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A subset of myeloid-derived suppressor cells (MDSCs) that express murine Schlafen4 (SLFN4) or its human ortholog SLFN12L polarize in the Helicobacter -inflamed stomach coincident with intestinal or spasmolytic polypeptide-expressing metaplasia. We propose that individuals with a more robust response to damage-activated molecular patterns and increased Toll-like receptor 9 (TLR9) expression are predisposed to the neoplastic complications of Helicobacter infection. A mouse or human Transwell co-culture system composed of dendritic cells (DCs), 2-dimensional gastric epithelial monolayers, and Helicobacter were used to dissect the cellular source of interferon-α (IFNα) in the stomach by flow cytometry. Conditioned media from the co-cultures polarized primary myeloid cells. MDSC activity was determined by T-cell suppression assays. In human subjects with intestinal metaplasia or gastric cancer, the rs5743836 TLR9 T>C variant was genotyped and linked to TLR9, IFNα, and SLFN12L expression by immunohistochemistry. Nuclear factor-κB binding to the TLR9 C allele was determined by electrophoretic mobility shift assays. Helicobacter infection induced gastric epithelial and plasmacytoid DC expression of TLR9 and IFNα. Co-culturing primary mouse or human cells with DCs and Helicobacter induced TLR9, IFNα secretion, and SLFN+-MDSC polarization. Neutralizing IFNα in vivo mitigated Helicobacter -induced spasmolytic polypeptide-expressing metaplasia. The TLR9 minor C allele creates a nuclear factor-κB binding site associated with higher levels of TLR9, IFNα, and SLFN12L in Helicobacter -infected stomachs that correlated with a greater incidence of metaplasias and cancer. TLR9 plays an essential role in the production of IFNα and polarization of SLFN+ MDSCs on Helicobacter infection. Subjects carrying the rs5743836 TLR9 minor C allele are predisposed to neoplastic complications if chronically infected. [Display omitted] Some Helicobacter pylori –infected subjects have a genetic tendency to develop an aggressive immune response to bacterial and cellular debris that accumulates in the inflamed microenvironment, ultimately leading to pre-neoplastic sequelae. [ABSTRACT FROM AUTHOR]

Background & Aims: Math1 (Atoh1) is a basic helix-loop-helix transcription factor important for intestinal secretory cell differentiation. We hypothesized that Math1 is important in cell fate commitment, and therefore mediates proliferative homeostasis and the adaptive response following intestinal resection in the adult intestine. Methods: We generated mice with an intestine-specific mosaic deletion of Math1 (Math1Δintestine ) using the Cre/loxP system. Histologic analysis in adult Math1Δintestine and wild-type littermates at baseline and following small bowel resection or sham surgery was performed. Results: We observed loss of Paneth, goblet, and enteroendocrine cells in Math1-null crypts. In addition, aberrant activation of the Math1 promoter occurred in absorptive enterocytes derived from Math1-null crypts, suggesting a change in cell fate. Proliferation was increased but apoptosis unchanged in Math1-mutant crypts compared to adjacent wild-type crypts. Math1Δintestine mice and wild-type littermates displayed similar physiologic adaptive responses to small bowel resection as measured by changes in body weight and ileal wet weight. In contrast, Math1-mutant crypts displayed a blunted adaptive response compared to adjacent wild-type crypts. Conclusions: We show that Math1 is essential for adult intestinal secretory cell production, and in its absence cells destined to a secretory phenotype instead adopt an absorptive phenotype. Subtle abnormalities of proliferation within Math1-null crypts in Math1Δintestine mice were identified, together with a substantial defect in the adaptive response of Math1-null crypts following small bowel resection. Our results suggest that Math1 is critical for both cell fate determination within the intestinal epithelium and for regulation of the response to intestinal resection. [Copyright &y& Elsevier]

Background & Aims Little is known about the prevalence of nonalcoholic fatty liver disease (NAFLD) among severely obese adolescents or factors that determine its development. We investigated the prevalence of NAFLD in a multicenter cohort of adolescents undergoing bariatric surgery and the factors associated with it. Methods We enrolled 242 adolescents undergoing bariatric surgery between March 2007 and February 2012 at 5 tertiary care centers into a multicenter, prospective observational cohort study. Intraoperative core liver biopsies were collected from 165 subjects; 17 were excluded because of insufficient liver tissue or use of hepatotoxic medications, so 148 remained in the study (mean age, 16.8 ± 1.6 years; median body mass index = 52 kg/m 2 ). Liver tissues were analyzed by histology using validated criteria. Hepatic gene expression was analyzed in 67 samples. Results NAFLD was present in 59% of this predominantly female (72%), white (68%), non-Hispanic (91%) cohort. Of subjects with NAFLD, 24% had borderline and 10% had definite nonalcoholic steatohepatitis (NASH). Mild fibrosis (stage 2 or lower) was observed in 18% of liver biopsies and stage 3 was observed in 0.7%, but cirrhosis was not detected. Dyslipidemia was present in 78% of subjects, hypertension in 44%, and diabetes in 14%. More severe NAFLD was associated with increasing levels of alanine aminotransferase, fasting glucose level, hypertension (each P < .01), and white blood cell count ( P = .04). Only diabetes was associated with detection of fibrosis (odds ratio = 3.56; 95% confidence interval: 1.93−6.56). Microarray analysis associated presence of NASH with altered expression of genes that regulate macrophage chemotaxis, cholesterol absorption, and fatty acid binding. Conclusions More than half of adolescents undergoing bariatric surgery in this cohort had NAFLD, yet the prevalence of severe or fibrotic NASH was low. Increasing severity of NAFLD was associated with level of alanine aminotransferase and cardiometabolic risk factors, but not body mass index. Based on gene expression analysis, borderline and definite NASH were associated with abnormal immune function, intestinal cholesterol absorption, and lipid metabolism. [ABSTRACT FROM AUTHOR]

Background & Aims: Identification of intestinal stem cells (ISCs) has relied heavily on the use of transgenic reporters in mice, but this approach is limited by mosaic expression patterns and difficult to directly apply to human tissues. We sought to identify reliable surface markers of ISCs and establish a robust functional assay to characterize ISCs from mouse and human tissues. Methods: We used immunohistochemistry, real-time reverse-transcription polymerase chain reaction, and fluorescence-activated cell sorting (FACS) to analyze intestinal epithelial cells isolated from mouse and human intestinal tissues. We compared different combinations of surface markers among ISCs isolated based on expression of Lgr5–green fluorescent protein. We developed a culture protocol to facilitate the identification of functional ISCs from mice and then tested the assay with human intestinal crypts and putative ISCs. Results: CD44+CD24loCD166+ cells, isolated by FACS from mouse small intestine and colon, expressed high levels of stem cell–associated genes. Transit-amplifying cells and progenitor cells were then excluded based on expression of GRP78 or c-Kit. CD44+CD24loCD166+ GRP78lo/– putative stem cells from mouse small intestine included Lgr5-GFPhi and Lgr5-GFPmed/lo cells. Incubation of these cells with the GSK inhibitor CHIR99021 and the E-cadherin stabilizer Thiazovivin resulted in colony formation by 25% to 30% of single-sorted ISCs. Conclusions: We developed a culture protocol to identify putative ISCs from mouse and human tissues based on cell surface markers. CD44+CD24loCD166+, GRP78lo/–, and c-Kit− facilitated identification of putative stem cells from the mouse small intestine and colon, respectively. CD44+CD24−/loCD166+ also identified putative human ISCs. These findings will facilitate functional studies of mouse and human ISCs. [Copyright &y& Elsevier]