Question: Urbanization is an important risk factor related to the transmission of dengue fever.

Spatial and Temporal Clustering of Dengue Virus Transmission in Thai Villages. 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.

Mutation spectrum in the nephrin gene (NPHS1) in congenital nephrotic syndrome.. Congenital nephrotic syndrome, Finnish type (CNF or NPHS1), is an autosomal recessive disease characterized by massive proteinuria and development of nephrotic syndrome shortly after birth. The disease is most common in Finland, but many patients have been identified in other populations. The disease is caused by mutations in the gene for nephrin which is a key component of the glomerual ultrafilter, the podocyte slit diaphragm. A total of 30 mutations have been reported in the nephrin gene in patients with congenital nephrotic syndrome worldwide. In the Finnish population, two main mutations have been found. These two nonsense mutations account for over 94% of all mutations in Finland. Most mutations found in non-Finnish patients are missense mutations, but they include also nonsense and splice site mutations, as well as deletions and insertions. This mutation update summarizes the nature of all previously reported nephrin mutations and, additionally, describes 20 novel mutations recently identified in our laboratory.


Question: Primary cervical cytology screening with HPV detection has higher longitudinal sensitivity to detect severe cervical intraepithelial neoplasia than conventional cytology.

Efficacy of human papillomavirus testing for the detection of invasive cervical cancers and cervical intraepithelial neoplasia: a randomised controlled trial.. BACKGROUND Human papillomavirus (HPV) testing is known to be more sensitive, but less specific than cytology for detecting cervical intraepithelial neoplasia (CIN). We assessed the efficacy of cervical-cancer screening policies that are based on HPV testing. METHODS Between March, 2004, and December, 2004, in two separate recruitment phases, women aged 25-60 years were randomly assigned to conventional cytology or to HPV testing in combination with liquid-based cytology (first phase) or alone (second phase). Randomisation was done by computer in two screening centres and by sequential opening of numbered sealed envelopes in the remaining seven centres. During phase one, women who were HPV-positive and aged 35-60 years were referred to colposcopy, whereas women aged 25-34 years were referred to colposcopy only if cytology was also abnormal or HPV testing was persistently positive. During phase two, women in the HPV group were referred for colposcopy if the HPV test was positive. Two rounds of screening occurred in each phase, and all women had cytology testing only at the second round. The primary endpoint was the detection of grade 2 and 3 CIN, and of invasive cervical cancers during the first and second screening rounds. Analysis was done by intention to screen. This trial is registered, number ISRCTN81678807. FINDINGS In total for both phases, 47,001 women were randomly assigned to the cytology group and 47,369 to HPV testing. 33,851 women from the cytology group and 32,998 from the HPV-testing group had a second round of screening. We also retrieved the histological diagnoses from screening done elsewhere. The detection of invasive cervical cancers was similar for the two groups in the first round of screening (nine in the cytology group vs seven in the HPV group, p=0.62); no cases were detected in the HPV group during round two, compared with nine in the cytology group (p=0.004). Overall, in the two rounds of screening, 18 invasive cancers were detected in the cytology group versus seven in the HPV group (p=0.028). Among women aged 35-60 years, at round one the relative detection (HPV vs cytology) was 2.00 (95% CI 1.44-2.77) for CIN2, 2.08 (1.47-2.95) for CIN3, and 2.03 (1.60-2.57) for CIN2 and 3 together. At round two the relative detection was 0.54 (0.23-1.28) for CIN2, 0.48 (0.21-1.11) for CIN3, and 0.51 (0.28-0.93) for CIN2 and 3 together. Among women aged 25-34 years, there was significant heterogeneity between phases in the relative detection of CIN3. At round one the relative detection was 0.93 (0.52-1.64) in phase one and 3.91 (2.02-7.57) in phase two. At round two the relative detection was 1.34 (0.46-3.84) in phase one and 0.20 (0.04-0.93) in phase two. Pooling both phases, the detection ratio of CIN2 for women aged 25-34 years was 4.09 (2.24-7.48) at round one and 0.64 (0.23-1.27) at round two. INTERPRETATION HPV-based screening is more effective than cytology in preventing invasive cervical cancer, by detecting persistent high-grade lesions earlier and providing a longer low-risk period. However, in younger women, HPV screening leads to over-diagnosis of regressive CIN2. FUNDING European Union, Italian Ministry of Health, Regional Health Administrations of Piemonte, Tuscany, Veneto and Emilia-Romagna, and Public Health Agency of Lazio.

Bed sharing when parents do not smoke: is there a risk of SIDS? An individual level analysis of five major case–control studies. OBJECTIVE To resolve uncertainty as to the risk of Sudden Infant Death Syndrome (SIDS) associated with sleeping in bed with your baby if neither parent smokes and the baby is breastfed. DESIGN Bed sharing was defined as sleeping with a baby in the parents' bed; room sharing as baby sleeping in the parents' room. Frequency of bed sharing during last sleep was compared between babies who died of SIDS and living control infants. Five large SIDS case-control datasets were combined. Missing data were imputed. Random effects logistic regression controlled for confounding factors. SETTING Home sleeping arrangements of infants in 19 studies across the UK, Europe and Australasia. PARTICIPANTS 1472 SIDS cases, and 4679 controls. Each study effectively included all cases, by standard criteria. Controls were randomly selected normal infants of similar age, time and place. RESULTS In the combined dataset, 22.2% of cases and 9.6% of controls were bed sharing, adjusted OR (AOR) for all ages 2.7; 95% CI (1.4 to 5.3). Bed sharing risk decreased with increasing infant age. When neither parent smoked, and the baby was less than 3 months, breastfed and had no other risk factors, the AOR for bed sharing versus room sharing was 5.1 (2.3 to 11.4) and estimated absolute risk for these room sharing infants was very low (0.08 (0.05 to 0.14)/1000 live-births). This increased to 0.23 (0.11 to 0.43)/1000 when bed sharing. Smoking and alcohol use greatly increased bed sharing risk. CONCLUSIONS Bed sharing for sleep when the parents do not smoke or take alcohol or drugs increases the risk of SIDS. Risks associated with bed sharing are greatly increased when combined with parental smoking, maternal alcohol consumption and/or drug use. A substantial reduction of SIDS rates could be achieved if parents avoided bed sharing.


Question: Sympathetic nerve activity is elevated throughout normal pregnancy.

Regulation of sympathetic nerve activity during the cold pressor test in normotensive pregnant and nonpregnant women.. Baseline neurovascular transduction is reduced in normotensive pregnancy; however, little is known about changes to neurovascular transduction during periods of heightened sympathetic activation. We tested the hypothesis that, despite an exacerbated muscle sympathetic nerve activity (microneurography) response to cold pressor stimulation, the blunting of neurovascular transduction in normotensive pregnant women would result in similar changes in vascular resistance and mean arterial pressure (Finometer) relative to nonpregnant controls. Baseline neurovascular transduction was reduced in pregnant women relative to controls when expressed as the quotient of both total resistance and mean arterial pressure and sympathetic burst frequency (0.32±0.07 versus 0.58±0.16 mm Hg/L/min/bursts/min, P<0.001 and 2.4±0.7 versus 3.6±0.8 mm Hg/bursts/min, P=0.001). Sympathetic activation was greater across all 3 minutes of cold pressor stimulation in the pregnant women relative to the nonpregnant controls. Peak sympathoexcitation was also greater in pregnant than in nonpregnant women, whether expressed as sympathetic burst frequency (+17±13 versus +7±8 bursts/min, P=0.049), burst incidence (+17±9 versus +6±11 bursts/100 hb, P=0.03), or total activity (+950±660 versus +363±414 arbitrary units, P=0.04). However, neurovascular transduction during peak cold pressor-induced sympathoexcitation remained blunted in pregnant women (0.25±0.11 versus 0.45±0.08 mm Hg/L/min/bursts/min, P<0.001 and 1.9±1.0 versus 3.2±0.9 mm Hg/bursts/min, P=0.006). Therefore, mean arterial pressure (93±21 versus 99±6 mm Hg, P=0.4) and total peripheral resistance (12±3 versus 14±3 mm Hg/L/min) were not different between pregnant and nonpregnant women during peak sympathoexcitation. These data indicate that the third trimester of normotensive pregnancy is associated with reductions in neurovascular transduction, which result in the dissociation of sympathetic outflow from hemodynamic outcomes, even during cold pressor-induced sympathoexcitation.

Metformin-associated lactic acidosis: a rare or very rare clinical entity?. AIMS Lactic acidosis is a well recognized complication of biguanide therapy which is potentially serious. Although the prevalence of metformin-associated lactic acidosis (MALA) is much lower than that associated with phenformin, it is still being reported sporadically which raises concerns for the practising clinicians. We review the currently available world-wide data of the prevalence of MALA, the risk factors for its development and the current practical guidelines on the use of metformin to minimize the risk of this potential hazard. METHODS An extensive literature search was conducted from both Medline and Ovid (1965-98) using the following keywords: 'Type 2 diabetes mellitus', 'oral hypoglycaemic drugs', 'biguanides', 'metformin-associated lactic acidosis' and 'renal impairment'. RESULTS MALA was found to be a very rare clinical entity, being 20 times less common than phenformin-associated lactic acidosis. Amongst all the risk factors, renal impairment appears to be the major precipitating factor for the development of MALA in metformin-treated patients. We also found cases of MALA where no precipitating factors were identified and the underlying mechanism in these cases remains unclear. Practical recommendations of metformin use to minimize the risk of MALA have been listed based on previous reports. CONCLUSIONS The low prevalence of MALA is comparable to the prevalence of sulphonylurea-induced hypoglycaemia. Metformin has many beneficial metabolic effects in the management of Type 2 diabetes mellitus. Provided that the recommended guidelines for metformin use are strictly adhered to, its widespread use would be safe and the incidence of MALA will be further reduced.


Question: The DESMOND program demonstrates no significant impact on biochemical outcomes.

Effectiveness of a diabetes education and self management programme (DESMOND) for people with newly diagnosed type 2 diabetes mellitus: three year follow-up of a cluster randomised controlled trial in primary care. OBJECTIVE To measure whether the benefits of a single education and self management structured programme for people with newly diagnosed type 2 diabetes mellitus are sustained at three years. DESIGN Three year follow-up of a multicentre cluster randomised controlled trial in primary care, with randomisation at practice level. SETTING 207 general practices in 13 primary care sites in the United Kingdom. PARTICIPANTS 731 of the 824 participants included in the original trial were eligible for follow-up. Biomedical data were collected on 604 (82.6%) and questionnaire data on 513 (70.1%) participants. INTERVENTION A structured group education programme for six hours delivered in the community by two trained healthcare professional educators compared with usual care. MAIN OUTCOME MEASURES The primary outcome was glycated haemoglobin (HbA(1c)) levels. The secondary outcomes were blood pressure, weight, blood lipid levels, smoking status, physical activity, quality of life, beliefs about illness, depression, emotional impact of diabetes, and drug use at three years. RESULTS HbA(1c) levels at three years had decreased in both groups. After adjusting for baseline and cluster the difference was not significant (difference -0.02, 95% confidence interval -0.22 to 0.17). The groups did not differ for the other biomedical and lifestyle outcomes and drug use. The significant benefits in the intervention group across four out of five health beliefs seen at 12 months were sustained at three years (P<0.01). Depression scores and quality of life did not differ at three years. CONCLUSION A single programme for people with newly diagnosed type 2 diabetes mellitus showed no difference in biomedical or lifestyle outcomes at three years although there were sustained improvements in some illness beliefs. TRIAL REGISTRATION Current Controlled Trials ISRCTN17844016.

Transglutaminase 2 Undergoes a Large Conformational Change upon Activation . Human transglutaminase 2 (TG2), a member of a large family of enzymes that catalyze protein crosslinking, plays an important role in the extracellular matrix biology of many tissues and is implicated in the gluten-induced pathogenesis of celiac sprue. Although vertebrate transglutaminases have been studied extensively, thus far all structurally characterized members of this family have been crystallized in conformations with inaccessible active sites. We have trapped human TG2 in complex with an inhibitor that mimics inflammatory gluten peptide substrates and have solved, at 2-A resolution, its x-ray crystal structure. The inhibitor stabilizes TG2 in an extended conformation that is dramatically different from earlier transglutaminase structures. The active site is exposed, revealing that catalysis takes place in a tunnel, bridged by two tryptophan residues that separate acyl-donor from acyl-acceptor and stabilize the tetrahedral reaction intermediates. Site-directed mutagenesis was used to investigate the acyl-acceptor side of the tunnel, yielding mutants with a marked increase in preference for hydrolysis over transamidation. By providing the ability to visualize this activated conformer, our results create a foundation for understanding the catalytic as well as the non-catalytic roles of TG2 in biology, and for dissecting the process by which the autoantibody response to TG2 is induced in celiac sprue patients.


Question: Human embryonic stem cells give rise to cell types from the outer embryonic germ layer, but not the other two layers.

Establishment and in vitro differentiation of a new embryonic stem cell line from human blastocyst.. Embryonic stem cells have the ability to remain undifferentiated and proliferate indefinitely in vitro while maintaining the potential to differentiate into derivatives of all three embryonic germ layers. These cells have, therefore, potential for in vitro differentiation studies, gene function, and so on. The aim of this study was to produce a human embryonic stem cell line. An inner cell mass of a human blastocyst was separated and cultured on mouse embryonic fibroblasts in embryonic stem cell medium with related additives. The established line was evaluated by morphology; passaging; freezing and thawing; alkaline phosphatase; Oct-4 expression; anti-surface markers including Tra-1-60 and Tra-1-81; and karyotype and spontaneous differentiation. Differentiated cardiomyocytes and neurons were evaluated by transmission electron microscopy and immunocytochemistry. Here, we report the derivation of a new embryonic stem cell line (Royan H1) from a human blastocyst that remains undifferentiated in morphology during continuous passaging for more than 30 passages, maintains a normal XX karyotype, is viable after freezing and thawing, and expresses alkaline phosphatase, Oct-4, Tra-1-60, and Tra-1-81. These cells remain undifferentiated when grown on mouse embryonic fibroblast feeder layers in the presence or absence of recombinant human leukemia inhibitory factor. Royan H1 cells can differentiate in vitro in the absence of feeder cells and can produce embryoid bodies that can further differentiate into beating cardiomyocytes as well as neurons. These results define Royan H1 cells as a new human embryonic stem cell line.

Metabolic shutdown in Escherichia coli cells lacking the outer membrane channel TolC.. The outer membrane channel TolC is a key component of multidrug efflux and type I secretion transporters in Escherichia coli. Mutational inactivation of TolC renders cells highly susceptible to antibiotics and leads to defects in secretion of protein toxins. Despite impairment of various transport functions, no growth defects were reported in cells lacking TolC. Unexpectedly, we found that the loss of TolC notably impairs cell division and growth in minimal glucose medium. The TolC-dependent phenotype was further exacerbated by the loss of ygiB and ygiC genes expressed in the same operon as tolC and their homologues yjfM and yjfC located elsewhere on the chromosome. Our results show that this growth deficiency is caused by depletion of the critical metabolite NAD(+) and high NADH/NAD(+) ratios. The increased amounts of PspA and decreased rates of NADH oxidation in Delta tolC membranes indicated stress on the membrane and dissipation of a proton motive force. We conclude that inactivation of TolC triggers metabolic shutdown in E. coli cells grown in minimal glucose medium. The Delta tolC phenotype is partially rescued by YgiBC and YjfMC, which have parallel functions independent from TolC.


Question: Active Ly49Q prevents neutrophil polarization.

The Ly49Q receptor plays a crucial role in neutrophil polarization and migration by regulating raft trafficking.. Neutrophils rapidly undergo polarization and directional movement to infiltrate the sites of infection and inflammation. Here, we show that an inhibitory MHC I receptor, Ly49Q, was crucial for the swift polarization of and tissue infiltration by neutrophils. During the steady state, Ly49Q inhibited neutrophil adhesion by preventing focal-complex formation, likely by inhibiting Src and PI3 kinases. However, in the presence of inflammatory stimuli, Ly49Q mediated rapid neutrophil polarization and tissue infiltration in an ITIM-domain-dependent manner. These opposite functions appeared to be mediated by distinct use of effector phosphatase SHP-1 and SHP-2. Ly49Q-dependent polarization and migration were affected by Ly49Q regulation of membrane raft functions. We propose that Ly49Q is pivotal in switching neutrophils to their polarized morphology and rapid migration upon inflammation, through its spatiotemporal regulation of membrane rafts and raft-associated signaling molecules.

Quantifying heterogeneity in a meta-analysis.. The extent of heterogeneity in a meta-analysis partly determines the difficulty in drawing overall conclusions. This extent may be measured by estimating a between-study variance, but interpretation is then specific to a particular treatment effect metric. A test for the existence of heterogeneity exists, but depends on the number of studies in the meta-analysis. We develop measures of the impact of heterogeneity on a meta-analysis, from mathematical criteria, that are independent of the number of studies and the treatment effect metric. We derive and propose three suitable statistics: H is the square root of the chi2 heterogeneity statistic divided by its degrees of freedom; R is the ratio of the standard error of the underlying mean from a random effects meta-analysis to the standard error of a fixed effect meta-analytic estimate, and I2 is a transformation of (H) that describes the proportion of total variation in study estimates that is due to heterogeneity. We discuss interpretation, interval estimates and other properties of these measures and examine them in five example data sets showing different amounts of heterogeneity. We conclude that H and I2, which can usually be calculated for published meta-analyses, are particularly useful summaries of the impact of heterogeneity. One or both should be presented in published meta-analyses in preference to the test for heterogeneity.


Question: Suboptimal nutrition is a leading cause of premature death

Global, regional, and national comparative risk assessment of 79 behavioural, environmental and occupational, and metabolic risks or clusters of risks, 1990–2015: a systematic analysis for the Global Burden of Disease Study 2015. 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.

Crystal structure of the first plant urease from jack bean: 83 years of journey from its first crystal to molecular structure.. Urease, a nickel-dependent metalloenzyme, is synthesized by plants, some bacteria, and fungi. It catalyzes the hydrolysis of urea into ammonia and carbon dioxide. Although the amino acid sequences of plant and bacterial ureases are closely related, some biological activities differ significantly. Plant ureases but not bacterial ureases possess insecticidal properties independent of its ureolytic activity. To date, the structural information is available only for bacterial ureases although the jack bean urease (Canavalia ensiformis; JBU), the best-studied plant urease, was the first enzyme to be crystallized in 1926. To better understand the biological properties of plant ureases including the mechanism of insecticidal activity, we initiated the structural studies on some of them. Here, we report the crystal structure of JBU, the first plant urease structure, at 2.05 A resolution. The active-site architecture of JBU is similar to that of bacterial ureases containing a bi-nickel center. JBU has a bound phosphate and covalently modified residue (Cys592) by beta-mercaptoethanol at its active site, and the concomitant binding of multiple inhibitors (phosphate and beta-mercaptoethanol) is not observed so far in bacterial ureases. By correlating the structural information of JBU with the available biophysical and biochemical data on insecticidal properties of plant ureases, we hypothesize that the amphipathic beta-hairpin located in the entomotoxic peptide region of plant ureases might form a membrane insertion beta-barrel as found in beta-pore-forming toxins.


Question: At least 85% of patients exposed to radiation have activated markers of myofibroblasts.

KLF4-dependent phenotypic modulation of smooth muscle cells has a key role in atherosclerotic plaque pathogenesis. Previous studies investigating the role of smooth muscle cells (SMCs) and macrophages in the pathogenesis of atherosclerosis have provided controversial results owing to the use of unreliable methods for clearly identifying each of these cell types. Here, using Myh11-CreERT2 ROSA floxed STOP eYFP Apoe−/− mice to perform SMC lineage tracing, we find that traditional methods for detecting SMCs based on immunostaining for SMC markers fail to detect >80% of SMC-derived cells within advanced atherosclerotic lesions. These unidentified SMC-derived cells exhibit phenotypes of other cell lineages, including macrophages and mesenchymal stem cells (MSCs). SMC-specific conditional knockout of Krüppel-like factor 4 (Klf4) resulted in reduced numbers of SMC-derived MSC- and macrophage-like cells, a marked reduction in lesion size, and increases in multiple indices of plaque stability, including an increase in fibrous cap thickness as compared to wild-type controls. On the basis of in vivo KLF4 chromatin immunoprecipitation–sequencing (ChIP-seq) analyses and studies of cholesterol-treated cultured SMCs, we identified >800 KLF4 target genes, including many that regulate pro-inflammatory responses of SMCs. Our findings indicate that the contribution of SMCs to atherosclerotic plaques has been greatly underestimated, and that KLF4-dependent transitions in SMC phenotype are critical in lesion pathogenesis.

p62/SQSTM1 by Binding to Vitamin D Receptor Inhibits Hepatic Stellate Cell Activity, Fibrosis, and Liver Cancer.. Hepatic stellate cells (HSCs) play critical roles in liver fibrosis and hepatocellular carcinoma (HCC). Vitamin D receptor (VDR) activation in HSCs inhibits liver inflammation and fibrosis. We found that p62/SQSTM1, a protein upregulated in liver parenchymal cells but downregulated in HCC-associated HSCs, negatively controls HSC activation. Total body or HSC-specific p62 ablation potentiates HSCs and enhances inflammation, fibrosis, and HCC progression. p62 directly interacts with VDR and RXR promoting their heterodimerization, which is critical for VDR:RXR target gene recruitment. Loss of p62 in HSCs impairs the repression of fibrosis and inflammation by VDR agonists. This demonstrates that p62 is a negative regulator of liver inflammation and fibrosis through its ability to promote VDR signaling in HSCs, whose activation supports HCC.


Question: PKG-la plays an essential role in expression of spinal long term potentiation in PGK-la knockout mice.

Presynaptically Localized Cyclic GMP-Dependent Protein Kinase 1 Is a Key Determinant of Spinal Synaptic Potentiation and Pain Hypersensitivity. Synaptic long-term potentiation (LTP) at spinal neurons directly communicating pain-specific inputs from the periphery to the brain has been proposed to serve as a trigger for pain hypersensitivity in pathological states. Previous studies have functionally implicated the NMDA receptor-NO pathway and the downstream second messenger, cGMP, in these processes. Because cGMP can broadly influence diverse ion-channels, kinases, and phosphodiesterases, pre- as well as post-synaptically, the precise identity of cGMP targets mediating spinal LTP, their mechanisms of action, and their locus in the spinal circuitry are still unclear. Here, we found that Protein Kinase G1 (PKG-I) localized presynaptically in nociceptor terminals plays an essential role in the expression of spinal LTP. Using the Cre-lox P system, we generated nociceptor-specific knockout mice lacking PKG-I specifically in presynaptic terminals of nociceptors in the spinal cord, but not in post-synaptic neurons or elsewhere (SNS-PKG-I(-/-) mice). Patch clamp recordings showed that activity-induced LTP at identified synapses between nociceptors and spinal neurons projecting to the periaqueductal grey (PAG) was completely abolished in SNS-PKG-I(-/-) mice, although basal synaptic transmission was not affected. Analyses of synaptic failure rates and paired-pulse ratios indicated a role for presynaptic PKG-I in regulating the probability of neurotransmitter release. Inositol 1,4,5-triphosphate receptor 1 and myosin light chain kinase were recruited as key phosphorylation targets of presynaptic PKG-I in nociceptive neurons. Finally, behavioural analyses in vivo showed marked defects in SNS-PKG-I(-/-) mice in several models of activity-induced nociceptive hypersensitivity, and pharmacological studies identified a clear contribution of PKG-I expressed in spinal terminals of nociceptors. Our results thus indicate that presynaptic mechanisms involving an increase in release probability from nociceptors are operational in the expression of synaptic LTP on spinal-PAG projection neurons and that PKG-I localized in presynaptic nociceptor terminals plays an essential role in this process to regulate pain sensitivity.

Krüppel-like factor 2 (KLF2) regulates B-cell reactivity, subset differentiation, and trafficking molecule expression.. The transcription factor Krüppel-like factor 2 (KLF2) is critical for normal trafficking of T lymphocytes, but its role in B cells is unclear. We report that B cell-specific KLF2 deficiency leads to decreased expression of the trafficking molecules CD62L and β7-integrin, yet expression of sphingosine-1 phosphate receptor 1 (which is a critical target of KLF2 in T cells) was, unexpectedly, minimally altered. Unexpectedly, Klf2 deletion led to a drastic reduction in the B1 B-cell pool and a substantial increase in transitional and marginal zone B-cell numbers. In addition, we observed that KLF2-deficient B cells showed increased apoptosis and impaired proliferation after B-cell receptor cross-linking. Gene expression analysis indicated that KLF2-deficient follicular B cells display numerous characteristics shared by normal marginal zone B cells, including reduced expression of several signaling molecules that may contribute to defective activation of these cells. Hence, our data indicate that KLF2 plays a critical role in dictating normal subset differentiation and functional reactivity of mature B cells.


Question: 1-1% of colorectal cancer patients are diagnosed with regional or distant metastases.

Relation between Medicare screening reimbursement and stage at diagnosis for older patients with colon cancer.. CONTEXT Medicare's reimbursement policy was changed in 1998 to provide coverage for screening colonoscopies for patients with increased colon cancer risk, and expanded further in 2001 to cover screening colonoscopies for all individuals. OBJECTIVE To determine whether the Medicare reimbursement policy changes were associated with an increase in either colonoscopy use or early stage colon cancer diagnosis. DESIGN, SETTING, AND PARTICIPANTS Patients in the Surveillance, Epidemiology, and End Results Medicare linked database who were 67 years of age and older and had a primary diagnosis of colon cancer during 1992-2002, as well as a group of Medicare beneficiaries who resided in Surveillance, Epidemiology, and End Results areas but who were not diagnosed with cancer. MAIN OUTCOME MEASURES Trends in colonoscopy and sigmoidoscopy use among Medicare beneficiaries without cancer were assessed using multivariate Poisson regression. Among the patients with cancer, stage was classified as early (stage I) vs all other (stages II-IV). Time was categorized as period 1 (no screening coverage, 1992-1997), period 2 (limited coverage, January 1998-June 2001), and period 3 (universal coverage, July 2001-December 2002). A multivariate logistic regression (outcome = early stage) was used to assess temporal trends in stage at diagnosis; an interaction term between tumor site and time was included. RESULTS Colonoscopy use increased from an average rate of 285/100,000 per quarter in period 1 to 889 and 1919/100,000 per quarter in periods 2 (P<.001) and 3 (P vs 2<.001), respectively. During the study period, 44,924 eligible patients were diagnosed with colorectal cancer. The proportion of patients diagnosed at an early stage increased from 22.5% in period 1 to 25.5% in period 2 and 26.3% in period 3 (P<.001 for each pairwise comparison). The changes in Medicare coverage were strongly associated with early stage at diagnosis for patients with proximal colon lesions (adjusted relative risk period 2 vs 1, 1.19; 95% confidence interval, 1.13-1.26; adjusted relative risk period 3 vs 2, 1.10; 95% confidence interval, 1.02-1.17) but weakly associated, if at all, for patients with distal colon lesions (adjusted relative risk period 2 vs 1, 1.07; 95% confidence interval, 1.01-1.13; adjusted relative risk period 3 vs 2, 0.97; 95% confidence interval, 0.90-1.05). CONCLUSIONS Expansion of Medicare reimbursement to cover colon cancer screening was associated with an increased use of colonoscopy for Medicare beneficiaries, and for those who were diagnosed with colon cancer, an increased probability of being diagnosed at an early stage. The selective effect of the coverage change on proximal colon lesions suggests that increased use of whole-colon screening modalities such as colonoscopy may have played a pivotal role.

Association Between ADHD and Obesity: A Systematic Review and Meta-Analysis.. OBJECTIVE Impulsivity and inattention related to attention deficit hyperactivity disorder (ADHD) may increase food intake and, consequently, weight gain. However, findings on the association between obesity/overweight and ADHD are mixed. The authors conducted a meta-analysis to estimate this association. METHOD A broad range of databases was searched through Aug. 31, 2014. Unpublished studies were also obtained. Study quality was rated with the Newcastle-Ottawa Scale. Random-effects models were used. RESULTS Forty-two studies that included a total of 728,136 individuals (48,161 ADHD subjects; 679,975 comparison subjects) were retained. A significant association between obesity and ADHD was found for both children (odds ratio=1.20, 95% CI=1.05-1.37) and adults (odds ratio=1.55, 95% CI=1.32-1.81). The pooled prevalence of obesity was increased by about 70% in adults with ADHD (28.2%, 95% CI=22.8-34.4) compared with those without ADHD (16.4%, 95% CI=13.4-19.9), and by about 40% in children with ADHD (10.3%, 95% CI=7.9-13.3) compared with those without ADHD (7.4%, 95% CI=5.4-10.1). The significant association between ADHD and obesity remained when limited to studies 1) reporting odds ratios adjusted for possible confounding factors; 2) diagnosing ADHD by direct interview; and 3) using directly measured height and weight. Gender, study setting, study country, and study quality did not moderate the association between obesity and ADHD. ADHD was also significantly associated with overweight. Individuals medicated for ADHD were not at higher risk of obesity. CONCLUSIONS This study provides meta-analytic evidence for a significant association between ADHD and obesity/overweight. Further research should address possible underlying mechanisms and the long-term effects of ADHD treatments on weight in individuals with both ADHD and obesity.


Question: Mice that lack Interferon-γ or its receptor are highly susceptible to experimental autoimmune myocarditis.

Lethal autoimmune myocarditis in interferon-gamma receptor-deficient mice: enhanced disease severity by impaired inducible nitric oxide synthase induction.. BACKGROUND Interferon-gamma (IFN-gamma) is an essential cytokine in the regulation of inflammatory responses in autoimmune diseases. Little is known about its role in inflammatory heart disease. METHODS AND RESULTS We showed that IFN-gamma receptor-deficient mice (IFN-gammaR(-/-)) on a BALB/c background immunized with a peptide derived from cardiac alpha-myosin heavy chain develop severe myocarditis with high mortality. Although myocarditis subsided in wild-type mice after 3 weeks, IFN-gammaR(-/-) mice showed persistent disease. The persistent inflammation was accompanied by vigorous in vitro CD4 T-cell responses and impaired inducible nitric oxide synthase expression, together with evidence of impaired nitric oxide production in IFN-gammaR(-/-) hearts. Treatment of wild-type mice with the nitric oxide synthetase inhibitor N:-nitro-l-arginine-methyl-ester enhanced in vitro CD4 T-cell proliferation and prevented healing of myocarditis. CONCLUSIONS Our data provide evidence that IFN-gamma protects mice from lethal autoimmune myocarditis by inducing the expression of inducible nitric oxide synthase followed by the downregulation of T-cell responses.

DNA Methylation in Major Depressive Disorder.. Epigenetic mechanisms regulate gene expression, influencing protein levels and ultimately shaping phenotypes during life. However, both stochastic epigenetic variations and environmental reprogramming of the epigenome might influence neurodevelopment and ageing, and this may contribute to the origins of mental ill-health. Studying the role of epigenetic mechanisms is challenging, as genotype-, tissue- and cell type-dependent epigenetic changes have to be taken into account, while the nature of mental disorders also poses significant challenges for linking them with biological profiles. In this chapter, we summarise the current evidence suggesting the role of DNA methylation as a key epigenetic mechanism in major depressive disorder.


Question: Citrullinated proteins externalized in neutrophil extracellular traps act indirectly to disrupt the inflammatory cycle.

Neutrophil extracellular chromatin traps connect innate immune response to autoimmunity. Autoantibodies to DNA and histones (chromatin) are the defining antigen specificity in systemic lupus erythematosus (SLE) and related musculoskeletal disorders but the mechanisms responsible for their induction remain mysterious. That situation rapidly changed once neutrophil extracellular chromatin traps (NETs) were discovered and observed to play a conserved role in innate immune responses to a broad variety of microbial pathogens. At the center of an infectious process, neutrophils exert various antimicrobial defenses, including the release of nuclear chromatin into the extracellular space. The externalized NETs, a complex meshwork of nuclear chromatin and antimicrobial proteins, serve to immobilize and degrade microbial pathogens. Here, we critically evaluate the evidence supporting NETs versus apoptotic bodies as a source for nuclear antigens in autoimmunity. We also discuss the possibility that NET chromatin forms an essential component of immune deposits in the pathogenesis of glomerulonephritis in SLE and other autoimmune immune complex diseases.

Genetic alterations and their relationship in the phosphatidylinositol 3-kinase/Akt pathway in thyroid cancer.. PURPOSE To investigate the overall occurrence and relationship of genetic alterations in the phosphatidylinositol 3-kinase (PI3K)/Akt pathway in thyroid tumors and explore the scope of this pathway as a therapeutic target for thyroid cancer. EXPERIMENTAL DESIGN We examined collectively the major genetic alterations and their relationship in this pathway, including PIK3CA copy number gain and mutation, Ras mutation, and PTEN mutation, in a large series of primary thyroid tumors. RESULTS Occurrence of any of these genetic alterations was found in 25 of 81 (31%) benign thyroid adenoma (BTA), 47 of 86 (55%) follicular thyroid cancer (FTC), 21 of 86 (24%) papillary thyroid cancer (PTC), and 29 of 50 (58%) anaplastic thyroid cancer (ATC), with FTC and ATC most frequently harboring these genetic alterations. PIK3CA copy gain was associated with increased PIK3CA protein expression. A mutual exclusivity among these genetic alterations was seen in BTA, FTC, and PTC, suggesting an independent role of each of them through the PI3K/Akt pathway in the tumorigenesis of the differentiated thyroid tumors. However, coexistence of these genetic alterations was increasingly seen with progression from differentiated tumor to undifferentiated ATC. Their coexistence with BRAF mutation was also frequent in PTC and ATC. CONCLUSIONS The data provide strong genetic implication that aberrant activation of PI3K/Akt pathway plays an extensive role in thyroid tumorigenesis, particularly in FTC and ATC, and promotes progression of BTA to FTC and to ATC as the genetic alterations of this pathway accumulate. Progression of PTC to ATC may be facilitated by coexistence of PI3K/Akt pathway-related genetic alterations and BRAF mutation. The PI3K/Akt pathway may thus be a major therapeutic target in thyroid cancers.


Question: G-CSF increases the expansion and infiltration of MDSCs into tumors.

Oncogenic mTOR signaling recruits myeloid-derived suppressor cells to promote tumor initiation. Myeloid-derived suppressor cells (MDSCs) play critical roles in primary and metastatic cancer progression. MDSC regulation is widely variable even among patients harbouring the same type of malignancy, and the mechanisms governing such heterogeneity are largely unknown. Here, integrating human tumour genomics and syngeneic mammary tumour models, we demonstrate that mTOR signalling in cancer cells dictates a mammary tumour's ability to stimulate MDSC accumulation through regulating G-CSF. Inhibiting this pathway or its activators (for example, FGFR) impairs tumour progression, which is partially rescued by restoring MDSCs or G-CSF. Tumour-initiating cells (TICs) exhibit elevated G-CSF. MDSCs reciprocally increase TIC frequency through activating Notch in tumour cells, forming a feedforward loop. Analyses of primary breast cancers and patient-derived xenografts corroborate these mechanisms in patients. These findings establish a non-canonical oncogenic role of mTOR signalling in recruiting pro-tumorigenic MDSCs and show how defined cancer subsets may evolve to promote and depend on a distinct immune microenvironment.

Cerebral palsy among term and postterm births.. CONTEXT Although preterm delivery is a well-established risk factor for cerebral palsy (CP), preterm deliveries contribute only a minority of affected infants. There is little information on the relation of CP risk to gestational age in the term range, where most CP occurs. OBJECTIVE To determine whether timing of birth in the term and postterm period is associated with risk of CP. DESIGN, SETTING, AND PARTICIPANTS Population-based follow-up study using the Medical Birth Registry of Norway to identify 1,682,441 singleton children born in the years 1967-2001 with a gestational age of 37 through 44 weeks and no congenital anomalies. The cohort was followed up through 2005 by linkage to other national registries. MAIN OUTCOME MEASURES Absolute and relative risk of CP for children surviving to at least 4 years of age. RESULTS Of the cohort of term and postterm children, 1938 were registered with CP in the National Insurance Scheme. Infants born at 40 weeks had the lowest risk of CP, with a prevalence of 0.99/1000 (95% confidence interval [CI], 0.90-1.08). Risk for CP was higher with earlier or later delivery, with a prevalence at 37 weeks of 1.91/1000 (95% CI, 1.58-2.25) and a relative risk (RR) of 1.9 (95% CI, 1.6-2.4), a prevalence at 38 weeks of 1.25/1000 (95% CI, 1.07-1.42) and an RR of 1.3 (95% CI, 1.1-1.6), a prevalence at 42 weeks of 1.36/1000 (95% CI, 1.19-1.53) and an RR of 1.4 (95% CI, 1.2-1.6), and a prevalence after 42 weeks of 1.44 (95% CI, 1.15-1.72) and an RR of 1.4 (95% CI, 1.1-1.8). These associations were even stronger in a subset with gestational age based on ultrasound measurements: at 37 weeks the prevalence was 1.17/1000 (95% CI, 0.30-2.04) and the relative risk was 3.7 (95% CI, 1.5-9.1). At 42 weeks the prevalence was 0.85/1000 (95% CI, 0.33-1.38) and the relative risk was 2.4 (95% CI, 1.1-5.3). Adjustment for infant sex, maternal age, and various socioeconomic measures had little effect. CONCLUSION Compared with delivery at 40 weeks' gestation, delivery at 37 or 38 weeks or at 42 weeks or later was associated with an increased risk of CP.


Question: Pre-mRNAs associated with spliceosomal components are more stable than unassociated splicing substrates.

Analyzing mechanisms of alternative pre-mRNA splicing using in vitro splicing assays.. The development of in vitro assays to analyze pre-mRNA splicing resulted in the discovery of many fundamental features characterizing splicing signals and the machinery that completes this process. Because in vitro assays can be manipulated by various biochemical approaches, the versatility of investigating alternative pre-mRNA splicing in the test tube appears endless. Importantly, modifications in reaction conditions can lead to the accumulation, isolation, and characterization of reaction intermediates, a prerequisite for gaining mechanistic insights into how the spliceosome carries out intron removal, and how regulatory elements assist the general splicing machinery in defining splice sites and alternative exons. These considerable experimental advantages have made the in vitro splicing system a standard assay, even though this approach is independent from RNA transcription and other RNA processing events, and in some respects deviates from the natural process of mRNA biogenesis. Here, we describe the tools and techniques necessary to carry out in vitro splicing assays. Analyses of various experimental designs are presented to highlight the approaches taken to gain insights into the mechanisms by which splice site recognition and activation are communicated with the general splicing machinery. Methods to measure the kinetics of splicing, to observe the formation of the pre-spliceosomal complexes, and to manipulate and modify the in vitro system to resolve the regulatory influences in alternative splicing are presented.

STAR: ultrafast universal RNA-seq aligner.. MOTIVATION Accurate alignment of high-throughput RNA-seq data is a challenging and yet unsolved problem because of the non-contiguous transcript structure, relatively short read lengths and constantly increasing throughput of the sequencing technologies. Currently available RNA-seq aligners suffer from high mapping error rates, low mapping speed, read length limitation and mapping biases. RESULTS To align our large (>80 billon reads) ENCODE Transcriptome RNA-seq dataset, we developed the Spliced Transcripts Alignment to a Reference (STAR) software based on a previously undescribed RNA-seq alignment algorithm that uses sequential maximum mappable seed search in uncompressed suffix arrays followed by seed clustering and stitching procedure. STAR outperforms other aligners by a factor of >50 in mapping speed, aligning to the human genome 550 million 2 × 76 bp paired-end reads per hour on a modest 12-core server, while at the same time improving alignment sensitivity and precision. In addition to unbiased de novo detection of canonical junctions, STAR can discover non-canonical splices and chimeric (fusion) transcripts, and is also capable of mapping full-length RNA sequences. Using Roche 454 sequencing of reverse transcription polymerase chain reaction amplicons, we experimentally validated 1960 novel intergenic splice junctions with an 80-90% success rate, corroborating the high precision of the STAR mapping strategy. AVAILABILITY AND IMPLEMENTATION STAR is implemented as a standalone C++ code. STAR is free open source software distributed under GPLv3 license and can be downloaded from http://code.google.com/p/rna-star/.


Question: PTEN is a regulator for the transcriptional activity of SRF

Nuclear PTEN functions as an essential regulator of SRF-dependent transcription to control smooth muscle differentiation. Vascular disease progression is associated with marked changes in vascular smooth muscle cell (SMC) phenotype and function. SMC contractile gene expression and, thus differentiation, is under direct transcriptional control by the transcription factor, serum response factor (SRF); however, the mechanisms dynamically regulating SMC phenotype are not fully defined. Here we report that the lipid and protein phosphatase, PTEN, has a novel role in the nucleus by functioning as an indispensible regulator with SRF to maintain the differentiated SM phenotype. PTEN interacts with the N-terminal domain of SRF and PTEN-SRF interaction promotes SRF binding to essential promoter elements in SM-specific genes. Factors inducing phenotypic switching promote loss of nuclear PTEN through nucleo-cytoplasmic translocation resulting in reduced myogenically active SRF, but enhanced SRF activity on target genes involved in proliferation. Overall decreased expression of PTEN was observed in intimal SMCs of human atherosclerotic lesions underlying the potential clinical importance of these findings.

Neuregulin 1 promotes excitatory synapse development and function in GABAergic interneurons.. Neuregulin 1 (NRG1) and its receptor ErbB4 are both susceptibility genes of schizophrenia. However, little is known about the underlying mechanisms of their malfunction. Although ErbB4 is enriched in GABAergic interneurons, the role of NRG1 in excitatory synapse formation in these neurons remains poorly understood. We showed that NRG1 increased both the number and size of PSD-95 puncta and the frequency and amplitude of miniature EPSCs (mEPSCs) in GABAergic interneurons, indicating that NRG1 stimulates the formation of new synapses and strengthens existing synapses. In contrast, NRG1 treatment had no effect on either the number or size of excitatory synapses in glutamatergic neurons, suggesting its synaptogenic effect is specific to GABAergic interneurons. Ecto-ErbB4 treatment diminished both the number and size of excitatory synapses, suggesting that endogenous NRG1 may be critical for basal synapse formation. NRG1 could stimulate the stability of PSD-95 in the manner that requires tyrosine kinase activity of ErbB4. Finally, deletion of ErbB4 in parvalbumin-positive interneurons led to reduced frequency and amplitude of mEPSCs, providing in vivo evidence that ErbB4 is important in excitatory synaptogenesis in interneurons. Together, our findings suggested a novel synaptogenic role of NRG1 in excitatory synapse development, possibly via stabilizing PSD-95, and this effect is specific to GABAergic interneurons. In light of the association of the genes of both NRG1 and ErbB4 with schizophrenia and dysfunction of GABAergic system in this disorder, these results provide insight into its potential pathological mechanism.


Question: Adult tissue-resident macrophages stem from the embryonal yolk sac and fetal liver.

Origin and functions of tissue macrophages.. Macrophages are distributed in tissues throughout the body and contribute to both homeostasis and disease. Recently, it has become evident that most adult tissue macrophages originate during embryonic development and not from circulating monocytes. Each tissue has its own composition of embryonically derived and adult-derived macrophages, but it is unclear whether macrophages of distinct origins are functionally interchangeable or have unique roles at steady state. This new understanding also prompts reconsideration of the function of circulating monocytes. Classical Ly6c(hi) monocytes patrol the extravascular space in resting organs, and Ly6c(lo) nonclassical monocytes patrol the vasculature. Inflammation triggers monocytes to differentiate into macrophages, but whether resident and newly recruited macrophages possess similar functions during inflammation is unclear. Here, we define the tools used for identifying the complex origin of tissue macrophages and discuss the relative contributions of tissue niche versus ontological origin to the regulation of macrophage functions during steady state and inflammation.

Update on technical issues concerning complementary feeding of young children in developing countries and implications for intervention programs.. This paper provides an update to the 1998 WHO/UNICEF report on complementary feeding. New research findings are generally consistent with the guidelines in that report, but the adoption of new energy and micronutrient requirements for infants and young children will result in lower recommendations regarding minimum meal frequency and energy density of complementary foods, and will alter the list of "problem nutrients. " Without fortification, the densities of iron, zinc, and vitamin B6 in complementary foods are often inadequate, and the intake of other nutrients may also be low in some populations. Strategies for obtaining the needed amounts of problem nutrients, as well as optimizing breastmilk intake when other foods are added to the diet, are discussed. The impact of complementary feeding interventions on child growth has been variable, which calls attention to the need for more comprehensive programs. A six-step approach to planning, implementing, and evaluating such programs is recommended.


Question: The number of unfertilized oocytes in mated hermaphrodites increases with age.

TGF-β and Insulin Signaling Regulate Reproductive Aging via Oocyte and Germline Quality Maintenance. Reproductive cessation is perhaps the earliest aging phenotype that humans experience. Similarly, reproduction of Caenorhabditis elegans ceases in mid-adulthood. Although somatic aging has been studied in both worms and humans, mechanisms regulating reproductive aging are not yet understood. Here, we show that TGF-β Sma/Mab and Insulin/IGF-1 signaling regulate C. elegans reproductive aging by modulating multiple aspects of the reproductive process, including embryo integrity, oocyte fertilizability, chromosome segregation fidelity, DNA damage resistance, and oocyte and germline morphology. TGF-β activity regulates reproductive span and germline/oocyte quality noncell-autonomously and is temporally and transcriptionally separable from its regulation of growth. Chromosome segregation, cell cycle, and DNA damage response genes are upregulated in TGF-β mutant oocytes, decline in aged mammalian oocytes, and are critical for oocyte quality maintenance. Our data suggest that C. elegans and humans share many aspects of reproductive aging, including the correlation between reproductive aging and declining oocyte quality and mechanisms determining oocyte quality.

Body-mass index and cause-specific mortality in 900 000 adults: collaborative analyses of 57 prospective studies.. BACKGROUND The main associations of body-mass index (BMI) with overall and cause-specific mortality can best be assessed by long-term prospective follow-up of large numbers of people. The Prospective Studies Collaboration aimed to investigate these associations by sharing data from many studies. METHODS Collaborative analyses were undertaken of baseline BMI versus mortality in 57 prospective studies with 894 576 participants, mostly in western Europe and North America (61% [n=541 452] male, mean recruitment age 46 [SD 11] years, median recruitment year 1979 [IQR 1975-85], mean BMI 25 [SD 4] kg/m(2)). The analyses were adjusted for age, sex, smoking status, and study. To limit reverse causality, the first 5 years of follow-up were excluded, leaving 66 552 deaths of known cause during a mean of 8 (SD 6) further years of follow-up (mean age at death 67 [SD 10] years): 30 416 vascular; 2070 diabetic, renal or hepatic; 22 592 neoplastic; 3770 respiratory; 7704 other. FINDINGS In both sexes, mortality was lowest at about 22.5-25 kg/m(2). Above this range, positive associations were recorded for several specific causes and inverse associations for none, the absolute excess risks for higher BMI and smoking were roughly additive, and each 5 kg/m(2) higher BMI was on average associated with about 30% higher overall mortality (hazard ratio per 5 kg/m(2) [HR] 1.29 [95% CI 1.27-1.32]): 40% for vascular mortality (HR 1.41 [1.37-1.45]); 60-120% for diabetic, renal, and hepatic mortality (HRs 2.16 [1.89-2.46], 1.59 [1.27-1.99], and 1.82 [1.59-2.09], respectively); 10% for neoplastic mortality (HR 1.10 [1.06-1.15]); and 20% for respiratory and for all other mortality (HRs 1.20 [1.07-1.34] and 1.20 [1.16-1.25], respectively). Below the range 22.5-25 kg/m(2), BMI was associated inversely with overall mortality, mainly because of strong inverse associations with respiratory disease and lung cancer. These inverse associations were much stronger for smokers than for non-smokers, despite cigarette consumption per smoker varying little with BMI. INTERPRETATION Although other anthropometric measures (eg, waist circumference, waist-to-hip ratio) could well add extra information to BMI, and BMI to them, BMI is in itself a strong predictor of overall mortality both above and below the apparent optimum of about 22.5-25 kg/m(2). The progressive excess mortality above this range is due mainly to vascular disease and is probably largely causal. At 30-35 kg/m(2), median survival is reduced by 2-4 years; at 40-45 kg/m(2), it is reduced by 8-10 years (which is comparable with the effects of smoking). The definite excess mortality below 22.5 kg/m(2) is due mainly to smoking-related diseases, and is not fully explained.


Question: Ectopic expression of Sall4, Nanog, Esrrb, and Lin28 generates induced pluripotent stem cells from mouse embryonic fibroblasts more efficiently than other factor combinations.

The developmental potential of iPSCs is greatly influenced by reprogramming factor selection.. Induced pluripotent stem cells (iPSCs) are commonly generated by transduction of Oct4, Sox2, Klf4, and Myc (OSKM) into cells. Although iPSCs are pluripotent, they frequently exhibit high variation in terms of quality, as measured in mice by chimera contribution and tetraploid complementation. Reliably high-quality iPSCs will be needed for future therapeutic applications. Here, we show that one major determinant of iPSC quality is the combination of reprogramming factors used. Based on tetraploid complementation, we found that ectopic expression of Sall4, Nanog, Esrrb, and Lin28 (SNEL) in mouse embryonic fibroblasts (MEFs) generated high-quality iPSCs more efficiently than other combinations of factors including OSKM. Although differentially methylated regions, transcript number of master regulators, establishment of specific superenhancers, and global aneuploidy were comparable between high- and low-quality lines, aberrant gene expression, trisomy of chromosome 8, and abnormal H2A.X deposition were distinguishing features that could potentially also be applicable to human.

Colorectal cancer survival in socioeconomic groups in England: variation is mainly in the short term after diagnosis.. UNLABELLED The objective of this study was to examine differences in cancer survival between socioeconomic groups in England, with particular attention to survival in the short term of follow-up. PATIENTS AND METHODS Individuals diagnosed with colorectal cancer between 1996 and 2004 in England were identified from cancer registry records. Five-year cumulative relative survival and excess death rates were computed. RESULTS For colon cancer there was a very high excess death rate in the first month of follow-up, and the excess death rate was highest in the socioeconomically deprived groups. In subsequent periods, excess mortality rates were much lower and there was less socioeconomic variation. The pattern of variation in excess death rates was generally similar in rectal cancer but the socioeconomic difference in death rates persisted several years longer. If the excess death rates in the entire colorectal cancer patient population were the same as those observed in the most affluent socioeconomic quintile, the annual reduction would be 360 deaths in colon cancer and 336 deaths in rectal cancer patients. These deaths occurred almost entirely in the first month and the first year after diagnosis. CONCLUSION Recent developments in the national cancer control agenda have included an increasing emphasis on outcome measures, with short-term cancer survival an operational measure of variation and progress in cancer control. In providing clues to the nature of the survival differences between socioeconomic groups, the results presented here give strong support for this strategy.


Question: Kir7.1 modulates channel activity by the G protein-coupled melanocortin-4 receptor (MC4R) in the paraventricular nucleus of the hypothalamus.

G-protein-independent coupling of MC4R to Kir7.1 in hypothalamic neurons. The regulated release of anorexigenic α-melanocyte stimulating hormone (α-MSH) and orexigenic Agouti-related protein (AgRP) from discrete hypothalamic arcuate neurons onto common target sites in the central nervous system has a fundamental role in the regulation of energy homeostasis. Both peptides bind with high affinity to the melanocortin-4 receptor (MC4R); existing data show that α-MSH is an agonist that couples the receptor to the Gαs signalling pathway, while AgRP binds competitively to block α-MSH binding and blocks the constitutive activity mediated by the ligand-mimetic amino-terminal domain of the receptor. Here we show that, in mice, regulation of firing activity of neurons from the paraventricular nucleus of the hypothalamus (PVN) by α-MSH and AgRP can be mediated independently of Gαs signalling by ligand-induced coupling of MC4R to closure of inwardly rectifying potassium channel, Kir7.1. Furthermore, AgRP is a biased agonist that hyperpolarizes neurons by binding to MC4R and opening Kir7.1, independently of its inhibition of α-MSH binding. Consequently, Kir7.1 signalling appears to be central to melanocortin-mediated regulation of energy homeostasis within the PVN. Coupling of MC4R to Kir7.1 may explain unusual aspects of the control of energy homeostasis by melanocortin signalling, including the gene dosage effect of MC4R and the sustained effects of AgRP on food intake.

Targeted blockade of TGF-β and IL-6/JAK2/STAT3 pathways inhibits lung cancer growth promoted by bone marrow-derived myofibroblasts. To investigate the role of TGF-β and IL-6 in myofibroblasts (MFs) - lung cancer cell interactions, lung cancer cells (Lewis and CTM-167 cell lines) were stimulated by IL-6, MF-conditioned medium (MF-CM) or MFs, with or without TGF-β signaling inhibitor - SB431542 and/or JAK2/STAT3 inhibitor - JSI-124. MFs were stimulated by TGF-β, cancer cell-CM or cancer cells, with or without SB431542 and JSI-124. Cell proliferation, the levels of cytokines, expression of mRNA and protein were determined. Mice bearing xenograft tumors were intraperitoneally treated with SB431542 or JSI-124 and monitored for up to 45 days. In co-culture systems, MFs secreted high levels of IL-6, while cancer cells produced high levels of TGF-β. Recombinant IL-6 and MF-CM activated STAT3 and upregulated TGF-β in cancer cells. In contrast, cancer cell-CM or TGF-β stimulated MFs to produce IL-6. Blockade of JAK2/STAT3 and TGF-β signaling by specific inhibitors significantly inhibited cell proliferation in vitro and tumor growth in vivo of lung cancer cells. Our study demontrated that the TGF-β and IL-6/JAK2/STAT3 signaling pathways form a positive feedback signaling loop that mediated the interactions between MFs and lung cancer cells. Targeted inhibiton of this signaling loop could be a new approach for lung cancer prevention and therapy.


Question: Reduced phosphorylation of PP2A suppresses HDAC4 dephosphorylation.

Nuclear accumulation of HDAC4 in ATM deficiency promotes neurodegeneration in ataxia-telangiectasia. Ataxia telangiectasia is a neurodegenerative disease caused by mutation of the Atm gene. Here we report that ataxia telangiectasia mutated (ATM) deficiency causes nuclear accumulation of histone deacetylase 4 (HDAC4) in neurons and promotes neurodegeneration. Nuclear HDAC4 binds to chromatin, as well as to myocyte enhancer factor 2A (MEF2A) and cAMP-responsive element binding protein (CREB), leading to histone deacetylation and altered neuronal gene expression. Blocking either HDAC4 activity or its nuclear accumulation blunts these neurodegenerative changes and rescues several behavioral abnormalities of ATM-deficient mice. Full rescue of the neurodegeneration, however, also requires the presence of HDAC4 in the cytoplasm, suggesting that the ataxia telangiectasia phenotype results both from a loss of cytoplasmic HDAC4 as well as its nuclear accumulation. To remain cytoplasmic, HDAC4 must be phosphorylated. The activity of the HDAC4 phosphatase, protein phosphatase 2A (PP2A), is downregulated by ATM-mediated phosphorylation. In ATM deficiency, enhanced PP2A activity leads to HDAC4 dephosphorylation and the nuclear accumulation of HDAC4. Our results define a crucial role of the cellular localization of HDAC4 in the events leading to ataxia telangiectasia neurodegeneration.

Disease-associated functions of IL-33: the new kid in the IL-1 family. Interleukin-33 (IL-33), a newly described member of the IL-1 family, is expressed by many cell types following pro-inflammatory stimulation and is thought to be released on cell lysis. The IL-33 receptor, consisting of ST2 and IL-1 receptor accessory protein, is also widely expressed, particularly by T helper 2 (TH2) cells and mast cells. IL-33 is host-protective against helminth infection and reduces atherosclerosis by promoting TH2-type immune responses. However, IL-33 can also promote the pathogenesis of asthma by expanding TH2 cells and mediate joint inflammation, atopic dermatitis and anaphylaxis by mast cell activation. Thus IL-33 could be a new target for therapeutic intervention across a range of diseases.


Question: LRBA prevents CTLA - 4 recycling.

Autosomal dominant immune dysregulation syndrome in humans with CTLA4 mutations. The protein cytotoxic T lymphocyte antigen-4 (CTLA-4) is an essential negative regulator of immune responses, and its loss causes fatal autoimmunity in mice. We studied a large family in which five individuals presented with a complex, autosomal dominant immune dysregulation syndrome characterized by hypogammaglobulinemia, recurrent infections and multiple autoimmune clinical features. We identified a heterozygous nonsense mutation in exon 1 of CTLA4. Screening of 71 unrelated patients with comparable clinical phenotypes identified five additional families (nine individuals) with previously undescribed splice site and missense mutations in CTLA4. Clinical penetrance was incomplete (eight adults of a total of 19 genetically proven CTLA4 mutation carriers were considered unaffected). However, CTLA-4 protein expression was decreased in regulatory T cells (Treg cells) in both patients and carriers with CTLA4 mutations. Whereas Treg cells were generally present at elevated numbers in these individuals, their suppressive function, CTLA-4 ligand binding and transendocytosis of CD80 were impaired. Mutations in CTLA4 were also associated with decreased circulating B cell numbers. Taken together, mutations in CTLA4 resulting in CTLA-4 haploinsufficiency or impaired ligand binding result in disrupted T and B cell homeostasis and a complex immune dysregulation syndrome.

Oxymetholone Therapy of Fanconi Anemia Suppresses Osteopontin Transcription and Induces Hematopoietic Stem Cell Cycling. Androgens are widely used for treating Fanconi anemia (FA) and other human bone marrow failure syndromes, but their mode of action remains incompletely understood. Aged Fancd2(-/-) mice were used to assess the therapeutic efficacy of oxymetholone (OXM) and its mechanism of action. Eighteen-month-old Fancd2(-/-) mice recapitulated key human FA phenotypes, including reduced bone marrow cellularity, red cell macrocytosis, and peripheral pancytopenia. As in humans, chronic OXM treatment significantly improved these hematological parameters and stimulated the proliferation of hematopoietic stem and progenitor cells. RNA-Seq analysis implicated downregulation of osteopontin as an important potential mechanism for the drug's action. Consistent with the increased stem cell proliferation, competitive repopulation assays demonstrated that chronic OXM therapy eventually resulted in stem cell exhaustion. These results expand our knowledge of the regulation of hematopoietic stem cell proliferation and have direct clinical implications for the treatment of bone marrow failure.


Question: COPI coatmer is involved in viral replication.

COPI Complex Is a Regulator of Lipid Homeostasis. Lipid droplets are ubiquitous triglyceride and sterol ester storage organelles required for energy storage homeostasis and biosynthesis. Although little is known about lipid droplet formation and regulation, it is clear that members of the PAT (perilipin, adipocyte differentiation related protein, tail interacting protein of 47 kDa) protein family coat the droplet surface and mediate interactions with lipases that remobilize the stored lipids. We identified key Drosophila candidate genes for lipid droplet regulation by RNA interference (RNAi) screening with an image segmentation-based optical read-out system, and show that these regulatory functions are conserved in the mouse. Those include the vesicle-mediated Coat Protein Complex I (COPI) transport complex, which is required for limiting lipid storage. We found that COPI components regulate the PAT protein composition at the lipid droplet surface, and promote the association of adipocyte triglyceride lipase (ATGL) with the lipid droplet surface to mediate lipolysis. Two compounds known to inhibit COPI function, Exo1 and Brefeldin A, phenocopy COPI knockdowns. Furthermore, RNAi inhibition of ATGL and simultaneous drug treatment indicate that COPI and ATGL function in the same pathway. These data indicate that the COPI complex is an evolutionarily conserved regulator of lipid homeostasis, and highlight an interaction between vesicle transport systems and lipid droplets.

ALDH1 is a marker of normal and malignant human mammary stem cells and a predictor of poor clinical outcome.. Application of stem cell biology to breast cancer research has been limited by the lack of simple methods for identification and isolation of normal and malignant stem cells. Utilizing in vitro and in vivo experimental systems, we show that normal and cancer human mammary epithelial cells with increased aldehyde dehydrogenase activity (ALDH) have stem/progenitor properties. These cells contain the subpopulation of normal breast epithelium with the broadest lineage differentiation potential and greatest growth capacity in a xenotransplant model. In breast carcinomas, high ALDH activity identifies the tumorigenic cell fraction, capable of self-renewal and of generating tumors that recapitulate the heterogeneity of the parental tumor. In a series of 577 breast carcinomas, expression of ALDH1 detected by immunostaining correlated with poor prognosis. These findings offer an important new tool for the study of normal and malignant breast stem cells and facilitate the clinical application of stem cell concepts.


Question: In British Men, haplogroup I increases risk of cardiovascular disease by 50%.

Inheritance of coronary artery disease in men: an analysis of the role of the Y chromosome. BACKGROUND A sexual dimorphism exists in the incidence and prevalence of coronary artery disease--men are more commonly affected than are age-matched women. We explored the role of the Y chromosome in coronary artery disease in the context of this sexual inequity. METHODS We genotyped 11 markers of the male-specific region of the Y chromosome in 3233 biologically unrelated British men from three cohorts: the British Heart Foundation Family Heart Study (BHF-FHS), West of Scotland Coronary Prevention Study (WOSCOPS), and Cardiogenics Study. On the basis of this information, each Y chromosome was tracked back into one of 13 ancient lineages defined as haplogroups. We then examined associations between common Y chromosome haplogroups and the risk of coronary artery disease in cross-sectional BHF-FHS and prospective WOSCOPS. Finally, we undertook functional analysis of Y chromosome effects on monocyte and macrophage transcriptome in British men from the Cardiogenics Study. FINDINGS Of nine haplogroups identified, two (R1b1b2 and I) accounted for roughly 90% of the Y chromosome variants among British men. Carriers of haplogroup I had about a 50% higher age-adjusted risk of coronary artery disease than did men with other Y chromosome lineages in BHF-FHS (odds ratio 1·75, 95% CI 1·20-2·54, p=0·004), WOSCOPS (1·45, 1·08-1·95, p=0·012), and joint analysis of both populations (1·56, 1·24-1·97, p=0·0002). The association between haplogroup I and increased risk of coronary artery disease was independent of traditional cardiovascular and socioeconomic risk factors. Analysis of macrophage transcriptome in the Cardiogenics Study revealed that 19 molecular pathways showing strong differential expression between men with haplogroup I and other lineages of the Y chromosome were interconnected by common genes related to inflammation and immunity, and that some of them have a strong relevance to atherosclerosis. INTERPRETATION The human Y chromosome is associated with risk of coronary artery disease in men of European ancestry, possibly through interactions of immunity and inflammation. FUNDING British Heart Foundation; UK National Institute for Health Research; LEW Carty Charitable Fund; National Health and Medical Research Council of Australia; European Union 6th Framework Programme; Wellcome Trust.

Sympathetic activation during early pregnancy in humans.. Sympathetic activity has been reported to increase in normotensive pregnant women, and to be even greater in women with gestational hypertension and preeclampsia at term. Whether sympathetic overactivity develops early during pregnancy, remaining high throughout gestation, or whether it only occurs at term providing the substrate for hypertensive disorders is unknown. We tested the hypothesis that sympathetic activation occurs early during pregnancy in humans. Eleven healthy women (29 ± 3 (SD) years) without prior hypertensive pregnancies were tested during the mid-luteal phase (PRE) and early pregnancy (EARLY; 6.2 ± 1.2 weeks of gestation). Muscle sympathetic nerve activity (MSNA) and haemodynamics were measured supine, at 30 deg and 60 deg upright tilt for 5 min each. Blood samples were drawn for catecholamines, direct renin, and aldosterone. MSNA was significantly greater during EARLY than PRE (supine: 25 ± 8 vs. 14 ± 8 bursts min(-1), 60 deg tilt: 49 ± 14 vs. 40 ± 10 bursts min(-1); main effect, P < 0.05). Resting diastolic pressure trended lower (P = 0.09), heart rate was similar, total peripheral resistance decreased (2172 ± 364 vs. 2543 ± 352 dyne s cm(-5); P < 0.05), sympathetic vascular transduction was blunted (0.10 ± 0.05 vs. 0.36 ± 0.47 units a.u.(-1) min(-1); P < 0.01), and both renin (supine: 27.9 ± 6.2 vs. 14.2 ± 8.7 pg ml(-1), P < 0.01) and aldosterone (supine: 16.7 ± 14.1 vs. 7.7 ± 6.8 ng ml(-1), P = 0.05) were higher during EARLY than PRE. These results suggest that sympathetic activation is a common characteristic of early pregnancy in humans despite reduced diastolic pressure and total peripheral resistance. These observations challenge conventional thinking about blood pressure regulation during pregnancy, showing marked sympathetic activation occurring within the first few weeks of conception, and may provide the substrate for pregnancy induced cardiovascular complications.


Question: Tuberculosis incidence occurs at higher rates in more sparsely populated areas in the UK.

Ecological analysis of ethnic differences in relation between tuberculosis and poverty.. OBJECTIVE To examine the effect of ethnicity on the relation between tuberculosis and deprivation. DESIGN Retrospective ecological study comparing incidence of tuberculosis in white and south Asian residents of the 39 electoral wards in Birmingham with ethnic specific indices of deprivation. SETTING Birmingham, 1989-93. SUBJECTS 1516 notified cases of tuberculosis. MAIN OUTCOME MEASURES Rates of tuberculosis and measures of deprivation. RESULTS Univariate analysis showed significant associations of tuberculosis rates for the whole population with several indices of deprivation (P<0.01) and with the proportion of the population of south Asian origin (P<0.01). All deprivation covariates were positively associated with each other but on multiple regression, higher level of overcrowding was independently associated with tuberculosis rates. For the white population, overcrowding was associated with tuberculosis rates independently of other variables (P=0.0036). No relation with deprivation was found for the south Asian population in either single or multivariable analyses. CONCLUSIONS Poverty is significantly associated with tuberculosis in the white population, but no such relation exists for those of Asian ethnicity. These findings suggest that causal factors, and therefore potential interventions, will also differ by ethnic group.

Dietary quality among men and women in 187 countries in 1990 and 2010: a systematic assessment. BACKGROUND Healthy dietary patterns are a global priority to reduce non-communicable diseases. Yet neither worldwide patterns of diets nor their trends with time are well established. We aimed to characterise global changes (or trends) in dietary patterns nationally and regionally and to assess heterogeneity by age, sex, national income, and type of dietary pattern. METHODS In this systematic assessment, we evaluated global consumption of key dietary items (foods and nutrients) by region, nation, age, and sex in 1990 and 2010. Consumption data were evaluated from 325 surveys (71·7% nationally representative) covering 88·7% of the global adult population. Two types of dietary pattern were assessed: one reflecting greater consumption of ten healthy dietary items and the other based on lesser consumption of seven unhealthy dietary items. The mean intakes of each dietary factor were divided into quintiles, and each quintile was assigned an ordinal score, with higher scores being equivalent to healthier diets (range 0-100). The dietary patterns were assessed by hierarchical linear regression including country, age, sex, national income, and time as exploratory variables. FINDINGS From 1990 to 2010, diets based on healthy items improved globally (by 2·2 points, 95% uncertainty interval (UI) 0·9 to 3·5), whereas diets based on unhealthy items worsened (-2·5, -3·3 to -1·7). In 2010, the global mean scores were 44·0 (SD 10·5) for the healthy pattern and 52·1 (18·6) for the unhealthy pattern, with weak intercorrelation (r=-0·08) between countries. On average, better diets were seen in older adults compared with younger adults, and in women compared with men (p<0·0001 each). Compared with low-income nations, high-income nations had better diets based on healthy items (+2·5 points, 95% UI 0·3 to 4·1), but substantially poorer diets based on unhealthy items (-33·0, -37·8 to -28·3). Diets and their trends were very heterogeneous across the world regions. For example, both types of dietary patterns improved in high-income countries, but worsened in some low-income countries in Africa and Asia. Middle-income countries showed the largest improvement in dietary patterns based on healthy items, but the largest deterioration in dietary patterns based on unhealthy items. INTERPRETATION Consumption of healthy items improved, while consumption of unhealthy items worsened across the world, with heterogeneity across regions and countries. These global data provide the best estimates to date of nutrition transitions across the world and inform policies and priorities for reducing the health and economic burdens of poor diet quality. FUNDING The Bill & Melinda Gates Foundation and Medical Research Council.


Question: The RANK-RANK-Ligand (RANKL) pathway is involved in the development of Aire-expressing medullary thymic epithelial cells (mTECs).

Rank Signaling Links the Development of Invariant γδ T Cell Progenitors and Aire+ Medullary Epithelium. The thymic medulla provides a specialized microenvironment for the negative selection of T cells, with the presence of autoimmune regulator (Aire)-expressing medullary thymic epithelial cells (mTECs) during the embryonic-neonatal period being both necessary and sufficient to establish long-lasting tolerance. Here we showed that emergence of the first cohorts of Aire(+) mTECs at this key developmental stage, prior to αβ T cell repertoire selection, was jointly directed by Rankl(+) lymphoid tissue inducer cells and invariant Vγ5(+) dendritic epidermal T cell (DETC) progenitors that are the first thymocytes to express the products of gene rearrangement. In turn, generation of Aire(+) mTECs then fostered Skint-1-dependent, but Aire-independent, DETC progenitor maturation and the emergence of an invariant DETC repertoire. Hence, our data attributed a functional importance to the temporal development of Vγ5(+) γδ T cells during thymus medulla formation for αβ T cell tolerance induction and demonstrated a Rank-mediated reciprocal link between DETC and Aire(+) mTEC maturation.

Insulin Signaling and Dietary Restriction Differentially Influence the Decline of Learning and Memory with Age. Of all the age-related declines, memory loss is one of the most devastating. While conditions that increase longevity have been identified, the effects of these longevity-promoting factors on learning and memory are unknown. Here we show that the C. elegans Insulin/IGF-1 receptor mutant daf-2 improves memory performance early in adulthood and maintains learning ability better with age but, surprisingly, demonstrates no extension in long-term memory with age. By contrast, eat-2 mutants, a model of Dietary Restriction (DR), exhibit impaired long-term memory in young adulthood but maintain this level of memory longer with age. We find that crh-1, the C. elegans homolog of the CREB transcription factor, is required for long-term associative memory, but not for learning or short-term memory. The expression of crh-1 declines with age and differs in the longevity mutants, and CREB expression and activity correlate with memory performance. Our results suggest that specific longevity treatments have acute and long-term effects on cognitive functions that decline with age through their regulation of rate-limiting genes required for learning and memory.


Question: Taking 400mg of α-tocopheryl acetate does not prevent prostate cancer.

Vitamins E and C in the prevention of prostate and total cancer in men: the Physicians' Health Study II randomized controlled trial.. CONTEXT Many individuals take vitamins in the hopes of preventing chronic diseases such as cancer, and vitamins E and C are among the most common individual supplements. A large-scale randomized trial suggested that vitamin E may reduce risk of prostate cancer; however, few trials have been powered to address this relationship. No previous trial in men at usual risk has examined vitamin C alone in the prevention of cancer. OBJECTIVE To evaluate whether long-term vitamin E or C supplementation decreases risk of prostate and total cancer events among men. DESIGN, SETTING, AND PARTICIPANTS The Physicians' Health Study II is a randomized, double-blind, placebo-controlled factorial trial of vitamins E and C that began in 1997 and continued until its scheduled completion on August 31, 2007. A total of 14,641 male physicians in the United States initially aged 50 years or older, including 1307 men with a history of prior cancer at randomization, were enrolled. INTERVENTION Individual supplements of 400 IU of vitamin E every other day and 500 mg of vitamin C daily. MAIN OUTCOME MEASURES Prostate and total cancer. RESULTS During a mean follow-up of 8.0 years, there were 1008 confirmed incident cases of prostate cancer and 1943 total cancers. Compared with placebo, vitamin E had no effect on the incidence of prostate cancer (active and placebo vitamin E groups, 9.1 and 9.5 events per 1000 person-years; hazard ratio [HR], 0.97; 95% confidence interval [CI], 0.85-1.09; P = .58) or total cancer (active and placebo vitamin E groups, 17.8 and 17.3 cases per 1000 person-years; HR, 1.04; 95% CI, 0.95-1.13; P = .41). There was also no significant effect of vitamin C on total cancer (active and placebo vitamin C groups, 17.6 and 17.5 events per 1000 person-years; HR, 1.01; 95% CI, 0.92-1.10; P = .86) or prostate cancer (active and placebo vitamin C groups, 9.4 and 9.2 cases per 1000 person-years; HR, 1.02; 95% CI, 0.90-1.15; P = .80). Neither vitamin E nor vitamin C had a significant effect on colorectal, lung, or other site-specific cancers. Adjustment for adherence and exclusion of the first 4 or 6 years of follow-up did not alter the results. Stratification by various cancer risk factors demonstrated no significant modification of the effect of vitamin E on prostate cancer risk or either agent on total cancer risk. CONCLUSIONS In this large, long-term trial of male physicians, neither vitamin E nor C supplementation reduced the risk of prostate or total cancer. These data provide no support for the use of these supplements for the prevention of cancer in middle-aged and older men. TRIAL REGISTRATION clinicaltrials.gov Identifier: NCT00270647.

Trophoblast differentiation defect in human embryonic stem cells lacking PIG-A and GPI-anchored cell-surface proteins.. Pluripotent human embryonic stem (hES) cells can differentiate into various cell types derived from the three embryonic germ layers and extraembryonic tissues such as trophoblasts. The mechanisms governing lineage choices of hES cells are largely unknown. Here, we report that we established two independent hES cell clones lacking a group of cell surface molecules, glycosyl-phosphatidyl-inositol-anchored proteins (GPI-APs). The GPI-AP deficiency in these two hES clones is due to the deficiency in the gene expression of PIG-A (phosphatidyl-inositol-glycan class A), which is required for the first step of GPI synthesis. GPI-AP-deficient hES cells were capable of forming embryoid bodies and initiating cell differentiation into the three embryonic germ layers. However, GPI-AP-deficient hES cells failed to form trophoblasts after differentiation induction by embryoid body formation or by adding exogenous BMP4. The defect in trophoblast formation was due to the lack of GPI-anchored BMP coreceptors, resulting in the impairment of full BMP4 signaling activation in the GPI-AP-deficient hES cells. These data reveal that GPI-AP-enhanced full activation of BMP signaling is required for human trophoblast formation.


Question: There is no known interaction between Pioneer factor OCT3/4 and major chromatin remodeling factors.

Oct4 links multiple epigenetic pathways to the pluripotency network. Oct4 is a well-known transcription factor that plays fundamental roles in stem cell self-renewal, pluripotency, and somatic cell reprogramming. However, limited information is available on Oct4-associated protein complexes and their intrinsic protein-protein interactions that dictate Oct4's critical regulatory activities. Here we employed an improved affinity purification approach combined with mass spectrometry to purify Oct4 protein complexes in mouse embryonic stem cells (mESCs), and discovered many novel Oct4 partners important for self-renewal and pluripotency of mESCs. Notably, we found that Oct4 is associated with multiple chromatin-modifying complexes with documented as well as newly proved functional significance in stem cell maintenance and somatic cell reprogramming. Our study establishes a solid biochemical basis for genetic and epigenetic regulation of stem cell pluripotency and provides a framework for exploring alternative factor-based reprogramming strategies.

Population dynamics of a pathogen: the conundrum of vivax malaria. Building a mathematical model of population dynamics of pathogens within their host involves considerations of factors similar to those in ecology, as pathogens can prey on cells in the host. But within the multicellular host, attacked cell types are integrated with other cellular systems, which in turn intervene in the infection. For example, immune responses attempt to sense and then eliminate or contain pathogens, and homeostatic mechanisms try to compensate for cell loss. This review focuses on modeling applied to malarias, diseases caused by single-cell eukaryote parasites that infect red blood cells, with special concern given to vivax malaria, a disease often thought to be benign (if sometimes incapacitating) because the parasite only attacks a small proportion of red blood cells, the very youngest ones. However, I will use mathematical modeling to argue that depletion of this pool of red blood cells can be disastrous to the host if growth of the parasite is not vigorously check by host immune responses. Also, modeling can elucidate aspects of new field observations that indicate that vivax malaria is more dangerous than previously thought. ELECTRONIC SUPPLEMENTARY MATERIAL: The online version of this article (doi:10.1007/s12551-010-0034-3) contains supplementary material, which is available to authorized users.


Question: ER-localized phosphatase Sac1 processes PI4P through coordination with OSBP and the endosome-localized protein sorting nexin 2.

Endosome-ER Contacts Control Actin Nucleation and Retromer Function through VAP-Dependent Regulation of PI4P. VAP (VAPA and VAPB) is an evolutionarily conserved endoplasmic reticulum (ER)-anchored protein that helps generate tethers between the ER and other membranes through which lipids are exchanged across adjacent bilayers. Here, we report that by regulating PI4P levels on endosomes, VAP affects WASH-dependent actin nucleation on these organelles and the function of the retromer, a protein coat responsible for endosome-to-Golgi traffic. VAP is recruited to retromer budding sites on endosomes via an interaction with the retromer SNX2 subunit. Cells lacking VAP accumulate high levels of PI4P, actin comets, and trans-Golgi proteins on endosomes. Such defects are mimicked by downregulation of OSBP, a VAP interactor and PI4P transporter that participates in VAP-dependent ER-endosomes tethers. These results reveal a role of PI4P in retromer-/WASH-dependent budding from endosomes. Collectively, our data show how the ER can control budding dynamics and association with the cytoskeleton of another membrane by direct contacts leading to bilayer lipid modifications.

An Excitatory Loop with Astrocytes Contributes to Drive Neurons to Seizure Threshold. Seizures in focal epilepsies are sustained by a highly synchronous neuronal discharge that arises at restricted brain sites and subsequently spreads to large portions of the brain. Despite intense experimental research in this field, the earlier cellular events that initiate and sustain a focal seizure are still not well defined. Their identification is central to understand the pathophysiology of focal epilepsies and to develop new pharmacological therapies for drug-resistant forms of epilepsy. The prominent involvement of astrocytes in ictogenesis was recently proposed. We test here whether a cooperation between astrocytes and neurons is a prerequisite to support ictal (seizure-like) and interictal epileptiform events. Simultaneous patch-clamp recording and Ca2+ imaging techniques were performed in a new in vitro model of focal seizures induced by local applications of N-methyl-D-aspartic acid (NMDA) in rat entorhinal cortex slices. We found that a Ca2+ elevation in astrocytes correlates with both the initial development and the maintenance of a focal, seizure-like discharge. A delayed astrocyte activation during ictal discharges was also observed in other models (including the whole in vitro isolated guinea pig brain) in which the site of generation of seizure activity cannot be precisely monitored. In contrast, interictal discharges were not associated with Ca2+ changes in astrocytes. Selective inhibition or stimulation of astrocyte Ca2+ signalling blocked or enhanced, respectively, ictal discharges, but did not affect interictal discharge generation. Our data reveal that neurons engage astrocytes in a recurrent excitatory loop (possibly involving gliotransmission) that promotes seizure ignition and sustains the ictal discharge. This neuron-astrocyte interaction may represent a novel target to develop effective therapeutic strategies to control seizures.