Beta-hydroxy beta-methylbutyrate/arginine/glutamine (HMB/Arg/Gln) supplementation to improve the management of cachexia in patients with advanced lung cancer: an open-label, multicentre, randomised, controlled phase II trial (NOURISH).

BACKGROUND: Cancer cachexia causes significant morbidity and mortality in advanced lung cancer patients. Clinical benefit of ß-hydroxy-ß-methylbutyrate, arginine, and glutamine (HMB/Arg/Gln) was assessed in newly diagnosed patients. METHODS: NOURISH, a prospective, two-arm, open-label, multi-centre, randomised controlled phase II trial compared cachexia in patients who received HMB/Arg/Gln with those who did not. All patients received structured nutritional, exercise and symptom control via a Macmillan Durham Cachexia Pack. Conducted in five UK centres, patients aged > 18 years, with newly diagnosed advanced small cell lung cancer (SCLC) or non-small cell lung cancer (NSCLC), who were able to take oral nutrition, with a performance status of 0-to-2 and a life expectancy > 4 months were eligible for trial entry. Patients suitable for treatment with curative intent were ineligible. The trial was designed as a signal-seeking pilot study with target recruitment of 96 patients. One-to-one randomisation was stratified by diagnosis (SCLC or NSCLC), stage of disease (locally advanced or metastatic) and performance status. The primary outcome measure was treatment success defined as a patient being alive without significant loss of lean body mass (not > 5%) by 12 weeks. Secondary outcome measures included quality of life. RESULTS: Between February-2012 and February-2013, 38 patients were recruited, 19 to each arm. Baseline characteristics were balanced. The trial was halted due to slow accrual and partial adherence. Trial data demonstrated no evidence of treatment benefit. No serious adverse events were reported during the trial. CONCLUSIONS: Further evaluation of HMB/Arg/Gln in this setting could not be recommended on the basis of this trial. CLINICAL TRIAL REGISTRATION: ISRCTN registry: 39911673; 14-Apr-2011 https://doi.org/10.1186/ISRCTN39911673 .

Integrating the Epigenome to Identify Drivers of Hepatocellular Carcinoma.

Disruption of epigenetic mechanisms has been intimately linked to the etiology of human cancer. Understanding how these epigenetic mechanisms (including DNA methylation [5mC], hydroxymethylation [5hmC], and histone post-translational modifications) work in concert to drive cancer initiation and progression remains unknown. Hepatocellular carcinoma (HCC) is increasing in frequency in Western countries but lacks efficacious treatments. The epigenome of HCC remains understudied. To better understand the epigenetic underpinnings of HCC, we performed a genome-wide assessment of 5mC, 5hmC, four histone modifications linked to promoter/enhancer function (H3K4me1, H3K27ac, H3K4me3, and H3K27me3), and transcription across normal, cirrhotic, and HCC liver tissue. Implementation of bioinformatic strategies integrated these epigenetic marks with each other and with transcription to provide a comprehensive epigenetic profile of how and when the liver epigenome is perturbed during progression to HCC. Our data demonstrate significant deregulation of epigenetic regulators combined with disruptions in the epigenome hallmarked by profound loss of 5hmC, locus-specific gains in 5mC and 5hmC, and markedly altered histone modification profiles, particularly remodeling of enhancers. Data integration demonstrates that these marks collaborate to influence transcription (e.g., hyper-5hmC in HCC-gained active enhancers is linked to elevated expression) of genes regulating HCC proliferation. Two such putative epigenetic driver loci identified through our integrative approach, COMT and FMO3, increase apoptosis and decrease cell viability in liver-derived cancer cell lines when ectopically re-expressed. Conclusion: Altogether, integration of multiple epigenetic parameters is a powerful tool for identifying epigenetically regulated drivers of HCC and elucidating how epigenome deregulation contributes to liver disease and HCC.

ZBTB24 is a transcriptional regulator that coordinates with DNMT3B to control DNA methylation.

The interplay between transcription factors and epigenetic writers like the DNA methyltransferases (DNMTs), and the role of this interplay in gene expression, is being increasingly appreciated. ZBTB24, a poorly characterized zinc-finger protein, or the de novo methyltransferase DNMT3B, when mutated, cause Immunodeficiency, Centromere Instability, and Facial anomalies (ICF) syndrome, suggesting an underlying mechanistic link. Chromatin immunoprecipitation coupled with loss-of-function approaches in model systems revealed common loci bound by ZBTB24 and DNMT3B, where they function to regulate gene body methylation. Genes coordinately regulated by ZBTB24 and DNMT3B are enriched for molecular mechanisms essential for cellular homeostasis, highlighting the importance of the ZBTB24-DNMT3B interplay in maintaining epigenetic patterns required for normal cellular function. We identify a ZBTB24 DNA binding motif, which is contained within the promoters of most of its transcriptional targets, including CDCA7, AXIN2, and OSTC. Direct binding of ZBTB24 at the promoters of these genes targets them for transcriptional activation. ZBTB24 binding at the promoters of RNF169 and CAMKMT, however, targets them for transcriptional repression. The involvement of ZBTB24 targets in diverse cellular programs, including the VDR/RXR and interferon regulatory pathways, suggest that ZBTB24's role as a transcriptional regulator is not restricted to immune cells.

Chromosomal aneuploidies induced upon Lamin B2 depletion are mislocalized in the interphase nucleus.

Chromosome territories assume non-random positions in the interphase nucleus with gene-rich chromosomes localized toward the nuclear interior and gene-poor chromosome territories toward the nuclear periphery. Lamins are intermediate filament proteins of the inner nuclear membrane required for the maintenance of nuclear structure and function. Here, we show using whole-genome expression profiling that Lamin A/C or Lamin B2 depletion in an otherwise diploid colorectal cancer cell line (DLD1) deregulates transcript levels from specific chromosomes. Further, three-dimensional fluorescence in situ hybridization (3D-FISH) analyses of a subset of these transcriptionally deregulated chromosome territories revealed that the diploid chromosome territories in Lamin-depleted cells largely maintain conserved positions in the interphase nucleus in a gene-density-dependent manner. In addition, chromosomal aneuploidies were induced in ~25 % of Lamin A/C or Lamin B2-depleted cells. Sub-populations of these aneuploid cells consistently showed a mislocalization of the gene-rich aneuploid chromosome 19 territory toward the nuclear periphery, while gene-poor aneuploid chromosome 18 territory was mislocalized toward the nuclear interior predominantly upon Lamin B2 than Lamin A/C depletion. In addition, a candidate gene locus ZNF570 (Chr.19q13.12) significantly overexpressed upon Lamin B2 depletion was remarkably repositioned away from the nuclear lamina. Taken together, our studies strongly implicate an overarching role for Lamin B2 in the maintenance of nuclear architecture since loss of Lamin B2 relieves the spatial positional constraints required for maintaining conserved localization of aneuploid chromosome territories in the interphase nucleus.

Asbestos-Induced Mesothelial to Fibroblastic Transition Is Modulated by the Inflammasome.

Despite the causal relationship established between malignant mesothelioma (MM) and asbestos exposure, the exact mechanism by which asbestos induces this neoplasm and other asbestos-related diseases is still not well understood. MM is characterized by chronic inflammation, which is believed to play an intrinsic role in the origin of this disease. We recently found that asbestos activates the nod-like receptor family member containing a pyrin domain 3 (NLRP3) inflammasome in a protracted manner, leading to an up-regulation of IL-1ß and IL-18 production in human mesothelial cells. Combined with biopersistence of asbestos fibers, we hypothesize that this creates an environment of chronic IL-1ß signaling in human mesothelial cells, which may promote mesothelial to fibroblastic transition (MFT) in an NLRP3-dependent manner. Using a series of experiments, we found that asbestos induces a fibroblastic transition of mesothelial cells with a gain of mesenchymal markers (vimentin and N-cadherin), whereas epithelial markers, such as E-cadherin, are down-regulated. Use of siRNA against NLRP3, recombinant IL-1ß, and IL-1 receptor antagonist confirmed the role of NLRP3 inflammasome-dependent IL-1ß in the process. In vivo studies using wild-type and various inflammasome component knockout mice also revealed the process of asbestos-induced mesothelial to fibroblastic transition and its amelioration in caspase-1 knockout mice. Taken together, our data are the first to suggest that asbestos induces mesothelial to fibroblastic transition in an inflammasome-dependent manner.

Neoadjuvant cisplatin and fluorouracil versus epirubicin, cisplatin, and capecitabine followed by resection in patients with oesophageal adenocarcinoma (UK MRC OE05): an open-label, randomised phase 3 trial.

BACKGROUND: Neoadjuvant chemotherapy before surgery improves survival compared with surgery alone for patients with oesophageal cancer. The OE05 trial assessed whether increasing the duration and intensity of neoadjuvant chemotherapy further improved survival compared with the current standard regimen. METHODS: OE05 was an open-label, phase 3, randomised clinical trial. Patients with surgically resectable oesophageal adenocarcinoma classified as stage cT1N1, cT2N1, cT3N0/N1, or cT4N0/N1 were recruited from 72 UK hospitals. Eligibility criteria included WHO performance status 0 or 1, adequate respiratory, cardiac, and liver function, white blood cell count at least 3 × 109 cells per L, platelet count at least 100 × 109 platelets per L, and a glomerular filtration rate at least 60 mL/min. Participants were randomly allocated (1:1) using a computerised minimisation program with a random element and stratified by centre and tumour stage, to receive two cycles of cisplatin and fluorouracil (CF; two 3-weekly cycles of cisplatin [80 mg/m2 intravenously on day 1] and fluorouracil [1 g/m2 per day intravenously on days 1-4]) or four cycles of epirubicin, cisplatin, and capecitabine (ECX; four 3-weekly cycles of epirubicin [50 mg/m2] and cisplatin [60 mg/m2] intravenously on day 1, and capecitabine [1250 mg/m2] daily throughout the four cycles) before surgery, stratified according to centre and clinical disease stage. Neither patients nor study staff were masked to treatment allocation. Two-phase oesophagectomy with two-field (abdomen and thorax) lymphadenectomy was done within 4-6 weeks of completion of chemotherapy. The primary outcome measure was overall survival, and primary and safety analyses were done in the intention-to-treat population. This trial is registered with the ISRCTN registry (number 01852072) and ClinicalTrials.gov (NCT00041262), and is completed. FINDINGS: Between Jan 13, 2005, and Oct 31, 2011, 897 patients were recruited and 451 were assigned to the CF group and 446 to the ECX group. By Nov 14, 2016, 327 (73%) of 451 patients in the CF group and 302 (68%) of 446 in the ECX group had died. Median survival was 23·4 months (95% CI 20·6-26·3) with CF and 26·1 months (22·5-29·7) with ECX (hazard ratio 0·90 (95% CI 0·77-1·05, p=0·19). No unexpected chemotherapy toxicity was seen, and neutropenia was the most commonly reported event (grade 3 or 4 neutropenia: 74 [17%] of 446 patients in the CF group vs 101 [23%] of 441 people in the ECX group). The proportions of patients with postoperative complications (224 [56%] of 398 people for whom data were available in the CF group and 233 [62%] of 374 in the ECX group; p=0·089) were similar between the two groups. One patient in the ECX group died of suspected treatment-related neutropenic sepsis. INTERPRETATION: Four cycles of neoadjuvant ECX compared with two cycles of CF did not increase survival, and cannot be considered standard of care. Our study involved a large number of centres and detailed protocol with comprehensive prospective assessment of health-related quality of life in a patient population confined to people with adenocarcinomas of the oesophagus and gastro-oesophageal junction (Siewert types 1 and 2). Alternative chemotherapy regimens and neoadjuvant chemoradiation are being investigated to improve outcomes for patients with oesophageal carcinoma. FUNDING: Cancer Research UK and Medical Research Council Clinical Trials Unit at University College London.

Differential Susceptibility of Human Pleural and Peritoneal Mesothelial Cells to Asbestos Exposure.

Malignant mesothelioma (MM) is an aggressive cancer of mesothelial cells of pleural and peritoneal cavities. In 85% of cases both pleural and peritoneal MM is caused by asbestos exposure. Although both are asbestos-induced cancers, the incidence of pleural MM is significantly higher (85%) than peritoneal MM (15%). It has been proposed that carcinogenesis is a result of asbestos-induced inflammation but it is not clear what contributes to the differences observed between incidences of these two cancers. We hypothesize that the observed differences in incidences of pleural and peritoneal MM are the result of differences in the direct response of these cell types to asbestos rather than to differences mediated by the in vivo microenvironment. To test this hypothesis we characterized cellular responses to asbestos in a controlled environment. We found significantly greater changes in genome-wide expression in response to asbestos exposure in pleural mesothelial cells as compared to peritoneal mesothelial cells. In particular, a greater response in many common genes (IL-8, ATF3, CXCL2, CXCL3, IL-6, GOS2) was seen in pleural mesothelial cells as compared to peritoneal mesothelial cells. Unique genes expressed in pleural mesothelial cells were mainly pro-inflammatory (G-CSF, IL-1ß, IL-1α, GREM1) and have previously been shown to be involved in development of MM. Our results are consistent with the hypothesis that differences in incidences of pleural and peritoneal MM upon exposure to asbestos are the result of differences in mesothelial cell physiology that lead to differences in the inflammatory response, which leads to cancer.

Assessment of provider competence and quality of maternal/newborn care in selected Latin American and Caribbean countries.

OBJECTIVE: To obtain a snapshot of the maternal and newborn care provided by different types of maternal and child health providers in Latin America and the Caribbean (LAC) to 1) better inform advocacy and programmatic strategies and interventions to improve the quality of those services in the region, and 2) determine the need for more rigorous study of the issues. METHODS: A rapid assessment of 83 health workers providing antepartum, intrapartum, and immediate postpartum and newborn care (within two hours of birth) in eight LAC countries was conducted in November and December of 2011. Health workers were observed by two-person expert maternal/newborn clinician teams using pretested forms based on international quality-of-care standards. A total of 105 care encounters were observed, primarily in urban, public, referral-level settings. Providers of care included obstetricians, midwives, generalist physicians, medical residents, registered nurses, auxiliary nurses, and students of medicine, midwifery, and nursing. RESULTS: Hand washing, as an indicator of quality of antepartum care, was observed in only 41% of the observed encounters. Labor management often lacked certain elements of respectful maternity care across all provider groups. Several clinical tasks of high importance in the identification and prevention of common complications of antepartum, intrapartum, and immediate postpartum/newborn care were not documented as performed during the observation periods. Providers self-reported limited competence (ability to perform to a defined level of proficiency) in manual removal of the placenta, bimanual compression of the uterus, and newborn resuscitation. CONCLUSIONS: The findings suggest that 1) the quality of maternal and newborn care and 2) the competence of maternal and child health providers in the diverse selection of LAC countries that were studied require substantial attention.

Gefitinib for oesophageal cancer progressing after chemotherapy (COG): a phase 3, multicentre, double-blind, placebo-controlled randomised trial.

BACKGROUND: Evidence is scarce for the effectiveness of therapies for oesophageal cancer progressing after chemotherapy, and no randomised trials have been reported. We aimed to compare gefitinib with placebo in previously treated advanced oesophageal cancer. METHODS: For this phase 3, parallel, randomised, placebo-controlled trial, eligible patients were adults with advanced oesophageal cancer or type I/II Siewert junctional tumours, histologically confirmed squamous-cell carcinoma or adenocarcinoma, who had progressed after chemotherapy, with WHO performance status 0-2, and with measurable or evaluable disease on CT scan. Participants were recruited from 48 UK centres and randomly assigned (1:1) to gefitinib (500 mg) or matching placebo by simple randomisation with no stratification factors. Patients, clinicians, and trial office staff were masked to treatment allocation. Treatment continued until disease progression, unacceptable toxicity, or patient choice. The primary outcome was overall survival, analysed by intention to treat. This trial is registered, number ISRCTN29580179. FINDINGS: Between March 30, 2009, and Nov 18, 2011, 450 patients were randomly assigned to treatment groups (one patient withdrew consent; 224 patients allocated gefitinib and 225 allocated placebo included in analyses). Overall survival did not differ between groups (median 3·73 months, 95% CI 3·23-4·50, for gefitinib vs 3·67 months, 95% CI 2·97-4·37, for placebo; hazard ratio [HR] 0·90, 95% CI 0·74-1·09, p=0·29). Among the prespecified patient-reported outcomes (110 patients on gefitinib and 121 on placebo completed both baseline and 4 week questionnaires and were included in analyses), odynophagia was significantly better in the gefitinib group (adjusted mean difference -8·61, 95% CI -14·49 to -2·73; n=227; p=0·004), whereas the other outcomes were not significantly improved compared with placebo: global quality of life (2·69, 95% CI -2·33 to 7·72, n=231, p=0·293), dysphagia (-3·18, 95% CI -8·36 to 2·00, n=231, p=0·228), and eating (-4·11, 95% CI -9·96 to 1·75, n=229, p=0·168). Median progression-free survival was marginally longer with gefitinib than it was with placebo (1·57 months, 95% CI 1·23-1·90 in the gefitinib group vs 1·17 months, 95% CI 1·07-1·37 in the placebo group; HR 0·80, 95% CI 0·66-0·96, p=0·020). The most common toxicities were diarrhoea (36 [16%] of 224 patients on gefitinib vs six [3%] of 225 on placebo) and skin toxicity (46 [21%] vs two [1%]), both mostly grade 2. The commonest grade 3-4 toxicities were fatigue (24 [11%] vs 13 [6%] patients) and diarrhoea (13 [6%] vs two [1%]). Serious adverse events were reported in 109 (49%) of 224 patients assigned to gefitinib and 101 (45%) of 225 on placebo. 54 (24%) of patients in the gefitinib group achieved disease control at 8 weeks, as did 35 (16%) of patients on placebo (p=0·023). INTERPRETATION: The use of gefitinib as a second-line treatment in oesophageal cancer in unselected patients does not improve overall survival, but has palliative benefits in a subgroup of these difficult-to-treat patients with short life expectancy. Future research should focus on identification of predictive biomarkers to identify this subgroup of benefiting patients. FUNDING: Cancer Research UK.

Extracellular signal-regulated kinase 5 and cyclic AMP response element binding protein are novel pathways inhibited by vandetanib (ZD6474) and doxorubicin in mesotheliomas.

Malignant mesothelioma (MM), lung cancers, and asbestosis are hyperproliferative diseases associated with exposures to asbestos. All have a poor prognosis; thus, the need to develop novel and effective therapies is urgent. Vandetanib (Van) (ZD6474, ZACTIMA) is a tyrosine kinase inhibitor that has shown equivocal results in clinical trials for advanced non-small cell lung cancer. However, tyrosine kinase inhibitors alone have shown no significant clinical activity in phase II trials of patients with unresectable MM. Using epithelioid (HMESO) and sarcomatoid (H2373) human MM lines, the efficacy of tumor cell killing and signaling pathways modulated by Van with and without doxorubicin (Dox) was examined. Van alone reduced total cell numbers in HMESO MM and synergistically increased the toxicity of Dox in HMESO and H2373 cells. Most importantly, we identified two novel cell survival/resistance pathways, ERK5 and cyclic AMP response element binding protein (CREB), that were inhibited by Van and Dox. After silencing of either ERK5 or CREB, significant decreases in cell numbers in the Dox-resistant sarcomatoid H2373 line were observed. Results suggest that a plethora of cell signaling pathways associated with cell survival are induced by Dox but inhibited by the addition of Van in MM. Data from our study support the combined efficacy of Van and Dox as a novel approach in the treatment of MM that is further enhanced by blocking ERK5 or CREB signaling cascades.

Malignant mesothelioma: development to therapy.

Malignant mesothelioma (MM) is an aggressive cancer of the mesothelium caused by asbestos. Asbestos use has been reduced but not completely stopped. In addition, natural or man-made disasters will continue to dislodge asbestos from old buildings into the atmosphere and as long as respirable asbestos is available, MM will continue to be a threat. Due to the long latency period of MM development, it would still take decades to eradicate this disease if asbestos was completely removed from our lives today. Therefore, there is a need for researchers and clinicians to work together to understand this deadly disease and find a solution for early diagnosis and treatment. This article focuses on developmental mechanisms as well as current therapies available for MM.

Asbestos modulates thioredoxin-thioredoxin interacting protein interaction to regulate inflammasome activation.

BACKGROUND: Asbestos exposure is related to various diseases including asbestosis and malignant mesothelioma (MM). Among the pathogenic mechanisms proposed by which asbestos can cause diseases involving epithelial and mesothelial cells, the most widely accepted one is the generation of reactive oxygen species and/or depletion of antioxidants like glutathione. It has also been demonstrated that asbestos can induce inflammation, perhaps due to activation of inflammasomes. METHODS: The oxidation state of thioredoxin was analyzed by redox Western blot analysis and ROS generation was assessed spectrophotometrically as a read-out of solubilized formazan produced by the reduction of nitrotetrazolium blue (NTB) by superoxide. Quantitative real time PCR was used to assess changes in gene transcription. RESULTS: Here we demonstrate that crocidolite asbestos fibers oxidize the pool of the antioxidant, Thioredoxin-1 (Trx1), which results in release of Thioredoxin Interacting Protein (TXNIP) and subsequent activation of inflammasomes in human mesothelial cells. Exposure to crocidolite asbestos resulted in the depletion of reduced Trx1 in human peritoneal mesothelial (LP9/hTERT) cells. Pretreatment with the antioxidant dehydroascorbic acid (a reactive oxygen species (ROS) scavenger) reduced the level of crocidolite asbestos-induced Trx1 oxidation as well as the depletion of reduced Trx1. Increasing Trx1 expression levels using a Trx1 over-expression vector, reduced the extent of Trx1 oxidation and generation of ROS by crocidolite asbestos, and increased cell survival. In addition, knockdown of TXNIP expression by siRNA attenuated crocidolite asbestos-induced activation of the inflammasome. CONCLUSION: Our novel findings suggest that extensive Trx1 oxidation and TXNIP dissociation may be one of the mechanisms by which crocidolite asbestos activates the inflammasome and helps in development of MM.

Structural perturbation of chromatin domains with multiple developmental regulators can severely impact gene regulation and development.

Chromatin domain boundaries delimited by CTCF motifs can restrict the range of enhancer action. However, disruption of domain structure often results in mild gene dysregulation and thus predicting the impact of boundary rearrangements on animal development remains challenging. Here, we tested whether structural perturbation of a chromatin domain with multiple developmental regulators can result in more acute gene dysregulation and severe developmental phenotypes. We targeted clusters of CTCF motifs in a domain of the mouse genome containing three FGF ligand genes-Fgf3, Fgf4, and Fgf15-that regulate several developmental processes. Deletion of the 23.9kb cluster that defines the centromeric boundary of this domain resulted in ectopic interactions of the FGF genes with enhancers located across the deleted boundary that are active in the developing brain. This caused strong induction of FGF expression and perinatal lethality with encephalocele and orofacial cleft phenotypes. Heterozygous boundary deletion was sufficient to cause these fully penetrant phenotypes, and strikingly, loss of a single CTCF motif within the cluster also recapitulated ectopic FGF expression and caused encephalocele. However, such phenotypic sensitivity to perturbation of domain structure did not extend to all CTCF clusters of this domain, nor to all developmental processes controlled by these three FGF genes-for example, the ability to undergo lineage specification in the blastocyst and pre-implantation development were not affected. By tracing the impact of different chromosomal rearrangements throughout mouse development, we start to uncover the determinants of phenotypic robustness and sensitivity to perturbation of chromatin boundaries. Our data show how small sequence variants at certain domain boundaries can have a surprisingly outsized effect and must be considered as potential sources of gene dysregulation during development and disease.

Therapeutic targeting of differentiation-state dependent metabolic vulnerabilities in diffuse midline glioma.

H3K27M diffuse midline gliomas (DMG), including diffuse intrinsic pontine gliomas (DIPG), exhibit cellular heterogeneity comprising less-differentiated oligodendrocyte precursors (OPC)-like stem cells and more differentiated astrocyte (AC)-like cells. Here, we establish in vitro models that recapitulate DMG-OPC-like and AC-like phenotypes and perform transcriptomics, metabolomics, and bioenergetic profiling to identify metabolic programs in the different cellular states. We then define strategies to target metabolic vulnerabilities within specific tumor populations. We show that AC-like cells exhibit a mesenchymal phenotype and are sensitized to ferroptotic cell death. In contrast, OPC-like cells upregulate cholesterol biosynthesis, have diminished mitochondrial oxidative phosphorylation (OXPHOS), and are accordingly more sensitive to statins and OXPHOS inhibitors. Additionally, statins and OXPHOS inhibitors show efficacy and extend survival in preclinical orthotopic models established with stem-like H3K27M DMG cells. Together, this study demonstrates that cellular subtypes within DMGs harbor distinct metabolic vulnerabilities that can be uniquely and selectively targeted for therapeutic gain.

Asbestos and erionite prime and activate the NLRP3 inflammasome that stimulates autocrine cytokine release in human mesothelial cells.

BACKGROUND: Pleural fibrosis and malignant mesotheliomas (MM) occur after exposures to pathogenic fibers, yet the mechanisms initiating these diseases are unclear. RESULTS: We document priming and activation of the NLRP3 inflammasome in human mesothelial cells by asbestos and erionite that is causally related to release of IL-1ß, IL-6, IL-8, and Vascular Endothelial Growth Factor (VEGF). Transcription and release of these proteins are inhibited in vitro using Anakinra, an IL-1 receptor antagonist that reduces these cytokines in a human peritoneal MM mouse xenograft model. CONCLUSIONS: These novel data show that asbestos-induced priming and activation of the NLRP3 inflammasome triggers an autocrine feedback loop modulated via the IL-1 receptor in mesothelial cell type targeted in pleural infection, fibrosis, and carcinogenesis.

Sputum matrix metalloproteinase-12 in patients with chronic obstructive pulmonary disease and asthma: relationship to disease severity.

BACKGROUND: Matrix metalloproteinase (MMP)-12 has been implicated in the pathogenesis of both chronic obstructive pulmonary disease (COPD) and asthma. The influence of disease severity on sputum MMP-12 concentrations and activity is not known. OBJECTIVES: We sought to examine the relationship between disease severity assessed by means of lung function and computed tomography (CT) and induced sputum MMP-12 concentrations and activity in patients with asthma and COPD. METHODS: In 208 subjects (109 asthmatic patients, smokers and never smokers, mild, moderate, and severe; 53 patients with COPD, smokers and exsmokers, mild, moderate, and severe; and 46 healthy control subjects, smokers and never smokers), we measured induced sputum MMP-12 concentrations (ELISA) and enzyme activity (fluorescence resonance energy transfer), sputum cell MMP12 mRNA expression (quantitative PCR [qPCR]), diffusing capacity for carbon monoxide (Dlco), and CT assessment of emphysema (percentage of low-attenuation areas at less -950 Hounsfield units). RESULTS: Sputum MMP-12 concentrations are greater in patients with COPD and smokers with asthma than in healthy nonsmokers (P = .003 and P = .035, respectively) but similar to those seen in healthy smokers. In patients with COPD, disease severity, when measured by means of CT-assessed emphysema, but not by means of spirometry or Dlco values, is directly associated with sputum MMP-12 concentrations and activity. In the asthma groups there is no significant association between disease severity and sputum MMP-12 concentrations or activity. CONCLUSIONS: Sputum MMP-12 concentrations and activity in patients with COPD are directly associated with the extent of emphysema measured by means of CT. This finding supports a role for MMP-12 in the pathogenesis of COPD and might suggest that blocking MMP-12 activity in patients with COPD could prevent the further development of emphysema.

Provision of weight management advice for obese women during pregnancy: a survey of current practice and midwives' views on future approaches.

A semi-structured, web-based questionnaire was developed to survey midwives (n = 241) employed by NHS Tayside, UK, to identify current practice and views on weight management of obese women during pregnancy and the puerperium. A total of 78 (32%) midwives submitted responses following email invitation. Most respondents (79%) reported always calculating women's body mass index (BMI) at booking, with 73% routinely explaining the BMI category. In terms of future practice for obese women, although few respondents (15%) currently offer personalised advice regarding weight management based on a woman's diet and physical activity levels, 77% of respondents thought such advice would be appropriate and 69% thought it could possibly be feasible to offer such advice. The respondents viewed weight management to be of importance and felt that universal advice is appropriate, but confidence in discussing weight management and knowledge of the subject was low. Strategies to improve midwife confidence and weight management services should include training, ongoing support and definition of the midwife's role within the multidisciplinary team to support practice in the future.

MLL3/MLL4 methyltransferase activities control early embryonic development and embryonic stem cell differentiation in a lineage-selective manner.

H3K4me1 methyltransferases MLL3 (KMT2C) and MLL4 (KMT2D) are critical for enhancer activation, cell differentiation and development. However, roles of MLL3/4 enzymatic activities and MLL3/4-mediated enhancer H3K4me1 in these processes remain unclear. Here we report that constitutive elimination of both MLL3 and MLL4 enzymatic activities prevents initiation of gastrulation and leads to early embryonic lethality in mice. However, selective elimination of MLL3/4 enzymatic activities in embryonic, but not extraembryonic, lineages leaves gastrulation largely intact. Consistent with this, embryonic stem cells (ESCs) lacking MLL3/4 enzymatic activities can differentiate toward the three embryonic germ layers but show aberrant differentiation to extraembryonic endoderm (ExEn) and trophectoderm. The failure in ExEn differentiation can be attributed to markedly reduced enhancer-binding of the lineage-determining transcription factor GATA6. Furthermore, we show that MLL3/4-catalyzed H3K4me1 is largely dispensable for enhancer activation during ESC differentiation. Together, our findings suggest a lineage-selective, but enhancer activation-independent, role of MLL3/4 methyltransferase activities in early embryonic development and ESC differentiation.

Extensive co-binding and rapid redistribution of NANOG and GATA6 during emergence of divergent lineages.

Fate-determining transcription factors (TFs) can promote lineage-restricted transcriptional programs from common progenitor states. The inner cell mass (ICM) of mouse blastocysts co-expresses the TFs NANOG and GATA6, which drive the bifurcation of the ICM into either the epiblast (Epi) or the primitive endoderm (PrE), respectively. Here, we induce GATA6 in embryonic stem cells-that also express NANOG-to characterize how a state of co-expression of opposing TFs resolves into divergent lineages. Surprisingly, we find that GATA6 and NANOG co-bind at the vast majority of Epi and PrE enhancers, a phenomenon we also observe in blastocysts. The co-bound state is followed by eviction and repression of Epi TFs, and quick remodeling of chromatin and enhancer-promoter contacts thus establishing the PrE lineage while repressing the Epi fate. We propose that co-binding of GATA6 and NANOG at shared enhancers maintains ICM plasticity and promotes the rapid establishment of Epi- and PrE-specific transcriptional programs.