Changes in in-hospital mortality in the first wave of COVID-19: a multicentre prospective observational cohort study using the WHO Clinical Characterisation Protocol UK.
Mortality rates in hospitalised patients with COVID-19 in the UK appeared to decline during the first wave of the pandemic. We aimed to quantify potential drivers of this change and identify groups of patients who remain at high risk of dying in hospital. In this multicentre prospective observational cohort study, the International Severe Acute Respiratory and Emerging Infections Consortium WHO Clinical Characterisation Protocol UK recruited a prospective cohort of patients with COVID-19 admitted to 247 acute hospitals in England, Scotland, and Wales during the first wave of the pandemic (between March 9 and Aug 2, 2020). We included all patients aged 18 years and older with clinical signs and symptoms of COVID-19 or confirmed COVID-19 (by RT-PCR test) from assumed community-acquired infection. We did a three-way decomposition mediation analysis using natural effects models to explore associations between week of admission and in-hospital mortality, adjusting for confounders (demographics, comorbidities, and severity of illness) and quantifying potential mediators (level of respiratory support and steroid treatment). The primary outcome was weekly in-hospital mortality at 28 days, defined as the proportion of patients who had died within 28 days of admission of all patients admitted in the observed week, and it was assessed in all patients with an outcome. This study is registered with the ISRCTN Registry, ISRCTN66726260. Between March 9, and Aug 2, 2020, we recruited 80 713 patients, of whom 63 972 were eligible and included in the study. Unadjusted weekly in-hospital mortality declined from 32·3% (95% CI 31·8-32·7) in March 9 to April 26, 2020, to 16·4% (15·0-17·8) in June 15 to Aug 2, 2020. Reductions in mortality were observed in all age groups, in all ethnic groups, for both sexes, and in patients with and without comorbidities. After adjustment, there was a 32% reduction in the risk of mortality per 7-week period (odds ratio [OR] 0·68 [95% CI 0·65-0·71]). The higher proportions of patients with severe disease and comorbidities earlier in the first wave (March and April) than in June and July accounted for 10·2% of this reduction. The use of respiratory support changed during the first wave, with gradually increased use of non-invasive ventilation over the first wave. Changes in respiratory support and use of steroids accounted for 22·2%, OR 0·95 (0·94-0·95) of the reduction in in-hospital mortality. The reduction in in-hospital mortality in patients with COVID-19 during the first wave in the UK was partly accounted for by changes in the case-mix and illness severity. A significant reduction in in-hospital mortality was associated with differences in respiratory support and critical care use, which could partly reflect accrual of clinical knowledge. The remaining improvement in in-hospital mortality is not explained by these factors, and could be associated with changes in community behaviour, inoculum dose, and hospital capacity strain. National Institute for Health Research and the Medical Research Council.


What level of domestic government health expenditure should we aspire to for universal health coverage?
Global discussions on universal health coverage (UHC) have focussed attention on the need for increased government funding for health care in many low- and middle-income countries. The objective of this paper is to explore potential targets for government spending on health to progress towards UHC. An explicit target for government expenditure on health care relative to gross domestic product (GDP) is a potentially powerful tool for holding governments to account in progressing to UHC, particularly in the context of UHC's inclusion in the Sustainable Development Goals. It is likely to be more influential than the Abuja target, which requires decreases in budget allocations to other sectors and is opposed by finance ministries for undermining their autonomy in making sectoral budget allocation decisions. International Monetary Fund and World Health Organisation data sets were used to analyse the relationship between government health expenditure and proxy indicators for the UHC goals of financial protection and access to quality health care, and triangulated with available country case studies estimating the resource requirements for a universal health system. Our analyses point towards a target of government spending on health of at least 5% of GDP for progressing towards UHC. This can be supplemented by a per capita target of $86 to promote universal access to primary care services in low-income countries.


Functional magnetic resonance imaging of human auditory cortex.
Magnetic resonance imaging methods recently demonstrated regional cerebral signal changes in response to limb movement and visual stimulation, attributed to blood flow enhancement. We studied 5 normal subjects scanned while listening to auditory stimuli including nonspeech noise, meaningless speech sounds, single words, and narrative text. Imaged regions included the lateral aspects of both hemispheres. Signal changes in the superior temporal gyrus and superior temporal sulcus were observed bilaterally in all subjects. Speech stimuli were associated with significantly more widespread signal changes than was the noise stimulus, while no consistent differences were observed between responses to different speech stimuli. Considerable intersubject variability in the topography of signal changes was observed. These observations confirm the utility of magnetic resonance imaging in the study of human brain structure-function relationships and emphasize the role of the superior temporal gyrus in perception of acoustic-phonetic features of speech, rather than processing of semantic features.


Combined proteomic and transcriptomic interrogation of the venom gland of Conus geographus uncovers novel components and functional compartmentalization.
Cone snails are highly successful marine predators that use complex venoms to capture prey. At any given time, hundreds of toxins (conotoxins) are synthesized in the secretory epithelial cells of the venom gland, a long and convoluted organ that can measure 4 times the length of the snail's body. In recent years a number of studies have begun to unveil the transcriptomic, proteomic and peptidomic complexity of the venom and venom glands of a number of cone snail species. By using a combination of DIGE, bottom-up proteomics and next-generation transcriptome sequencing the present study identifies proteins involved in envenomation and conotoxin maturation, significantly extending the repertoire of known (poly)peptides expressed in the venom gland of these remarkable animals. We interrogate the molecular and proteomic composition of different sections of the venom glands of 3 specimens of the fish hunter Conus geographus and demonstrate regional variations in gene expression and protein abundance. DIGE analysis identified 1204 gel spots of which 157 showed significant regional differences in abundance as determined by biological variation analysis. Proteomic interrogation identified 342 unique proteins including those that exhibited greatest fold change. The majority of these proteins also exhibited significant changes in their mRNA expression levels validating the reliability of the experimental approach. Transcriptome sequencing further revealed a yet unknown genetic diversity of several venom gland components. Interestingly, abundant proteins that potentially form part of the injected venom mixture, such as echotoxins, phospholipase A2 and con-ikots-ikots, classified into distinct expression clusters with expression peaking in different parts of the gland. Our findings significantly enhance the known repertoire of venom gland polypeptides and provide molecular and biochemical evidence for the compartmentalization of this organ into distinct functional entities.


[Clinical evaluation of postoperative urinary incontinence after laparoscopic radical prostatectomy: "cross sectional study based on the original questionnaire"].
A cross-sectional study for certain symptoms of postoperative incontinence was done for patients who underwent laparoscopic radical prostatectomy (LRP) using our original self-administered questionnaire. A total of 104 patients, who underwent LRP from April 2002 to March 2005 in our institute and related hospitals, participated in our questionnaire study. Mean age and median observation period was 64.6 years old (range 51-74) and 18 months (range 3-36). The patients were classified into "D group" (disappearance of incontinence during 3 months after LRP) and "C group" (continuous incontinence more than 3 months). Then age, body mass index, preoperative PSA level, status of voiding, defecation and potency were compared. Patients with incontinence were divided according to observation period into "short period group" (S group; 3-11 months), "intermediate period group" (IM group; 12-23 months) and "long period group" (L group; more than 23 months). Incitant factors of incontinence, status of taking pads and QOL score were compared. There were no differences between the D and C groups at any measurements. About 40% of incontinence patients were pad-free, although most of these patients did not satisfy the status of voiding. The incitant factor in 90% of the S group was "cough or sneeze", but that in the L group was "without notice" (about 40%) and "too late to toilet (voiding)" (about 25%). After all, postoperative QOL score was still lower in the patients with incontinence regardless of its volume. Further study to revolutionary improve incontinence is required.


COVID-19 vaccine acceptance and hesitancy in low- and middle-income countries.
Widespread acceptance of COVID-19 vaccines is crucial for achieving sufficient immunization coverage to end the global pandemic, yet few studies have investigated COVID-19 vaccination attitudes in lower-income countries, where large-scale vaccination is just beginning. We analyze COVID-19 vaccine acceptance across 15 survey samples covering 10 low- and middle-income countries (LMICs) in Asia, Africa and South America, Russia (an upper-middle-income country) and the United States, including a total of 44,260 individuals. We find considerably higher willingness to take a COVID-19 vaccine in our LMIC samples (mean 80.3%; median 78%; range 30.1 percentage points) compared with the United States (mean 64.6%) and Russia (mean 30.4%). Vaccine acceptance in LMICs is primarily explained by an interest in personal protection against COVID-19, while concern about side effects is the most common reason for hesitancy. Health workers are the most trusted sources of guidance about COVID-19 vaccines. Evidence from this sample of LMICs suggests that prioritizing vaccine distribution to the Global South should yield high returns in advancing global immunization coverage. Vaccination campaigns should focus on translating the high levels of stated acceptance into actual uptake. Messages highlighting vaccine efficacy and safety, delivered by healthcare workers, could be effective for addressing any remaining hesitancy in the analyzed LMICs.


Brief report: incidence of selected opportunistic infections among children with juvenile idiopathic arthritis.
To compare incidence rates of selected opportunistic infections among children with and children without juvenile idiopathic arthritis (JIA). Using U.S. national Medicaid administrative claims data from 2000 through 2005, we identified a cohort of children with JIA based on physician diagnosis codes and dispensed medications. We also identified a non-JIA comparator cohort of children diagnosed as having attention deficit hyperactivity disorder (ADHD). We defined 15 types of opportunistic infection using physician diagnosis or hospital discharge codes; criteria for 7 of these types also included evidence of treatment with specific antimicrobial agents. We calculated infection incidence rates. The rates in the ADHD comparator cohort were standardized to the age, sex, and race distribution of the JIA cohort. We calculated incidence rate ratios (IRRs) with 95% confidence intervals (95% CIs) to compare infection rates. The JIA cohort included 8,503 children with 13,990 person-years of followup. The ADHD comparator cohort included 360,362 children with 477,050 person-years of followup. When all opportunistic infections were considered together as a single outcome, there were 42 infections in the JIA cohort (incidence rate 300 per 100,000 person-years; IRR 2.4 [95% CI 1.7-3.3] versus ADHD). The most common opportunistic infections among children with JIA were 3 cases of Coccidioides (incidence rate 21 per 100,000 person-years; IRR 101 [95% CI 8.1-5,319] versus ADHD), 5 cases of Salmonella (incidence rate 35 per 100,000 person-years; IRR 3.8 [95% CI 1.2-9.5]), and 32 cases of herpes zoster (incidence rate 225 per 100,000 person-years; IRR 2.1 [95% CI 1.4-3.0]). Opportunistic infections are rare among children with JIA. Nevertheless, children with JIA had a higher rate of opportunistic infections, including an increased rate of Coccidioides, Salmonella, and herpes zoster compared to children with ADHD.


2001 SCCM/ESICM/ACCP/ATS/SIS International Sepsis Definitions Conference.
In 1991, the American College of Chest Physicians (ACCP) and the Society of Critical Care Medicine (SCCM) convened a "Consensus Conference," the goals of which were "to provide a conceptual and a practical framework to define the systemic inflammatory response to infection, which is a progressive injurious process that falls under the generalized term 'sepsis' and includes sepsis-associated organ dysfunction as well." The general definitions introduced as a result of that conference have been widely used in practice and have served as the foundation for inclusion criteria for numerous clinical trials of therapeutic interventions. Nevertheless, there has been an impetus from experts in the field to modify these definitions to reflect our current understanding of the pathophysiology of these syndromes. Several North American and European intensive care societies agreed to revisit the definitions for sepsis and related conditions. This conference was sponsored by the SCCM, The European Society of Intensive Care Medicine (ESICM), The American College of Chest Physicians (ACCP), the American Thoracic Society (ATS), and the Surgical Infection Society (SIS). The conference was attended by 29 participants from Europe and North America. In advance of the conference, five subgroups were formed to evaluate the following areas: signs and symptoms of sepsis, cell markers, cytokines, microbiologic data, and coagulation parameters. The subgroups corresponded electronically before the conference and met in person during the conference. A spokesperson for each group presented the deliberation of each group to all conference participants during a plenary session. A writing committee was formed at the conference and developed the current article based on executive summary documents generated by each group and the plenary group presentations. The present article serves as the final report of the 2001 International Sepsis Definitions Conference. This document reflects a process whereby a group of experts and opinion leaders revisited the 1992 sepsis guidelines and found that apart from expanding the list of signs and symptoms of sepsis to reflect clinical bedside experience, no evidence exists to support a change to the definitions. This lack of evidence serves to underscore the challenge still present in diagnosing sepsis in 2003 for clinicians and researchers and also provides the basis for introducing PIRO as a hypothesis-generating model for future research.


Whole-genome sequencing of bladder cancers reveals somatic CDKN1A mutations and clinicopathological associations with mutation burden.
Bladder cancers are a leading cause of death from malignancy. Molecular markers might predict disease progression and behaviour more accurately than the available prognostic factors. Here we use whole-genome sequencing to identify somatic mutations and chromosomal changes in 14 bladder cancers of different grades and stages. As well as detecting the known bladder cancer driver mutations, we report the identification of recurrent protein-inactivating mutations in CDKN1A and FAT1. The former are not mutually exclusive with TP53 mutations or MDM2 amplification, showing that CDKN1A dysfunction is not simply an alternative mechanism for p53 pathway inactivation. We find strong positive associations between higher tumour stage/grade and greater clonal diversity, the number of somatic mutations and the burden of copy number changes. In principle, the identification of sub-clones with greater diversity and/or mutation burden within early-stage or low-grade tumours could identify lesions with a high risk of invasive progression.


Characterization of mitral valve annular dynamics in the beating heart.
The objective of this study is to establish a mathematical characterization of the mitral valve annulus that allows a precise qualitative and quantitative assessment of annular dynamics in the beating heart. We define annular geometry through 16 miniature markers sewn onto the annuli of 55 sheep. Using biplane videofluoroscopy, we record marker coordinates in vivo. By approximating these 16 marker coordinates through piecewise cubic splines, we generate a smooth mathematical representation of the 55 mitral annuli. We time-align these 55 annulus representations with respect to characteristic hemodynamic time points to generate an averaged baseline annulus representation. To characterize annular physiology, we extract classical clinical metrics of annular form and function throughout the cardiac cycle. To characterize annular dynamics, we calculate displacements, strains, and curvature from the discrete mathematical representations. To illustrate potential future applications of this approach, we create rapid prototypes of the averaged mitral annulus at characteristic hemodynamic time points. In summary, this study introduces a novel mathematical model that allows us to identify temporal, regional, and inter-subject variations of clinical and mechanical metrics that characterize mitral annular form and function. Ultimately, this model can serve as a valuable tool to optimize both surgical and interventional approaches that aim at restoring mitral valve competence.


Anaortic, total-arterial, off-pump coronary artery bypass surgery: how to do it.
Several large series have demonstrated that performing off-pump coronary artery bypass surgery without manipulating the ascending aorta (anaortic) utilising all-arterial grafts provides superior protection against neurological injury. Recent series comparing percutaneous coronary intervention (PCI) with surgical revascularisation have demonstrated superior results for surgery over PCI. However, a significant criticism of coronary artery surgery remains the higher incidence of neurological injury when compared to PCI. We present a simple and reproducible technique for anaortic, total-arterial off-pump coronary artery revascularisation.


INFLUENCE OF DIETARY FACTORS AND SEX ON THE TOXICITY OF CARBON TETRACHLORIDE IN RATS.
Six groups of rats on different diets were exposed to the inhalation of carbon tetrachloride (about 300 p. p. m.) for 150 days. Food intake and changes in weight were followed throughout the experiment. Animals fed a diet low in protein showed greater susceptibility than rats on a diet high in protein. Methionine was a good substitute for protein (casein) in the diet. Increase in fat intake with correspondingly lower carbohydrate intake exerted a harmful effect, especially evident in combination with a low protein diet. In this change of the fat: carbohydrate ratio, whether the increased fat or the lowered carbohydrate is the specific factor must remain unanswered at the present time. Necrotizing nephrosis was the presenting sign of the intoxication caused by carbon tetrachloride, in addition to hepatic changes, such as hydropic degeneration, necrosis, and cirrhosis. Dietary factors (methionine and methionine-containing protein, as well as low fat intake) more consistently prevented renal injury than cirrhosis of the liver. Under identical dietary conditions, especially with higher fat intake, male rats appeared to evince greater susceptibility to carbon tetrachloride than female rats. The significance of this observation and its wider applicability has been discussed.


E-cadherin reactivity of 95 noninvasive ductal and lobular lesions of the breast. Implications for the interpretation of problematic lesions.
Studies suggest that E-cadherin is useful to classify epithelial breast lesions as ductal or lobular, but extensive experience with this antibody is lacking. We studied reactivity of lesions with classic and indeterminate morphologic features. We reviewed 95 lesions and divided them into unanimous and nonunanimous diagnosis groups; the unanimous group served as benchmark lesions to which E-cadherin reactivity could be standardized and compared. All 37 ductal lesions in the unanimous group had strong, diffuse E-cadherin reactivity. Two of 22 classic lobular carcinoma in situ (LCIS) lesions had sparse E-cadherin-reactive lobular cells within a few terminal duct lobular units. Neither displayed transition from nonreactive to reactive cells. Of 36 lesions in the nonunanimous group, 19 had insufficient morphologic features for definitive classification. Only 6 of 19 were E-cadherin reactive, including several minimally proliferative lesions. The other 17 lesions in the nonunanimous group had LCIS and ductal carcinoma in situ (DCIS) features. All had no E-cadherin, or strong membrane reactivity of constituent cells in varying proportions, without a transition between reactive and nonreactive cells. Results suggest that the majority of morphologically nondiagnostic atypical lesions are lobular, including those associated with DCIS. E-cadherin seems to be absent in most lobular lesions.


Design of short membrane selective antimicrobial peptides containing tryptophan and arginine residues for improved activity, salt-resistance, and biocompatibility.
Antimicrobial peptides (AMPs) kill microbes by non-specific membrane permeabilization, making them ideal templates for designing novel peptide-based antibiotics that can combat multi-drug resistant pathogens. For maximum efficacy in vivo and in vitro, AMPs must be biocompatible, salt-tolerant and possess broad-spectrum antimicrobial activity. These attributes can be obtained by rational design of peptides guided by good understanding of peptide structure-function. Toward this end, this study investigates the influence of charge and hydrophobicity on the activity of tryptophan and arginine rich decamer peptides engineered from a salt resistant human β-defensin-28 variant. Mechanistic investigations of the decamers with detergents mimicking the composition of bacterial and mammalian membrane, reveal a correlation between improved antibacterial activity and the increase in tryptophan and positive residue content, while keeping hemolysis low. The potent antimicrobial activity and high cell membrane selective behavior of the two most active decamers, D5 and D6, are attributed to an optimum peptide charge to hydrophobic ratio bestowed by systematic arginine and tryptophan substitution. D5 and D6 show surface localization behavior with binding constants of 1.86 × 10(8) and 2.6 × 10(8) M(-1) , respectively, as determined by isothermal calorimetry measurements. NMR derived structures of D5 and D6 in SDS detergent micelles revealed proximity of Trp and Arg residues in an extended structural scaffold. Such potential cation-π interactions may be critical in cell permeabilization of the AMPs. The fundamental characterization of the engineered decamers provided in this study improves the understanding of structure-activity relationship of short arginine tryptophan rich AMPs, which will pave the way for future de novo design of potent AMPs for therapeutic and biomedical applications.


Aberrant p53, mdm2, and proliferation differ in glioblastomas from long-term compared with typical survivors.
Glioblastoma multiforme (GBM) is a highly lethal neoplasm with a median survival of approximately 1 year. Only 2-5% of patients originally diagnosed with GBM will survive > or = 3 years. Whether tumors from these long-term survivors (LTSs) exhibit molecular genetic differences compared with typical GBM survivors is not known. Tumors from 41 patients initially diagnosed with GBM and having survival > or = 3 years (LTS) was compared with 48 GBMs from short-term survivors (STSs, survival < or = 1.5 years) for p53 aberrations (expression/mutation), epidermal growth factor receptor overexpression, mdm2 overexpression, and proliferation index. Nuclear p53 expression was significantly more frequent in the LTS group. However, no difference in the rate of p53 mutation was evident. Overexpression of epidermal growth factor receptor was slightly more frequent in the STS patients, but this is not statistically different. mdm2 overexpression was significantly more frequent in the STSs, and this group had a significantly higher median proliferation index. Long-term GBM survivors were more likely to have p53-overexpressing tumors, although a difference in p53 mutation rate could not be detected. They were less likely to exhibit mdm2 overexpression and had a lower proliferation rate compared with typical GBM survivors.


Deep sequencing of small RNAs from human skin reveals major alterations in the psoriasis miRNAome.
Psoriasis is a chronic and complex inflammatory skin disease with lesions displaying dramatically altered mRNA expression profiles. However, much less is known about the expression of small RNAs. Here, we describe a comprehensive analysis of the normal and psoriatic skin miRNAome with next-generation sequencing in a large patient cohort. We generated 6.7 × 10(8) small RNA reads representing 717 known and 284 putative novel microRNAs (miRNAs). We also observed widespread expression of isomiRs and miRNA*s derived from known and novel miRNA loci, and a low frequency of miRNA editing in normal and psoriatic skin. The expression and processing of selected novel miRNAs were confirmed with qRT-PCR in skin and other human tissues or cell lines. Eighty known and 18 novel miRNAs were 2-42-fold differentially expressed in psoriatic skin. Of particular significance was the 2.7-fold upregulation of a validated novel miRNA derived from the antisense strand of the miR-203 locus, which plays a role in epithelial differentiation. Other differentially expressed miRNAs included hematopoietic-specific miRNAs such as miR-142-3p and miR-223/223*, and angiogenic miRNAs such as miR-21, miR-378, miR-100 and miR-31, which was the most highly upregulated miRNA in psoriatic skin. The functions of these miRNAs are consistent with the inflammatory and hyperproliferative phenotype of psoriatic lesions. In situ hybridization of differentially expressed miRNAs revealed stratified epidermal expression of an uncharacterized keratinocyte-derived miRNA, miR-135b, as well as the epidermal infiltration of the hematopoietic-specific miRNA, miR-142-3p, in psoriatic lesions. This study lays a critical framework for functional characterization of miRNAs in healthy and diseased skin.


Neuropeptide Y deficiency attenuates responses to fasting and high-fat diet in obesity-prone mice.
Neuropeptide Y (NPY) stimulates feeding and weight gain, but deletion of the NPY gene does not affect food intake and body weight in mice bred on a mixed genetic background. We reasoned that the orexigenic action of NPY would be evident in C57Bl/6J mice susceptible to obesity. NPY deficiency has no significant effect in mice fed a normal rodent diet. However, energy expenditure is elevated during fasting, and hyperphagia and weight gain are blunted during refeeding. Expression of agouti-related peptide (AGRP) in the hypothalamus is increased in NPY knockout (NPYko) than wild-type mice, but unlike wild type there is no further increase in AGRP when NPYko mice are fasted. Moreover, NPYko mice have higher oxygen consumption and uncoupling protein-1 expression in brown adipose tissue during fasting. The failure of an increase in orexigenic peptides and higher thermogenesis may contribute to attenuation of weight gain when NPYko mice are refed. C57Bl/6J mice lacking NPY are also less susceptible to diet-induced obesity (DIO) as a result of reduced feeding and increased energy expenditure. The resistance to DIO in NPYko mice is associated with a reduction in nocturnal feeding and increased expression of anorexigenic hypothalamic peptides. Insulin, leptin, and triglyceride levels increase with adiposity in both wild-type and NPYko mice.


Calcium plus vitamin D supplementation and health outcomes five years after active intervention ended: the Women's Health Initiative.
Clinical outcomes of the Women's Health Initiative (WHI) calcium plus vitamin D supplementation trial have been reported during 7.0 years of active intervention. We now report outcomes 4.9 years after the intervention stopped and cumulative findings. Postmenopausal women (N=36,282) were randomized; postintervention follow-up continued among 29,862 (86%) of surviving participants. Primary outcomes were hip fracture and colorectal cancer. Breast cancer, all cancers, cardiovascular disease (CVD), and total mortality were predetermined major study outcomes. Hip fracture incidence was comparable in the supplement and the placebo groups, postintervention hazard ratio (HR)=0.95, 95% confidence interval (95% CI: 0.78, 1.15) and overall HR=0.91 (95% CI: 0.79, 1.05). Overall, colorectal cancer incidence did not differ between randomization groups, HR=0.95 (95% CI: 0.80, 1.13). Throughout, there also was no difference in invasive breast cancer, CVD, and all-cause mortality between groups. In subgroup analyses, the invasive breast cancer effect varied by baseline vitamin D intake (p=0.03 for interaction). Women with vitamin D intakes >600 IU/d, had an increased risk of invasive breast cancer, HR=1.28 (95% CI; 1.03, 1.60). Over the entire study period, in post hoc analyses, the incidence of vertebral fractures, HR=0.87 (95% CI: 0.76, 0.98) and in situ breast cancers, HR=0.82 (95% CI: 0.68, 0.99) were lower among women randomized to supplementation. After an average of 11 years, calcium and vitamin D supplementation did not decrease hip fracture or colorectal cancer incidence. Exploratory analyses found lower vertebral fracture and in situ breast cancer incidence in the supplement users. There was no effect on CVD or all-cause mortality.


Effects of climate change and variability on population dynamics in a long-lived shorebird.
Climate change affects both the mean and variability of climatic variables, but their relative impact on the dynamics of populations is still largely unexplored. Based on a long-term study of the demography of a declining Eurasian Oystercatcher (Haematopus ostralegus) population, we quantify the effect of changes in mean and variance of winter temperature on different vital rates across the life cycle. Subsequently, we quantify, using stochastic stage-structured models, how changes in the mean and variance of this environmental variable affect important characteristics of the future population dynamics, such as the time to extinction. Local mean winter temperature is predicted to strongly increase, and we show that this is likely to increase the population's persistence time via its positive effects on adult survival that outweigh the negative effects that higher temperatures have on fecundity. Interannual variation in winter temperature is predicted to decrease, which is also likely to increase persistence time via its positive effects on adult survival that outweigh the negative effects that lower temperature variability has on fecundity. Overall, a 0.1 degrees C change in mean temperature is predicted to alter median time to extinction by 1.5 times as many years as would a 0.1 degrees C change in the standard deviation in temperature, suggesting that the dynamics of oystercatchers are more sensitive to changes in the mean than in the interannual variability of this climatic variable. Moreover, as climate models predict larger changes in the mean than in the standard deviation of local winter temperature, the effects of future climatic variability on this population's time to extinction are expected to be overwhelmed by the effects of changes in climatic means. We discuss the mechanisms by which climatic variability can either increase or decrease population viability and how this might depend both on species' life histories and on the vital rates affected. This study illustrates that, for making reliable inferences about population consequences in species in which life history changes with age or stage, it is crucial to investigate the impact of climate change on vital rates across the entire life cycle. Disturbingly, such data are unavailable for most species of conservation concern.


Dynamic on-demand defragmentation in flexible bandwidth elastic optical networks.
While flexible bandwidth elastic optical networking is a promising direction for future networks, the spectral fragmentation problem in such a network inevitably raises the blocking probability and significantly degrades network performance. This paper addresses the spectral defragmentation problem using an auxiliary graph based approach, which transforms the problem into a matter of finding the maximum independent set (MIS) in the constructed auxiliary graph. The enabling technologies and defragmentation-capable node architectures, together with heuristic defragmentation algorithms are proposed and evaluated. Simulation results show that the proposed min-cost defragmentation algorithms can significantly reduce the blocking probability of incoming requests in a spectrally fragmented flexible bandwidth optical network, while substantially minimizing the number of disrupted connections.


Genome-wide association scan of quantitative traits for attention deficit hyperactivity disorder identifies novel associations and confirms candidate gene associations.
Attention deficit hyperactivity disorder (ADHD) is a complex condition with environmental and genetic etiologies. Up to this point, research has identified genetic associations with candidate genes from known biological pathways. In order to identify novel ADHD susceptibility genes, 600,000 SNPs were genotyped in 958 ADHD proband-parent trios. After applying data cleaning procedures we examined 429,981 autosomal SNPs in 909 family trios. We generated six quantitative phenotypes from 18 ADHD symptoms to be used in genome-wide association analyses. With the PBAT screening algorithm, we identified 2 SNPs, rs6565113 and rs552655 that met the criteria for significance within a specified phenotype. These SNPs are located in intronic regions of genes CDH13 and GFOD1, respectively. CDH13 has been implicated previously in substance use disorders. We also evaluated the association of SNPs from a list of 37 ADHD candidate genes that was specified a priori. These findings, along with association P-values with a magnitude less than 10(-5), are discussed in this manuscript. Seventeen of these candidate genes had association P-values lower then 0.01: SLC6A1, SLC9A9, HES1, ADRB2, HTR1E, DDC, ADRA1A, DBH, DRD2, BDNF, TPH2, HTR2A, SLC6A2, PER1, CHRNA4, SNAP25, and COMT. Among the candidate genes, SLC9A9 had the strongest overall associations with 58 association test P-values lower than 0.01 and multiple association P-values at a magnitude of 10(-5) in this gene. In sum, these findings identify novel genetic associations at viable ADHD candidate genes and provide confirmatory evidence for associations at previous candidate genes. Replication of these results is necessary in order to confirm the proposed genetic variants for ADHD.


Leukotriene and purinergic receptors are involved in the hyperpolarizing effect of glucagon in liver cells.
The pancreatic hormone glucagon hyperpolarizes the liver cell membrane. In the present study, we investigated the cellular signalling pathway of glucagon-induced hyperpolarization of liver cells by using the conventional microelectrode method. The membrane potential was recorded in superficial liver cells of superfused mouse liver slices. In the presence of the K+ channel blockers tetraethylammonium (TEA, 1 mmol/l) and Ba2+ (BaCl2, 5 mmol/l) and the blocker of the Na+/K+ ATPase, ouabain (1 mmol/l), no glucagon-induced hyperpolarization was observed confirming previous findings. The hyperpolarizing effect of glucagon was abolished by the leukotriene B4 receptor antagonist CP 195543 (0.1 mmol/l) and the purinergic receptor antagonist PPADS (5 micromol/l). ATPgammaS (10 micromol/l), a non-hydrolyzable ATP analogue, induced a hyperpolarization of the liver cell membrane similar to glucagon. U 73122 (1 micromol/l), a blocker of phospholipase C, prevented both the glucagon- and ATPgammaS-induced hyperpolarization. These findings suggest that glucagon affects the hepatic membrane potential partly by inducing the formation and release of leukotrienes and release of ATP acting on purinergic receptors of the liver cell membrane.


Alcohol consumption and gastric cancer risk-A pooled analysis within the StoP project consortium.
An association between heavy alcohol drinking and gastric cancer risk has been recently reported, but the issue is still open to discussion and quantification. We investigated the role of alcohol drinking on gastric cancer risk in the "Stomach cancer Pooling (StoP) Project," a consortium of epidemiological studies. A total of 9,669 cases and 25,336 controls from 20 studies from Europe, Asia and North America were included. We estimated summary odds-ratios (ORs) and the corresponding 95% confidence intervals (CIs) by pooling study-specific ORs using random-effects meta-regression models. Compared with abstainers, drinkers of up to 4 drinks/day of alcohol had no increase in gastric cancer risk, while the ORs were 1.26 (95% CI, 1.08-1.48) for heavy (>4 to 6 drinks/day) and 1.48 (95% CI 1.29-1.70) for very heavy (>6 drinks/day) drinkers. The risk for drinkers of >4 drinks/day was higher in never smokers (OR 1.87, 95% CI 1.35-2.58) as compared with current smokers (OR 1.14, 95% CI 0.93-1.40). Somewhat stronger associations emerged with heavy drinking in cardia (OR 1.61, 95% CI 1.11-2.34) than in non-cardia (OR 1.28, 95% CI 1.13-1.45) gastric cancers, and in intestinal-type (OR 1.54, 95% CI 1.20-1.97) than in diffuse-type (OR 1.29, 95% CI 1.05-1.58) cancers. The association was similar in strata of H. pylori infected (OR = 1.52, 95% CI 1.16-2.00) and noninfected subjects (OR = 1.69, 95% CI 0.95-3.01). Our collaborative pooled-analysis provides definite, more precise quantitative evidence than previously available of an association between heavy alcohol drinking and gastric cancer risk.


Real-Time Monitoring: A Key Element in Personalized Health and Precision Health.
Current management of psychiatric disorders relies heavily on retrospective, subjective reports provided by patients and their families. Consequently, psychiatric services are often provisioned inefficiently and with suboptimal outcomes. Recent advances in computing and sensor technologies have enabled the development of real-time monitoring systems for the diagnosis and management of psychiatric disorders. The state of these technologies is rapidly evolving, with passive monitoring and predictive modeling as two areas that have great potential to affect psychiatric care. Although outpatient psychiatry probably stands to benefit the most from the use of real-time monitoring technologies, there are also several ways in which inpatient psychiatry may also benefit. As the capabilities of these technologies increase and their use becomes more common, many ethical and legal issues will need to be considered. The role of governmental regulatory bodies and nongovernmental organizations in providing oversight of the implementation of these technologies is an active area of discussion.


Attitudes of osteopathic physicians toward physician-assisted suicide.
The use of physician-assisted suicide (PAS) in the care of terminally ill patients is controversial. While there are published surveys assessing the attitudes and views of physicians in certain specialties, both nationally and in targeted geographic areas, no such data are available specifically pertaining to osteopathic physicians, who deliver 9% of the primary care in the United States. This article describes a study whose purpose was to identify the views and perspectives of osteopathic physicians on PAS through a national survey. Questionnaires were forwarded to all board-certified internists who are members of the American College of Osteopathic Internists and all surgeons and a random sample of family physicians who are members of the American Osteopathic Association. Physicians were surveyed with initial and follow-up mailings. The 49-item questionnaire was a modified version of the Physician Survey, Center for Ethics in Health Care: Oregon Health Sciences University. Of the 2910 questionnaires that were mailed, 1028 were returned in a usable form, for a 35% response rate. Respondents were predominantly (81%) male and from all parts of the country. Most (58%) would not be willing to prescribe a medication for a competent, terminally ill patient to use with the primary intention of ending his or her own life, and most (55%) would oppose the legalization of PAS. Thirty-three percent of osteopathic physicians admit to receiving requests by terminally ill patients for PAS. Most (53%) osteopathic physicians indicate that the osteopathic principle of holistic care has influenced their decision-making on this issue, and a significant minority (44%) responded that the osteopathic oath influenced their view on PAS. Religion and the degree of prayer frequency were important predictors of the decisions to support or oppose PAS. When compared to other physician surveys, this survey of osteopathic physicians demonstrates some similarities in response but also significant differences. These data suggest that osteopathic education and training may result in a perspective on PAS that reflects the distinctiveness of the osteopathic profession.


Contemporary Women's Perceptions of Childbirth Education.
The purpose of this grounded theory study was to describe and understand contemporary childbearing women's perceptions of the role of childbirth education in preparing for birth. Participants were interviewed three times: prior to beginning classes, at the end of classes, and within 2 weeks after giving birth. Constant comparative analysis of the data was concurrent with data collection. The core process that emerged was "Negotiating the Journey," with supporting categories of "Exploring the Unknown," "Making It Real," and "Sensing the Readiness." The findings indicated that, for contemporary women, the value of childbirth education may not be in affecting physiological birth outcomes but rather in helping them to be ready for childbirth and, thereby, completing an important developmental milestone.


In vitro selection of a neutralization-resistant hepatitis C virus escape mutant.
Effective immunization against hepatitis C virus (HCV) infections is likely to require the induction of both robust T and B cell immunity. Although neutralizing antibodies may play an important role in control of infection, there is little understanding of the structure of the HCV envelope glycoproteins and how they interact with such antibodies. An additional challenge for vaccine design is the genetic diversity of HCV and the rapid evolution of viral quasispecies that escape antibody-mediated neutralization. We used a cell culture-infectious, chimeric HCV with the structural proteins of genotype 1a virus to identify envelope residues contributing to the epitope recognized by a broadly neutralizing, murine monoclonal antibody, AP33. By repetitive rounds of neutralization followed by amplification, we selected a population of viral escape mutants that resist stringent neutralization with AP33 and no longer bind the antibody. Two amino acid substitutions, widely separated in the linear sequence of the E2 envelope protein (N415Y and E655G), were identified by sequencing of cloned cDNA and shown by reverse genetics analysis to contribute jointly to the AP33 resistance phenotype. The N415Y mutation substantially lowered virus fitness, most likely because of a defect in viral entry, but did not reduce binding of soluble CD81 to immobilized HCV-pseudotyped retrovirus particles. The in vitro selection of an HCV escape mutant recapitulates the ongoing evolution of antigenic variants that contributes to viral persistence in humans and reveals information concerning the conformational structure of the AP33 epitope, its role in viral replication, and constraints on its molecular evolution.


Linking soil, water, and honey composition to assess the geographical origin of argentinean honey by multielemental and isotopic analyses.
The objective of this research was to investigate the development of a reliable fingerprint from elemental and isotopic signatures of Argentinean honey to assess its geographical provenance. Honey, soil, and water from three regions (Córdoba, Buenos Aires, and Entre Rı́os) were collected. The multielemental composition was determined by ICP-MS. δ(13)C was measured by isotopic ratio mass spectrometry, whereas the (87)Sr/(86)Sr ratio was determined using thermal ionization mass spectrometry. The data were analyzed by chemometrics looking for the association between the elements, stable isotopes, and honey samples from the three studied areas. Honey samples were differentiated by classification trees and discriminant analysis using a combination of eight key variables (Rb, K/Rb, B, U, (87)Sr/(86)Sr, Na, La, and Zn) presenting differences among the studied regions. The application of canonical correlation analysis and generalized procrustes analysis showed 91.5% consensus between soil, water, and honey samples, in addition to clear differences between studied areas. To the authors' knowledge, this is the first report demonstrating the correspondence between soil, water, and honey samples using different statistical methods, showing that elemental and isotopic honey compositions are related to soil and water characteristics of the site of origin.


Presaccadic perceptual facilitation effects depend on saccade execution: evidence from the stop-signal paradigm.
Prior to the onset of a saccadic eye movement, perception is facilitated at the saccade target location. This has been attributed to a shift of attention. To test whether presaccadic attention shifts are strictly dependent on saccade execution, we examined whether they are found when observers are required to cancel the eye movement. We combined a dual task with the stop-signal paradigm: Subjects made saccades as quickly as possible to a cued location while discriminating a stimulus either at the saccade target or at the opposite location. A stop signal was presented on a subset of trials, asking subjects to cancel the eye movement. The delay of the stop signal was adjusted to yield successful inhibition of the saccade in 50% of trials. Results show similar perceptual facilitation at the saccade target for saccades with or without a stop signal, suggesting that presaccadic attention shifts are obligatory for all saccades. However, there was facilitation only when saccades were actually performed, not when observers successfully inhibited them. Thus, preparing an eye movement without subsequently executing it does not result in an attention shift. The results speak to a difference between saccade preparation and saccade programming. In light of the strong dependence on saccade execution, we discuss the functional role and causes of presaccadic attention shifts.


Thirty-Year follow-up of an African American family with macular dystrophy of the retina, locus 1 (North Carolina macular dystrophy).
To describe clinical characteristics, including visual acuity (VA), genetic analysis, and management of complications, over a 30-year period in an African American family with macular dystrophy of the retina, locus 1 (MCDR1), commonly referred to as "North Carolina macular dystrophy." Observational, cohort study. Twelve family members from a 4-generation pedigree. A total of 12 African American patients in an affected family were examined. Clinical examination was documented during 2 different follow-up periods from 1979 to 1982 in 10 patients and from 2005 to 2009 in 11 patients. Genetic analysis was performed in 4 affected members during this time. Foveal microperimetry, fundus autofluorescence, and spectral domain optical coherence tomography (OCT) data were also obtained. Change in VA of 8 members followed over 3 decades and clinical data and management of complications for all patients. Nine of 11 living family members had classic findings ranging from disease grade 2 (confluent foveal drusen, 8 eyes) to grade 3 (central coloboma-like lesion, 10 eyes). Two members developed choroidal neovascularization (CNV) requiring laser ablation, and 1 member developed non-clearing vitreous hemorrhage and underwent 25-gauge pars plana vitrectomy. Another family member developed exotropia and amblyopia in 1 eye by age 7 years. Those without CNV had no significant change in VA over 30 years. Linkage studies of 4 affected family members showed the same short tandem repeats on markers spanning D6S249 and D6S283 within the MCDR1 region of chromosome 6q16. Microperimetry analysis of an affected member with grade 3 MCDR1 revealed absent function in the region of the central coloboma-like lesions, corresponding to photoreceptor absence on OCT, although there were preserved foveal function and intact photoreceptors adjacent to the lesion. This African American family shares similar clinical findings as other MCDR1 pedigrees and the same haplotype as the originally described family from North Carolina. Clinical characteristics, including retinal features and stable VA in the absence of amblyopia and CNV, are similar to those in other reports. Eccentric viewing around impaired photoreceptors may explain good VA in patients with clinically severe-appearing macular lesions. Sequencing of the MCDR1 interval may help identify a protein responsible for early macular development. Proprietary or commercial disclosure may be found after the references.


Inbred and furious: negative association between aggression and genetic diversity in highly inbred fish.
Aggressive behaviour plays an important role in securing resources, defending against predators and shaping social interactions. Although aggression can have positive effects on growth and reproductive success, it is also energetically costly and may increase injury and compromise survival. Individual genetic diversity has been positively associated with aggression, but the cause for such an association is not clear, and it might be related to the ability to recognize kin. To disentangle the relationships between genetic diversity, kinship and aggression, we quantified aggressive behaviour in a wild, self-fertilizing fish (Kryptolebias marmoratus) with naturally variable degrees of genetic diversity, relatedness and familiarity. We found that in contrast to captive fish, levels of aggression among wild K. marmoratus are positively associated with individual homozygosity, but not with relatedness or familiarity. We suggest that the higher aggression shown by homozygous fish could be related to better kin discrimination and may be facilitated by hermaphrodite competition for scarce males, given the fitness advantages provided by outcrossing in terms of parasite resistance. It seems likely that the relationship between aggression and genetic diversity is largely influenced by both the environment and population history.


Evaluation of mannitol effect in patients with acute hepatic failure and acute-on-chronic liver failure using conventional MRI, diffusion tensor imaging and in-vivo proton MR spectroscopy.
To evaluate the effect of an intravenous bolus of mannitol in altering brain metabolites, brain water content, brain parenchyma volume, cerebrospinal fluid (CSF) volume and clinical signs in controls and in patients with acute liver failure (ALF) and acute-on-chronic liver failure (ACLF), by comparing changes in conventional magnetic resonance imaging (MRI), in vivo proton magnetic resonance spectroscopy (PMRS) and diffusion tensor imaging (DTI) before and after its infusion. Five patients each with ALF and ACLF in grade 3 or 4 hepatic encephalopathy and with clinical signs of raised intracranial pressure were studied along with five healthy volunteers. After baseline MRI, an intravenous bolus of 20% mannitol solution was given over 10 min in controls as well as in patients with ALF and ACLF. Repeat MRI for the same position was acquired 30 min after completing the mannitol injection. No statistically significant difference was observed between controls and patients with ALF and ACLF in metabolite ratios, DTI metrics and brain volume or CSF volume following 45 min of mannitol infusion. There was no change in clinical status at the end of post-mannitol imaging. The osmotic effect of mannitol did not result in significant reduction of brain water content, alteration in metabolite ratios or any change in the clinical status of these patients during or within 45 min of mannitol infusion.


Goblet cell associated antigen passages are inhibited during Salmonella typhimurium infection to prevent pathogen dissemination and limit responses to dietary antigens.
Dietary antigen acquisition by lamina propria (LP) dendritic cells (DCs) is crucial to induce oral tolerance and maintain homeostasis. However, encountering innocuous antigens during infection can lead to inflammatory responses, suggesting processes may limit steady-state luminal antigen capture during infection. We observed that goblet cell (GC) associated antigen passages (GAPs), a steady-state pathway delivering luminal antigens to LP-DCs, are inhibited during Salmonella infection. GAP inhibition was mediated by IL-1β. Infection abrogated luminal antigen delivery and antigen-specific T cell proliferation in the mesenteric lymph node (MLN). Antigen-specific T cell proliferation to dietary antigen was restored by overriding GAP suppression; however, this did not restore regulatory T cell induction, but induced inflammatory T cell responses. Salmonella translocation to the MLN required GCs and correlated with GAPs. Genetic manipulations overriding GAP suppression, or antibiotics inducing colonic GAPs, but not antibiotics that do not, increased dissemination and worsened outcomes independent of luminal pathogen burden. Thus, steady-state sampling pathways are suppressed during infection to prevent responses to dietary antigens, limit pathogen entry, and lessen the disease. Moreover, antibiotics may worsen Salmonella infection by means beyond blunting gut microbiota colonization resistance, providing new insight into how precedent antibiotic use aggravates enteric infection.


Association between air pollution and coronary artery calcification within six metropolitan areas in the USA (the Multi-Ethnic Study of Atherosclerosis and Air Pollution): a longitudinal cohort study.
Long-term exposure to fine particulate matter less than 2.5 μm in diameter (PM2.5) and traffic-related air pollutant concentrations are associated with cardiovascular risk. The disease process underlying these associations remains uncertain. We aim to assess association between long-term exposure to ambient air pollution and progression of coronary artery calcium and common carotid artery intima-media thickness. In this prospective 10-year cohort study, we repeatedly measured coronary artery calcium by CT in 6795 participants aged 45-84 years enrolled in the Multi-Ethnic Study of Atherosclerosis and Air Pollution (MESA Air) in six metropolitan areas in the USA. Repeated scans were done for nearly all participants between 2002 and 2005, for a subset of participants between 2005 and 2007, and for half of all participants between 2010 and 2012. Common carotid artery intima-media thickness was measured by ultrasound in all participants at baseline and in 2010-12 for 3459 participants. Residence-specific spatio-temporal pollution concentration models, incorporating community-specific measurements, agency monitoring data, and geographical predictors, estimated concentrations of PM2.5 and nitrogen oxides (NOX) between 1999 and 2012. The primary aim was to examine the association between both progression of coronary artery calcium and mean carotid artery intima-media thickness and long-term exposure to ambient air pollutant concentrations (PM2.5, NOX, and black carbon) between examinations and within the six metropolitan areas, adjusting for baseline age, sex, ethnicity, socioeconomic characteristics, cardiovascular risk factors, site, and CT scanner technology. In this population, coronary calcium increased on average by 24 Agatston units per year (SD 58), and intima-media thickness by 12 μm per year (10), before adjusting for risk factors or air pollutant exposures. Participant-specific pollutant concentrations averaged over the years 2000-10 ranged from 9.2-22.6 μg PM2.5/m(3) and 7.2-139.2 parts per billion (ppb) NOX. For each 5 μg PM2.5/m(3) increase, coronary calcium progressed by 4.1 Agatston units per year (95% CI 1.4-6.8) and for each 40 ppb NOX coronary calcium progressed by 4.8 Agatston units per year (0.9-8.7). Pollutant exposures were not associated with intima-media thickness change. The estimate for the effect of a 5 μg/m(3) higher long-term exposure to PM2.5 in intima-media thickness was -0.9 μm per year (95% CI -3.0 to 1.3). For 40 ppb higher NOX, the estimate was 0.2 μm per year (-1.9 to 2.4). Increased concentrations of PM2.5 and traffic-related air pollution within metropolitan areas, in ranges commonly encountered worldwide, are associated with progression in coronary calcification, consistent with acceleration of atherosclerosis. This study supports the case for global efforts of pollution reduction in prevention of cardiovascular diseases. US Environmental Protection Agency and US National Institutes of Health.


Structural basis for TetM-mediated tetracycline resistance.
Ribosome protection proteins (RPPs) confer tetracycline resistance by binding to the ribosome and chasing the drug from its binding site. The current model for the mechanism of action of RPPs proposes that drug release is indirect and achieved via conformational changes within the drug-binding site induced upon binding of the RPP to the ribosome. Here we report a cryo-EM structure of the RPP TetM in complex with the 70S ribosome at 7.2-Å resolution. The structure reveals the contacts of TetM with the ribosome, including interaction between the conserved and functionally critical C-terminal extension of TetM and the decoding center of the small subunit. Moreover, we observe direct interaction between domain IV of TetM and the tetracycline binding site and identify residues critical for conferring tetracycline resistance. A model is presented whereby TetM directly dislodges tetracycline to confer resistance.


Resveratrol trimer enhances gene delivery to hematopoietic stem cells by reducing antiviral restriction at endosomes.
Therapeutic gene delivery to hematopoietic stem cells (HSCs) holds great potential as a life-saving treatment of monogenic, oncologic, and infectious diseases. However, clinical gene therapy is severely limited by intrinsic HSC resistance to modification with lentiviral vectors (LVs), thus requiring high doses or repeat LV administration to achieve therapeutic gene correction. Here we show that temporary coapplication of the cyclic resveratrol trimer caraphenol A enhances LV gene delivery efficiency to human and nonhuman primate hematopoietic stem and progenitor cells with integrating and nonintegrating LVs. Although significant ex vivo, this effect was most dramatically observed in human lineages derived from HSCs transplanted into immunodeficient mice. We further show that caraphenol A relieves restriction of LV transduction by altering the levels of interferon-induced transmembrane (IFITM) proteins IFITM2 and IFITM3 and their association with late endosomes, thus augmenting LV core endosomal escape. Caraphenol A-mediated IFITM downregulation did not alter the LV integration pattern or bias lineage differentiation. Taken together, these findings compellingly demonstrate that the pharmacologic modification of intrinsic immune restriction factors is a promising and nontoxic approach for improving LV-mediated gene therapy.


US Intergroup Trial of Response-Adapted Therapy for Stage III to IV Hodgkin Lymphoma Using Early Interim Fluorodeoxyglucose-Positron Emission Tomography Imaging: Southwest Oncology Group S0816.
Four US National Clinical Trials Network components (Southwest Oncology Group, Cancer and Leukemia Group B/Alliance, Eastern Cooperative Oncology Group, and the AIDS Malignancy Consortium) conducted a phase II Intergroup clinical trial that used early interim fluorodeoxyglucose positron emission tomography (FDG-PET) imaging to determine the utility of response-adapted therapy for stage III to IV classic Hodgkin lymphoma. The Southwest Oncology Group S0816 (Fludeoxyglucose F 18-PET/CT Imaging and Combination Chemotherapy With or Without Additional Chemotherapy and G-CSF in Treating Patients With Stage III or Stage IV Hodgkin Lymphoma) trial enrolled 358 HIV-negative patients between July 1, 2009, and December 2, 2012. A PET scan was performed after two initial cycles of doxorubicin, bleomycin, vinblastine, and dacarbazine (ABVD) and was labeled PET2. PET2-negative patients (Deauville score 1 to 3) received an additional four cycles of ABVD, whereas PET2-positive patients (Deauville score 4 to 5) were switched to escalated bleomycin, etoposide, doxorubicin, cyclophosphamide, vincristine, procarbazine, and prednisone (eBEACOPP) for six cycles. Among 336 eligible and evaluable patients, the median age was 32 years (range, 18 to 60 years), with 52% stage III, 48% stage IV, 49% International Prognostic Score 0 to 2, and 51% score 3 to 7. Three hundred thirty-six of the enrolled patients were evaluable. Central review of the interim PET2 scan was performed in 331 evaluable patients, with 271 (82%) PET2-negative and 60 (18%) PET2-positive. Of 60 eligible PET2-positive patients, 49 switched to eBEACOPP as planned and 11 declined. With a median follow-up of 39.7 months, the Kaplan-Meier estimate for 2-year overall survival was 98% (95% CI, 95% to 99%), and the 2-year estimate for progression-free survival (PFS) was 79% (95% CI, 74% to 83%). The 2-year estimate for PFS in the subset of patients who were PET2-positive after two cycles of ABVD was 64% (95% CI, 50% to 75%). Both nonhematologic and hematologic toxicities were greater in the eBEACOPP arm than in the continued ABVD arm. Response-adapted therapy based on interim PET imaging after two cycles of ABVD seems promising with a 2-year PFS of 64% for PET2-positive patients, which is much higher than the expected 2-year PFS of 15% to 30%.


Development of several organs that require inductive epithelial-mesenchymal interactions is impaired in LEF-1-deficient mice.
Lymphoid enhancer factor 1 (LEF-1) is a sequence-specific DNA-binding protein that is expressed in pre-B and T lymphocytes of adult mice, and in the neural crest, mesencephalon, tooth germs, whisker follicles, and other sites during embryogenesis. We have generated mice carrying a homozygous germ-line mutation in the LEF-1 gene that eliminates its protein expression and causes postnatal lethality. The mutant mice lack teeth, mammary glands, whiskers, and hair but show no obvious defects in lymphoid cell populations at birth. The LEF-1-deficient mice also lack the mesencephalic nucleus of the trigeminal nerve, the only neural crest-derived neuronal populations. Together, the pattern of these defects suggest an essential role for LEF-1 in the formation of several organs and structures that require inductive tissue interactions.


Evaluation of colonic myoelectrical activity in health and functional disorders.
Electrical activity of the colon was investigated in six healthy volunteers and 22 patients presenting functional colonic disorders associated with pain, constipation, or diarrhoea. Myoelectrical activity during 10 hours, including two daily meals, was recorded from eight groups of nickel-chrome electrodes using a 1.5 m length probe introduced by the rectal route. Electromyograms of volunteers consisted of slow waves at two distinct rhythms, approximately 3 and 10 cycles per minute, during 16 to 28% of the recording time respectively for each site. Action potentials were also recorded as 11 to 80 short spike bursts (SSB) per hour lasting 1.5 to 3.5 s at any site and 20-26 long spike bursts (LSB) per hour, lasting 17 to 21 s, occurring in series of four to eight propagated bursts. Additional high-velocity propagated bursts were recorded during the three hour postprandial period. Three typical changes in spike activity patterns were detected: an increase by 170-420% of the number of SSB associated to a high spiking activity was recorded in 13 patients (group I), the absence of SSB, a low spiking activity level and only 3 to 8 LSB per hour (group II), in six patients; and the postprandial response was absent (group III) in three other investigated patients. Evidence for three groups of motor disturbances and their possible relation to clinical manifestations of functional disorders is presented.


Application of Population Sequencing (POPSEQ) for Ordering and Imputing Genotyping-by-Sequencing Markers in Hexaploid Wheat.
The advancement of next-generation sequencing technologies in conjunction with new bioinformatics tools enabled fine-tuning of sequence-based, high-resolution mapping strategies for complex genomes. Although genotyping-by-sequencing (GBS) provides a large number of markers, its application for association mapping and genomics-assisted breeding is limited by a large proportion of missing data per marker. For species with a reference genomic sequence, markers can be ordered on the physical map. However, in the absence of reference marker order, the use and imputation of GBS markers is challenging. Here, we demonstrate how the population sequencing (POPSEQ) approach can be used to provide marker context for GBS in wheat. The utility of a POPSEQ-based genetic map as a reference map to create genetically ordered markers on a chromosome for hexaploid wheat was validated by constructing an independent de novo linkage map of GBS markers from a Synthetic W7984 × Opata M85 recombinant inbred line (SynOpRIL) population. The results indicated that there is strong agreement between the independent de novo linkage map and the POPSEQ mapping approach in mapping and ordering GBS markers for hexaploid wheat. After ordering, a large number of GBS markers were imputed, thus providing a high-quality reference map that can be used for QTL mapping for different traits. The POPSEQ-based reference map and whole-genome sequence assemblies are valuable resources that can be used to order GBS markers and enable the application of highly accurate imputation methods to leverage the application GBS markers in wheat.


Evidence of multiple pyrethroid resistance mechanisms in the malaria vector Anopheles gambiae sensu stricto from Nigeria.
Pyrethroid insecticide resistance in Anopheles gambiae sensu stricto is a major concern to malaria vector control programmes. Resistance is mainly due to target-site insensitivity arising from a single point mutation, often referred to as knockdown resistance (kdr). Metabolic-based resistance mechanisms have also been implicated in pyrethroid resistance in East Africa and are currently being investigated in West Africa. Here we report the co-occurrence of both resistance mechanisms in a population of An. gambiae s.s. from Nigeria. Bioassay, synergist and biochemical analysis carried out on resistant and susceptible strains of An. gambiae s.s. from the same geographical area revealed >50% of the West African kdr mutation in the resistant mosquitoes but <3% in the susceptible mosquitoes. Resistant mosquitoes synergized using pyperonyl butoxide before permethrin exposure showed a significant increase in mortality compared with the non-synergized. Biochemical assays showed an increased level of monooxygenase but not glutathione-S-transferase or esterase activities in the resistant mosquitoes. Microarray analysis using the An. gambiae detox-chip for expression of detoxifying genes showed five over-expressed genes in the resistant strain when compared with the susceptible one. Two of these, CPLC8 and CPLC#, are cuticular genes not implicated in pyrethroid metabolism in An. gambiae s.s, and could constitute a novel set of candidate genes that warrant further investigation.


Abnormal calcium handling properties underlie familial hypertrophic cardiomyopathy pathology in patient-specific induced pluripotent stem cells.
Familial hypertrophic cardiomyopathy (HCM) is a prevalent hereditary cardiac disorder linked to arrhythmia and sudden cardiac death. While the causes of HCM have been identified as genetic mutations in the cardiac sarcomere, the pathways by which sarcomeric mutations engender myocyte hypertrophy and electrophysiological abnormalities are not understood. To elucidate the mechanisms underlying HCM development, we generated patient-specific induced pluripotent stem cell cardiomyocytes (iPSC-CMs) from a ten-member family cohort carrying a hereditary HCM missense mutation (Arg663His) in the MYH7 gene. Diseased iPSC-CMs recapitulated numerous aspects of the HCM phenotype including cellular enlargement and contractile arrhythmia at the single-cell level. Calcium (Ca(2+)) imaging indicated dysregulation of Ca(2+) cycling and elevation in intracellular Ca(2+) ([Ca(2+)](i)) are central mechanisms for disease pathogenesis. Pharmacological restoration of Ca(2+) homeostasis prevented development of hypertrophy and electrophysiological irregularities. We anticipate that these findings will help elucidate the mechanisms underlying HCM development and identify novel therapies for the disease.


The Dictyostelium mitogen-activated protein kinase ERK2 is regulated by Ras and cAMP-dependent protein kinase (PKA) and mediates PKA function.
The chemoattractant cAMP, acting through serpentine cAMP receptors, results in a rapid and transient stimulation of the Dictyostelium mitogen-activated protein kinase ERK2 activity (). In this study we show that other pathways required for aggregation, including Ras and cAMP-dependent protein kinase (PKA), are important regulators of ERK2 activation and adaptation. By examining both the level and kinetics of activation and adaptation of ERK2, we show that Ras is a negative regulator of ERK2. Activated Ras or disruption of a Ras GAP gene results in reduced ERK2 activation whereas disruption of putative Ras GEF or expression of dominant negative Ras proteins have a more rapid, higher, and extended activation. CRAC, a PH domain-containing protein required for adenylyl cyclase activation, is also required for proper ERK2 adaptation. PKA overexpression results in a more rapid, higher level of activation, whereas pka null cells show a lower level but more extended ERK2 activation. Furthermore, we show that constitutive expression of PKA catalytic subunit bypasses the requirement of ERK2 for aggregation and later development, indicating that PKA lies downstream from ERK2 and that ERK2 may regulate one or more components of the signaling pathway required for mediating PKA function, possibly by directly regulating PKA R or a protein controlling the intracellular level of cAMP.


Reliability and Validity Assessment of the Observation of Human-Animal Interaction for Research (OHAIRE) Behavior Coding Tool.
The Observation of Human-Animal Interaction for Research (OHAIRE) is a coding tool developed to capture the behavior of children when interacting with social partners and animals in naturalistic settings. The OHAIRE behavioral categories of focus are emotional displays, social communication behaviors toward adults and peers, behaviors directed toward animals or experimental control objects, and interfering behaviors. To date, the OHAIRE has been used by 14 coders to code 2,732 min of video across four studies with a total of 201 participants ages 5 to 18 years (M = 10.1, SD = 2.5). Studies involved animal-assisted intervention with three species (i.e., dogs, horses, and guinea pigs) and three populations (i.e., autism spectrum disorder, attention-deficit hyperactivity disorder, and typically developing children) in a school, a therapeutic horseback riding program, a group therapy program, and the hospital setting. We explored the psychometric properties of the OHAIRE through analyses of its inter-rater reliability, intra-rater reliability, convergent and divergent validity, and internal structure, using data from these four human-animal interaction studies. The average inter-rater reliability was excellent (kappa = 0.81), with good reliability in most of the behavioral categories coded. Intra-rater reliability was consistently excellent (0.87 ≤ kappa ≤0.96). Internal structure analyses with Cronbach's alpha supported the exploratory use of subscales to measure social communication behaviors toward peers (α = 0.638) and adults (α = 0.605), and interactions experimental control objects (α = 0.589), and the use of a subscale to measure interactions with animals (α = 0.773). Correlation analyses with multiple questionnaires showed a convergence between positive emotional display and social behaviors as assessed by the OHAIRE and social skills as assessed by the Social Skills Rating System (SSRS) and the Social Communication Questionnaires (SCQ). Little concordance was found between the OHAIRE and the Social Responsiveness Scale (SRS) or the Aberrant Behavior Checklist-Community (ABC). The OHAIRE shows promise for wider use in the field of Human-Animal Interaction, with a need for generalization across more settings and ages.


Impaired skeletal muscle microcirculation in systemic sclerosis.
Muscle symptoms in systemic sclerosis (SSc) may originate from altered skeletal muscle microcirculation, which can be investigated by means of blood oxygenation level dependent (BOLD) magnetic resonance imaging (MRI). After ethics committee approval and written consent, 11 consecutive SSc patients (5 men, mean age 52.6 years, mean SSc disease duration 5.4 years) and 12 healthy volunteers (4 men, mean age 45.1 years) were included. Subjects with peripheral arterial occlusive disease were excluded. BOLD MRI was performed on calf muscles during cuff-induced ischemia and reactive hyperemia, using a 3-T whole-body scanner (Verio, Siemens, Erlangen, Germany) and fat-suppressed single-short multi-echo echo planar imaging (EPI) with four different effective echo times. Muscle BOLD signal time courses were obtained for gastrocnemius and soleus muscles: minimal hemoglobin oxygen saturation (T2*min) and maximal T2* values (T2*max), time to T2* peak (TTP), and slopes of oxygen normalization after T2* peaking. The vast majority of SSc patients lacked skeletal muscle atrophy, weakness or serum creatine kinase elevation. Nevertheless, more intense oxygen desaturation during ischemia was observed in calf muscles of SSc patients (mean T2*min -15.0%), compared with controls (-9.1%, P = 0.02). SSc patients also had impaired oxygenation during hyperemia (median T2*max 9.2% vs. 20.1%, respectively, P = 0.007). The slope of muscle oxygen normalization was significantly less steep and prolonged (TTP) in SSc patients (P<0.001 for both). Similar differences were found at a separate analysis of gastrocnemius and soleus muscles, with most pronounced impairment in the gastrocnemius. BOLD MRI demonstrates a significant impairment of skeletal muscle microcirculation in SSc.


Genome-wide studies to identify risk factors for kidney disease with a focus on patients with diabetes.
Chronic kidney disease (CKD) affects 10-13% of the general population and diabetic nephropathy (DN) is the leading cause of end-stage renal disease (ESRD). In addition to known demographic, biochemical and lifestyle risk factors, genetics is also contributing to CKD risk. In recent years, genome-wide association studies (GWAS) have provided a hypothesis-free approach to identify common genetic variants that could account for the genetic risk component of common diseases such as CKD. The identification of these variants might reveal the biological processes underlying renal impairment and could aid in improving risk estimates for CKD. This review aims to describe the methods as well as strengths and limitations of GWAS in CKD and to summarize the findings of recent GWAS in DN. Several loci and SNPs have been found to be associated with distinct CKD traits such as eGFR and albuminuria. For diabetic kidney disease, several loci were identified in different populations. Subsequent functional studies provided insights into the mechanism of action of some of these variants, such as UMOD or CERS2. However, overall, the results were ambiguous, and a few of the variants were not consistently replicated. In addition, the slow progression from albuminuria to ESRD could limit the power of longitudinal studies. The typically small effect size associated with genetic variants as well as the small portion of the variability of the phenotype explained by these variants limits the utility of genetic variants in improving risk prediction. Nevertheless, identifying these variants could give a deeper understanding of the molecular pathways underlying CKD, which in turn, could potentially lead to the development of new diagnostic and therapeutic tools.


Adopting best practices: lessons learned in the Collaborative Initiative to Help End Chronic Homelessness (CICH).
The Collaborative Initiative to Help End Chronic Homelessness (CICH) was established to provide housing and supportive services for individuals experiencing chronic homelessness. As part of this initiative, 11 projects across the country received funding to apply models of best practices to support their clients in housing. This paper reports on the experiences of the CICH projects in their use of Assertive Community Treatment (ACT) and Motivational Interviewing (MI), clinical practice models commonly used by CICH projects. A qualitative analysis identified five areas of challenge for the projects: (1) Incomplete and underdeveloped staff teams; (2) Incomplete understanding of the practice models; (3) Using the elements of the practice models; (4) Interagency teaming; and (5) Competing expectations of multiple federal agencies. The paper describes the specific challenges in each of the five areas as well as training approaches and gaps in training and the perceived benefits of the practice models as reported by project staff.


Mutation rate and dominance of genes affecting viability in Drosophila melanogaster.
Spontaneous mutations were allowed to accumulate in a second chromosome that was transmitted only through heterozygous males for 40 generations. At 10-generation intervals the chromosomes were assayed for homozygous effects of the accumulated mutants. From the regression of homozygous viability on the number of generations of mutant accumulation and from the increase in genetic variance between replicate chromosomes it is possible to estimate the mutation rate and average effect of the individual mutants. Lethal mutations arose at a rate of 0.0060 per chromosome per generation. The mutants having small effects on viability are estimated to arise with a frequency at least 10 times as high as lethals, more likely 20 times as high, and possibly many more times as high if there is a large class of very nearly neutral mutations.-The dominance of such mutants was measured for chromosomes extracted from a natural population. This was determined from the regression of heterozygous viability on that of the sum of the two constituent homozygotes. The average dominance for minor viability genes in an equilibrium population was estimated to be 0.21. This is lower than the value for new mutants, as expected since those with the greatest heterozygous effect are most quickly eliminated from the population. That these mutants have a disproportionately large heterozygous effect on total fitness (as well as on the viability component thereof) is shown by the low ratio of the genetic load in equilibrium homozygotes to that of new mutant homozygotes.


All-electron real-time and imaginary-time time-dependent density functional theory within a numeric atom-centered basis function framework.
Real-time time-dependent density functional theory (RT-TDDFT) is an attractive tool to model quantum dynamics by real-time propagation without the linear response approximation. Sharing the same technical framework of RT-TDDFT, imaginary-time time-dependent density functional theory (it-TDDFT) is a recently developed robust-convergence ground state method. Presented here are high-precision all-electron RT-TDDFT and it-TDDFT implementations within a numerical atom-centered orbital (NAO) basis function framework in the FHI-aims code. We discuss the theoretical background and technical choices in our implementation. First, RT-TDDFT results are validated against linear-response TDDFT results. Specifically, we analyze the NAO basis sets' convergence for Thiel's test set of small molecules and confirm the importance of the augmentation basis functions for adequate convergence. Adopting a velocity-gauge formalism, we next demonstrate applications for systems with periodic boundary conditions. Taking advantage of the all-electron full-potential implementation, we present applications for core level spectra. For it-TDDFT, we confirm that within the all-electron NAO formalism, it-TDDFT can successfully converge systems that are difficult to converge in the standard self-consistent field method. We finally benchmark our implementation for systems up to ∼500 atoms. The implementation exhibits almost linear weak and strong scaling behavior.


Clearance of adenovirus-infected hepatocytes by MHC class I-restricted CD4+ CTLs in vivo.
E1-deleted recombinant adenoviruses have been developed for liver-directed gene therapy because efficient gene transfer to hepatocytes can be achieved in vivo. However, these viruses also express viral proteins in hepatocytes, leading to the development of destructive immune responses. Our previous studies indicated that MHC class I-restricted C D8+CTLs are major effectors in eliminating virus-infected cells, and CD4+ cells are also necessary in developing a fully competent CTL response by secretion of IFN-gamma, which sensitizes the virus-infected hepatocytes to CTLs through up-regulation of MHC class I expression. In this study, we have used adoptive transfer techniques in combination with mice deficient in immune functions to further define the role of CD4+ cells in the primary response to adenovirus-mediated gene transfer to the liver. Our studies indicate that CD4+ cells alone are capable of destroying virus-infected hepatocytes. Adoptive transfer experiments with beta 2m- mice along with in vitro CTL assays suggest that these CD4+ can act as CTL effectors, which are MHC class I-restricted. Depletion of these CD4+ effectors in vivo leads to prolongation of adenovirus-mediated transgene expression in hepatocytes. These results suggest that class I-restricted CD4+ CTLs contribute to elimination of adenovirus-transduced hepatocytes and extend our understanding of functional importance of CD4+ cells in viral pathogenesis.


Results of the Meso-ORIGINS feasibility study regarding collection of matched benign-mesothelioma tissue pairs by longitudinal surveillance.
To assess key elements of the design for Meso-ORIGINS (Mesothelioma Observational study of RIsk prediction and Generation of paired benign-meso tissue samples, Including a Nested MRI Substudy), an ambitious, UK-wide, prospective study that will collect ≥63 matched benign-mesothelioma tissue pairs through longitudinal surveillance and repeat biopsy of patients with asbestos-associated pleural inflammation (AAPI). A multicentre, mixed-methods feasibility study, comprising a prospective observational element, evaluating recruitment feasibility, technical feasibility of repeat local anaesthetic thoracoscopy (LAT) and patient acceptability, and a retrospective cohort study focused on AAPI-mesothelioma evolution rate, informing sample size. 4 UK pleural disease centres (February 2019-January 2020). Patients with AAPI (history or typical imaging plus appropriate pleural histology) were eligible for both elements. In August 2019, eligibility for the prospective element was broadened, including addition of radiological AAPI for technical feasibility and patient acceptability endpoints only. Retrospective cases required ≥2 years follow-up. A prospective recruitment target was set a priori at 27 histological AAPI cases (or 14 in any 6 months). Technical feasibility and patient acceptability were determined at 6-month follow-up by thoracic ultrasound surrogates and questionnaires, respectively. Retrospective malignant pleural mesothelioma evolution rate was defined by proportion (95% CI). Baseline predictors of evolution were identified using logistic regression. 296 patients with AAPI (39 prospective, 257 retrospective) were recruited/selected. 21/39 prospective recruits were histologically diagnosed (target n=27). Repeat LAT was technically feasible and acceptable in 13/28 (46%) and 24/36 (67%) cases with complete follow-up data. Mesothelioma evolution was confirmed histologically in 36/257 retrospective cases (14% (95% CI 10.3% to 18.8%)) and associated with malignant CT features (OR 4.78 (95% CI 2.36 to 9.86)) and age (OR 1.06 (95% CI 1.02 to 1.12)). Our initial eligibility criteria were too narrow. Meso-ORIGINS will recruit a broader cohort, including prevalent cases, any biopsy type and patients with malignant CT features. A range of rebiopsy techniques will be allowed, accounting for technical and patient factors. The sample size has been reduced to 500. ISRCTN12840870.


Reptile Toll-like receptor 5 unveils adaptive evolution of bacterial flagellin recognition.
Toll-like receptors (TLR) are ancient innate immune receptors crucial for immune homeostasis and protection against infection. TLRs are present in mammals, birds, amphibians and fish but have not been functionally characterized in reptiles despite the central position of this animal class in vertebrate evolution. Here we report the cloning, characterization, and function of TLR5 of the reptile Anolis carolinensis (Green Anole lizard). The receptor (acTLR5) displays the typical TLR protein architecture with 22 extracellular leucine rich repeats flanked by a N- and C-terminal leucine rich repeat domain, a membrane-spanning region, and an intracellular TIR domain. The receptor is phylogenetically most similar to TLR5 of birds and most distant to fish TLR5. Transcript analysis revealed acTLR5 expression in multiple lizard tissues. Stimulation of acTLR5 with TLR ligands demonstrated unique responsiveness towards bacterial flagellin in both reptile and human cells. Comparison of acTLR5 and human TLR5 using purified flagellins revealed differential sensitivity to Pseudomonas but not Salmonella flagellin, indicating development of species-specific flagellin recognition during the divergent evolution of mammals and reptiles. Our discovery of reptile TLR5 fills the evolutionary gap regarding TLR conservation across vertebrates and provides novel insights in functional evolution of host-microbe interactions.


COVID-19 and the gender health paradox.
This article examines gender-based health inequalities arising from the COVID-19 pandemic by drawing on insights from research into the 'gender health paradox'. Decades of international research shows that, across Europe, men have shorter life expectancies and higher mortality rates than women, and yet, women report higher morbidity. These gender-based health inequalities also appear to be evident within the pandemic and its aftermath. The article starts by providing an overview of the 'gender health paradox' and the biological, social, economic and political explanations for it. It then outlines the international estimates of gender-based inequalities in COVID-19 morbidity and mortality rates - where emerging data suggests that women are more likely to be diagnosed with COVID-19 but that men have a higher mortality rate. It then explores the longer term consequences for gender-based health inequalities of the aftermath of the COVID-19 pandemic, focusing on the impacts of government policy responses and the emerging economic crisis, suggesting that this might lead to increased mortality amongst men and increased morbidity amongst women. The essay concludes by reflecting on the pathways shaping gender-based health inequalities in the COVID-19 pandemic and the responses needed to ensure that it does not exacerbate gender-based health inequalities into the future.


Characterization of the bacterial magnetosome membrane.
Intact magnetosomes of Aquaspirillum magnetotacticum were purified from broken cells by a magnetic separation technique. Electron microscopic and chemical analyses revealed the magnetite to be enclosed by a lipid bilayer admixed with proteins. Lipids were recovered in fractions expected to contain (i) neutral lipids and free fatty acids, (ii) glycolipids and sulfolipids, and (iii) phospholipids (in a weight ratio of 1:4:6). Phospholipids included phosphatidylserine and phosphatidylethanolamine. Two of the numerous proteins detected in the magnetosome membrane were not found in other cell membranes or soluble fractions.


Duty-hour restrictions and the work of surgical faculty: results of a multi-institutional study.
To examine whether duty-hour restrictions have been consequential for various aspects of the work of surgical faculty and if those consequences differ for faculty in academic and nonacademic general surgery residency programs. Questionnaires were distributed in 2004 to 233 faculty members in five academic and four nonacademic U.S. residency programs in general surgery. Participation was restricted to those who had been faculty for at least one year. Ten items on the questionnaire probed faculty work experiences. Results include means, percentages, and t-tests on mean differences. Of the 146 faculty members (63%) who completed the questionnaire, 101 volunteered to be interviewed. Of these, 28 were randomly chosen for follow-up interviews that probed experiences and rationales underlying items on the questionnaire. Interview transcripts (187 single-spaced pages) were analyzed for main themes. Questionnaire respondents and interviewees associated duty-hour restrictions with lowered faculty expectations and standards for residents, little change in the supervision of residents, a loss of time for teaching, increased work and stress, and less satisfaction. No significant differences in these perceptions (p < or = .05) were found for faculty in academic and nonacademic programs. Main themes from the interviews included a shift of routine work from residents to faculty, a transfer of responsibility to faculty, more frequent skill gaps at night, a loss of time for research, and the challenges of controlling residents' hours. Duty-hour restrictions have been consequential for the work of surgical faculty. Faculty should not be overlooked in future studies of duty-hour restrictions.


Motivation, effort and life circumstances as predictors of foot ulcers and amputations in people with Type 2 diabetes mellitus.
To investigate the predictive value of both patients' motivation and effort in their management of Type 2 diabetes and their life circumstances for the development of foot ulcers and amputations. This study was based on the Diabetes Care in General Practice study and Danish population and health registers. The associations between patient motivation, effort and life circumstances and foot ulcer prevalence 6 years after diabetes diagnosis and the incidence of amputation in the following 13 years were analysed using odds ratios from logistic regression and hazard ratios from Cox regression models, respectively. Foot ulcer prevalence 6 years after diabetes diagnosis was 2.93% (95% CI 1.86-4.00) among 956 patients. General practitioners' indication of 'poor' vs 'very good' patient motivation for diabetes management was associated with higher foot ulcer prevalence (odds ratio 6.11, 95% CI 1.22-30.61). The same trend was seen for 'poor' vs 'good' influence of the patient's own effort in diabetes treatment (odds ratio 7.06, 95% CI 2.65-18.84). Of 1058 patients examined at 6-year follow-up, 45 experienced amputation during the following 13 years. 'Poor' vs 'good' influence of the patients' own effort was associated with amputation (hazard ratio 7.12, 95% CI 3.40-14.92). When general practitioners assessed the influence of patients' life circumstances as 'poor' vs 'good', the amputation incidence increased (hazard ratio 2.97, 95% CI 1.22-7.24). 'Poor' vs 'very good' patient motivation was also associated with a higher amputation incidence (hazard ratio 7.57, 95% CI 2.43-23.57), although not in fully adjusted models. General practitioners' existing knowledge of patients' life circumstances, motivation and effort in diabetes management should be included in treatment strategies to prevent foot complications.


Potential Therapies Using Myogenic Stem Cells Combined with Bio-Engineering Approaches for Treatment of Muscular Dystrophies.
Muscular dystrophies (MDs) are a group of heterogeneous genetic disorders caused by mutations in the genes encoding the structural components of myofibres. The current state-of-the-art treatment is oligonucleotide-based gene therapy that restores disease-related protein. However, this therapeutic approach has limited efficacy and is unlikely to be curative. While the number of studies focused on cell transplantation therapy has increased in the recent years, this approach remains challenging due to multiple issues related to the efficacy of engrafted cells, source of myogenic cells, and systemic injections. Technical innovation has contributed to overcoming cell source challenges, and in recent studies, a combination of muscle resident stem cells and gene editing has shown promise as a novel approach. Furthermore, improvement of the muscular environment both in cultured donor cells and in recipient MD muscles may potentially facilitate cell engraftment. Artificial skeletal muscle generated by myogenic cells and muscle resident cells is an alternate approach that may enable the replacement of damaged tissues. Here, we review the current status of myogenic stem cell transplantation therapy, describe recent advances, and discuss the remaining obstacles that exist in the search for a cure for MD patients.


Identification of a cis-acting element in the class I major histocompatibility complex gene promoter responsive to activation by retroviral sequences.
The infection of cells with Moloney murine leukemia virus (M-MuLV) causes an increase in specific cellular gene products, including the major histocompatibility complex (MHC) class I antigens. This upregulation occurs through a transactivation process mediated by the long terminal repeat (LTR) of M-MuLV, and we show here that the gene activation response to the LTR requires at least one specific cis element within the MHC proximal promoter region. Nested deletions of MHC class I H-2Kb gene promoter sequence were subcloned into a chloramphenicol acetyltransferase (CAT) reporter vector and then transiently introduced into BALB/c-3T3 cells expressing M-MuLV or cotransfected into BALB/c-3T3 cells with a vector containing subgenomic portions of the virus, including the LTR. CAT activity assays demonstrated that a minimal H-2Kb gene promoter (-64 to +12) contained elements sufficient for this transactivation. DNase I footprinting assays located a protein-binding site in the region of -64 to -34 bp from the transcriptional start site, and point mutation analysis confirmed the location of this cis-acting element, designated the let response element (LRE), and defined a binding motif. This LRE is distinct from binding sites for currently known transcription factors in the class I MHC gene promoter and is conserved in the promoters of human and murine MHC class I genes. Mutation of the LRE resulted in dramatic reduction in both DNA-protein binding activity in electrophoretic mobility shift assay and in the ability of the mutated promoter to respond to retroviral transactivation. Addition of the LRE to a heterologous promoter conferred the ability to respond to retroviral transactivation.


Fullerenol C60(OH)24 effects on antioxidative enzymes activity in irradiated human erythroleukemia cell line.
Radiotherapy-induced toxicity is a major dose-limiting factor in anti-cancer treatment. Ionizing radiation leads to the formation of reactive oxygen and nitrogen species (ROS/RNS) that are associated with radiation-induced cell death. Investigations of biological effects of fullerenol have provided evidence for its ROS/RNS scavenger properties in vitro and radioprotective efficiency in vivo. Therefore we were interested to evaluate its radioprotective properties in vitro in the human erythroleukemia cell line. Pre-treatment of irradiated cells by fullerenol exerted statistically significant effects on cell numbers and the response of antioxidative enzymes to X-ray irradiation-induced oxidative stress in cells. Our study provides evidence that the pre-treatment with fullerenol enhanced the enzymatic activity of superoxide dismutase and glutathione peroxidase in irradiated K562 cells.