitalization or OS of both groups of patients, however, those whose lesions did not mandate stent placement had significantly lower odds of hospitalization compared to patients with CAO requiring a stent (OR: 15.913, 95% CI: 1.211-209.068, P = 0.0352). Patients with advanced NSCLC and CAO that required stent placement had an OS of 13.9 m [3.9-19.9 m] compared to an OS of 23.9 m for patients with CAO not requiring a stent. We found out that patients with less severe CAO have lower odds of hospitalization and better OS compared to patients with CAO mandating stent placement. CAO patients with interventional pulmonology (IP) evaluation and management in addition, may have improved OS suggesting that IP consultation might offer both improvement in quality of life and overall survival to patients with advanced NSCLC and CAO. <br />.


Outbreak of hepatitis C virus infection during sclerotherapy of varicose veins: long-term follow-up of 196 patients (4535 patient-years).
The aim of this study was to describe the natural history of a HCV infection outbreak in 196 patients who had sclerotherapy by a same physician and to confirm patient-to-patient transmission using phylogenetic analysis in a large series of patients. Demographic information included clinical and biological parameters. Fibrosis evaluation was performed using liver biopsy or transient elastography. Follow-up was maintained until death, or the end of the observation period. In order to determine if the virus had been transmitted between the HCV genotype 2 patients, sequence analysis was undertaken of a part of the NS5b region of the genomes in samples of patients. The mean duration of follow-up was 23.1+/-6.7 years (4535 patient-years). In patients with fibrosis evaluation, 55.7% had no or mild fibrosis and 44.3% had significant fibrosis. No patient died from HCV-related disease. Nucleotide sequence analysis of a part of the NS5b region revealed that patients were all infected with the same HCV subtype (genotype 2d). The most evident feature of the tree is the clustering of all patients involved in the outbreak without any unrelated isolates. This study emphasizes the risk for nosocomial spread of HCV during intravenous therapy.


Cavitation in Poly(4-methyl-1-pentene) during Tensile Deformation.
The poly(4-methyl-1-pentene) sample was used to investigate the cavitation-induced stress-whitening phenomenon during stretching at different temperatures via the ultrasmall-angle X-ray scattering technique. Two modes of cavitation were found that mode I cavitation activated around yield point followed by mode II cavitation generated in highly oriented state. The critical strain for initiating the mode II cavitation increases with the increase of the stretching temperature, whereas the critical stress grew steadily in the lower temperature regime (30-60 °C) and reached a plateau at 70 °C. The appearance of mode II cavitation at large strains was independent of the mode I cavitation. The mode I cavitation was attributed to the competitive process between the formation of cavities and shearing yield of lamellae, whereas the mode II cavitation was proven to be related to the failure of the whole highly oriented entangled amorphous network because of the breaking of interfibrillar load-bearing tie molecules. Size distribution of cavities has been successfully calculated using a model fitting procedure. The results showed that the quantity of cavities increased heavily while the size was kept nearly constant during the propagation of the mode II cavitation.


Spine expansion and stabilization associated with long-term potentiation.
Stable expression of long-term synaptic plasticity is critical for the developmental refinement of neural circuits and for some forms of learning and memory. Although structural remodeling of dendritic spines is associated with the stable expression of long-term potentiation (LTP), the relationship between structural and physiological plasticity remains unclear. To define whether these two processes are related or distinct, we simultaneously monitored EPSPs and dendritic spines, using combined patch-clamp recording and two-photon time-lapse imaging in the same CA1 pyramidal neurons in acute hippocampal slices. We found that theta burst stimulation paired with postsynaptic spiking, which reliably induced LTP, also induced a rapid and persistent expansion of dendritic spines. Like LTP, this expansion was NMDA receptor dependent. Spine expansion occurred even when LTP was inhibited by postsynaptic inhibition of exocytosis or PKA (protein kinase A); however, under these conditions, the spine expansion was unstable and collapsed spontaneously. Furthermore, similar changes in LTP and spine expansion were observed when hippocampal neurons were treated with protein synthesis inhibitors. Like LTP, spine expansion was reversed by low-frequency stimulation (LFS) via a phosphatase-dependent mechanism, but only if the LFS was applied in a critical time window after induction. These results indicate that the initial expression of LTP and spine expansion is dissociable, but there is a high degree of mechanistic overlap between the stabilization of structural plasticity and LTP.


Hepatocyte-based flow analytical bioreactor for online xenobiotics metabolism bioprediction.
The research for new in vitro screening tools for predictive metabolic profiling of drug candidates is of major interest in the pharmaceutical field. The main motivation is to avoid late rejection in drug development and to deliver safer drugs to the market. Thanks to the superparamagnetic properties of iron oxide nanoparticles, a flow bioreactor has been developed which is able to perform xenobiotic metabolism studies. The selected cell line (HepaRG) maintained its metabolic competencies once iron oxide nanoparticles were internalized. Based on magnetically trapped cells in a homemade immobilization chamber, through which a flow of circulating phase was injected to transport nutrients and/or the studied xenobiotic, off-line and online (when coupled to a high-performance liquid chromatography chain) metabolic assays were developed using diclofenac as a reference compound. The diclofenac demonstrated a similar metabolization profile chromatogram, both with the newly developed setup and with the control situation. Highly versatile, this pioneering and innovative instrumental design paves the way for a new approach in predictive metabolism studies.


Patient use of an online triage platform: a mixed-methods retrospective exploration in UK primary care.
Recent years have seen the introduction of online triage allowing patients to describe their problem via an online form. Subsequently, a GP telephones the patient, conducting a telephone consultation or arranging a face-to-face consultation. This study aimed to explore patterns-of-use and patients' experiences of using an online triage system. This retrospective study analysed routinely collected data (from all practices using the 'askmyGP' platform for the duration of the study period, 19 May 2017 to 31 July 2017), using both quantitative and qualitative approaches. Data originated from an online triaging platform used by patients in nine general practices across the UK. Data from 5447 patients were quantitatively analysed to describe characteristics of users, patterns-of-use, and reasons given by patients for using the platform. Free-text comments left by patients (n = 569) on their experience of use were qualitatively analysed. Highest levels of use were observed in females (65.5%, n = 3570) and those aged 25-34 years. Patterns of use were high between 0800 and 0959, and on Mondays and Tuesdays. Use outside of GP practice opening hours was low. Common reasons for using the platform were for medication-related enquiries, for administrative requests, and to report a specific symptom. Comments left by patients suggested advantages to using the platform, for example, convenience and the written format, but these did not extend to all users. Patterns-of-use and patient types were in line with typical contacts to GP practices. Though the age of users was broad, highest levels of use were from younger patients. The perceived advantages to using online triage, such as convenience and ease of use, are often context dependent.


Determination of L-ephedrine, pseudoephedrine, and caffeine in rat plasma by liquid chromatography-tandem mass spectrometry.
A rapid and simple liquid chromatography tandem mass spectrometry method was developed and validated for the simultaneous determination of L-ephedrine, pseudoephedrine, and caffeine in male Fisher-344 rat plasma at nanogram-per-milliliter concentrations for use in support of toxicology studies. Only 25 μL of plasma is required, and extraction is performed using a simple, single-step protein precipitation. The method was validated over a range of 2.09 to 5460 ng/mL for L-ephedrine, 2.09 to 5050 ng/mL for pseudoephedrine and 2.03 to 5340 ng/mL for caffeine. A binary gradient elution at 0.3 mL/min was used with a Waters XBridge Phenyl (2.1 × 150 mm, 3.5 μm) column and a Waters XBridge Phenyl 2.1- × 10-mm guard column at ambient temperature. The mobile phase consisted of 10 mM ammonium acetate in water (pH 5.0) and methanol. Caffeine trimethyl-(13)C(3) was used as the internal standard. The method was evaluated for linearity, recovery, precision, accuracy, and stability, and it was successfully applied in toxicokinetic studies of ephedrine, administered alone, in combination with caffeine, and in the herbal source Ma Huang.


Using Genetic Variants to Assess the Relationship Between Circulating Lipids and Type 2 Diabetes.
The effects of dyslipidemia on the risk of type 2 diabetes (T2D) and related traits are not clear. We used regression models and 140 lipid-associated genetic variants to estimate associations between circulating HDL cholesterol (HDL-C), LDL cholesterol (LDL-C), and triglycerides and T2D and related traits. Each genetic test was corrected for effects of variants on the other two lipid types and surrogates of adiposity. We used the largest data sets available: 34,840 T2D case and 114,981 control subjects from the DIAGRAM (DIAbetes Genetics Replication And Meta-analysis) consortium and up to 133,010 individuals without diabetes for insulin secretion and sensitivity from the MAGIC (Meta-Analyses of Glucose and Insulin-related traits Consortium) and GENESIS (GENEticS of Insulin Sensitivity) studies. Eight of 21 associations between groups of variants and diabetes traits were significant at the nominal level, including those between genetically determined lower HDL-C (β = -0.12, P = 0.03) and T2D and genetically determined lower LDL-C (β = -0.21, P = 5 × 10(-6)) and T2D. Although some of these may represent causal associations, we discuss why caution must be used when using Mendelian randomization in the context of circulating lipid levels and diabetes traits. In conclusion, we found evidence of links between genetic variants associated with lipids and T2D, but deeper knowledge of the underlying genetic mechanisms of specific lipid variants is needed before drawing definite conclusions about causality based on Mendelian randomization methodology.