Question:
Broadly HIV-1 Neutralizing Antibodies (bnAb) 10EB have no affinity for phospholipids.

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.

Neoadjuvant chemotherapy (NAC) induces a pathological complete response (pCR) in ∼30% of patients with breast cancer. However, many patients have residual cancer after chemotherapy, which correlates with a higher risk of metastatic recurrence and poorer outcome than those who achieve a pCR. We hypothesized that molecular profiling of tumors after NAC would identify genes associated with drug resistance. Digital transcript counting was used to profile surgically resected breast cancers after NAC. Low concentrations of dual specificity protein phosphatase 4 (DUSP4), an ERK phosphatase, correlated with high post-NAC tumor cell proliferation and with basal-like breast cancer (BLBC) status. BLBC had higher DUSP4 promoter methylation and gene expression patterns of Ras-ERK pathway activation relative to other breast cancer subtypes. DUSP4 overexpression increased chemotherapy-induced apoptosis, whereas DUSP4 depletion dampened the response to chemotherapy. Reduced DUSP4 expression in primary tumors after NAC was associated with treatment-refractory high Ki-67 scores and shorter recurrence-free survival. Finally, inhibition of mitogen-activated protein kinase kinase (MEK) synergized with docetaxel treatment in BLBC xenografts. Thus, DUSP4 downregulation activates the Ras-ERK pathway in BLBC, resulting in an attenuated response to anti-cancer chemotherapy.

IMPORTANCE Limited information about the relationship between specific mutations in BRCA1 or BRCA2 (BRCA1/2) and cancer risk exists. OBJECTIVE To identify mutation-specific cancer risks for carriers of BRCA1/2. DESIGN, SETTING, AND PARTICIPANTS Observational study of women who were ascertained between 1937 and 2011 (median, 1999) and found to carry disease-associated BRCA1 or BRCA2 mutations. The international sample comprised 19,581 carriers of BRCA1 mutations and 11,900 carriers of BRCA2 mutations from 55 centers in 33 countries on 6 continents. We estimated hazard ratios for breast and ovarian cancer based on mutation type, function, and nucleotide position. We also estimated RHR, the ratio of breast vs ovarian cancer hazard ratios. A value of RHR greater than 1 indicated elevated breast cancer risk; a value of RHR less than 1 indicated elevated ovarian cancer risk. EXPOSURES Mutations of BRCA1 or BRCA2. MAIN OUTCOMES AND MEASURES Breast and ovarian cancer risks. RESULTS Among BRCA1 mutation carriers, 9052 women (46%) were diagnosed with breast cancer, 2317 (12%) with ovarian cancer, 1041 (5%) with breast and ovarian cancer, and 7171 (37%) without cancer. Among BRCA2 mutation carriers, 6180 women (52%) were diagnosed with breast cancer, 682 (6%) with ovarian cancer, 272 (2%) with breast and ovarian cancer, and 4766 (40%) without cancer. In BRCA1, we identified 3 breast cancer cluster regions (BCCRs) located at c.179 to c.505 (BCCR1; RHR = 1.46; 95% CI, 1.22-1.74; P = 2 × 10(-6)), c.4328 to c.4945 (BCCR2; RHR = 1.34; 95% CI, 1.01-1.78; P = .04), and c. 5261 to c.5563 (BCCR2', RHR = 1.38; 95% CI, 1.22-1.55; P = 6 × 10(-9)). We also identified an ovarian cancer cluster region (OCCR) from c.1380 to c.4062 (approximately exon 11) with RHR = 0.62 (95% CI, 0.56-0.70; P = 9 × 10(-17)). In BRCA2, we observed multiple BCCRs spanning c.1 to c.596 (BCCR1; RHR = 1.71; 95% CI, 1.06-2.78; P = .03), c.772 to c.1806 (BCCR1'; RHR = 1.63; 95% CI, 1.10-2.40; P = .01), and c.7394 to c.8904 (BCCR2; RHR = 2.31; 95% CI, 1.69-3.16; P = .00002). We also identified 3 OCCRs: the first (OCCR1) spanned c.3249 to c.5681 that was adjacent to c.5946delT (6174delT; RHR = 0.51; 95% CI, 0.44-0.60; P = 6 × 10(-17)). The second OCCR spanned c.6645 to c.7471 (OCCR2; RHR = 0.57; 95% CI, 0.41-0.80; P = .001). Mutations conferring nonsense-mediated decay were associated with differential breast or ovarian cancer risks and an earlier age of breast cancer diagnosis for both BRCA1 and BRCA2 mutation carriers. CONCLUSIONS AND RELEVANCE Breast and ovarian cancer risks varied by type and location of BRCA1/2 mutations. With appropriate validation, these data may have implications for risk assessment and cancer prevention decision making for carriers of BRCA1 and BRCA2 mutations.

DNA double-strand breaks (DSBs), which are formed by the Spo11 protein, initiate meiotic recombination. Previous DSB-mapping studies have used rad50S or sae2Δ mutants, which are defective in break processing, to accumulate Spo11-linked DSBs, and report large (≥ 50 kb) “DSB-hot” regions that are separated by “DSB-cold” domains of similar size. Substantial recombination occurs in some DSB-cold regions, suggesting that DSB patterns are not normal in rad50S or sae2Δ mutants. We therefore developed a novel method to map genome-wide, single-strand DNA (ssDNA)–associated DSBs that accumulate in processing-capable, repair-defective dmc1Δ and dmc1Δ rad51Δ mutants. DSBs were observed at known hot spots, but also in most previously identified “DSB-cold” regions, including near centromeres and telomeres. Although approximately 40% of the genome is DSB-cold in rad50S mutants, analysis of meiotic ssDNA from dmc1Δ shows that most of these regions have substantial DSB activity. Southern blot assays of DSBs in selected regions in dmc1Δ, rad50S, and wild-type cells confirm these findings. Thus, DSBs are distributed much more uniformly than was previously believed. Comparisons of DSB signals in dmc1, dmc1 rad51, and dmc1 spo11 mutant strains identify Dmc1 as a critical strand-exchange activity genome-wide, and confirm previous conclusions that Spo11-induced lesions initiate all meiotic recombination.

Characterization of human monoclonal antibodies is providing considerable insight into mechanisms of broad HIV-1 neutralization. Here we report an HIV-1 gp41 membrane-proximal external region (MPER)-specific antibody, named 10E8, which neutralizes ∼98% of tested viruses. An analysis of sera from 78 healthy HIV-1-infected donors demonstrated that 27% contained MPER-specific antibodies and 8% contained 10E8-like specificities. In contrast to other neutralizing MPER antibodies, 10E8 did not bind phospholipids, was not autoreactive, and bound cell-surface envelope. The structure of 10E8 in complex with the complete MPER revealed a site of vulnerability comprising a narrow stretch of highly conserved gp41-hydrophobic residues and a critical arginine or lysine just before the transmembrane region. Analysis of resistant HIV-1 variants confirmed the importance of these residues for neutralization. The highly conserved MPER is a target of potent, non-self-reactive neutralizing antibodies, suggesting that HIV-1 vaccines should aim to induce antibodies to this region of HIV-1 envelope glycoprotein.


The tissue surrounding the granuloma in an immune cell induces an anti-inflammatory immune response.

CONTEXT Dexamethasone is widely used to prevent postoperative nausea and vomiting (PONV) in pediatric tonsillectomy. OBJECTIVE To assess whether dexamethasone dose-dependently reduces the risk of PONV at 24 hours after tonsillectomy. DESIGN, SETTING, AND PATIENTS Randomized placebo-controlled trial conducted among 215 children undergoing elective tonsillectomy at a major public teaching hospital in Switzerland from February 2005 to December 2007. INTERVENTIONS Children were randomly assigned to receive dexamethasone (0.05, 0.15, or 0.5 mg/kg) or placebo intravenously after induction of anesthesia. Acetaminophen-codeine and ibuprofen were given as postoperative analgesia. Follow-up continued until the 10th postoperative day. MAIN OUTCOME MEASURES The primary end point was prevention of PONV at 24 hours; secondary end points were decrease in the need for ibuprofen at 24 hours and evaluation of adverse effects. RESULTS At 24 hours, 24 of 54 participants who received placebo (44%; 95% confidence interval [CI], 31%-59%) had experienced PONV compared with 20 of 53 (38%; 95% CI, 25%-52%), 13 of 54 (24%; 95% CI, 13%-38%), and 6 of 52 (12%; 95% CI, 4%-23%) who received dexamethasone at 0.05, 0.15, and 0.5 mg/kg, respectively (P<.001 for linear trend). Children who received dexamethasone received significantly less ibuprofen. There were 26 postoperative bleeding episodes in 22 children. Two of 53 (4%; 95% CI, 0.5%-13%) children who received placebo had bleeding compared with 6 of 53 (11%; 95% CI, 4%-23%), 2 of 51 (4%; 95% CI, 0.5%-13%), and 12 of 50 (24%; 95% CI, 13%-38%) who received dexamethasone at 0.05, 0.15, and 0.5 mg/kg, respectively (P = .003). Dexamethasone, 0.5 mg/kg, was associated with the highest bleeding risk (adjusted relative risk, 6.80; 95% CI, 1.77-16.5). Eight children had to undergo emergency reoperation because of bleeding, all of whom had received dexamethasone. The trial was stopped early for safety reasons. CONCLUSION In this study of children undergoing tonsillectomy, dexamethasone decreased the risk of PONV dose dependently but was associated with an increased risk of postoperative bleeding. TRIAL REGISTRATION clinicaltrials.gov Identifier: NCT00403806.

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.

Tumor necrosis factor (TNF), a key effector in controlling tuberculosis, is thought to exert protection by directing formation of granulomas, organized aggregates of macrophages and other immune cells. Loss of TNF signaling causes progression of tuberculosis in humans, and the increased mortality of Mycobacterium tuberculosis-infected mice is associated with disorganized necrotic granulomas, although the precise roles of TNF signaling preceding this endpoint remain undefined. We monitored transparent Mycobacterium marinum-infected zebrafish live to conduct a stepwise dissection of how TNF signaling operates in mycobacterial pathogenesis. We found that loss of TNF signaling caused increased mortality even when only innate immunity was operant. In the absence of TNF, intracellular bacterial growth and granuloma formation were accelerated and was followed by necrotic death of overladen macrophages and granuloma breakdown. Thus, TNF is not required for tuberculous granuloma formation, but maintains granuloma integrity indirectly by restricting mycobacterial growth within macrophages and preventing their necrosis.

In eukaryotes, accurate protein synthesis relies on a family of translational GTPases that pair with specific decoding factors to decipher the mRNA code on ribosomes. We present structures of the mammalian ribosome engaged with decoding factor⋅GTPase complexes representing intermediates of translation elongation (aminoacyl-tRNA⋅eEF1A), termination (eRF1⋅eRF3), and ribosome rescue (Pelota⋅Hbs1l). Comparative analyses reveal that each decoding factor exploits the plasticity of the ribosomal decoding center to differentially remodel ribosomal proteins and rRNA. This leads to varying degrees of large-scale ribosome movements and implies distinct mechanisms for communicating information from the decoding center to each GTPase. Additional structural snapshots of the translation termination pathway reveal the conformational changes that choreograph the accommodation of decoding factors into the peptidyl transferase center. Our results provide a structural framework for how different states of the mammalian ribosome are selectively recognized by the appropriate decoding factor⋅GTPase complex to ensure translational fidelity.

Blood monocytes are well-characterized precursors for macrophages and dendritic cells. Subsets of human monocytes with differential representation in various disease states are well known. In contrast, mouse monocyte subsets have been characterized minimally. In this study we identify three subpopulations of mouse monocytes that can be distinguished by differential expression of Ly-6C, CD43, CD11c, MBR, and CD62L. The subsets share the characteristics of extensive phagocytosis, similar expression of M-CSF receptor (CD115), and development into macrophages upon M-CSF stimulation. By eliminating blood monocytes with dichloromethylene-bisphosphonate-loaded liposomes and monitoring their repopulation, we showed a developmental relationship between the subsets. Monocytes were maximally depleted 18 h after liposome application and subsequently reappeared in the circulation. These cells were exclusively of the Ly-6C(high) subset, resembling bone marrow monocytes. Serial flow cytometric analyses of newly released Ly-6C(high) monocytes showed that Ly-6C expression on these cells was down-regulated while in circulation. Under inflammatory conditions elicited either by acute infection with Listeria monocytogenes or chronic infection with Leishmania major, there was a significant increase in immature Ly-6C(high) monocytes, resembling the inflammatory left shift of granulocytes. In addition, acute peritoneal inflammation recruited preferentially Ly-6C(med-high) monocytes. Taken together, these data identify distinct subpopulations of mouse blood monocytes that differ in maturation stage and capacity to become recruited to inflammatory sites.

G-quadruplex (G4)-forming genomic sequences, including telomeres, represent natural replication fork barriers. Stalled replication forks can be stabilized and restarted by homologous recombination (HR), which also repairs DNA double-strand breaks (DSBs) arising at collapsed forks. We have previously shown that HR facilitates telomere replication. Here, we demonstrate that the replication efficiency of guanine-rich (G-rich) telomeric repeats is decreased significantly in cells lacking HR. Treatment with the G4-stabilizing compound pyridostatin (PDS) increases telomere fragility in BRCA2-deficient cells, suggesting that G4 formation drives telomere instability. Remarkably, PDS reduces proliferation of HR-defective cells by inducing DSB accumulation, checkpoint activation, and deregulated G2/M progression and by enhancing the replication defect intrinsic to HR deficiency. PDS toxicity extends to HR-defective cells that have acquired olaparib resistance through loss of 53BP1 or REV7. Altogether, these results highlight the therapeutic potential of G4-stabilizing drugs to selectively eliminate HR-compromised cells and tumors, including those resistant to PARP inhibition.


Question:
A breast cancer patient's capacity to metabolize tamoxifen influences treatment outcome.

CONTEXT The growth inhibitory effect of tamoxifen, which is used for the treatment of hormone receptor-positive breast cancer, is mediated by its metabolites, 4-hydroxytamoxifen and endoxifen. The formation of active metabolites is catalyzed by the polymorphic cytochrome P450 2D6 (CYP2D6) enzyme. OBJECTIVE To determine whether CYP2D6 variation is associated with clinical outcomes in women receiving adjuvant tamoxifen. DESIGN, SETTING, AND PATIENTS Retrospective analysis of German and US cohorts of patients treated with adjuvant tamoxifen for early stage breast cancer. The 1325 patients had diagnoses between 1986 and 2005 of stage I through III breast cancer and were mainly postmenopausal (95.4%). Last follow-up was in December 2008; inclusion criteria were hormone receptor positivity, no metastatic disease at diagnosis, adjuvant tamoxifen therapy, and no chemotherapy. DNA from tumor tissue or blood was genotyped for CYP2D6 variants associated with reduced (*10, *41) or absent (*3, *4, *5) enzyme activity. Women were classified as having an extensive (n=609), heterozygous extensive/intermediate (n=637), or poor (n=79) CYP2D6 metabolism. MAIN OUTCOME MEASURES Time to recurrence, event-free survival, disease-free survival, and overall survival. RESULTS Median follow-up was 6.3 years. At 9 years of follow-up, the recurrence rates were 14.9% for extensive metabolizers, 20.9% for heterozygous extensive/intermediate metabolizers, and 29.0% for poor metabolizers, and all-cause mortality rates were 16.7%, 18.0%, and 22.8%, respectively. Compared with extensive metabolizers, there was a significantly increased risk of recurrence for heterozygous extensive/intermediate metabolizers (time to recurrence adjusted hazard ratio [HR], 1.40; 95% confidence interval [CI], 1.04-1.90) and for poor metabolizers (time to recurrence HR, 1.90; 95% CI, 1.10-3.28). Compared with extensive metabolizers, those with decreased CYP2D6 activity (heterozygous extensive/intermediate and poor metabolism) had worse event-free survival (HR, 1.33; 95% CI, 1.06-1.68) and disease-free survival (HR, 1.29; 95% CI, 1.03-1.61), but there was no significant difference in overall survival (HR, 1.15; 95% CI, 0.88-1.51). CONCLUSION Among women with breast cancer treated with tamoxifen, there was an association between CYP2D6 variation and clinical outcomes, such that the presence of 2 functional CYP2D6 alleles was associated with better clinical outcomes and the presence of nonfunctional or reduced-function alleles with worse outcomes.


F4/80+ macrophages regulate mature osteoblast maintenance.

OBJECTIVE To determine the effectiveness and cost effectiveness of using information from circulating biomarkers to inform the prioritisation process of patients with stable angina awaiting coronary artery bypass graft surgery. DESIGN Decision analytical model comparing four prioritisation strategies without biomarkers (no formal prioritisation, two urgency scores, and a risk score) and three strategies based on a risk score using biomarkers: a routinely assessed biomarker (estimated glomerular filtration rate), a novel biomarker (C reactive protein), or both. The order in which to perform coronary artery bypass grafting in a cohort of patients was determined by each prioritisation strategy, and mean lifetime costs and quality adjusted life years (QALYs) were compared. DATA SOURCES Swedish Coronary Angiography and Angioplasty Registry (9935 patients with stable angina awaiting coronary artery bypass grafting and then followed up for cardiovascular events after the procedure for 3.8 years), and meta-analyses of prognostic effects (relative risks) of biomarkers. RESULTS The observed risk of cardiovascular events while on the waiting list for coronary artery bypass grafting was 3 per 10,000 patients per day within the first 90 days (184 events in 9935 patients). Using a cost effectiveness threshold of pound20,000- pound30,000 (euro22,000-euro33,000; $32,000-$48,000) per additional QALY, a prioritisation strategy using a risk score with estimated glomerular filtration rate was the most cost effective strategy (cost per additional QALY was < pound410 compared with the Ontario urgency score). The impact on population health of implementing this strategy was 800 QALYs per 100,000 patients at an additional cost of pound 245,000 to the National Health Service. The prioritisation strategy using a risk score with C reactive protein was associated with lower QALYs and higher costs compared with a risk score using estimated glomerular filtration rate. CONCLUSION Evaluating the cost effectiveness of prognostic biomarkers is important even when effects at an individual level are small. Formal prioritisation of patients awaiting coronary artery bypass grafting using a routinely assessed biomarker (estimated glomerular filtration rate) along with simple, routinely collected clinical information was cost effective. Prioritisation strategies based on the prognostic information conferred by C reactive protein, which is not currently measured in this context, or a combination of C reactive protein and estimated glomerular filtration rate, is unlikely to be cost effective. The widespread practice of using only implicit or informal means of clinically ordering the waiting list may be harmful and should be replaced with formal prioritisation approaches.

Blind individuals often demonstrate enhanced nonvisual perceptual abilities. However, the neural substrate that underlies this improved performance remains to be fully understood. An earlier behavioral study demonstrated that some early-blind people localize sounds more accurately than sighted controls using monaural cues. In order to investigate the neural basis of these behavioral differences in humans, we carried out functional imaging studies using positron emission tomography and a speaker array that permitted pseudo-free-field presentations within the scanner. During binaural sound localization, a sighted control group showed decreased cerebral blood flow in the occipital lobe, which was not seen in early-blind individuals. During monaural sound localization (one ear plugged), the subgroup of early-blind subjects who were behaviorally superior at sound localization displayed two activation foci in the occipital cortex. This effect was not seen in blind persons who did not have superior monaural sound localization abilities, nor in sighted individuals. The degree of activation of one of these foci was strongly correlated with sound localization accuracy across the entire group of blind subjects. The results show that those blind persons who perform better than sighted persons recruit occipital areas to carry out auditory localization under monaural conditions. We therefore conclude that computations carried out in the occipital cortex specifically underlie the enhanced capacity to use monaural cues. Our findings shed light not only on intermodal compensatory mechanisms, but also on individual differences in these mechanisms and on inhibitory patterns that differ between sighted individuals and those deprived of vision early in life.

Objective To explore the in vitro maintenance and characterization of human embryonic stem cells(hESCs).Methods hESCs were cultured on feeder layer with ES culture medium,which consists of 20% Knockout Serum Replacement,Knockout DMEM and 10 ng/mL bFGF.Undifferentiated status of hESCs was identified by cell morphology,and the expressions of cell surface marker SSEA-1,SSEA-3 and TRA-1-60.G banding technique was employed for cell karyotype analysis. Pluropotency of cells were analyzed via in vitro embyoid body(EB) formation and in vivo terotoma formation. Results Most of cells showed undifferentiated properties in cell morphology and normal karyotype throughout extended culture periods. They maintained undifferentiated status with positive immunoreactivity to SSEA-3,SSEA-4 and TRA-1-60.in vitro EB formation and in vivo teratoma formation demonstrated the pluripotency of human ES cells. Conclusion The fundamental requirement to hESCs for research and clinical application were their undifferentiated status and pluropotency in culture. Our result demonstrated their potential for these purposes.

Exosomes are emerging mediators of intercellular communication; whether the release of exosomes has an effect on the exosome donor cells in addition to the recipient cells has not been investigated to any extent. Here, we examine different exosomal miRNA expression profiles in primary mouse colon tumour, liver metastasis of colon cancer and naive colon tissues. In more advanced disease, higher levels of tumour suppressor miRNAs are encapsulated in the exosomes. miR-193a interacts with major vault protein (MVP). Knockout of MVP leads to miR-193a accumulation in the exosomal donor cells instead of exosomes, inhibiting tumour progression. Furthermore, miR-193a causes cell cycle G1 arrest and cell proliferation repression through targeting of Caprin1, which upregulates Ccnd2 and c-Myc. Human colon cancer patients with more advanced disease show higher levels of circulating exosomal miR-193a. In summary, our data demonstrate that MVP-mediated selective sorting of tumour suppressor miRNA into exosomes promotes tumour progression.

BACKGROUND High body-mass index (BMI) predisposes to several site-specific cancers, but a large-scale systematic and detailed characterisation of patterns of risk across all common cancers adjusted for potential confounders has not previously been undertaken. We aimed to investigate the links between BMI and the most common site-specific cancers. METHODS With primary care data from individuals in the Clinical Practice Research Datalink with BMI data, we fitted Cox models to investigate associations between BMI and 22 of the most common cancers, adjusting for potential confounders. We fitted linear then non-linear (spline) models; investigated effect modification by sex, menopausal status, smoking, and age; and calculated population effects. FINDINGS 5·24 million individuals were included; 166,955 developed cancers of interest. BMI was associated with 17 of 22 cancers, but effects varied substantially by site. Each 5 kg/m(2) increase in BMI was roughly linearly associated with cancers of the uterus (hazard ratio [HR] 1·62, 99% CI 1·56-1·69; p<0·0001), gallbladder (1·31, 1·12-1·52; p<0·0001), kidney (1·25, 1·17-1·33; p<0·0001), cervix (1·10, 1·03-1·17; p=0·00035), thyroid (1·09, 1·00-1·19; p=0·0088), and leukaemia (1·09, 1·05-1·13; p≤0·0001). BMI was positively associated with liver (1·19, 1·12-1·27), colon (1·10, 1·07-1·13), ovarian (1·09, 1.04-1.14), and postmenopausal breast cancers (1·05, 1·03-1·07) overall (all p<0·0001), but these effects varied by underlying BMI or individual-level characteristics. We estimated inverse associations with prostate and premenopausal breast cancer risk, both overall (prostate 0·98, 0·95-1·00; premenopausal breast cancer 0·89, 0·86-0·92) and in never-smokers (prostate 0·96, 0·93-0·99; premenopausal breast cancer 0·89, 0·85-0·94). By contrast, for lung and oral cavity cancer, we observed no association in never smokers (lung 0·99, 0·93-1·05; oral cavity 1·07, 0·91-1·26): inverse associations overall were driven by current smokers and ex-smokers, probably because of residual confounding by smoking amount. Assuming causality, 41% of uterine and 10% or more of gallbladder, kidney, liver, and colon cancers could be attributable to excess weight. We estimated that a 1 kg/m(2) population-wide increase in BMI would result in 3790 additional annual UK patients developing one of the ten cancers positively associated with BMI. INTERPRETATION BMI is associated with cancer risk, with substantial population-level effects. The heterogeneity in the effects suggests that different mechanisms are associated with different cancer sites and different patient subgroups. FUNDING National Institute for Health Research, Wellcome Trust, and Medical Research Council.

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.


Question:
CCL19 is predominantly present within dLNs.

Defective brain hormonal signaling has been associated with Alzheimer's disease (AD), a disorder characterized by synapse and memory failure. Irisin is an exercise-induced myokine released on cleavage of the membrane-bound precursor protein fibronectin type III domain-containing protein 5 (FNDC5), also expressed in the hippocampus. Here we show that FNDC5/irisin levels are reduced in AD hippocampi and cerebrospinal fluid, and in experimental AD models. Knockdown of brain FNDC5/irisin impairs long-term potentiation and novel object recognition memory in mice. Conversely, boosting brain levels of FNDC5/irisin rescues synaptic plasticity and memory in AD mouse models. Peripheral overexpression of FNDC5/irisin rescues memory impairment, whereas blockade of either peripheral or brain FNDC5/irisin attenuates the neuroprotective actions of physical exercise on synaptic plasticity and memory in AD mice. By showing that FNDC5/irisin is an important mediator of the beneficial effects of exercise in AD models, our findings place FNDC5/irisin as a novel agent capable of opposing synapse failure and memory impairment in AD.

Chemokines orchestrate immune cell trafficking by eliciting either directed or random migration and by activating integrins in order to induce cell adhesion. Analyzing dendritic cell (DC) migration, we showed that these distinct cellular responses depended on the mode of chemokine presentation within tissues. The surface-immobilized form of the chemokine CCL21, the heparan sulfate-anchoring ligand of the CC-chemokine receptor 7 (CCR7), caused random movement of DCs that was confined to the chemokine-presenting surface because it triggered integrin-mediated adhesion. Upon direct contact with CCL21, DCs truncated the anchoring residues of CCL21, thereby releasing it from the solid phase. Soluble CCL21 functionally resembles the second CCR7 ligand, CCL19, which lacks anchoring residues and forms soluble gradients. Both soluble CCR7 ligands triggered chemotactic movement, but not surface adhesion. Adhesive random migration and directional steering cooperate to produce dynamic but spatially restricted locomotion patterns closely resembling the cellular dynamics observed in secondary lymphoid organs.

CONTEXT Physical inactivity contributes to weight gain in adults, but whether this relationship is true for children of different ethnic groups is not well established. OBJECTIVE To assess participation in vigorous activity and television watching habits and their relationship to body weight and fatness in US children. DESIGN Nationally representative cross-sectional survey with an in-person interview and medical examination. SETTING AND PARTICIPANTS Between 1988 and 1994, 4063 children aged 8 through 16 years were examined as part of the National Health and Nutrition Examination Survey III. Mexican Americans and non-Hispanic blacks were oversampled to produce reliable estimates for these groups. MAIN OUTCOME MEASURES Episodes of weekly vigorous activity and daily hours of television watched, and their relationship to body mass index and body fatness. RESULTS Eighty percent of US children reported performing 3 or more bouts of vigorous activity each week. This rate was lower in non-Hispanic black and Mexican American girls (69% and 73%, respectively). Twenty percent of US children participated in 2 or fewer bouts of vigorous activity perweek, and the rate was higher in girls (26%) than in boys (17%). Overall, 26% of US children watched 4 or more hours of television per day and 67% watched at least 2 hours per day. Non-Hispanic black children had the highest rates of watching 4 or more hours of television per day (42%). Boys and girls who watch 4 or more hours of television each day had greater body fat (P<.001) and had a greater body mass index (P<.001) than those who watched less than 2 hours per day. CONCLUSIONS Many US children watch a great deal of television and are inadequately vigorously active. Vigorous activity levels are lowest among girls, non-Hispanic blacks, and Mexican Americans. Intervention strategies to promote lifelong physical activity among US children are needed to stem the adverse health consequences of inactivity.

During fasting, mammals maintain normal glucose homeostasis by stimulating hepatic gluconeogenesis. Elevations in circulating glucagon and epinephrine, two hormones that activate hepatic gluconeogenesis, trigger the cAMP-mediated phosphorylation of cAMP response element-binding protein (Creb) and dephosphorylation of the Creb-regulated transcription coactivator-2 (Crtc2)--two key transcriptional regulators of this process. Although the underlying mechanism is unclear, hepatic gluconeogenesis is also regulated by the circadian clock, which coordinates glucose metabolism with changes in the external environment. Circadian control of gene expression is achieved by two transcriptional activators, Clock and Bmal1, which stimulate cryptochrome (Cry1 and Cry2) and Period (Per1, Per2 and Per3) repressors that feed back on Clock-Bmal1 activity. Here we show that Creb activity during fasting is modulated by Cry1 and Cry2, which are rhythmically expressed in the liver. Cry1 expression was elevated during the night-day transition, when it reduced fasting gluconeogenic gene expression by blocking glucagon-mediated increases in intracellular cAMP concentrations and in the protein kinase A-mediated phosphorylation of Creb. In biochemical reconstitution studies, we found that Cry1 inhibited accumulation of cAMP in response to G protein-coupled receptor (GPCR) activation but not to forskolin, a direct activator of adenyl cyclase. Cry proteins seemed to modulate GPCR activity directly through interaction with G(s)α. As hepatic overexpression of Cry1 lowered blood glucose concentrations and improved insulin sensitivity in insulin-resistant db/db mice, our results suggest that compounds that enhance cryptochrome activity may provide therapeutic benefit to individuals with type 2 diabetes.

Combinatorial transcription factor (TF) interactions control cellular phenotypes and, therefore, underpin stem cell formation, maintenance, and differentiation. Here, we report the genome-wide binding patterns and combinatorial interactions for ten key regulators of blood stem/progenitor cells (SCL/TAL1, LYL1, LMO2, GATA2, RUNX1, MEIS1, PU.1, ERG, FLI-1, and GFI1B), thus providing the most comprehensive TF data set for any adult stem/progenitor cell type to date. Genome-wide computational analysis of complex binding patterns, followed by functional validation, revealed the following: first, a previously unrecognized combinatorial interaction between a heptad of TFs (SCL, LYL1, LMO2, GATA2, RUNX1, ERG, and FLI-1). Second, we implicate direct protein-protein interactions between four key regulators (RUNX1, GATA2, SCL, and ERG) in stabilizing complex binding to DNA. Third, Runx1(+/-)::Gata2(+/-) compound heterozygous mice are not viable with severe hematopoietic defects at midgestation. Taken together, this study demonstrates the power of genome-wide analysis in generating novel functional insights into the transcriptional control of stem and progenitor cells.


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

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.

Pluripotent stem cells exist in naive and primed states, epitomized by mouse embryonic stem cells (ESCs) and the developmentally more advanced epiblast stem cells (EpiSCs; ref. ). In the naive state of ESCs, the genome has an unusual open conformation and possesses a minimum of repressive epigenetic marks. In contrast, EpiSCs have activated the epigenetic machinery that supports differentiation towards the embryonic cell types. The transition from naive to primed pluripotency therefore represents a pivotal event in cellular differentiation. But the signals that control this fundamental differentiation step remain unclear. We show here that paracrine and autocrine Wnt signals are essential self-renewal factors for ESCs, and are required to inhibit their differentiation into EpiSCs. Moreover, we find that Wnt proteins in combination with the cytokine LIF are sufficient to support ESC self-renewal in the absence of any undefined factors, and support the derivation of new ESC lines, including ones from non-permissive mouse strains. Our results not only demonstrate that Wnt signals regulate the naive-to-primed pluripotency transition, but also identify Wnt as an essential and limiting ESC self-renewal factor.

The tastes of sugars (sweet) and glutamate (umami) are thought to be detected by T1r receptors expressed in taste cells. Molecular genetics and heterologous expression implicate T1r2 plus T1r3 as a sweet-responsive receptor,and T1r1 plus T1r3,as well as a truncated form of the type 4 metabotropic glutamate receptor (taste-mGluR4),as umami-responsive receptors. Here,we show that mice lacking T1r3 showed no preference for artificial sweeteners and had diminished but not abolished behavioral and nerve responses to sugars and umami compounds. These results indicate that T1r3-independent sweet- and umami-responsive receptors and/or pathways exist in taste cells.

This paper seeks to establish the strength of association between contemporary tuberculosis (TB) in England and Wales and several potential aetiological factors. It presents an ecological analysis of standardised annual TB mortality rates for the 403 local authority districts between 1982 and 1992, disaggregated by age and sex. Social, demographic and ethnicity measures from the 1981 and 1991 censuses and standardised annual AIDS-related mortality rates for young men are used to calculate Poisson regression models. A strong association was found between all TB mortality groups and overcrowding at the household level. For women, no other measures improved the explanatory power of the models. In multiple regressions, both poverty and AIDS-related mortality explained additional variation in the model for younger men. The link between ethnicity and tuberculosis notifications was not reflected in this analysis of mortality. For all groups no evidence of a positive relationship with ethnicity was found, once overcrowding had been accounted for. The significance of household as opposed to district level crowding suggests that prolonged contact is required for disease transmission. Regression analysis indicates that it is the overcrowding and poverty among ethnic populations that is significant for their tuberculosis mortality. The fact that the relationship between AIDS and TB is confined to the group most typical of AIDS patients provides evidence that AIDS has little influence on the level of tuberculosis mortality in the wider population. Explanations for the observed relationship include preferential certification, migration for treatment and shortcomings in health care provision.


Allogeneic mechanical circulatory support is not as effective as autologous mechanical circulatory support for treating acute myocardial infarction.

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.

The purpose of this study was to examine differences in resting, submaximal, and maximal (VO2max) oxygen consumption (VO2) in African-American (n = 44) and Caucasian (n = 31) prepubertal children aged 5-10 yr. Resting VO2 was measured via indirect calorimetry in the fasted state. Submaximal VO2 and VO2max were determined during an all out, progressive treadmill exercise test appropriate for children. Dual-energy X-ray absorptiometry was used to determine total fat mass (FM), soft lean tissue mass (LTM), and leg soft LTM. Doubly labeled water was used to determine total energy expenditure (TEE) and activity energy expenditure (AEE). A significant effect of ethnicity (P < 0.01) was found for VO2max but not resting or submaximal VO2, with African-American children having absolute VO2max approximately 15% lower than Caucasian children (1.21 +/- 0.032 vs. 1.43 +/- 0.031 l/min, respectively). The lower VO2max persisted in African-American children after adjustment for soft LTM (1.23 +/- 0.025 vs. 1.39 +/- 0.031 l/min; P < 0.01), leg soft LTM (1.20 +/- 0.031 vs. 1.43 +/- 0.042 l/min; P < 0.01), and soft LTM and FM (1.23 +/- 0.025 vs. 1.39 +/- 0.031 l/min; P < 0.01). The lower VO2max persisted also after adjustment for TEE (1.20 +/- 0.02 vs. 1.38 +/- 0.0028 l/min P < 0.001) and AEE (1.20 +/- 0.024 vs. 1.38 +/- 0.028 l/min; P < 0.001). In conclusion, our data indicate that African-American and Caucasian children have similar rates of VO2 at rest and during submaximal exercise, but VO2max is approximately 15% lower in African-American children, independent of soft LTM, FM, leg LTM, TEE, and AEE.

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.

CONTEXT Mesenchymal stem cells (MSCs) are under evaluation as a therapy for ischemic cardiomyopathy (ICM). Both autologous and allogeneic MSC therapies are possible; however, their safety and efficacy have not been compared. OBJECTIVE To test whether allogeneic MSCs are as safe and effective as autologous MSCs in patients with left ventricular (LV) dysfunction due to ICM. DESIGN, SETTING, AND PATIENTS A phase 1/2 randomized comparison (POSEIDON study) in a US tertiary-care referral hospital of allogeneic and autologous MSCs in 30 patients with LV dysfunction due to ICM between April 2, 2010, and September 14, 2011, with 13-month follow-up. INTERVENTION Twenty million, 100 million, or 200 million cells (5 patients in each cell type per dose level) were delivered by transendocardial stem cell injection into 10 LV sites. MAIN OUTCOME MEASURES Thirty-day postcatheterization incidence of predefined treatment-emergent serious adverse events (SAEs). Efficacy assessments included 6-minute walk test, exercise peak VO2, Minnesota Living with Heart Failure Questionnaire (MLHFQ), New York Heart Association class, LV volumes, ejection fraction (EF), early enhancement defect (EED; infarct size), and sphericity index. RESULTS Within 30 days, 1 patient in each group (treatment-emergent SAE rate, 6.7%) was hospitalized for heart failure, less than the prespecified stopping event rate of 25%. The 1-year incidence of SAEs was 33.3% (n = 5) in the allogeneic group and 53.3% (n = 8) in the autologous group (P = .46). At 1 year, there were no ventricular arrhythmia SAEs observed among allogeneic recipients compared with 4 patients (26.7%) in the autologous group (P = .10). Relative to baseline, autologous but not allogeneic MSC therapy was associated with an improvement in the 6-minute walk test and the MLHFQ score, but neither improved exercise VO2 max. Allogeneic and autologous MSCs reduced mean EED by −33.21% (95% CI, −43.61% to −22.81%; P < .001) and sphericity index but did not increase EF. Allogeneic MSCs reduced LV end-diastolic volumes. Low-dose concentration MSCs (20 million cells) produced greatest reductions in LV volumes and increased EF. Allogeneic MSCs did not stimulate significant donor-specific alloimmune reactions. CONCLUSIONS In this early-stage study of patients with ICM, transendocardial injection of allogeneic and autologous MSCs without a placebo control were both associated with low rates of treatment-emergent SAEs, including immunologic reactions. In aggregate, MSC injection favorably affected patient functional capacity, quality of life, and ventricular remodeling. TRIAL REGISTRATION clinicaltrials.gov Identifier: NCT01087996.

Fever is commonly used to diagnose disease and is consistently associated with increased mortality in critically ill patients. However, the molecular controls of elevated body temperature are poorly understood. We discovered that the expression of RNA-binding motif protein 3 (RBM3), known to respond to cold stress and to modulate microRNA (miRNA) expression, was reduced in 30 patients with fever, and in THP-1-derived macrophages maintained at a fever-like temperature (40 °C). Notably, RBM3 expression is reduced during fever whether or not infection is demonstrable. Reduced RBM3 expression resulted in increased expression of RBM3-targeted temperature-sensitive miRNAs, we termed thermomiRs. ThermomiRs such as miR-142-5p and miR-143 in turn target endogenous pyrogens including IL-6, IL6ST, TLR2, PGE2 and TNF to complete a negative feedback mechanism, which may be crucial to prevent pathological hyperthermia. Using normal PBMCs that were exogenously exposed to fever-like temperature (40 °C), we further demonstrate the trend by which decreased levels of RBM3 were associated with increased levels of miR-142-5p and miR-143 and vice versa over a 24 h time course. Collectively, our results indicate the existence of a negative feedback loop that regulates fever via reduced RBM3 levels and increased expression of miR-142-5p and miR-143.

Cardiovascular disease is a leading cause of death worldwide. The limited capability of heart tissue to regenerate has prompted methodological developments for creating de novo cardiomyocytes, both in vitro and in vivo. Beyond uses in cell replacement therapy, patient-specific cardiomyocytes may find applications in drug testing, drug discovery, and disease modeling. Recently, approaches for generating cardiomyocytes have expanded to encompass three major sources of starting cells: human pluripotent stem cells (hPSCs), adult heart-derived cardiac progenitor cells (CPCs), and reprogrammed fibroblasts. We discuss state-of-the-art methods for generating de novo cardiomyocytes from hPSCs and reprogrammed fibroblasts, highlighting potential applications and future challenges.


Formation of N-terminal pyroglutamate by glutamine cyclase (GC) competes with NTAQ1 for Nt-Gln substrates.

Combinatorial transcription factor (TF) interactions control cellular phenotypes and, therefore, underpin stem cell formation, maintenance, and differentiation. Here, we report the genome-wide binding patterns and combinatorial interactions for ten key regulators of blood stem/progenitor cells (SCL/TAL1, LYL1, LMO2, GATA2, RUNX1, MEIS1, PU.1, ERG, FLI-1, and GFI1B), thus providing the most comprehensive TF data set for any adult stem/progenitor cell type to date. Genome-wide computational analysis of complex binding patterns, followed by functional validation, revealed the following: first, a previously unrecognized combinatorial interaction between a heptad of TFs (SCL, LYL1, LMO2, GATA2, RUNX1, ERG, and FLI-1). Second, we implicate direct protein-protein interactions between four key regulators (RUNX1, GATA2, SCL, and ERG) in stabilizing complex binding to DNA. Third, Runx1(+/-)::Gata2(+/-) compound heterozygous mice are not viable with severe hematopoietic defects at midgestation. Taken together, this study demonstrates the power of genome-wide analysis in generating novel functional insights into the transcriptional control of stem and progenitor cells.

Cigarette smoking promotes body weight reduction in humans while paradoxically also promoting insulin resistance (IR) and hyperinsulinemia. However, the mechanisms behind these effects are unclear. Here we show that nicotine, a major constituent of cigarette smoke, selectively activates AMP-activated protein kinase α2 (AMPKα2) in adipocytes, which in turn phosphorylates MAP kinase phosphatase-1 (MKP1) at serine 334, initiating its proteasome-dependent degradation. The nicotine-dependent reduction of MKP1 induces the aberrant activation of both p38 mitogen-activated protein kinase and c-Jun N-terminal kinase, leading to increased phosphorylation of insulin receptor substrate 1 (IRS1) at serine 307. Phosphorylation of IRS1 leads to its degradation, protein kinase B inhibition, and the loss of insulin-mediated inhibition of lipolysis. Consequently, nicotine increases lipolysis, which results in body weight reduction, but this increase also elevates the levels of circulating free fatty acids and thus causes IR in insulin-sensitive tissues. These results establish AMPKα2 as an essential mediator of nicotine-induced whole-body IR in spite of reductions in adiposity.

Organ-specific functions of tissue-resident macrophages in the steady-state heart are unknown. Here, we show that cardiac macrophages facilitate electrical conduction through the distal atrioventricular node, where conducting cells densely intersperse with elongated macrophages expressing connexin 43. When coupled to spontaneously beating cardiomyocytes via connexin-43-containing gap junctions, cardiac macrophages have a negative resting membrane potential and depolarize in synchrony with cardiomyocytes. Conversely, macrophages render the resting membrane potential of cardiomyocytes more positive and, according to computational modeling, accelerate their repolarization. Photostimulation of channelrhodopsin-2-expressing macrophages improves atrioventricular conduction, whereas conditional deletion of connexin 43 in macrophages and congenital lack of macrophages delay atrioventricular conduction. In the Cd11bDTR mouse, macrophage ablation induces progressive atrioventricular block. These observations implicate macrophages in normal and aberrant cardiac conduction.

BACKGROUND The MTHFR 677C→T polymorphism has been associated with raised homocysteine concentration and increased risk of stroke. A previous overview showed that the effects were greatest in regions with low dietary folate consumption, but differentiation between the effect of folate and small-study bias was difficult. A meta-analysis of randomised trials of homocysteine-lowering interventions showed no reduction in coronary heart disease events or stroke, but the trials were generally set in populations with high folate consumption. We aimed to reduce the effect of small-study bias and investigate whether folate status modifies the association between MTHFR 677C→T and stroke in a genetic analysis and meta-analysis of randomised controlled trials. METHODS We established a collaboration of genetic studies consisting of 237 datasets including 59,995 individuals with data for homocysteine and 20,885 stroke events. We compared the genetic findings with a meta-analysis of 13 randomised trials of homocysteine-lowering treatments and stroke risk (45,549 individuals, 2314 stroke events, 269 transient ischaemic attacks). FINDINGS The effect of the MTHFR 677C→T variant on homocysteine concentration was larger in low folate regions (Asia; difference between individuals with TT versus CC genotype, 3·12 μmol/L, 95% CI 2·23 to 4·01) than in areas with folate fortification (America, Australia, and New Zealand, high; 0·13 μmol/L, -0·85 to 1·11). The odds ratio (OR) for stroke was also higher in Asia (1·68, 95% CI 1·44 to 1·97) than in America, Australia, and New Zealand, high (1·03, 0·84 to 1·25). Most randomised trials took place in regions with high or increasing population folate concentrations. The summary relative risk (RR) of stroke in trials of homocysteine-lowering interventions (0·94, 95% CI 0·85 to 1·04) was similar to that predicted for the same extent of homocysteine reduction in large genetic studies in populations with similar folate status (predicted RR 1·00, 95% CI 0·90 to 1·11). Although the predicted effect of homocysteine reduction from large genetic studies in low folate regions (Asia) was larger (RR 0·78, 95% CI 0·68 to 0·90), no trial has evaluated the effect of lowering of homocysteine on stroke risk exclusively in a low folate region. INTERPRETATION In regions with increasing levels or established policies of population folate supplementation, evidence from genetic studies and randomised trials is concordant in suggesting an absence of benefit from lowering of homocysteine for prevention of stroke. Further large-scale genetic studies of the association between MTHFR 677C→T and stroke in low folate settings are needed to distinguish effect modification by folate from small-study bias. If future randomised trials of homocysteine-lowering interventions for stroke prevention are undertaken, they should take place in regions with low folate consumption. FUNDING Full funding sources listed at end of paper (see Acknowledgments).

OBJECTIVE To investigate the long term effect of radioactive iodine on thyroid function and size in patients with non-toxic multinodular goitre. DESIGN Consecutive patients with multinodular non-toxic goitre selected for radioactive iodine treatment and followed for a minimum of 12 months (median 48 months) after an intended dose of 3.7 MBq/g thyroid tissue corrected to a 100% uptake of iodine-131 in 24 hours. PATIENTS 69 patients with a growing multinodular non-toxic goitre causing local compression symptoms or cosmetic inconveniences. The treatment was chosen because of a high operative risk, previous thyroidectomy, or refusal to be operated on. MAIN OUTCOME MEASUREMENTS Standard thyroid function variables and ultrasonically determined thyroid volume before treatment as well as 1, 2, 3, 6, and 12 months after treatment and then once a year. RESULTS 56 patients were treated with a single dose of 131I, 12 with two doses, and one with four doses. In 45 patients treated with one dose and remaining euthyroid the median thyroid volume was reduced from 73 (interquartile range 50-106) ml to 29 (23-48) ml at 24 months in the 39 patients in whom this was measured during follow up. The median reduction was 40 (22-48) ml (60% reduction, p < 0.0001), half of which occurred within three months. Patients treated with two doses as well as those developing hypothyroidism and hyperthyroidism had a significant reduction in thyroid volume. Eleven patients developed hypothyroidism (cumulative five year risk 22%, 95% confidence interval 4.8% to 38.4%). Side effects were few: three cases of hyperthyroidism and two cases of radiation thyroiditis. Only one patient was dissatisfied with the result; she was referred for operation six months after treatment. CONCLUSIONS A substantial reduction in thyroid volume accompanied by a low incidence of hypothyroidism and few side effects makes the use of radioactive iodine an attractive alternative to surgery in selected cases of non-toxic multinodular goitre.

OBJECTIVE To determine whether there are risk factors in a doctor's time at medical school that are associated with subsequent professional misconduct. DESIGN Matched case-control study. Setting Records from medical schools and the General Medical Council (GMC). PARTICIPANTS 59 doctors who had graduated from any one of eight medical schools in the United Kingdom in 1958-97 and had a proved finding of serious professional misconduct in GMC proceedings in 1999-2004 (cases); 236 controls (four for each case) were selected by systematic sampling from matching graduation cohorts. Case-control status was revealed by the GMC after completion of data entry. MAIN OUTCOME MEASURE Odds ratios for being a "case," with multivariable conditional logistic regression of potential risk factors including pre-admission characteristics and progress during the course. These data were obtained from anonymised copies of the students' progress files held by their original medical schools. RESULTS Univariate conditional logistic regression analysis found that cases were more likely to be men, to be of lower estimated social class, and to have had academic difficulties during their medical course, especially in the early years. Multivariable analysis showed that male sex (odds ratio 9.80, 95% confidence interval 2.43 to 39.44, P=0.001), lower social class (4.28, 1.52 to 12.09, P=0.006), and failure of early or preclinical examinations (5.47, 2.17 to 13.79, P<0.001) were independently associated with being a case. CONCLUSIONS This small study suggests that male sex, a lower socioeconomic background, and early academic difficulties at medical school could be risk factors for subsequent professional misconduct. The findings are preliminary and should be interpreted with caution. Most doctors with risk factors will not come before the GMC's disciplinary panels.


Cognitive behavioral therapy is an ineffective treatment for insomnia

CONTEXT Insomnia is a common condition in older adults and is associated with a number of adverse medical, social, and psychological consequences. Previous research has suggested beneficial outcomes of both psychological and pharmacological treatments, but blinded placebo-controlled trials comparing the effects of these treatments are lacking. OBJECTIVE To examine short- and long-term clinical efficacy of cognitive behavioral therapy (CBT) and pharmacological treatment in older adults experiencing chronic primary insomnia. DESIGN, SETTING, AND PARTICIPANTS A randomized, double-blinded, placebo-controlled trial of 46 adults (mean age, 60.8 y; 22 women) with chronic primary insomnia conducted between January 2004 and December 2005 in a single Norwegian university-based outpatient clinic for adults and elderly patients. INTERVENTION CBT (sleep hygiene, sleep restriction, stimulus control, cognitive therapy, and relaxation; n = 18), sleep medication (7.5-mg zopiclone each night; n = 16), or placebo medication (n = 12). All treatment duration was 6 weeks, and the 2 active treatments were followed up at 6 months. MAIN OUTCOME MEASURES Ambulant clinical polysomnographic data and sleep diaries were used to determine total wake time, total sleep time, sleep efficiency, and slow-wave sleep (only assessed using polysomnography) on all 3 assessment points. RESULTS CBT resulted in improved short- and long-term outcomes compared with zopiclone on 3 out of 4 outcome measures. For most outcomes, zopiclone did not differ from placebo. Participants receiving CBT improved their sleep efficiency from 81.4% at pretreatment to 90.1% at 6-month follow-up compared with a decrease from 82.3% to 81.9% in the zopiclone group. Participants in the CBT group spent much more time in slow-wave sleep (stages 3 and 4) compared with those in other groups, and spent less time awake during the night. Total sleep time was similar in all 3 groups; at 6 months, patients receiving CBT had better sleep efficiency using polysomnography than those taking zopiclone. CONCLUSION These results suggest that interventions based on CBT are superior to zopiclone treatment both in short- and long-term management of insomnia in older adults. TRIAL REGISTRATION clinicaltrials.gov Identifier: NCT00295386.

Circadian rhythm sleep disorders are characterized by complaints of insomnia and excessive sleepiness that are primarily due to alterations in the internal circadian timing system or a misalignment between the timing of sleep and the 24-h social and physical environment. In addition to physiological and environmental factors, maladaptive behaviors often play an important role in the development of many of the circadian rhythm sleep disorders. This review will focus on the clinical approach to the diagnosis and management of the various circadian rhythm sleep disorders, including delayed sleep phase disorder, advanced sleep phase disorder, non-entrained type, irregular sleep-wake rhythm, shift work sleep disorder and jet lag disorder. Diagnostic tools such as sleep diaries and wrist activity monitoring are often useful in confirming the diagnosis. Because behavioral and environmental factors often are involved in the development of these conditions, a multimodal approach is usually necessary. Interventions include sleep hygiene education, timed exposure to bright light as well as avoidance of bright light at the wrong time of the day and pharmacologic approaches, such as melatonin. However, it should be noted that the use of melatonin is not an FDA-approved indication for the treatment of circadian rhythm sleep disorders.


Smc5/6 engagment halts the activation of SUMO E3 ligase Mms21 by ATP-dependent remolding.

Defined genetic models based on human pluripotent stem cells have opened new avenues for understanding disease mechanisms and drug screening. Many of these models assume cell-autonomous mechanisms of disease but it is possible that disease phenotypes or drug responses will only be evident if all cellular and extracellular components of a tissue are present and functionally mature. To derive optimal benefit from such models, complex multicellular structures with vascular components that mimic tissue niches will thus likely be necessary. Here we consider emerging research creating human tissue mimics and provide some recommendations for moving the field forward.

CONTEXT Findings from previous studies of the effects of exercise training on patient-reported health status have been inconsistent. OBJECTIVE To test the effects of exercise training on health status among patients with heart failure. DESIGN, SETTING, AND PATIENTS Multicenter, randomized controlled trial among 2331 medically stable outpatients with heart failure with left ventricular ejection fraction of 35% or less. Patients were randomized from April 2003 through February 2007. INTERVENTIONS Usual care plus aerobic exercise training (n = 1172), consisting of 36 supervised sessions followed by home-based training, vs usual care alone (n = 1159). Randomization was stratified by heart failure etiology, which was a covariate in all models. MAIN OUTCOME MEASURES Kansas City Cardiomyopathy Questionnaire (KCCQ) overall summary scale and key subscales at baseline, every 3 months for 12 months, and annually thereafter for up to 4 years. The KCCQ is scored from 0 to 100 with higher scores corresponding to better health status. Treatment group effects were estimated using linear mixed models according to the intention-to-treat principle. RESULTS Median follow-up was 2.5 years. At 3 months, usual care plus exercise training led to greater improvement in the KCCQ overall summary score (mean, 5.21; 95% confidence interval, 4.42 to 6.00) compared with usual care alone (3.28; 95% confidence interval, 2.48 to 4.09). The additional 1.93-point increase (95% confidence interval, 0.84 to 3.01) in the exercise training group was statistically significant (P < .001). After 3 months, there were no further significant changes in KCCQ score for either group (P = .85 for the difference between slopes), resulting in a sustained, greater improvement overall for the exercise group (P < .001). Results were similar on the KCCQ subscales, and no subgroup interactions were detected. CONCLUSIONS Exercise training conferred modest but statistically significant improvements in self-reported health status compared with usual care without training. Improvements occurred early and persisted over time. TRIAL REGISTRATION clinicaltrials.gov Identifier: NCT00047437.

Modification of proteins by SUMO is essential for the maintenance of genome integrity. During DNA replication, the Mms21-branch of the SUMO pathway counteracts recombination intermediates at damaged replication forks, thus facilitating sister chromatid disjunction. The Mms21 SUMO ligase docks to the arm region of the Smc5 protein in the Smc5/6 complex; together, they cooperate during recombinational DNA repair. Yet how the activity of the SUMO ligase is controlled remains unknown. Here we show that the SUMO ligase and the chromosome disjunction functions of Mms21 depend on its docking to an intact and active Smc5/6 complex, indicating that the Smc5/6-Mms21 complex operates as a large SUMO ligase in vivo. In spite of the physical distance separating the E3 and the nucleotide-binding domains in Smc5/6, Mms21-dependent sumoylation requires binding of ATP to Smc5, a step that is part of the ligase mechanism that assists Ubc9 function. The communication is enabled by the presence of a conserved disruption in the coiled coil domain of Smc5, pointing to potential conformational changes for SUMO ligase activation. In accordance, scanning force microscopy of the Smc5-Mms21 heterodimer shows that the molecule is physically remodeled in an ATP-dependent manner. Our results demonstrate that the ATP-binding activity of the Smc5/6 complex is coordinated with its SUMO ligase, through the coiled coil domain of Smc5 and the physical remodeling of the molecule, to promote sumoylation and chromosome disjunction during DNA repair.

Regenerative medicine is predicated on understanding the mechanisms regulating development and applying these conditions to direct stem cell fate. Embryogenesis is guided by cell-cell and cell-matrix interactions, but it is unclear how these physical cues influence stem cells in culture. We used human embryonic stem cells (hESCs) to examine whether mechanical features of the extracellular microenvironment could differentially modulate mesoderm specification. We found that, on a hydrogel-based compliant matrix, hESCs accumulate β-catenin at cell-cell adhesions and show enhanced Wnt-dependent mesoderm differentiation. Mechanistically, Src-driven ubiquitination of E-cadherin by Cbl-like ubiquitin ligase releases P120-catenin to facilitate transcriptional activity of β-catenin, which initiates and reinforces mesoderm differentiation. By contrast, on a stiff hydrogel matrix, hESCs show elevated integrin-dependent GSK3 and Src activity that promotes β-catenin degradation and inhibits differentiation. Thus, we found that mechanical features of the microenvironmental matrix influence tissue-specific differentiation of hESCs by altering the cellular response to morphogens.


Question:
Combination nicotine replacement therapies with varenicline or bupropion are more effective after 12 weeks of reatment compared to varenicline monotherapy.

IMPORTANCE Combining pharmacotherapies for tobacco-dependence treatment may increase smoking abstinence. OBJECTIVE To determine efficacy and safety of varenicline and bupropion sustained-release (SR; combination therapy) compared with varenicline (monotherapy) in cigarette smokers. DESIGN, SETTING, AND PARTICIPANTS Randomized, blinded, placebo-controlled multicenter clinical trial with a 12-week treatment period and follow-up through week 52 conducted between October 2009 and April 2013 at 3 midwestern clinical research sites. Five hundred six adult (≥18 years) cigarette smokers were randomly assigned and 315 (62%) completed the study. INTERVENTIONS Twelve weeks of varenicline and bupropion SR or varenicline and placebo. MAIN OUTCOMES AND MEASURES Primary outcome was abstinence rates at week 12, defined as prolonged (no smoking from 2 weeks after the target quit date) abstinence and 7-day point-prevalence (no smoking past 7 days) abstinence. Secondary outcomes were prolonged and point-prevalence smoking abstinence rates at weeks 26 and 52. Outcomes were biochemically confirmed. RESULTS At 12 weeks, 53.0% of the combination therapy group achieved prolonged smoking abstinence and 56.2% achieved 7-day point-prevalence smoking abstinence compared with 43.2% and 48.6% in varenicline monotherapy (odds ratio [OR], 1.49; 95% CI, 1.05-2.12; P = .03 and OR, 1.36; 95% CI, 0.95-1.93; P = .09, respectively). At 26 weeks, 36.6% of the combination therapy group achieved prolonged and 38.2% achieved 7-day point-prevalence smoking abstinence compared with 27.6% and 31.9% in varenicline monotherapy (OR, 1.52; 95% CI, 1.04-2.22; P = .03 and OR, 1.32; 95% CI, 0.91-1.91; P = .14, respectively). At 52 weeks, 30.9% of the combination therapy group achieved prolonged and 36.6% achieved 7-day point-prevalence smoking abstinence compared with 24.5% and 29.2% in varenicline monotherapy (OR, 1.39; 95% CI, 0.93-2.07; P = .11 and OR, 1.40; 95% CI, 0.96-2.05; P = .08, respectively). Participants receiving combination therapy reported more anxiety (7.2% vs 3.1%; P = .04) and depressive symptoms (3.6% vs 0.8%; P = .03). CONCLUSIONS AND RELEVANCE Among cigarette smokers, combined use of varenicline and bupropion, compared with varenicline alone, increased prolonged abstinence but not 7-day point prevalence at 12 and 26 weeks. Neither outcome was significantly different at 52 weeks. Further research is required to determine the role of combination therapy in smoking cessation. TRIAL REGISTRATION clinicaltrials.gov Identifier: http://clinicaltrials.gov/show/NCT00935818.

The pharmacological inhibition of general transcriptional regulators has the potential to block growth through targeting multiple tumorigenic signalling pathways simultaneously. Here, using an innovative cell-based screen, we identify a structurally unique small molecule (named JIB-04) that specifically inhibits the activity of the Jumonji family of histone demethylases in vitro, in cancer cells, and in tumours in vivo. Unlike known inhibitors, JIB-04 is not a competitive inhibitor of α-ketoglutarate. In cancer, but not in patient-matched normal cells, JIB-04 alters a subset of transcriptional pathways and blocks viability. In mice, JIB-04 reduces tumour burden and prolongs survival. Importantly, we find that patients with breast tumours that overexpress Jumonji demethylases have significantly lower survival. Thus, JIB-04, a novel inhibitor of Jumonji demethylases in vitro and in vivo, constitutes a unique potential therapeutic and research tool against cancer, and validates the use of unbiased cellular screens to discover chemical modulators with disease relevance.

OBJECTIVES We investigated the role of cytokines tumor necrosis factor-alpha (TNF-alpha) and interleukin-6 (IL-6) in cachexia development in newly diagnosed nonsmall cell lung cancer (NSCLC) patients. METHODS : We evaluated 44 (M/F:41/3) NSCLC patients and 12 (M/F:10/2) age matched healthy smokers. NSCLC cases with a weight loss of > or =10% consisted the cachectic group (n:23, M/F:21/2) and the ones with <10% weight loss consisted the noncachectic group (n:21, M/F:19/2). RESULTS Body mass index (BMI) of cachectics was significantly lower than that of noncachectics (21.0 +/- 2.9 versus 24.5 +/- 3.6, P = 0.02) and controls (21.0 +/- 2.9 versus 25.5 +/- 2.6, P = 0.01). Serum TNF-alpha level did not differ between cachectic and noncachectics (37.3 +/- 39.1 and 51.6 +/- 84.2 pg/mL, respectively). However, it was significantly higher in NSCLC patients compared with controls (44.1 +/- 64.3 and 15.1 +/- 14.3 pg/mL, P = 0.03). Serum IL-6 level was not different between 3 groups (6.4 +/- 4.1, 8.9 +/- 16.3, and 4.1 +/- 3.5 pg/mL, respectively) but it correlated significantly with TNF-alpha (r = 0.4, P = 0.006) and BMI (r = -0.3, P = 0.03). Erythrocyte sedimentation rate (ESR) correlated significantly with TNF-alpha (r = 0.4, P = 0.003) and BMI (r = -0.3, P = 0.03). Among 44 cases, survival of 12 and 17 patients was recorded in cachectics and noncachectics, with no statistical difference (12.2 +/- 3.7 and 11.2 +/- 1.0 months, respectively). CONCLUSIONS TNF-alpha and IL-6 levels did not differ significantly between cachectics and noncachectics. However, significant correlations between IL-6, BMI, and TNF-alpha suggested that these cytokines acted as cofactors in weight loss. Survival was neither influenced by BMI, nor the cytokine levels in the present study. The significant correlation of ESR with TNF-alpha suggested that ESR could provide valuable clue for considerable weight loss in the follow-up of NSCLC patients.


Reducing H3k4me3 methylation induces mouse epiblast stem cells to naive pluripotency efficiently.

The interconversion between naive and primed pluripotent states is accompanied by drastic epigenetic rearrangements. However, it is unclear whether intrinsic epigenetic events can drive reprogramming to naive pluripotency or if distinct chromatin states are instead simply a reflection of discrete pluripotent states. Here, we show that blocking histone H3K4 methyltransferase MLL1 activity with the small-molecule inhibitor MM-401 reprograms mouse epiblast stem cells (EpiSCs) to naive pluripotency. This reversion is highly efficient and synchronized, with more than 50% of treated EpiSCs exhibiting features of naive embryonic stem cells (ESCs) within 3 days. Reverted ESCs reactivate the silenced X chromosome and contribute to embryos following blastocyst injection, generating germline-competent chimeras. Importantly, blocking MLL1 leads to global redistribution of H3K4me1 at enhancers and represses lineage determinant factors and EpiSC markers, which indirectly regulate ESC transcription circuitry. These findings show that discrete perturbation of H3K4 methylation is sufficient to drive reprogramming to naive pluripotency.


Thiopurine active metabolites can be catabolized through dephosphorylation of thioguanine nucleotides.

OBJECTIVE To determine whether dietary n-3 long chain polyunsaturated fatty acid (LCPUFA) supplementation of pregnant women with a fetus at high risk of allergic disease reduces immunoglobulin E associated eczema or food allergy at 1 year of age. DESIGN Follow-up of infants at high hereditary risk of allergic disease in the Docosahexaenoic Acid to Optimise Mother Infant Outcome (DOMInO) randomised controlled trial. SETTING Adelaide, South Australia. PARTICIPANTS 706 infants at high hereditary risk of developing allergic disease whose mothers were participating in the DOMInO trial. INTERVENTIONS The intervention group (n=368) was randomly allocated to receive fish oil capsules (providing 900 mg of n-3 LCPUFA daily) from 21 weeks' gestation until birth; the control group (n=338) received matched vegetable oil capsules without n-3 LCPUFA. MAIN OUTCOME MEASURE Immunoglobulin E associated allergic disease (eczema or food allergy with sensitisation) at 1 year of age. RESULTS No differences were seen in the overall percentage of infants with immunoglobulin E associated allergic disease between the n-3 LCPUFA and control groups (32/368 (9%) v 43/338 (13%); unadjusted relative risk 0.68, 95% confidence interval 0.43 to 1.05, P=0.08; adjusted relative risk 0.70, 0.45 to 1.09, P=0.12), although the percentage of infants diagnosed as having atopic eczema (that is, eczema with associated sensitisation) was lower in the n-3 LCPUFA group (26/368 (7%) v 39/338 (12%); unadjusted relative risk 0.61, 0.38 to 0.98, P=0.04; adjusted relative risk 0.64, 0.40 to 1.02, P=0.06). Fewer infants were sensitised to egg in the n-3 LCPUFA group (34/368 (9%) v 52/338 (15%); unadjusted relative risk 0.61, 0.40 to 0.91, P=0.02; adjusted relative risk 0.62, 0.41 to 0.93, P=0.02), but no difference between groups in immunoglobulin E associated food allergy was seen. CONCLUSION n-3 LCPUFA supplementation in pregnancy did not reduce the overall incidence of immunoglobulin E associated allergies in the first year of life, although atopic eczema and egg sensitisation were lower. Longer term follow-up is needed to determine if supplementation has an effect on respiratory allergic diseases and aeroallergen sensitisation in childhood. TRIAL REGISTRATION Australian New Zealand Clinical Trials Registry ACTRN12610000735055 (DOMInO trial: ACTRN12605000569606).

Importance The Sepsis-3 Criteria emphasized the value of a change of 2 or more points in the Sequential [Sepsis-related] Organ Failure Assessment (SOFA) score, introduced quick SOFA (qSOFA), and removed the systemic inflammatory response syndrome (SIRS) criteria from the sepsis definition. Objective Externally validate and assess the discriminatory capacities of an increase in SOFA score by 2 or more points, 2 or more SIRS criteria, or a qSOFA score of 2 or more points for outcomes among patients who are critically ill with suspected infection. Design, Setting, and Participants Retrospective cohort analysis of 184 875 patients with an infection-related primary admission diagnosis in 182 Australian and New Zealand intensive care units (ICUs) from 2000 through 2015. Exposures SOFA, qSOFA, and SIRS criteria applied to data collected within 24 hours of ICU admission. Main Outcomes and Measures The primary outcome was in-hospital mortality. In-hospital mortality or ICU length of stay (LOS) of 3 days or more was a composite secondary outcome. Discrimination was assessed using the area under the receiver operating characteristic curve (AUROC). Adjusted analyses were performed using a model of baseline risk determined using variables independent of the scoring systems. Results Among 184 875 patients (mean age, 62.9 years [SD, 17.4]; women, 82 540 [44.6%]; most common diagnosis bacterial pneumonia, 32 634 [17.7%]), a total of 34 578 patients (18.7%) died in the hospital, and 102 976 patients (55.7%) died or experienced an ICU LOS of 3 days or more. SOFA score increased by 2 or more points in 90.1%; 86.7% manifested 2 or more SIRS criteria, and 54.4% had a qSOFA score of 2 or more points. SOFA demonstrated significantly greater discrimination for in-hospital mortality (crude AUROC, 0.753 [99% CI, 0.750-0.757]) than SIRS criteria (crude AUROC, 0.589 [99% CI, 0.585-0.593]) or qSOFA (crude AUROC, 0.607 [99% CI, 0.603-0.611]). Incremental improvements were 0.164 (99% CI, 0.159-0.169) for SOFA vs SIRS criteria and 0.146 (99% CI, 0.142-0.151) for SOFA vs qSOFA (P <.001). SOFA (AUROC, 0.736 [99% CI, 0.733-0.739]) outperformed the other scores for the secondary end point (SIRS criteria: AUROC, 0.609 [99% CI, 0.606-0.612]; qSOFA: AUROC, 0.606 [99% CI, 0.602-0.609]). Incremental improvements were 0.127 (99% CI, 0.123-0.131) for SOFA vs SIRS criteria and 0.131 (99% CI, 0.127-0.134) for SOFA vs qSOFA (P <.001). Findings were consistent for both outcomes in multiple sensitivity analyses. Conclusions and Relevance Among adults with suspected infection admitted to an ICU, an increase in SOFA score of 2 or more had greater prognostic accuracy for in-hospital mortality than SIRS criteria or the qSOFA score. These findings suggest that SIRS criteria and qSOFA may have limited utility for predicting mortality in an ICU setting.

Acute lymphoblastic leukemia (ALL) is an aggressive hematological tumor resulting from the malignant transformation of lymphoid progenitors. Despite intensive chemotherapy, 20% of pediatric patients and over 50% of adult patients with ALL do not achieve a complete remission or relapse after intensified chemotherapy, making disease relapse and resistance to therapy the most substantial challenge in the treatment of this disease. Using whole-exome sequencing, we identify mutations in the cytosolic 5'-nucleotidase II gene (NT5C2), which encodes a 5'-nucleotidase enzyme that is responsible for the inactivation of nucleoside-analog chemotherapy drugs, in 20/103 (19%) relapse T cell ALLs and 1/35 (3%) relapse B-precursor ALLs. NT5C2 mutant proteins show increased nucleotidase activity in vitro and conferred resistance to chemotherapy with 6-mercaptopurine and 6-thioguanine when expressed in ALL lymphoblasts. These results support a prominent role for activating mutations in NT5C2 and increased nucleoside-analog metabolism in disease progression and chemotherapy resistance in ALL.

The signals and molecular mechanisms that regulate the replication of terminally differentiated beta cells are unknown. Here, we report the identification and characterization of transmembrane protein 27 (Tmem27, collectrin) in pancreatic beta cells. Expression of Tmem27 is reduced in Tcf1(-/-) mice and is increased in islets of mouse models with hypertrophy of the endocrine pancreas. Tmem27 forms dimers and its extracellular domain is glycosylated, cleaved and shed from the plasma membrane of beta cells. This cleavage process is beta cell specific and does not occur in other cell types. Overexpression of full-length Tmem27, but not the truncated or soluble protein, leads to increased thymidine incorporation, whereas silencing of Tmem27 using RNAi results in a reduction of cell replication. Furthermore, transgenic mice with increased expression of Tmem27 in pancreatic beta cells exhibit increased beta cell mass. Our results identify a pancreatic beta cell transmembrane protein that regulates cell growth of pancreatic islets.