Detection of epithelial apoptosis in pelvic ileal pouches for ulcerative colitis and familial adenomatous polyposis

<p>Abstract</p> <p>Background</p> <p>Ileal pouch-anal anastomosis (IPAA) is the surgical procedure of choice for patients with refractory ulcerative colitis (UC) and for familial adenomatous polyposis (FAP) with many rectal polyps. Pouchitis is one of the more frequent complications after IPAA in UC patients; however, it is rare in FAP.</p> <p>Objective</p> <p>Evaluate pro-apoptotic activity in endoscopically and histological normal mucosa of the ileal pouch in patients with UC and FAP.</p> <p>Methods</p> <p>Eighteen patients (nine with UC and nine with FAP) with J pouch after total rectocolectomy were studied. Biopsies were obtained from the mucosa of the pouch and from normal ileum. The specimens were snap-frozen and the expressions of Bax and Bcl-2 were determined by immunoblot of protein extracts and by immunohistochemistry analysis. FADD, Caspase-8, APAF-1 and Caspase-9 were evaluated by immunoprecipitation and immunoblot.</p> <p>Results</p> <p>Patients with UC had significantly higher protein levels of Bax and APAF-1, Caspase-9 than patients with FAP, but were similar to controls. The expressions of Bcl-2 and FADD, Caspase-8 were similar in the groups. Immunohistochemistry for Bax showed less intensity of immunoreactions in FAP than in UC and Controls. Bcl-2 immunostaining was similar among the groups.</p> <p>Conclusion</p> <p>Patients with FAP present lower levels of pro-apoptotic proteins in all methods applied, even in the absence of clinical and endoscopic pouchitis and dysplasia in the histological analysis. These findings may explain a tendency of up-regulation of apoptosis in UC patients, resulting in higher rates of progression to pouchitis in these patients, which could correlate with mucosal atrophy that occurs in inflamed tissue. However, FAP patients had low pro-apoptotic activity in the mucosa, and it could explain the tendency to low cell turn over and presence of adenomas in this syndrome.</p

The predictive value of microRNA-126 in relation to first line treatment with capecitabine and oxaliplatin in patients with metastatic colorectal cancer

<p>Abstract</p> <p>Background</p> <p>MicroRNA-126 is the only microRNA (miRNA) known to be endothelial cell-specific influencing angiogenesis in several ways. The aim of the present study was to analyse the possible predictive value of miRNA-126 in relation to first line capecitabine and oxaliplatin (XELOX) in patients with metastatic colorectal cancer (mCRC).</p> <p>Methods</p> <p>The study included 89 patients with mCRC. <it>In situ </it>hybridization (ISH) was performed to detect miRNA-126 in formalin-fixed paraffin embedded tissue from primary tumours. The expression of miRNA-126, area per image (μm²), was measured using image analysis. Clinical response was evaluated according to RECIST. Progression free survival (PFS) was compared using the Kaplan-Meier method and the log rank test. Tumours were classified as low or high miRNA-126 expressing tumours using the median value from the patients with response as cut-off.</p> <p>Results</p> <p>The median miRNA-126 expression level was significantly higher in patients responding to XELOX, 3629 μm²(95% CI, 2566-4846), compared to the patients not responding, 1670 μm²(95% CI, 1436-2041), <it>p </it>< 0.0001. The positive predictive value was 90%, and the negative predictive value was 71%. The median PFS of patients with high expressing tumours was 11.5 months (95% CI, 9.0-12.7 months) compared to 6.0 months (95% CI, 4.8-6.9 months) for patients with low expressing tumours, <it>p </it>< 0.0001.</p> <p>Conclusions</p> <p>Angiogenesis quantified by ISH of miRNA-126 was related to response to first line XELOX in patients with mCRC, translating to a significant difference in PFS. The predictive value of miRNA-126 remains to be further elucidated in prospective studies.</p

Serum microrna biomarkers for detection of non-small cell lung cancer

Non small cell lung cancer (NSCLC) is the leading cause of cancer-related mortality world-wide and the majority of cases are diagnosed at late stages of disease. There is currently no cost-effective screening test for NSCLC, and the development of such a test is a public health imperative. Recent studies have suggested that chest computed tomography screening of patients at high risk of lung cancer can increase survival from disease, however, the cost effectiveness of such screening has not been established. In this Phase I/II biomarker study we examined the feasibility of using serum miRNA as biomarkers of NSCLC using RT-qPCR to examine the expression of 180 miRNAs in sera from 30 treatment naive NSCLC patients and 20 healthy controls. Receiver operating characteristic curves (ROC) and area under the curve were used to identify differentially expressed miRNA pairs that could distinguish NSCLC from healthy controls. Selected miRNA candidates were further validated in sera from an additional 55 NSCLC patients and 75 healthy controls. Examination of miRNA expression levels in serum from a multi-institutional cohort of 50 subjects (30 NSCLC patients and 20 healthy controls) identified differentially expressed miRNAs. A combination of two differentially expressed miRNAs miR-15b and miR-27b, was able to discriminate NSCLC from healthy controls with sensitivity, specificity, positive predictive value (PPV) and negative predictive value (NPV) of 100% in the training set. Upon further testing on additional 130 subjects (55 NSCLC and 75 healthy controls), this miRNA pair predicted NSCLC with a specificity of 84% (95% CI 0.73-0.91), sensitivity of 100% (95% CI; 0.93-1.0), NPV of 100%, and PPV of 82%. These data provide evidence that serum miRNAs have the potential to be sensitive, cost-effective biomarkers for the early detection of NSCLC. Further testing in a Phase III biomarker study in is necessary for validation of these results. © 2012 Hennessey et al

MicroRNA-21 Exhibits Antiangiogenic Function by Targeting RhoB Expression in Endothelial Cells

BACKGROUND: MicroRNAs (miRNAs) are endogenously expressed small non-coding RNAs that regulate gene expression at post-transcriptional level. The recent discovery of the involvement of these RNAs in the control of angiogenesis renders them very attractive in the development of new approaches for restoring the angiogenic balance. Whereas miRNA-21 has been demonstrated to be highly expressed in endothelial cells, the potential function of this miRNA in angiogenesis has never been investigated. METHODOLOGY/PRINCIPAL FINDINGS: We first observed in endothelial cells a negative regulation of miR-21 expression by serum and bFGF, two pro-angiogenic factors. Then using in vitro angiogenic assays, we observed that miR-21 acts as a negative modulator of angiogenesis. miR-21 overexpression reduced endothelial cell proliferation, migration and the ability of these cells to form tubes whereas miR-21 inhibition using a LNA-anti-miR led to opposite effects. Expression of miR-21 in endothelial cells also led to a reduction in the organization of actin into stress fibers, which may explain the decrease in cell migration. Further mechanistic studies showed that miR-21 targets RhoB, as revealed by a decrease in RhoB expression and activity in miR-21 overexpressing cells. RhoB silencing impairs endothelial cell migration and tubulogenesis, thus providing a possible mechanism for miR-21 to inhibit angiogenesis. Finally, the therapeutic potential of miR-21 as an angiogenesis inhibitor was demonstrated in vivo in a mouse model of choroidal neovascularization. CONCLUSIONS/SIGNIFICANCE: Our results identify miR-21 as a new angiogenesis inhibitor and suggest that inhibition of cell migration and tubulogenesis is mediated through repression of RhoB

In Search of an Uncultured Human-Associated TM7 Bacterium in the Environment

We have identified an environmental bacterium in the Candidate Division TM7 with ≥98.5% 16S rDNA gene homology to a group of TM7 bacteria associated with the human oral cavity and skin. The environmental TM7 bacterium (referred to as TM7a-like) was readily detectable in wastewater with molecular techniques over two years of sampling. We present the first images of TM7a-like cells through FISH technique and the first images of any TM7 as viable cells through the STARFISH technique. In situ quantification showed TM7 concentration in wastewater up to five times greater than in human oral sites. We speculate that upon further characterization of the physiology and genetics of the TM7a-like bacterium from environmental sources and confirmation of its genomic identity to human-associated counterparts it will serve as model organisms to better understand its role in human health. The approach proposed circumvents difficulties imposed by sampling humans, provides an alternative strategy to characterizing some diseases of unknown etiology, and renders a much needed understanding of the ecophysiological role hundreds of unique Bacteria and Archaea strains play in mixed microbial communities

Intestinal Microbiota Composition of Interleukin-10 Deficient C57BL/6J Mice and Susceptibility to Helicobacter hepaticus-Induced Colitis

The mouse pathobiont Helicobacter hepaticus can induce typhlocolitis in interleukin-10-deficient mice, and H. hepaticus infection of immunodeficient mice is widely used as a model to study the role of pathogens and commensal bacteria in the pathogenesis of inflammatory bowel disease. C57BL/6J Il10[superscript −/−] mice kept under specific pathogen-free conditions in two different facilities (MHH and MIT), displayed strong differences with respect to their susceptibilities to H. hepaticus-induced intestinal pathology. Mice at MIT developed robust typhlocolitis after infection with H. hepaticus, while mice at MHH developed no significant pathology after infection with the same H. hepaticus strain. We hypothesized that the intestinal microbiota might be responsible for these differences and therefore performed high resolution analysis of the intestinal microbiota composition in uninfected mice from the two facilities by deep sequencing of partial 16S rRNA amplicons. The microbiota composition differed markedly between mice from both facilities. Significant differences were also detected between two groups of MHH mice born in different years. Of the 119 operational taxonomic units (OTUs) that occurred in at least half the cecum or colon samples of at least one mouse group, 24 were only found in MIT mice, and another 13 OTUs could only be found in MHH samples. While most of the MHH-specific OTUs could only be identified to class or family level, the MIT-specific set contained OTUs identified to genus or species level, including the opportunistic pathogen, Bilophila wadsworthia. The susceptibility to H. hepaticus-induced colitis differed considerably between Il10[superscript −/−] mice originating from the two institutions. This was associated with significant differences in microbiota composition, highlighting the importance of characterizing the intestinal microbiome when studying murine models of IBD.National Institutes of Health (U.S.) (Grant NIH P01-CA26731)National Institutes of Health (U.S.) (Grant NIH P30ES0026731)National Institutes of Health (U.S.) (Grant NIH R01-OD011141

miRNAs at the heart of the matter

Cardiovascular disease is among the main causes of morbidity and mortality in developed countries. The pathological process of the heart is associated with altered expression profile of genes that are important for cardiac function. MicroRNAs (miRNAs) have emerged as one of the central players of gene expression regulation. The implications of miRNAs in the pathological process of cardiovascular system have recently been recognized, representing the most rapidly evolving research field. Here, we summarize and analyze the currently available data from our own laboratory and other groups, providing a comprehensive overview of miRNA function in the heart, including a brief introduction of miRNA biology, expression profile of miRNAs in cardiac tissue, role of miRNAs in cardiac hypertrophy and heart failure, the arrhythmogenic potential of miRNAs, the involvement of miRNAs in vascular angiogenesis, and regulation of cardiomyocyte apoptosis by miRNAs. The target genes and signaling pathways linking the miRNAs to cardiovascular disease are highlighted. The applications of miRNA interference technologies for manipulating miRNA expression, stability, and function as new strategies for molecular therapy of human disease are evaluated. Finally, some specific issues related to future directions of the research on miRNAs relevant to cardiovascular disease are pinpointed and speculated

Bone marrow niche trafficking of miR-126 controls the self-renewal of leukemia stem cells in chronic myelogenous leukemia

Leukemia stem cells (LSCs) in individuals with chronic myelogenous leukemia (CML) (hereafter referred to as CML LSCs) are responsible for initiating and maintaining clonal hematopoiesis. These cells persist in the bone marrow (BM) despite effective inhibition of BCR–ABL kinase activity by tyrosine kinase inhibitors (TKIs). Here we show that although the microRNA (miRNA) miR-126 supported the quiescence, self-renewal and engraftment capacity of CML LSCs, miR-126 levels were lower in CML LSCs than in long-term hematopoietic stem cells (LT-HSCs) from healthy individuals. Downregulation of miR-126 levels in CML LSCs was due to phosphorylation of Sprouty-related EVH1-domain-containing 1 (SPRED1) by BCR–ABL, which led to inhibition of the RAN–exportin-5–RCC1 complex that mediates miRNA maturation. Endothelial cells (ECs) in the BM supply miR-126 to CML LSCs to support quiescence and leukemia growth, as shown using mouse models of CML in which Mir126a (encoding miR-126) was conditionally knocked out in ECs and/or LSCs. Inhibition of BCR–ABL by TKI treatment caused an undesired increase in endogenous miR-126 levels, which enhanced LSC quiescence and persistence. Mir126a knockout in LSCs and/or ECs, or treatment with a miR-126 inhibitor that targets miR-126 expression in both LSCs and ECs, enhanced the in vivo anti-leukemic effects of TKI treatment and strongly diminished LSC leukemia-initiating capacity, providing a new strategy for the elimination of LSCs in individuals with CML

MicroRNA and Target Protein Patterns Reveal Physiopathological Features of Glioma Subtypes

Gliomas such as oligodendrogliomas (ODG) and glioblastomas (GBM) are brain tumours with different clinical outcomes. Histology-based classification of these tumour types is often difficult. Therefore the first aim of this study was to gain microRNA data that can be used as reliable signatures of oligodendrogliomas and glioblastomas. We investigated the levels of 282 microRNAs using membrane-array hybridisation and real-time PCR in ODG, GBM and control brain tissues. In comparison to these control tissues, 26 deregulated microRNAs were identified in tumours and the tissue levels of seven microRNAs (miR-21, miR-128, miR-132, miR-134, miR-155, miR-210 and miR-409-5p) appropriately discriminated oligodendrogliomas from glioblastomas. Genomic, epigenomic and host gene expression studies were conducted to investigate the mechanisms involved in these deregulations. Another aim of this study was to better understand glioma physiopathology looking for targets of deregulated microRNAs. We discovered that some targets of these microRNAs such as STAT3, PTBP1 or SIRT1 are differentially expressed in gliomas consistent with deregulation of microRNA expression. Moreover, MDH1, the target of several deregulated microRNAs, is repressed in glioblastomas, making an intramitochondrial-NAD reduction mediated by the mitochondrial aspartate-malate shuttle unlikely. Understanding the connections between microRNAs and bioenergetic pathways in gliomas may lead to identification of novel therapeutic targets

The gut microbiome: scourge, sentinel or spectator?

The gut microbiota consists of trillions of prokaryotes that reside in the intestinal mucosa. This long-established commensalism indicates that these microbes are an integral part of the eukaryotic host. Recent research findings have implicated the dynamics of microbial function in setting thresholds for many physiological parameters. Conversely, it has been convincingly argued that dysbiosis, representing microbial imbalance, may be an important underlying factor that contributes to a variety of diseases, inside and outside the gut. This review discusses the latest findings, including enterotype classification, changes brought on by dysbiosis, gut inflammation, and metabolic mediators in an attempt to underscore the importance of the gut microbiota for human health. A cautiously optimistic idea is taking hold, invoking the gut microbiota as a medium to track, target and treat a plethora of diseases