Question: The existence of secure locations for studying does not efficiently reduce homelessness.

Longitudinal Analysis of Mental Health Issues in Homeless Children and Families Post-Rehousing.
AIM: To assess the mental health requirements of homeless children and families before and after being provided with housing. DESIGN: A longitudinal, cross-sectional study. LOCATION: Birmingham City. PARTICIPANTS: 58 families who were rehoused, consisting of 103 children aged between 2 and 16 years, and 21 control families of low socioeconomic status with stable housing, comprising 54 children. KEY OUTCOME MEASURES: Children's mental health issues and communication levels; mothers' mental health issues and social support network one year post-rehousing. FINDINGS: Mental health issues remained significantly higher in rehoused mothers and their children compared to the control group (mothers 26% vs. 5%, P = 0.04; children 39% vs. 11%, P = 0.0003). Homeless mothers continued to have significantly reduced social support at the follow-up stage. Mothers with a history of abuse and poor social integration were more prone to have children with ongoing mental health issues. CONCLUSION: Homeless families possess a high level of intricate needs that cannot be addressed by standard health services and systems. Local strategies should be developed and implemented to ensure swift rehousing into permanent dwellings, effective social support and healthcare for both parents and children, and protection from violence and intimidation. This study highlights that the provision of housing alone is insufficient to mitigate the mental health challenges faced by homeless families.

BACKGROUND Blindness due to trachoma is avoidable through Surgery, Antibiotics, Facial hygiene and Environmental improvements (SAFE). Recent surveys have shown trachoma to be a serious cause of blindness in Southern Sudan. We conducted this survey in Ayod County of Jonglei State to estimate the need for intervention activities to eliminate blinding trachoma. METHODOLOGY AND FINDINGS A cross-sectional two-stage cluster random survey was conducted in November 2006. All residents of selected households were clinically assessed for trachoma using the World Health Organization (WHO) simplified grading scheme. A total of 2,335 people from 392 households were examined, of whom 1,107 were over 14 years of age. Prevalence of signs of active trachoma in children 1-9 years of age was: trachomatous inflammation follicular (TF) = 80.1% (95% confidence interval [CI], 73.9-86.3); trachomatous inflammation intense (TI) = 60.7% (95% CI, 54.6-66.8); and TF and/or TI (active trachoma) = 88.3% (95% CI, 83.7-92.9). Prevalence of trachomatous trichiasis (TT) was 14.6% (95% CI, 10.9-18.3) in adults over 14 years of age; 2.9% (95% CI, 0.4-5.3) in children 1-14 years of age; and 8.4% (95% CI, 5.5-11.3) overall. The prevalence of corneal opacity in persons over 14 years of age with TT was 6.4% (95% CI, 4.5-8.3). No statistically significant difference was observed in the prevalence of trachoma signs between genders. Trachoma affected almost all households surveyed: 384/392 (98.0%) had at least one person with active trachoma and 130 (33.2%) had at least one person with trichiasis. CONCLUSIONS Trachoma is an unnecessary public health problem in Ayod. The high prevalence of active trachoma and trichiasis confirms the severe burden of blinding trachoma found in other post-conflict areas of Southern Sudan. Based on WHO recommended thresholds, all aspects of the SAFE strategy are indicated to eliminate blinding trachoma in Ayod.

OBJECTIVE To determine whether exposure to low doses of ionising radiation in infancy affects cognitive function in adulthood. DESIGN Population based cohort study. SETTING Sweden. PARTICIPANTS 3094 men who had received radiation for cutaneous haemangioma before age 18 months during 1930-59. MAIN OUTCOME MEASURES Radiation dose to frontal and posterior parts of the brain, and association between dose and intellectual capacity at age 18 or 19 years based on cognitive tests (learning ability, logical reasoning, spatial recognition) and high school attendance. RESULTS The proportion of boys who attended high school decreased with increasing doses of radiation to both the frontal and the posterior parts of the brain from about 32% among those not exposed to around 17% in those who received > 250 mGy. For the frontal dose, the multivariate odds ratio was 0.47 (95% confidence interval 0.26 to 0.85, P for trend 0.0003) and for the posterior dose it was 0.59 (0.23 to 1.47, 0.0005). A negative dose-response relation was also evident for the three cognitive tests for learning ability and logical reasoning but not for the test of spatial recognition. CONCLUSIONS Low doses of ionising radiation to the brain in infancy influence cognitive abilities in adulthood.

CONTEXT Many smokers are unable or unwilling to completely quit smoking. A proposed means of harm reduction is to reduce the number of cigarettes smoked per day. However, it is not clear whether this strategy decreases the risk for tobacco-related diseases. OBJECTIVE To assess the effects of smoking reduction on lung cancer incidence. DESIGN, SETTING, AND PARTICIPANTS Observational population-based cohort study with up to 31 years of follow-up from the Copenhagen Centre for Prospective Population Studies, which administrates data from 3 longitudinal studies conducted in Copenhagen and suburbs, the Copenhagen City Heart Study, the Copenhagen Male Study, and the Glostrup Population Studies, Denmark. Participants were 11,151 men and 8563 women (N = 19,714) aged 20 to 93 years, who attended 2 consecutive examinations with a 5- to 10-year interval between 1964 and 1988. Participants underwent a physical examination and completed self-filled questionnaires about lifestyle habits. The study population was divided into 6 groups according to smoking habits: continued heavy smokers (> or =15 cigarettes/d), reducers (reduced from > or =15 cigarettes/d by minimum of 50% without quitting), continued light smokers (1-14 cigarettes/d), quitters (stopped between first and second examination), stable ex-smokers, and never smokers. MAIN OUTCOME MEASURE Incident primary lung cancer cases assessed by record linkage with the National Cancer Registry until December 31, 2003. RESULTS There were 864 incident lung cancers during follow-up. Using Cox regression, the adjusted hazard ratio (HR) for lung cancer in reducers was 0.73 (95% confidence interval [CI], 0.54-0.98) compared with persistent heavy smokers. The HR for light smokers was 0.44 (95% CI, 0.35-0.56); for quitters, HR 0.50 (95% CI, 0.36-0.69), for stable ex-smokers, HR 0.17 (95% CI, 0.13-0.23), and for never smokers, HR 0.09 (95% CI, 0.06-0.13). CONCLUSION Among individuals who smoke 15 or more cigarettes per day, smoking reduction by 50% significantly reduces the risk of lung cancer.

CONTEXT One key target of the United Nations Millennium Development goals is to reduce the prevalence of underweight among children younger than 5 years by half between 1990 and 2015. OBJECTIVE To estimate trends in childhood underweight by geographic regions of the world. DESIGN, SETTING, AND PARTICIPANTS Time series study of prevalence of underweight, defined as weight 2 SDs below the mean weight for age of the National Center for Health Statistics and World Health Organization (WHO) reference population. National prevalence rates derived from the WHO Global Database on Child Growth and Malnutrition, which includes data on approximately 31 million children younger than 5 years who participated in 419 national nutritional surveys in 139 countries from 1965 through 2002. MAIN OUTCOME MEASURES Linear mixed-effects modeling was used to estimate prevalence rates and numbers of underweight children by region in 1990 and 2015 and to calculate the changes (ie, increase or decrease) to these values between 1990 and 2015. RESULTS Worldwide, underweight prevalence was projected to decline from 26.5% in 1990 to 17.6% in 2015, a change of -34% (95% confidence interval [CI], -43% to -23%). In developed countries, the prevalence was estimated to decrease from 1.6% to 0.9%, a change of -41% (95% CI, -92% to 343%). In developing regions, the prevalence was forecasted to decline from 30.2% to 19.3%, a change of -36% (95% CI, -45% to -26%). In Africa, the prevalence of underweight was forecasted to increase from 24.0% to 26.8%, a change of 12% (95% CI, 8%-16%). In Asia, the prevalence was estimated to decrease from 35.1% to 18.5%, a change of -47% (95% CI, -58% to -34%). Worldwide, the number of underweight children was projected to decline from 163.8 million in 1990 to 113.4 million in 2015, a change of -31% (95% CI, -40% to -20%). Numbers are projected to decrease in all subregions except the subregions of sub-Saharan, Eastern, Middle, and Western Africa, which are expected to experience substantial increases in the number of underweight children. CONCLUSIONS An overall improvement in the global situation is anticipated; however, neither the world as a whole, nor the developing regions, are expected to achieve the Millennium Development goals. This is largely due to the deteriorating situation in Africa where all subregions, except Northern Africa, are expected to fail to meet the goal.

BACKGROUND Self-harm and suicide are common in prisoners, yet robust information on the full extent and characteristics of people at risk of self-harm is scant. Furthermore, understanding how frequently self-harm is followed by suicide, and in which prisoners this progression is most likely to happen, is important. We did a case-control study of all prisoners in England and Wales to ascertain the prevalence of self-harm in this population, associated risk factors, clustering effects, and risk of subsequent suicide after self-harm. METHODS Records of self-harm incidents in all prisons in England and Wales were gathered routinely between January, 2004, and December, 2009. We did a case-control comparison of prisoners who self-harmed and those who did not between January, 2006, and December, 2009. We also used a Bayesian approach to look at clustering of people who self-harmed. Prisoners who self-harmed and subsequently died by suicide in prison were compared with other inmates who self-harmed. FINDINGS 139,195 self-harm incidents were recorded in 26,510 individual prisoners between 2004 and 2009; 5-6% of male prisoners and 20-24% of female inmates self-harmed every year. Self-harm rates were more than ten times higher in female prisoners than in male inmates. Repetition of self-harm was common, particularly in women and teenage girls, in whom a subgroup of 102 prisoners accounted for 17,307 episodes. In both sexes, self-harm was associated with younger age, white ethnic origin, prison type, and a life sentence or being unsentenced; in female inmates, committing a violent offence against an individual was also a factor. Substantial evidence was noted of clustering in time and location of prisoners who self-harmed (adjusted intra-class correlation 0·15, 95% CI 0·11-0·18). 109 subsequent suicides in prison were reported in individuals who self-harmed; the risk was higher in those who self-harmed than in the general prison population, and more than half the deaths occurred within a month of self-harm. Risk factors for suicide after self-harm in male prisoners were older age and a previous self-harm incident of high or moderate lethality; in female inmates, a history of more than five self-harm incidents within a year was associated with subsequent suicide. INTERPRETATION The burden of self-harm in prisoners is substantial, particularly in women. Self-harm in prison is associated with subsequent suicide in this setting. Prevention and treatment of self-harm in prisoners is an essential component of suicide prevention in prisons. FUNDING Wellcome Trust, National Institute for Health Research, National Offender Management Service, and Department of Health.

Spatial Release from Masking (SRM) was measured for identification of a female target word spoken in the presence of male masker words. Target words from a single loudspeaker located at midline were presented when two, four, or six masker words were presented either from the same source as the target or from spatially separated masker sources. All masker words were presented from loudspeakers located symmetrically around the centered target source in the front azimuth hemifield. Three masking conditions were employed: speech-in-speech masking (involving both informational and energetic masking), speech-in-noise masking (involving energetic masking), and filtered speech-in-filtered speech masking (involving informational masking). Psychophysical results were summarized as three-point psychometric functions relating proportion of correct word identification to target-to-masker ratio (in decibels) for both the co-located and spatially separated target and masker sources cases. SRM was then calculated by comparing the slopes and intercepts of these functions. SRM decreased as the number of symmetrically placed masker sources increased from two to six. This decrease was independent of the type of masking, with almost no SRM measured for six masker sources. These results suggest that when SRM is dependent primarily on binaural processing, SRM is effectively limited to fewer than six sound sources.

The paper offers first a brief historical overview of disaster management planning. Second, it reviews Australian and American research findings and show that they urge the field of disaster management to shift its focus from response and recovery to sustainable hazard mitigation. It is argued that in order for this shift to occur, it is necessary to integrate disaster management and community planning. Current practice seldom reflects such a synthesis, and this is one of the reasons why hazard awareness is absent from local decision-making processes. Third, it is asserted that if mitigative strategies are to be successfully implemented, then the disaster management process must incorporate public participation at the local decision-making level. The paper concludes with a case study of California's Portola Valley, which demonstrates that when public participation is integrated into disaster management planning and community planning, the result is sustainable hazard mitigation.

Factors associated with primary care utilization and health insurance coverage were examined among 511 women leaving jail in New York City from 1997-2001. One year after release, roughly half of the sample reported primary care utilization (47%) and health insurance coverage (56%). Neither outcome was more likely among those reporting diabetes, asthma, or depression. Primary care utilization was more likely among those reporting receipt of public benefits, health insurance coverage, moderate social support, avoidance of illegal activity, and HIV seropositivity. Health insurance coverage was associated with receipt of public benefits, hospitalization, primary care, and avoiding re-arrest. This study demonstrated that a majority of women leaving jail, including those with chronic diseases, lack primary care. These data highlight the need to plan for continuity of care from corrections to the community and suggest further that this can be facilitated with provision of health benefits and social support.

BACKGROUND Transmission of dengue viruses (DENV), the leading cause of arboviral disease worldwide, is known to vary through time and space, likely owing to a combination of factors related to the human host, virus, mosquito vector, and environment. An improved understanding of variation in transmission patterns is fundamental to conducting surveillance and implementing disease prevention strategies. To test the hypothesis that DENV transmission is spatially and temporally focal, we compared geographic and temporal characteristics within Thai villages where DENV are and are not being actively transmitted. METHODS AND FINDINGS Cluster investigations were conducted within 100 m of homes where febrile index children with (positive clusters) and without (negative clusters) acute dengue lived during two seasons of peak DENV transmission. Data on human infection and mosquito infection/density were examined to precisely (1) define the spatial and temporal dimensions of DENV transmission, (2) correlate these factors with variation in DENV transmission, and (3) determine the burden of inapparent and symptomatic infections. Among 556 village children enrolled as neighbors of 12 dengue-positive and 22 dengue-negative index cases, all 27 DENV infections (4.9% of enrollees) occurred in positive clusters (p < 0.01; attributable risk [AR] = 10.4 per 100; 95% confidence interval 1-19.8 per 100]. In positive clusters, 12.4% of enrollees became infected in a 15-d period and DENV infections were aggregated centrally near homes of index cases. As only 1 of 217 pairs of serologic specimens tested in positive clusters revealed a recent DENV infection that occurred prior to cluster initiation, we attribute the observed DENV transmission subsequent to cluster investigation to recent DENV transmission activity. Of the 1,022 female adult Ae. aegypti collected, all eight (0.8%) dengue-infected mosquitoes came from houses in positive clusters; none from control clusters or schools. Distinguishing features between positive and negative clusters were greater availability of piped water in negative clusters (p < 0.01) and greater number of Ae. aegypti pupae per person in positive clusters (p = 0.04). During primarily DENV-4 transmission seasons, the ratio of inapparent to symptomatic infections was nearly 1:1 among child enrollees. Study limitations included inability to sample all children and mosquitoes within each cluster and our reliance on serologic rather than virologic evidence of interval infections in enrollees given restrictions on the frequency of blood collections in children. CONCLUSIONS Our data reveal the remarkably focal nature of DENV transmission within a hyperendemic rural area of Thailand. These data suggest that active school-based dengue case detection prompting local spraying could contain recent virus introductions and reduce the longitudinal risk of virus spread within rural areas. Our results should prompt future cluster studies to explore how host immune and behavioral aspects may impact DENV transmission and prevention strategies. Cluster methodology could serve as a useful research tool for investigation of other temporally and spatially clustered infectious diseases.

OBJECTIVE Methylphenidate (MPH), the most commonly prescribed drug for attention-deficit/hyperactivity disorder (ADHD), has a short half-life, which necessitates multiple daily doses. The need for multiple doses produces problems with medication administration during school and after-school hours, and therefore with compliance. Previous long-acting stimulants and preparations have shown effects equivalent to twice-daily dosing of MPH. This study tests the efficacy and duration of action, in natural and laboratory settings, of an extended-release MPH preparation designed to last 12 hours and therefore be equivalent to 3-times-daily dosing. METHODS Sixty-eight children with ADHD, 6 to 12 years old, participated in a within-subject, double-blind comparison of placebo, immediate-release (IR) MPH 3 times a day (tid), and Concerta, a once-daily MPH formulation. Three dosing levels of medication were used: 5 mg IR MPH tid/18 mg Concerta once a day (qd); 10 mg IR MPH tid/36 mg Concerta qd; and 15 mg IR MPH tid/54 mg Concerta qd. All children were currently medicated with MPH at enrollment, and each child's dose level was based on that child's MPH dosing before the study. The doses of Concerta were selected to be comparable to the daily doses of MPH that each child received. To achieve the ascending rate of MPH delivery determined by initial investigations to provide the necessary continuous coverage, Concerta doses were 20% higher on a daily basis than a comparable tid regimen of IR MPH. Children received each medication condition for 7 days. The investigation was conducted in the context of a background clinical behavioral intervention in both the natural environment and the laboratory setting. Parents received behavioral parent training and teachers were taught to establish a school-home daily report card (DRC). A DRC is a list of individual target behaviors that represent a child's most salient areas of impairment. Teachers set daily goals for each child's impairment targets, and parents provided rewards at home for goal attainment. Each weekday, teachers completed the DRC, and it was used as a dependent measure of individualized medication response. Teachers and parents also completed weekly standardized ratings of behavior and treatment effectiveness. To evaluate the time course of medication effects, children spent 12 hours in a laboratory setting on Saturdays and medication effects were measured using procedures and methods adapted from our summer treatment program. Measures of classroom behavior and academic productivity/accuracy were taken in a laboratory classroom setting during which children completed independent math and reading worksheets. Measures of social behavior were taken in structured, small-group board game settings and unstructured recess settings. Measures included behavior frequency counts, academic problems completed and accuracy, independent observations, teacher and counselor ratings, and individualized behavioral target goals. Reports of adverse events, sleep quality, and appetite were collected. RESULTS On virtually all measures in all settings, both drug conditions were significantly different from placebo, and the 2 drugs were not different from each other. In children's regular school settings, both medications improved behavior as measured by teacher ratings and individualized target behaviors (the DRC); these effects were seen into the evening as measured by parent ratings. In the laboratory setting, effects of Concerta were equivalent to tid MPH and lasted at least through 12 hours after dosing. Concerta was significantly superior to tid MPH on 2 parent rating scores, and when asked, more parents preferred Concerta than preferred tid IR MPH or placebo. Side effects on children's sleep and appetite were similar for the 2 preparations. In the lab setting, both medications improved productivity and accuracy on arithmetic seatwork assignments, disruptive and on-task behavior, and classroom rule following. Both medications improved children's rule following and negative behavior in small group board games, as well as in unstructured recess settings. Individual target behaviors also showed significant improvement with medication across domains in the laboratory setting. Children's behavior across settings deteriorated across the laboratory day, and the primary effect of medication was to prevent this deterioration as the day wore on. Results support the use of background behavioral treatment in clinical trials of stimulant medication, and illustrate the utility of a measure of individualized daily target goals (ie, the DRC) as an objective measure of medication response in both the laboratory and natural school settings. CONCLUSION This investigation clearly supports the efficacy of the Concerta long-acting formulation of MPH for parents who desire to have medication benefits for their child throughout the day and early evening. (ABSTRACT TRUNCATED)


Question: The advantages of colchicine were attained through the efficient broad implementation of secondary prevention tactics, including high-dose statins.

Comparative Risk Factors for Cardiovascular Events Over Four Years in Stable Outpatients Prone to Atherothrombosis.

Background: Identifying patients at the highest risk for cardiovascular events poses a challenge for both clinicians and researchers. Prior ischemic events, polyvascular disease, and diabetes mellitus are established predictors, but their relative impact on future cardiovascular risk remains uncertain. This study aims to categorize cardiovascular event risk in stable outpatients with varying initial signs of atherothrombosis using straightforward clinical indicators.

Methodology: Outpatients with coronary artery disease, cerebrovascular disease, peripheral arterial disease, or multiple risk factors for atherothrombosis were enrolled in the global Reduction of Atherothrombosis for Continued Health (REACH) Registry. The patients were followed for up to 4 years. The study involved 45,227 participants from 3647 centers across 29 countries, with enrollment between 2003 and 2004, and the final follow-up in 2008. The primary outcomes were rates of cardiovascular death, myocardial infarction, and stroke.

Results: A total of 5481 patients experienced at least one event during the follow-up period, including 2315 cardiovascular deaths, 1228 myocardial infarctions, and 1898 strokes. Patients with a prior history of ischemic events had the highest subsequent ischemic event rate (18.3%; 95% CI, 17.4%-19.1%). Those with stable coronary, cerebrovascular, or peripheral artery disease had a lower risk (12.2%; 95% CI, 11.4%-12.9%), and patients without established atherothrombosis but with risk factors only had the lowest risk (9.1%; 95% CI, 8.3%-9.9%) (P < .001 for all comparisons). In multivariate analysis, diabetes (HR, 1.44; 95% CI, 1.36-1.53; P < .001), an ischemic event within the past year (HR, 1.71; 95% CI, 1.57-1.85; P < .001), and polyvascular disease (HR, 1.99; 95%

BACKGROUND AND PURPOSE Soluble corin was decreased in coronary heart disease. Given the connections between cardiac dysfunction and stroke, circulating corin might be a candidate marker of stroke risk. However, the association between circulating corin and stroke has not yet been studied in humans. Here, we aimed to examine the association in patients wtith stroke and community-based healthy controls. METHODS Four hundred eighty-one patients with ischemic stroke, 116 patients with hemorrhagic stroke, and 2498 healthy controls were studied. Serum soluble corin and some conventional risk factors of stroke were examined. Because circulating corin was reported to be varied between men and women, the association between serum soluble corin and stroke was evaluated in men and women, respectively. RESULTS Patients with ischemic and hemorrhagic stroke had a significantly lower level of serum soluble corin than healthy controls in men and women (all P values, <0.05). In multivariate analysis, men in the lowest quartile of serum soluble corin were more likely to have ischemic (odds ratio [OR], 4.90; 95% confidence interval, 2.99-8.03) and hemorrhagic (OR, 17.57; 95% confidence interval, 4.85-63.71) stroke than men in the highest quartile. Women in the lowest quartile of serum soluble corin were also more likely to have ischemic (OR, 3.10; 95% confidence interval, 1.76-5.44) and hemorrhagic (OR, 8.54; 95% confidence interval, 2.35-31.02) stroke than women in the highest quartile. ORs of ischemic and hemorrhagic stroke were significantly increased with the decreasing levels of serum soluble corin in men and women (all P values for trend, <0.001). CONCLUSIONS Serum soluble corin was decreased in patients with stroke compared with healthy controls. Our findings raise the possibility that serum soluble corin may have a pathogenic role in stroke.

BACKGROUND The Global Burden of Diseases, Injuries, and Risk Factors Study 2015 provides an up-to-date synthesis of the evidence for risk factor exposure and the attributable burden of disease. By providing national and subnational assessments spanning the past 25 years, this study can inform debates on the importance of addressing risks in context. METHODS We used the comparative risk assessment framework developed for previous iterations of the Global Burden of Disease Study to estimate attributable deaths, disability-adjusted life-years (DALYs), and trends in exposure by age group, sex, year, and geography for 79 behavioural, environmental and occupational, and metabolic risks or clusters of risks from 1990 to 2015. This study included 388 risk-outcome pairs that met World Cancer Research Fund-defined criteria for convincing or probable evidence. We extracted relative risk and exposure estimates from randomised controlled trials, cohorts, pooled cohorts, household surveys, census data, satellite data, and other sources. We used statistical models to pool data, adjust for bias, and incorporate covariates. We developed a metric that allows comparisons of exposure across risk factors-the summary exposure value. Using the counterfactual scenario of theoretical minimum risk level, we estimated the portion of deaths and DALYs that could be attributed to a given risk. We decomposed trends in attributable burden into contributions from population growth, population age structure, risk exposure, and risk-deleted cause-specific DALY rates. We characterised risk exposure in relation to a Socio-demographic Index (SDI). FINDINGS Between 1990 and 2015, global exposure to unsafe sanitation, household air pollution, childhood underweight, childhood stunting, and smoking each decreased by more than 25%. Global exposure for several occupational risks, high body-mass index (BMI), and drug use increased by more than 25% over the same period. All risks jointly evaluated in 2015 accounted for 57·8% (95% CI 56·6-58·8) of global deaths and 41·2% (39·8-42·8) of DALYs. In 2015, the ten largest contributors to global DALYs among Level 3 risks were high systolic blood pressure (211·8 million [192·7 million to 231·1 million] global DALYs), smoking (148·6 million [134·2 million to 163·1 million]), high fasting plasma glucose (143·1 million [125·1 million to 163·5 million]), high BMI (120·1 million [83·8 million to 158·4 million]), childhood undernutrition (113·3 million [103·9 million to 123·4 million]), ambient particulate matter (103·1 million [90·8 million to 115·1 million]), high total cholesterol (88·7 million [74·6 million to 105·7 million]), household air pollution (85·6 million [66·7 million to 106·1 million]), alcohol use (85·0 million [77·2 million to 93·0 million]), and diets high in sodium (83·0 million [49·3 million to 127·5 million]). From 1990 to 2015, attributable DALYs declined for micronutrient deficiencies, childhood undernutrition, unsafe sanitation and water, and household air pollution; reductions in risk-deleted DALY rates rather than reductions in exposure drove these declines. Rising exposure contributed to notable increases in attributable DALYs from high BMI, high fasting plasma glucose, occupational carcinogens, and drug use. Environmental risks and childhood undernutrition declined steadily with SDI; low physical activity, high BMI, and high fasting plasma glucose increased with SDI. In 119 countries, metabolic risks, such as high BMI and fasting plasma glucose, contributed the most attributable DALYs in 2015. Regionally, smoking still ranked among the leading five risk factors for attributable DALYs in 109 countries; childhood underweight and unsafe sex remained primary drivers of early death and disability in much of sub-Saharan Africa. INTERPRETATION Declines in some key environmental risks have contributed to declines in critical infectious diseases. Some risks appear to be invariant to SDI. Increasing risks, including high BMI, high fasting plasma glucose, drug use, and some occupational exposures, contribute to rising burden from some conditions, but also provide opportunities for intervention. Some highly preventable risks, such as smoking, remain major causes of attributable DALYs, even as exposure is declining. Public policy makers need to pay attention to the risks that are increasingly major contributors to global burden. FUNDING Bill & Melinda Gates Foundation.

OBJECTIVE Serum soluble corin has been associated with stroke. However, whether it is associated with stroke prognosis has not yet been studied. Therefore, we aimed to study the association of serum soluble corin with risk of poor outcomes within 3 months after stroke. METHODS We followed 522 stroke patients for 3 months to identify major disability, death and vascular events. Serum soluble corin was measured at baseline for all participants. Logistic regression was used to examine the associations of baseline serum soluble corin with outcomes of stroke, adjusting for age, sex, baseline NIHSS score, hours from onset to hospitalization, smoking, drinking, hypertension, diabetes, coronary heart disease, atrial fibrillation, family history of stroke, and stroke subtype. RESULTS Patients with high corin had a significantly lower crude risk for the composite outcome of major disability or death (OR = 0.64, 95%CI: 0.43-0.96) than patients with low corin (the lowest tertile). After adjustment for age and baseline NIHSS score, patients with high corin still had a significantly lower risk for the composite outcome of major disability or death (OR = 0.60, 95%CI: 0.36-0.99). This association became bottom line significant after additionally adjusting for other conventional factors (OR = 0.61, P = 0.058). No association was found between serum soluble corin and other composite outcomes. CONCLUSION Serum soluble corin deficiency predicted risk for major disability within 3 months after stroke, independent of baseline neurological deficient. Our results may indicate a probable role of corin in stroke prognosis.

Coronary heart disease rates have been reported in several parts of the world to be unusually high in people originating from the Indian subcontinent. High coronary disease rates appear to be common to South Asian groups of different geographical origin, religion, and language. This presents a challenge to the understanding of coronary heart disease: the high rates in South Asians are not explained on the basis of elevated serum cholesterol, smoking or hypertension. Low plasma HDL cholesterol, high plasma triglyceride levels and high prevalence of non-insulin-dependent diabetes have been consistently found in South Asians overseas: this probably reflects an underlying state of insulin resistance. Further studies are needed to determine whether this metabolic disturbance can account for the high rates of coronary heart disease in South Asians, and to identify possibilities for prevention.

BACKGROUND Homocysteine is a risk factor for coronary artery disease (CAD), although a causal relation remains to be proven. The importance of determining direct causality rests in the fact that plasma homocysteine can be safely and inexpensively reduced by 25% with folic acid. This reduction is maximally achieved by doses of 0.4 mg/d. High-dose folic acid (5 mg/d) improves endothelial function in CAD, although the mechanism is controversial. It has been proposed that improvement occurs through reduction in total (tHcy) or free (non-protein bound) homocysteine (fHcy). We investigated the effects of folic acid on endothelial function before a change in homocysteine in patients with CAD. METHODS AND RESULTS A randomized, placebo-controlled study of folic acid (5 mg/d) for 6 weeks was undertaken in 33 patients. Endothelial function, assessed by flow-mediated dilatation (FMD), was measured before, at 2 and 4 hours after the first dose of folic acid, and after 6 weeks of treatment. Plasma folate increased markedly by 1 hour (200 compared with 25.8 nmol/L; P<0.001). FMD improved at 2 hours (83 compared with 47 microm; P<0.001) and was largely complete by 4 hours (101 compared with 51 microm; P<0.001). tHcy did not significantly differ acutely (4-hour tHcy, 9.56 compared with 9.79 micromol/L; P=NS). fHcy did not differ at 3 hours but was slightly reduced at 4 hours (1.55 compared with 1.78 micromol/L; P=0.02). FMD improvement did not correlate with reductions in either fHcy or tHcy at any time. CONCLUSIONS These data suggest that folic acid improves endothelial function in CAD acutely by a mechanism largely independent of homocysteine.

OBJECTIVE To determine the effects of smoking, plasma lipids, lipoproteins, apolipoproteins, and fibrinogen on the patency of saphenous vein femoropopliteal bypass grafts at one year. DESIGN Prospective study of patients with saphenous vein femoropopliteal bypass grafts entered into a multicentre trial. SETTING Surgical wards, outpatient clinics, and home visits coordinated by two tertiary referral centres in London and Birmingham. PATIENTS 157 Patients (mean age 66.6 (SD 8.2) years), 113 with patent grafts and 44 with occluded grafts one year after bypass. MAIN OUTCOME MEASURE Cumulative percentage patency at one year. RESULTS Markers for smoking (blood carboxyhaemoglobin concentration (p less than 0.05) and plasma thiocyanate concentration (p less than 0.01) and plasma concentrations of fibrinogen (p less than 0.001) and apolipoproteins AI (p less than 0.04) and (a) (p less than 0.05) were significantly higher in patients with occluded grafts. Serum cholesterol concentrations were significantly higher in patients with grafts that remained patent one year after bypass (p less than 0.005). Analysis of the smoking markers indicated that a quarter of patients (40) were untruthful in their claims to have stopped smoking. Based on smoking markers, patency of grafts in smokers was significantly lower at one year by life table analysis than in non-smokers (63% v 84%, p less than 0.02). Patency was significantly higher by life table analysis in patients with a plasma fibrinogen concentration below the median than in those with a concentration above (90% v 57%, p less than 0.0002). Surprisingly, increased plasma low density lipoprotein cholesterol concentration was significantly associated with improved patency at one year (85%) at values above the median compared with patency (only 68%) at values in the lower half of the range (p less than 0.02). CONCLUSIONS Plasma fibrinogen concentration was the most important variable predicting graft occlusion, followed by smoking markers. A more forceful approach is needed to stop patients smoking; therapeutic measures to improve patency of vein grafts should focus on decreasing plasma fibrinogen concentration rather than serum cholesterol concentration.

Unstable angina is a critical phase of coronary heart disease with widely variable symptoms and prognosis. A decade ago, a classification of unstable angina based on clinical symptoms was introduced. This system was then validated by prospective clinical studies to correlate with the prognosis and was linked to angiographic and histological findings. It has been used to categorize patients in many large clinical trials. In recent years, the pathophysiological roles of platelet activation and inflammation in unstable angina have been elucidated. Subsequently, improved markers of myocardial injury, acute-phase proteins, and hemostatic markers that may be associated with clinical outcomes have been identified. Particularly, cardiac-specific troponin T and troponin I have been shown to represent the best predictors of early risk in patients with angina at rest. Accordingly, it is suggested that the original classification be extended by subclassifying one large group of unstable angina patients, ie, those with angina at rest within the past 48 hours (class IIIB), into troponin-positive (T(pos)) and troponin-negative (T(neg)) patients. The 30-days risk for death and myocardial infarction is considered to be up to 20% in class IIIB-T(pos) but <2% in class IIIB-T(neg) patients. Initial results suggest that troponins may function as surrogate markers for thrombus formation and can effectively guide therapy with glycoprotein IIb/IIIa antagonists or low-molecular-weight heparins. These observations provide additional impetus for adding the measurement of these markers to the clinical classification and represent a novel concept of treating these high-risk patients.

IMPORTANCE Exacerbations of respiratory symptoms in chronic obstructive pulmonary disease (COPD) have profound and long-lasting adverse effects on patients. OBJECTIVE To test the hypothesis that elevated levels of inflammatory biomarkers in individuals with stable COPD are associated with an increased risk of having exacerbations. DESIGN, SETTING, AND PARTICIPANTS Prospective cohort study examining 61,650 participants with spirometry measurements from the Copenhagen City Heart Study (2001-2003) and the Copenhagen General Population Study (2003-2008). Of these, 6574 had COPD, defined as a ratio between forced expiratory volume in 1 second (FEV1) and forced vital capacity below 0.7. MAIN OUTCOMES AND MEASURES Baseline levels of C-reactive protein (CRP) and fibrinogen and leukocyte count were measured in participants at a time when they were not experiencing symptoms of exacerbations. Exacerbations were recorded and defined as short-course treatment with oral corticosteroids alone or in combination with an antibiotic or as a hospital admission due to COPD. Levels of CRP and fibrinogen and leukocyte count were defined as high or low according to cut points of 3 mg/L, 14 μmol/L, and 9 ×10(9)/L, respectively. RESULTS During follow-up, 3083 exacerbations were recorded (mean, 0.5/participant). In the first year of follow-up, multivariable-adjusted odds ratios for having frequent exacerbations were 1.2 (95% CI, 0.7-2.2; 17 events/1000 person-years) for individuals with 1 high biomarker, 1.7 (95% CI, 0.9-3.2; 32 events/1000 person-years) for individuals with 2 high biomarkers, and 3.7 (95% CI, 1.9-7.4; 81 events/1000 person-years) for individuals with 3 high biomarkers compared with individuals who had no elevated biomarkers (9 events/1000 person-years; trend: P = 2 × 10(-5)). Corresponding hazard ratios using maximum follow-up time were 1.4 (95% CI, 1.1-1.8), 1.6 (95% CI, 1.3-2.2), and 2.5 (95% CI, 1.8-3.4), respectively (trend: P = 1 × 10(-8)). The addition of inflammatory biomarkers to a basic model including age, sex, FEV1 percent predicted, smoking, use of any inhaled medication, body mass index, history of previous exacerbations, and time since most recent prior exacerbation improved the C statistics from 0.71 to 0.73 (comparison: P = 9 × 10(-5)). Relative risks were consistent in those with milder COPD, in those with no history of frequent exacerbations, and in the 2 studies separately. The highest 5-year absolute risks of having frequent exacerbations in those with 3 high biomarkers (vs no high biomarkers) were 62% (vs 24%) for those with Global Initiative for Chronic Obstructive Lung Disease (GOLD) grades C-D (n = 558), 98% (vs 64%) in those with a history of frequent exacerbations (n = 127), and 52% (vs 15%) for those with GOLD grades 3-4 (n = 465). CONCLUSIONS AND RELEVANCE Simultaneously elevated levels of CRP and fibrinogen and leukocyte count in individuals with COPD were associated with increased risk of having exacerbations, even in those with milder COPD and in those without previous exacerbations. Further investigation is needed to determine the clinical value of these biomarkers for risk stratification.

Over the past three decades great efforts have been made in search of cancer chemoprevention strategies. The increase in knowledge of the long process from normal to cancer cell has enabled interventions in terms of lifestyle modifications, natural compounds or drugs to block or reverse the process. Great successes have been achieved, especially for breast and colorectal cancer. However, these strategies have yet to find clinical application on a large scale. In this article we identify the achievements, the pitfalls and the next steps to be taken to improve the efficacy and applicability of chemoprevention strategies. Among the crucial key points to be implemented are educational activities for physicians to appropriately disseminate the aim and indeed the culture of chemoprevention. It is essential to improve the risk-benefit balance, seeking the minimal active doses, intermittent schedules, a better characterization of the risk categories via a more personalized intervention based on individual characteristics, and ensure the containment of costs of public and private health prevention programs.

Observational clinical and ex vivo studies have established a strong association between atrial fibrillation and inflammation. However, whether inflammation is the cause or the consequence of atrial fibrillation and which specific inflammatory mediators may increase the atria's susceptibility to fibrillation remain elusive. Here we provide experimental and clinical evidence for the mechanistic involvement of myeloperoxidase (MPO), a heme enzyme abundantly expressed by neutrophils, in the pathophysiology of atrial fibrillation. MPO-deficient mice pretreated with angiotensin II (AngII) to provoke leukocyte activation showed lower atrial tissue abundance of the MPO product 3-chlorotyrosine, reduced activity of matrix metalloproteinases and blunted atrial fibrosis as compared to wild-type mice. Upon right atrial electrophysiological stimulation, MPO-deficient mice were protected from atrial fibrillation, which was reversed when MPO was restored. Humans with atrial fibrillation had higher plasma concentrations of MPO and a larger MPO burden in right atrial tissue as compared to individuals devoid of atrial fibrillation. In the atria, MPO colocalized with markedly increased formation of 3-chlorotyrosine. Our data demonstrate that MPO is a crucial prerequisite for structural remodeling of the myocardium, leading to an increased vulnerability to atrial fibrillation.


Question: The binding direction of the ML-SA1 activator to hTRPML2 is distinct from the binding direction of the ML-SA1 activator to hTRPML1.

Structural Insights into Human TRPML1 Channel Regulation in Open and Closed States

Transient receptor potential mucolipin 1 (TRPML1) is a crucial Ca2+-conducting cation channel that governs calcium signaling and lysosomal homeostasis. Genetic variations in TRPML1 can cause mucolipidosis type IV, a severe lysosomal storage disease. In this study, we present two cryo-electron microscopy structures of the full-length human TRPML1 channel: a 3.72-Å resolution structure in the apo, closed state at pH 7.0, and a 3.49-Å resolution structure in the open state at pH 6.0, bound to an agonist. The agonist is accommodated within a hydrophobic pocket formed by specific aromatic and hydrophobic residues from pore helix 1, helices S5 and S6, and helix S6 of an adjacent subunit, indicating a unique agonist-binding site distinct from that observed in TRPV1, a TRP channel belonging to a different subfamily. The activation of TRPML1 involves a distinct widening of the lower gate and a subtle rearrangement of pore helix 1. This research elucidates the regulatory mechanisms of TRPML channels, deepens our understanding of TRP channel activation, and offers valuable insights into the molecular pathogenesis of mucolipidosis type IV.

The enteric nervous system (ENS) autonomously controls gut muscle activity. Mechanosensitive enteric neurons (MEN) initiate reflex activity by responding to mechanical deformation of the gastrointestinal wall. MEN throughout the gut primarily respond to compression or stretch rather than to shear force. Some MEN are multimodal as they respond to compression and stretch. Depending on the region up to 60% of the entire ENS population responds to mechanical stress. MEN fire action potentials after mechanical stimulation of processes or soma although they are more sensitive to process deformation. There are at least two populations of MEN based on their sensitivity to different modalities of mechanical stress and on their firing pattern. (1) Rapidly, slowly and ultra-slowly adapting neurons which encode compressive forces. (2) Ultra-slowly adapting stretch-sensitive neurons encoding tensile forces. Rapid adaptation of firing is typically observed after compressive force while slow adaptation or ongoing spike discharge occurs often during tensile stress (stretch). All MEN have some common properties: they receive synaptic input, are low fidelity mechanoreceptors and are multifunctional in that some serve interneuronal others even motor functions. Consequently, MEN possess processes with mechanosensitive as well as efferent functions. This raises the intriguing hypothesis that MEN sense and control muscle activity at the same time as servo-feedback loop. The mechanosensitive channel(s) or receptor(s) expressed by the different MEN populations are unknown. Future concepts have to incorporate compressive and tensile-sensitive MEN into neural circuits that controls muscle activity. They may interact to control various forms of a particular motor pattern or regulate different motor patterns independently from each other.

The proteasome is a major cytosolic proteolytic assembly, essential for the physiology of eukaryotic cells. Both the architecture and enzymatic properties of the 20S proteasome are relatively well understood. However, despite longstanding interest, the integration of structural and functional properties of the proteasome into a coherent model explaining the mechanism of its enzymatic actions has been difficult. Recently, we used tapping mode atomic force microscopy (AFM) in liquid to demonstrate that the alpha-rings of the proteasome imaged in a top-view position repeatedly switched between their open and closed conformations, apparently to control access to the central channel. Here, we show with AFM that the molecules in a side-view position acquired two stable conformations. The overall shapes of the 20S particles were classified as either barrel-like or cylinder-like. The relative abundance of the two conformers depended on the nature of their interactions with ligands. Similarly to the closed molecules in top view, the barrels predominated in control or inhibited molecules. The cylinders and open molecules prevailed when the proteasome was observed in the presence of peptide substrates. Based on these data, we developed the two-state model of allosteric transitions to explain the dynamics of proteasomal structure. This model helps to better understand the observed properties of the 20S molecule, and sets foundations for further studies of the structural dynamics of the proteasome.

Leucine-rich repeat kinase 2 (LRRK2) mutations are the most common genetic cause of Parkinson's disease. LRRK2 is a multifunctional protein affecting many cellular processes and has been described to bind microtubules. Defective microtubule-based axonal transport is hypothesized to contribute to Parkinson's disease, but whether LRRK2 mutations affect this process to mediate pathogenesis is not known. Here we find that LRRK2 containing pathogenic Roc-COR domain mutations (R1441C, Y1699C) preferentially associates with deacetylated microtubules, and inhibits axonal transport in primary neurons and in Drosophila, causing locomotor deficits in vivo. In vitro, increasing microtubule acetylation using deacetylase inhibitors or the tubulin acetylase αTAT1 prevents association of mutant LRRK2 with microtubules, and the deacetylase inhibitor trichostatin A (TSA) restores axonal transport. In vivo knockdown of the deacetylases HDAC6 and Sirt2, or administration of TSA rescues both axonal transport and locomotor behavior. Thus, this study reveals a pathogenic mechanism and a potential intervention for Parkinson's disease.

Single-molecule tracking and fluorescence correlation spectroscopy (FCS) applied to the plasma membrane in living cells have allowed a number of unprecedented observations, thus fostering a new basic understanding of molecular diffusion, interaction, and signal transduction in the plasma membrane. It is becoming clear that the plasma membrane is a heterogeneous entity, containing diverse structures on nano-meso-scales (2-200 nm) with a variety of lifetimes, where certain membrane molecules stay together for limited durations. Molecular interactions occur in the time-dependent inhomogeneous two-dimensional liquid of the plasma membrane, which might be a key for plasma membrane functions.

Neuromodulation underlies the flexibility of neural circuit operation and behavior. Individual neuromodulators can have divergent actions in a neuron by targeting multiple physiological mechanisms. Conversely, multiple neuromodulators may have convergent actions through overlapping targets. The divergent and convergent neuromodulator actions can be unambiguously synergistic or antagonistic, but neuromodulation often entails balanced adjustment of nonlinear membrane and synaptic properties by targeting ion channel and synaptic dynamics rather than just excitability or synaptic strength. In addition, neuromodulators can exert effects at multiple timescales, from short-term adjustments of neuron and synapse function to persistent long-term regulation. This short review summarizes some highlights of the diverse actions of neuromodulators on ion channel and synaptic properties.

Ionotropic glutamate receptor (iGluR) subunits contain a large N-terminal domain (NTD) that precedes the agonist-binding domain (ABD) and participates in subunit oligomerization. In NMDA receptors (NMDARs), the NTDs of NR2A and NR2B subunits also form binding sites for the endogenous inhibitor Zn(2+) ion. Although these allosteric sites have been characterized in detail, the molecular mechanisms by which the NTDs communicate with the rest of the receptor to promote its inhibition remain unknown. Here, we identify the ABD dimer interface as a major structural determinant that permits coupling between the NTDs and the channel gate. The strength of this interface also controls proton inhibition, another form of allosteric modulation of NMDARs. Conformational rearrangements at the ABD dimer interface thus appear to be a key mechanism conserved in all iGluR subfamilies, but have evolved to fulfill different functions: fast desensitization at AMPA and kainate receptors, allosteric inhibition at NMDARs.

The virally encoded chemokine receptors US28 from human cytomegalovirus and ORF74 from human herpesvirus 8 are both constitutively active. We show that both receptors constitutively activate the transcription factors nuclear factor of activated T cells (NFAT) and cAMP response element binding protein (CREB) and that both pathways are modulated by their respective endogenous receptor ligands. By addition of specific pathway modulators against the G protein subunit Galphai, phospholipase C, protein kinase C, calcineurin, p38 MAP kinase, and MEK1, we find that the constitutive and ligand-dependent inductions are mediated by multiple yet similar pathways in both receptors. The NFAT and CREB transcription factors and their upstream activators are known inducers of host and virally encoded genes. We propose that the activity of these virally encoded chemokine receptors coordinates host and potentially viral gene expression similarly. As ORF74 is a known inducer of neoplasia, these findings may have important implications for cytomegalovirus-associated pathogenicity.

Secreted Wnt proteins influence neural connectivity by regulating axon guidance, dendritic morphogenesis and synapse formation. We report a new role for Wnt and Frizzled proteins in establishing the anteroposterior polarity of the mechanosensory neurons ALM and PLM in C. elegans. Disruption of Wnt signaling leads to a complete inversion of ALM and PLM polarity: the anterior process adopts the length, branching pattern and synaptic properties of the wild-type posterior process, and vice versa. Different but overlapping sets of Wnt proteins regulate neuronal polarity in different body regions. Wnts act directly on PLM via the Frizzled LIN-17. In addition, we show that they are needed for axon branching and anteriorly directed axon growth. We also find that the retromer, a conserved protein complex that mediates transcytosis and endosome-to-Golgi protein trafficking, plays a key role in Wnt signaling. Deletion mutations of retromer subunits cause ALM and PLM polarity, and other Wnt-related defects. We show that retromer protein VPS-35 is required in Wnt-expressing cells and propose that retromer activity is needed to generate a fully active Wnt signal.

A novel mammalian regulatory system was created by using the Escherichia coli lac repressor. The lac repressor was converted into a mammalian transcriptional activator by modifying the lac repressor coding region to include a nuclear localization signal from the simian virus 40 (SV40) large tumor antigen and the transcription activation domain from the herpes simplex virus type 1 virion protein 16. The lac activator protein (LAP) fusions were potent activators of several promoters containing lac operator sequences positioned either upstream or downstream of the transcription unit. A single lac operator allowed for transactivation, whereas multiple operators acted synergistically when separated by a small distance. Promoters containing 14 or 21 operator sequences were induced at least 1,000-fold in response to LAP, reaching levels of activity 20 to 30 times greater than that of the SV40 early promoter in HeLa cells. Activation was strongly inhibited by isopropyl-beta-D-thiogalactoside (IPTG), indicating that LAP retained the functions needed for allosteric regulation. LAP was bifunctional, also acting as a repressor of expression of an SV40 promoter containing an operator immediately downstream of the TATA box. Finally, genetic selection schemes were developed such that LAP-expressing cell lines can be generated at high frequency from either established or primary cells in culture.

Half the world's population is chronically infected with Helicobacter pylori, causing gastritis, gastric ulcers and an increased incidence of gastric adenocarcinoma. Its proton-gated inner-membrane urea channel, HpUreI, is essential for survival in the acidic environment of the stomach. The channel is closed at neutral pH and opens at acidic pH to allow the rapid access of urea to cytoplasmic urease. Urease produces NH(3) and CO(2), neutralizing entering protons and thus buffering the periplasm to a pH of roughly 6.1 even in gastric juice at a pH below 2.0. Here we report the structure of HpUreI, revealing six protomers assembled in a hexameric ring surrounding a central bilayer plug of ordered lipids. Each protomer encloses a channel formed by a twisted bundle of six transmembrane helices. The bundle defines a previously unobserved fold comprising a two-helix hairpin motif repeated three times around the central axis of the channel, without the inverted repeat of mammalian-type urea transporters. Both the channel and the protomer interface contain residues conserved in the AmiS/UreI superfamily, suggesting the preservation of channel architecture and oligomeric state in this superfamily. Predominantly aromatic or aliphatic side chains line the entire channel and define two consecutive constriction sites in the middle of the channel. Mutation of Trp 153 in the cytoplasmic constriction site to Ala or Phe decreases the selectivity for urea in comparison with thiourea, suggesting that solute interaction with Trp 153 contributes specificity. The previously unobserved hexameric channel structure described here provides a new model for the permeation of urea and other small amide solutes in prokaryotes and archaea.


Question: The core of the granuloma in a defense cell triggers a pro-inflammatory immune reaction.

Spatial Organization of Inflammatory Signals in Tuberculosis Granulomas Revealed. Tuberculosis (TB) is characterized by the formation of granulomas, whose functional significance and formation mechanisms are not fully elucidated. To gain insights into the role of granulomas in TB, we conducted a comprehensive analysis of the proteomes of granulomas from TB patients without bias. Employing techniques such as laser-capture microdissection, mass spectrometry, and confocal microscopy, we created detailed molecular profiles of human granulomas. Our findings indicate that the granuloma centers exhibit a pro-inflammatory milieu, marked by the presence of antimicrobial peptides, reactive oxygen species, and pro-inflammatory eicosanoids. In contrast, the periphery of the caseum displays a relatively anti-inflammatory profile. These observations are consistent across a cohort of six human subjects and in a rabbit model. Given the critical importance of the balance between pro- and anti-inflammatory signals in determining TB outcomes, our data reveal that these signals are spatially segregated within individual granulomas. Based on the proteomic and lipidomic profiles of human and rabbit lesions analyzed herein, we propose that the pathological response to TB is influenced by the precise anatomical distribution of these inflammatory pathways during granuloma development.

The presence of DNA in the cytoplasm of mammalian cells is a danger signal that triggers host immune responses such as the production of type I interferons. Cytosolic DNA induces interferons through the production of cyclic guanosine monophosphate-adenosine monophosphate (cyclic GMP-AMP, or cGAMP), which binds to and activates the adaptor protein STING. Through biochemical fractionation and quantitative mass spectrometry, we identified a cGAMP synthase (cGAS), which belongs to the nucleotidyltransferase family. Overexpression of cGAS activated the transcription factor IRF3 and induced interferon-β in a STING-dependent manner. Knockdown of cGAS inhibited IRF3 activation and interferon-β induction by DNA transfection or DNA virus infection. cGAS bound to DNA in the cytoplasm and catalyzed cGAMP synthesis. These results indicate that cGAS is a cytosolic DNA sensor that induces interferons by producing the second messenger cGAMP.

Summary Neutrophils are specialized innate cells that require constant replenishment from proliferative bone marrow (BM) precursors as a result of their short half‐life. Although it is established that neutrophils are derived from the granulocyte‐macrophage progenitor (GMP), the differentiation pathways from GMP to functional mature neutrophils are poorly defined. Using mass cytometry (CyTOF) and cell‐cycle‐based analysis, we identified three neutrophil subsets within the BM: a committed proliferative neutrophil precursor (preNeu) which differentiates into non‐proliferating immature neutrophils and mature neutrophils. Transcriptomic profiling and functional analysis revealed that preNeu require the C/EBP&egr; transcription factor for their generation from the GMP, and their proliferative program is substituted by a gain of migratory and effector function as they mature. preNeus expand under microbial and tumoral stress, and immature neutrophils are recruited to the periphery of tumor‐bearing mice. In summary, our study identifies specialized BM granulocytic populations that ensure supply under homeostasis and stress responses. Graphical Abstract Figure. No Caption available. HighlightsProliferation activity identifies committed neutrophil precursor in mice and humansNeutrophil subsets possess distinct transcriptomic and functional signaturesDefect in neutrophil development leads to impaired neutrophil‐mediated responsesIncreased circulating immature neutrophils are associated with cancer progression &NA; The neutrophil differentiation pathway is poorly defined. Evrard et. al. demonstrate a workflow of characterizing bone marrow neutrophil subsets on the basis of their proliferative capacity and molecular signatures and thereby define the developmental trajectory and functional properties of neutrophils.

The Drosophila lymph gland is a haematopoietic organ in which progenitor cells, which are most akin to the common myeloid progenitor in mammals, proliferate and differentiate into three types of mature cell--plasmatocytes, crystal cells and lamellocytes--the functions of which are reminiscent of mammalian myeloid cells. During the first and early second instars of larval development, the lymph gland contains only progenitors, whereas in the third instar, a medial region of the primary lobe of the lymph gland called the medullary zone contains these progenitors, and maturing blood cells are found juxtaposed in a peripheral region designated the cortical zone. A third group of cells referred to as the posterior signalling centre functions as a haematopoietic niche. Similarly to mammalian myeloid cells, Drosophila blood cells respond to multiple stresses including hypoxia, infection and oxidative stress. However, how systemic signals are sensed by myeloid progenitors to regulate cell-fate determination has not been well described. Here, we show that the haematopoietic progenitors of Drosophila are direct targets of systemic (insulin) and nutritional (essential amino acid) signals, and that these systemic signals maintain the progenitors by promoting Wingless (WNT in mammals) signalling. We expect that this study will promote investigation of such possible direct signal sensing mechanisms by mammalian myeloid progenitors.

Antigen targeting and adjuvancy schemes that respectively facilitate delivery of antigen to dendritic cells and elicit their activation have been explored in vaccine development. Here we investigate whether nanoparticles can be used as a vaccine platform by targeting lymph node–residing dendritic cells via interstitial flow and activating these cells by in situ complement activation. After intradermal injection, interstitial flow transported ultra-small nanoparticles (25 nm) highly efficiently into lymphatic capillaries and their draining lymph nodes, targeting half of the lymph node–residing dendritic cells, whereas 100-nm nanoparticles were only 10% as efficient. The surface chemistry of these nanoparticles activated the complement cascade, generating a danger signal in situ and potently activating dendritic cells. Using nanoparticles conjugated to the model antigen ovalbumin, we demonstrate generation of humoral and cellular immunity in mice in a size- and complement-dependent manner.

T cell memory induced by prior infection or vaccination provides enhanced protection against subsequent microbial infections. The processes involved in generating and maintaining T cell memory are becoming better understood due to recent technological advances in identifying memory T cells and monitoring their behavior and function in vivo. Memory T cells develop in response to a progressive set of cues-starting with signals from antigen-loaded, activated antigen-presenting cells (APCs) and inflammatory mediators induced by the innate immune response, to the poorly defined subsequent signals triggered as the immune response wanes toward homeostasis. The persistence of the resting memory T cells that eventually develop is regulated by cytokines. This chapter discusses recent findings on how memory T cells develop to confer long-term protective immunity.

Studying the metabolism of immune cells in recent years has emphasized the tight link existing between the metabolic state and the phenotype of these cells. Macrophages in particular are a good example of this phenomenon. Whether the macrophage obtains its energy through glycolysis or through oxidative metabolism can give rise to different phenotypes. Classically activated or M1 macrophages are key players of the first line of defense against bacterial infections and are known to obtain energy through glycolysis. Alternatively activated or M2 macrophages on the other hand are involved in tissue repair and wound healing and use oxidative metabolism to fuel their longer-term functions. Metabolic intermediates, however, are not just a source of energy but can be directly implicated in a particular macrophage phenotype. In M1 macrophages, the Krebs cycle intermediate succinate regulates HIF1α, which is responsible for driving the sustained production of the pro-inflammatory cytokine IL1β. In M2 macrophages, the sedoheptulose kinase carbohydrate kinase-like protein is critical for regulating the pentose phosphate pathway. The potential to target these events and impact on disease is an exciting prospect.

To explore the mechanism by which herpes simplex virus (HSV)-2 infection is related to HIV-1 acquisition, we conducted in situ analysis of the cellular infiltrate from sequential biopsies of HSV-2 lesions from patients on and off antiviral therapy. CD4(+) and CD8(+) T cells and a mixed population of plasmacytoid and myeloid dendritic cells (DCs), including cells expressing the C-type lectin receptor DC-SIGN, persisted at sites of HSV-2 reactivation for months after healing, even with daily antiviral therapy. The CD4(+) T cells that persisted reacted to HSV-2 antigen, were enriched for expression of the chemokine receptor CCR5, and were contiguous to DCs expressing the interleukin-3 receptor CD123 or DC-SIGN. Ex vivo infection with a CCR5-tropic strain of HIV-1 revealed greater concentrations of integrated HIV-1 DNA in cells derived from healed genital lesion biopsies than in cells from control skin biopsies. The persistence and enrichment of HIV receptor-positive inflammatory cells in the genitalia help explain the inability of anti-HSV-2 therapy to reduce HIV acquisition.

Although most cytokines are studied for biological effects after engagement of their specific cell surface membrane receptors, increasing evidence suggests that some function in the nucleus. In the present study, the precursor form of IL-1alpha was overexpressed in various cells and assessed for activity in the presence of saturating concentrations of IL-1 receptor antagonist to prevent receptor signaling. Initially diffusely present in the cytoplasm of resting cells, IL-1alpha translocated to the to nucleus after activation by endotoxin, a Toll-like receptor ligand. The IL-1alpha precursor, but not the C-terminal mature form, activated the transcriptional machinery in the GAL4 system by 90-fold; a 50-fold increase was observed using only the IL-1alpha propiece, suggesting that transcriptional activation was localized to the N terminus where the nuclear localization sequence resides. Under conditions of IL-1 receptor blockade, intracellular overexpression of the precursor and propiece forms of IL-1alpha were sufficient to activate NF-kappaB and AP-1. Stable transfectants overproducing precursor IL-1alpha released the cytokines IL-8 and IL-6 but also exhibited a significantly lower threshold of activation to subpicomolar concentrations of tumor necrosis factor alpha or IFN-gamma. Thus, intracellular functions of IL-1alpha might play an unforeseen role in the genesis of inflammation. During disease-driven events, the cytosolic precursor moves to the nucleus, where it augments transcription of proinflammatory genes. Because this mechanism of action is not affected by extracellular inhibitors, reducing intracellular functions of IL-1alpha might prove beneficial in some inflammatory conditions.

Crohn's disease and ulcerative colitis, the two main types of chronic inflammatory bowel disease, are multifactorial conditions of unknown aetiology. A susceptibility locus for Crohn's disease has been mapped to chromosome 16. Here we have used a positional-cloning strategy, based on linkage analysis followed by linkage disequilibrium mapping, to identify three independent associations for Crohn's disease: a frameshift variant and two missense variants of NOD2, encoding a member of the Apaf-1/Ced-4 superfamily of apoptosis regulators that is expressed in monocytes. These NOD2 variants alter the structure of either the leucine-rich repeat domain of the protein or the adjacent region. NOD2 activates nuclear factor NF-kB; this activating function is regulated by the carboxy-terminal leucine-rich repeat domain, which has an inhibitory role and also acts as an intracellular receptor for components of microbial pathogens. These observations suggest that the NOD2 gene product confers susceptibility to Crohn's disease by altering the recognition of these components and/or by over-activating NF-kB in monocytes, thus documenting a molecular model for the pathogenic mechanism of Crohn's disease that can now be further investigated.

CD8 tissue-resident memory T (TRM) cells provide efficient local control of viral infection, but the role of CD4 TRM is less clear. Here, by using parabiotic mice, we show that a preexisting pool of CD4 TRM cells in the genital mucosa was required for full protection from a lethal herpes simplex virus 2 (HSV-2) infection. Chemokines secreted by a local network of macrophages maintained vaginal CD4 TRM in memory lymphocyte clusters (MLCs), independently of circulating memory T cells. CD4 TRM cells within the MLCs were enriched in clones that expanded in response to HSV-2. Our results highlight the need for vaccine strategies that enable establishment of TRM cells for protection from a sexually transmitted virus and provide insights as to how such a pool might be established.


Question: The co-occurrence of H3K4me3 and H3K79me2 is observed in dormant hair follicle stem cells.

Genome-wide landscapes of histone modifications reveal in vivo chromatin dynamics in hair follicle stem cell lineage. In a comprehensive study utilizing mouse skin, a rich source of synchronized hair follicle stem cells (HF-SCs) that drive cycles of regeneration, we investigate the chromatin remodeling strategies employed by adult stem cells in response to activation signals. By conducting global mRNA and chromatin profiling in quiescent and activated HF-SCs, as well as their committed, transit-amplifying (TA) progeny, we uncover a distinct role for polycomb-group (PcG)-mediated H3K27-trimethylation in the progression of the hair follicle lineage. Specifically, in HF-SCs, PcG represses non-hair follicle lineages and differentiation, while in TA progeny, non-hair follicle regulators remain under PcG repression, HF-SC regulators gain H3K27me3 marks, and lineage regulators shed these marks. Strikingly, genes that are poised in embryonic stem cells, active in HF-SCs, and PcG-repressed in TA progeny, not only include key transcription factors but also signaling regulators, whose significance in maintaining HF-SC quiescence is highlighted. These findings underscore the efficacy of chromatin mapping in elucidating stem cell behavior and delineate the functions of PcG-mediated repression in orchestrating an irreversible fate switch, elucidating how HF-SCs navigate between quiescent and activated states without compromising their stemness.

BACKGROUND Age-associated epigenetic changes are implicated in aging. Notably, age-associated DNA methylation changes comprise a so-called aging "clock", a robust biomarker of aging. However, while genetic, dietary and drug interventions can extend lifespan, their impact on the epigenome is uncharacterised. To fill this knowledge gap, we defined age-associated DNA methylation changes at the whole-genome, single-nucleotide level in mouse liver and tested the impact of longevity-promoting interventions, specifically the Ames dwarf Prop1 df/df mutation, calorie restriction and rapamycin. RESULTS In wild-type mice fed an unsupplemented ad libitum diet, age-associated hypomethylation was enriched at super-enhancers in highly expressed genes critical for liver function. Genes harbouring hypomethylated enhancers were enriched for genes that change expression with age. Hypermethylation was enriched at CpG islands marked with bivalent activating and repressing histone modifications and resembled hypermethylation in liver cancer. Age-associated methylation changes are suppressed in Ames dwarf and calorie restricted mice and more selectively and less specifically in rapamycin treated mice. CONCLUSIONS Age-associated hypo- and hypermethylation events occur at distinct regulatory features of the genome. Distinct longevity-promoting interventions, specifically genetic, dietary and drug interventions, suppress some age-associated methylation changes, consistent with the idea that these interventions exert their beneficial effects, in part, by modulation of the epigenome. This study is a foundation to understand the epigenetic contribution to healthy aging and longevity and the molecular basis of the DNA methylation clock.

NuA4, the major H4 lysine acetyltransferase (KAT) complex in Saccharomyces cerevisiae, is recruited to promoters and stimulates transcription initiation. NuA4 subunits contain domains that bind methylated histones, suggesting that histone methylation should target NuA4 to coding sequences during transcription elongation. We show that NuA4 is cotranscriptionally recruited, dependent on its physical association with elongating polymerase II (Pol II) phosphorylated on the C-terminal domain by cyclin-dependent kinase 7/Kin28, but independently of subunits (Eaf1 and Tra1) required for NuA4 recruitment to promoters. Whereas histone methylation by Set1 and Set2 is dispensable for NuA4's interaction with Pol II and targeting to some coding regions, it stimulates NuA4-histone interaction and H4 acetylation in vivo. The NuA4 KAT, Esa1, mediates increased H4 acetylation and enhanced RSC occupancy and histone eviction in coding sequences and stimulates the rate of transcription elongation. Esa1 cooperates with the H3 KAT in SAGA, Gcn5, to enhance these functions. Our findings delineate a pathway for acetylation-mediated nucleosome remodeling and eviction in coding sequences that stimulates transcription elongation by Pol II in vivo.

Histone variants help specialize chromatin regions; however, their impact on transcriptional regulation is largely unknown. Here, we determined the genome-wide localization and dynamics of Htz1, the yeast histone H2A variant. Htz1 localizes to hundreds of repressed/basal Pol II promoters and prefers TATA-less promoters. Specific Htz1 deposition requires the SWR1 complex, which largely colocalizes with Htz1. Htz1 occupancy correlates with particular histone modifications, and Htz1 deposition is partially reliant on Gcn5 (a histone acetyltransferase) and Bdf1, an SWR1 complex member that binds acetylated histones. Changes in growth conditions cause a striking redistribution of Htz1 from activated to repressed/basal promoters. Furthermore, Htz1 promotes full gene activation but does not generally impact repression. Importantly, Htz1 releases from purified chromatin in vitro under conditions where H2A and H3 remain associated. We suggest that Htz1-bearing nucleosomes are deposited at repressed/basal promoters but facilitate activation through their susceptibility to loss, thereby helping to expose promoter DNA.

Trypanosoma brucei, the causative agent of African sleeping sickness, is transmitted to its mammalian host by the tsetse. In the fly, the parasite's surface is covered with invariant procyclin, while in the mammal it resides extracellularly in its bloodstream form (BF) and is densely covered with highly immunogenic Variant Surface Glycoprotein (VSG). In the BF, the parasite varies this highly immunogenic surface VSG using a repertoire of ~2500 distinct VSG genes. Recent reports in mammalian systems point to a role for histone acetyl-lysine recognizing bromodomain proteins in the maintenance of stem cell fate, leading us to hypothesize that bromodomain proteins may maintain the BF cell fate in trypanosomes. Using small-molecule inhibitors and genetic mutants for individual bromodomain proteins, we performed RNA-seq experiments that revealed changes in the transcriptome similar to those seen in cells differentiating from the BF to the insect stage. This was recapitulated at the protein level by the appearance of insect-stage proteins on the cell surface. Furthermore, bromodomain inhibition disrupts two major BF-specific immune evasion mechanisms that trypanosomes harness to evade mammalian host antibody responses. First, monoallelic expression of the antigenically varied VSG is disrupted. Second, rapid internalization of antibodies bound to VSG on the surface of the trypanosome is blocked. Thus, our studies reveal a role for trypanosome bromodomain proteins in maintaining bloodstream stage identity and immune evasion. Importantly, bromodomain inhibition leads to a decrease in virulence in a mouse model of infection, establishing these proteins as potential therapeutic drug targets for trypanosomiasis. Our 1.25Å resolution crystal structure of a trypanosome bromodomain in complex with I-BET151 reveals a novel binding mode of the inhibitor, which serves as a promising starting point for rational drug design.

E2F proteins can either activate or repress transcription. Following mitogenic stimulation, repressive E2F4-p130-histone deacetylase complexes dissociate from, while activating species (E2F1, -2, and -3) associate with, target promoters. Histones H3 and H4 simultaneously become hyperacetylated, but it remains unclear whether this is a prerequisite or a consequence of E2F binding. Here, we show that activating E2F species are required for hyperacetylation of target chromatin in human cells. Overexpression of a dominant-negative (DN) E2F1 mutant in serum-stimulated T98G cells blocked all E2F binding, H4 acetylation, and, albeit partially, H3 acetylation. Target gene activation and S-phase entry were also blocked by DN E2F1. Conversely, ectopic activation of E2F1 rapidly induced H3 and H4 acetylation, demonstrating a direct role for E2F in these events. E2F1 was previously shown to bind the histone acetyltransferases (HATs) p300/CBP and PCAF/GCN5. In our hands, ectopically expressed E2F1 also bound the unrelated HAT Tip60 and induced recruitment of five subunits of the Tip60 complex (Tip60, TRRAP, p400, Tip48, and Tip49) to target promoters in vivo. Moreover, E2F-dependent recruitment of Tip60 to chromatin occurred in late G(1) following serum stimulation. We speculate that the activities of multiple HAT complexes account for E2F-dependent acetylation, transcription, and S-phase entry.

Pluripotent embryonic stem cells (ESCs) maintain self-renewal and the potential for rapid response to differentiation cues. Both ESC features are subject to epigenetic regulation. Here we show that the histone acetyltransferase Mof plays an essential role in the maintenance of ESC self-renewal and pluripotency. ESCs with Mof deletion lose characteristic morphology, alkaline phosphatase (AP) staining, and differentiation potential. They also have aberrant expression of the core transcription factors Nanog, Oct4, and Sox2. Importantly, the phenotypes of Mof null ESCs can be partially suppressed by Nanog overexpression, supporting the idea that Mof functions as an upstream regulator of Nanog in ESCs. Genome-wide ChIP-sequencing and transcriptome analyses further demonstrate that Mof is an integral component of the ESC core transcriptional network and that Mof primes genes for diverse developmental programs. Mof is also required for Wdr5 recruitment and H3K4 methylation at key regulatory loci, highlighting the complexity and interconnectivity of various chromatin regulators in ESCs.

Human skin copes with harmful environmental factors that are circadian in nature, yet how circadian rhythms modulate the function of human epidermal stem cells is mostly unknown. Here we show that in human epidermal stem cells and their differentiated counterparts, core clock genes peak in a successive and phased manner, establishing distinct temporal intervals during the 24 hr day period. Each of these successive clock waves is associated with a peak in the expression of subsets of transcripts that temporally segregate the predisposition of epidermal stem cells to respond to cues that regulate their proliferation or differentiation, such as TGFβ and calcium. Accordingly, circadian arrhythmia profoundly affects stem cell function in culture and in vivo. We hypothesize that this intricate mechanism ensures homeostasis by providing epidermal stem cells with environmentally relevant temporal functional cues during the course of the day and that its perturbation may contribute to aging and carcinogenesis.

Epigenetic processes play a key role in orchestrating transcriptional regulation during development. The importance of DNA methylation in fetal brain development is highlighted by the dynamic expression of de novo DNA methyltransferases during the perinatal period and neurodevelopmental deficits associated with mutations in the methyl-CpG binding protein 2 (MECP2) gene. However, our knowledge about the temporal changes to the epigenome during fetal brain development has, to date, been limited. We quantified genome-wide patterns of DNA methylation at ∼ 400,000 sites in 179 human fetal brain samples (100 male, 79 female) spanning 23 to 184 d post-conception. We identified highly significant changes in DNA methylation across fetal brain development at >7% of sites, with an enrichment of loci becoming hypomethylated with fetal age. Sites associated with developmental changes in DNA methylation during fetal brain development were significantly underrepresented in promoter regulatory regions but significantly overrepresented in regions flanking CpG islands (shores and shelves) and gene bodies. Highly significant differences in DNA methylation were observed between males and females at a number of autosomal sites, with a small number of regions showing sex-specific DNA methylation trajectories across brain development. Weighted gene comethylation network analysis (WGCNA) revealed discrete modules of comethylated loci associated with fetal age that are significantly enriched for genes involved in neurodevelopmental processes. This is, to our knowledge, the most extensive study of DNA methylation across human fetal brain development to date, confirming the prenatal period as a time of considerable epigenomic plasticity.

PURPOSE Acute myeloid leukemia (AML) is a heterogeneous disease with poor outcomes. Despite increased evidence shows that dysregulation of histone modification contributes to AML, specific drugs targeting key histone modulators are not applied in the clinical treatment of AML. Here, we investigated whether targeting KDM6B, the demethylase of tri-methylated histone H3 lysine 27 (H3K27me3), has a therapeutic potential for AML. METHODS A KDM6B-specific inhibitor, GSK-J4, was applied to treat the primary cells from AML patients and AML cell lines in vitro and in vivo. RNA-sequencing was performed to reveal the underlying mechanisms of inhibiting KDM6B for the treatment of AML. RESULTS Here we observed that the mRNA expression of KDM6B was up-regulated in AML and positively correlated with poor survival. Treatment with GSK-J4 increased the global level of H3K27me3 and reduced the proliferation and colony-forming ability of primary AML cells and AML cell lines. GSK-J4 treatment significantly induced cell apoptosis and cell-cycle arrest in Kasumi-1 cells, and displayed a synergistic effect with cytosine arabinoside. Notably, injection of GSK-J4 attenuated the disease progression in a human AML xenograft mouse model in vivo. Treatment with GSK-J4 predominantly resulted in down-regulation of DNA replication and cell-cycle-related pathways, as well as abrogated the expression of critical cancer-promoting HOX genes. ChIP-qPCR validated an increased enrichment of H3K27me3 in the transcription start sites of these HOX genes. CONCLUSIONS In summary, our findings suggest that targeting KDM6B with GSK-J4 has a therapeutic potential for the treatment of AML.

Imprinting is an epigenetic modification leading to monoallelic expression of some genes, and disrupted imprinting is believed to be a barrier to human stem cell transplantation, based on studies that suggest that epigenetic marks are unstable in mouse embryonic germ (EG) and embryonic stem (ES) cells. However, stem cell imprinting has not previously been examined directly in humans. We found that three imprinted genes, TSSC5, H19, and SNRPN, show monoallelic expression in in vitro differentiated human EG-derived cells, and a fourth gene, IGF2, shows partially relaxed imprinting at a ratio from 4:1 to 5:1, comparable to that found in normal somatic cells. In addition, we found normal methylation of an imprinting control region (ICR) that regulates H19 and IGF2 imprinting, suggesting that imprinting may not be a significant epigenetic barrier to human EG cell transplantation. Finally, we were able to construct an in vitro mouse model of genomic imprinting, by generating EG cells from 8.5-day embryos of an interspecific cross, in which undifferentiated cells show biallelic expression and acquire preferential parental allele expression after differentiation. This model should allow experimental manipulation of epigenetic modifications of cultured EG cells that may not be possible in human stem cell studies.