Yoshihide Matsumoto

Deletion of nardilysin prevents the development of steatohepatitis and liver fibrotic changes

Nonalcoholic steatohepatitis (NASH) is an inflammatory form of nonalcoholic fatty liver disease that progresses to liver cirrhosis. It is still unknown how only limited patients with fatty liver develop NASH. Tumor necrosis factor (TNF)-α is one of the key molecules in initiating the vicious circle of inflammations. Nardilysin (N-arginine dibasic convertase; Nrd1), a zinc metalloendopeptidase of the M16 family, enhances ectodomain shedding of TNF-α, resulting in the activation of inflammatory responses. In this study, we aimed to examine the role of Nrd1 in the development of NASH. Nrd1+/+ and Nrd1−/− mice were fed a control choline-supplemented amino acid-defined (CSAA) diet or a choline-deficient amino acid-defined (CDAA) diet. Fatty deposits were accumulated in the livers of both Nrd1+/+ and Nrd1−/− mice by the administration of the CSAA or CDAA diets, although the amount of liver triglyceride in Nrd1−/− mice was lower than that in Nrd1+/+ mice. Serum alanine aminotransferase levels were increased in Nrd1+/+ mice but not in Nrd1−/− mice fed the CDAA diet. mRNA expression of inflammatory cytokines were decreased in Nrd1−/− mice than in Nrd1+/+ mice fed the CDAA diet. While TNF-α protein was detected in both Nrd1+/+ and Nrd1−/− mouse livers fed the CDAA diet, secretion of TNF-α in Nrd1−/− mice was significantly less than that in Nrd1+/+ mice, indicating the decreased TNF-α shedding in Nrd1−/− mouse liver. Notably, fibrotic changes of the liver, accompanied by the increase of fibrogenic markers, were observed in Nrd1+/+ mice but not in Nrd1−/− mice fed the CDAA diet. Similar to the CDAA diet, fibrotic changes were not observed in Nrd1−/− mice fed a high-fat diet. Thus, deletion of nardilysin prevents the development of diet-induced steatohepatitis and liver fibrogenesis. Nardilysin could be an attractive target for anti-inflammatory therapy against NASH.

The BRG1/SOX9 axis is critical for acinar cell–derived pancreatic tumorigenesis

Chromatin remodeler Brahma related gene 1 (BRG1) is silenced in approximately 10% of human pancreatic ductal adenocarcinomas (PDAs). We previously showed that BRG1 inhibits the formation of intraductal pancreatic mucinous neoplasm (IPMN) and that IPMN-derived PDA originated from ductal cells. However, the role of BRG1 in pancreatic intraepithelial neoplasia–derived (PanIN-derived) PDA that originated from acinar cells remains elusive. Here, we found that exclusive elimination of Brg1 in acinar cells of Ptf1a-CreER; KrasG12D; Brg1fl/fl mice impaired the formation of acinar-to-ductal metaplasia (ADM) and PanIN independently of p53 mutation, while PDA formation was inhibited in the presence of p53 mutation. BRG1 bound to regions of the Sox9 promoter to regulate its expression and was critical for recruitment of upstream regulators, including PDX1, to the Sox9 promoter and enhancer in acinar cells. SOX9 expression was downregulated in BRG1-depleted ADMs/PanINs. Notably, Sox9 overexpression canceled this PanIN-attenuated phenotype in KBC mice. Furthermore, Brg1 deletion in established PanIN by using a dual recombinase system resulted in regression of the lesions in mice. Finally, BRG1 expression correlated with SOX9 expression in human PDAs. In summary, BRG1 is critical for PanIN initiation and progression through positive regulation of SOX9. Thus, the BRG1/SOX9 axis is a potential target for PanIN-derived PDA.

Nardilysin controls intestinal tumorigenesis through HDAC1/p53–dependent transcriptional regulation

Colon cancer is a complex disease affected by a combination of genetic and epigenetic factors. Here we demonstrate that nardilysin (N-arginine dibasic convertase; NRDC), a metalloendopeptidase of the M16 family, regulates intestinal tumorigenesis via its nuclear functions. NRDC is highly expressed in human colorectal cancers. Deletion of the Nrdc gene in ApcMin mice crucially suppressed intestinal tumor development. In ApcMin mice, epithelial cell-specific deletion of Nrdc recapitulated the tumor suppression observed in Nrdc-null mice. Moreover, epithelial cell-specific overexpression of Nrdc significantly enhanced tumor formation in ApcMin mice. Notably, epithelial NRDC controlled cell apoptosis in a gene dosage-dependent manner. In human colon cancer cells, nuclear NRDC directly associated with HDAC1, and controlled both acetylation and stabilization of p53, with alterations of p53 target apoptotic factors. These findings demonstrate that NRDC is critically involved in intestinal tumorigenesis through its epigenetic regulatory function, and targeting NRDC may lead to a novel prevention or therapeutic strategy against colon cancer.

Nardilysin inhibits pancreatitis and suppresses pancreatic ductal adenocarcinoma initiation in mice

Objective Nardilysin (NRDC), a zinc peptidase, exhibits multiple localisation-dependent functions including as an enhancer of ectodomain shedding in the extracellular space and a transcriptional coregulator in the nucleus. In this study, we investigated its functional role in exocrine pancreatic development, homeostasis and the formation of pancreatic ductal adenocarcinoma (PDA). Design We analysed Ptf1a-Cre; Nrdcflox/flox mice to investigate the impact of Nrdc deletion. Pancreatic acinar cells were isolated from Nrdcflox/flox mice and infected with adenovirus expressing Cre recombinase to examine the impact of Nrdc inactivation. Global gene expression in Nrdc-cKO pancreas was analysed compared with wild-type pancreas by microarray analysis. We also analysed Ptf1a-Cre; KrasG12D; Nrdcflox/flox mice to investigate the impact of Nrdc deletion in the context of oncogenic Kras. A total of 51 human samples of pancreatic intraepithelial lesions (PanIN) and PDA were examined by immunohistochemistry for NRDC. Results We found that pancreatic deletion of Nrdc leads to spontaneous chronic pancreatitis concomitant with acinar-to-ductal conversion, increased apoptosis and atrophic pancreas in mice. Acinar-to-ductal conversion was observed mainly through a non-cell autonomous mechanism, and the expression of several chemokines was significantly increased in Nrdc-null pancreatic acinar cells. Furthermore, pancreatic deletion of Nrdc dramatically accelerated KrasG12D -driven PanIN and subsequent PDA formation in mice. These data demonstrate a previously unappreciated anti-inflammatory and tumour suppressive functions of Nrdc in the pancreas in mice. Finally, absence of NRDC expression was observed in a subset of human PanIN and PDA. Conclusion Nrdc inhibits pancreatitis and suppresses PDA initiation in mice.

Gene expression profile of Dclk1+ cells in intestinal tumors

BACKGROUND Accumulating evidence has shown the existence of tumor stem cells with therapeutic potential. Previously, we reported that doublecortin like kinase 1 (Dclk1) marks tumor stem cells but not normal stem cells in the intestine of ApcMin/+ mice, and that Dclk1- and Lgr5-double positive tumor cells are the tumor stem cells of intestinal tumors. AIM To investigate molecules highly expressed in the Dclk1+ normal intestinal and Dclk1+ tumor cells in ApcMin/+ mice. METHODS We used microarray analyses to examine the gene expression profile of Dclk1+ cells in both mouse normal intestinal epithelium and ApcMin/+ mouse intestinal tumors. We also performed immunofluorescence analyses. RESULTS Genes related to microtubules and the actin cytoskeleton (e.g., Rac2), and members of the Src family kinases (i.e., Hck, Lyn, Csk, and Ptpn6) were highly expressed in both Dclk1+ normal intestinal and Dclk1+ tumor cells. Phosphorylated Hck and phosphorylated Lyn were expressed in Lgr5+ cells in the intestinal tumors of Lgr5EGFP-IRES-CreERT2/+; ApcMin/+ mice. CONCLUSION We revealed factors that are highly expressed in Dclk1+ intestinal tumor cells, which may help to develop cancer stem cell-targeted therapy in future.

An Irregular-Shaped Stenosis of the Sigmoid Colon

Gastroent Question: A 77year-old woman was admitted to our hospital for lower abdominal pain and diarrhea. She had undergone endoscopic mucosal resection to remove a 5-mm diameter rectal neuroendocrine tumor (NET) 10 years earlier. The endoscopic mucosal resection had been successfully performed with a negative surgical margin, and histopathology of the resected specimens suggested no vascular or lymphatic invasion. Blood tests on admission were normal, including tumor markers. Computed tomography showed wall thickening of the sigmoid colon. Colonoscopy revealed an irregular-shaped stenosis with edematous mucosa (Figure A). Multiple biopsies obtained from the stricture showed no significant findings. A gastrograffin enema revealed a stricture, which was approximately 20 cm (Figure B). What is the diagnosis? See the Gastroenterology web site (www.gastrojournal.org) for more information on submitting your favorite image to Clinical Challenges and Images in GI.