Establishment of mouse intestinal myofibroblast cell lines
Establishment of mouse intestinal myofibroblast cell lines作者机构：Laboratory of Veterinary Pharmacology Joint Faculty of Veterinary Medicine Yamaguchi University Department of Veterinary Pharmacology Graduate School of Agriculture and Life Sciences the University of Tokyo
出 版 物：《World Journal of Gastroenterology》 (世界胃肠病学杂志（英文版）)
年 卷 期：2013年第19卷第17期
页 面：2629-2637页
核心收录：
学科分类：090601[农学-基础兽医学] 1002[医学-临床医学] 09[农学] 0906[农学-兽医学]
基 金：Supported by Uehara Memorial Foundation Mishima Kaiun Memorial Foundation A Grant-in-Aid for Scientific Research from the Japanese Ministry of Education,Culture,Sports,Science and Technology
主 题：Cell line Colon Lipopolysaccharide Mouse Myofibroblast
摘 要：AIM:To establish novel intestinal myofibroblast (IMF) cell lines from mouse colonic mucosa and investigate their biological characters. METHODS:Primary IMFs were isolated from mucosal tissues of mouse colon that was denuded of epithelial cells and smooth muscle layer. For immortalization, primary IMFs were transfected with simian virus 40 large T antigen (designated as LmcMF). We also isolated some primary IMFs that spontaneously became immortalized without transfection (designated as SmcMF). To check immortality and normality of these cells, we examined their proliferative ability and contact inhibition. Moreover, the expression levels of proteins characterizing IMFs [including α-smooth muscle actin (α-SMA), vimentin, desmin, and type Ⅰ collagen] and proteins associated with the immune response [such as toll-like receptor 4 (TLR-4), CD14, MD2, IκBα, and p-p38] were determined by Western blotting. The localization of several myofibroblast protein markers was also detected by immunofluorescence staining. RESULTS:The cell growth assay results show that both LmcMF and SmcMF cells proliferated logarithmically at least up to passage 20. In addition, the contact inhibition assays show that LmcMF and SmcMF stopped growing after the cells reached confluence. These data suggest that these 2 types of cells were immortalized without losing contact inhibition of growth. Moreover, both LmcMF and SmcMF, like primary IMFs, showed spindle-shaped appearance. The expression levels of key myofibroblast protein markers, including α-SMA, vimentin, and desmin, were also examined by the Western blotting and immunofluorescence analyses. Our results show that these cells were positive for α-SMA and vimentin, but not desmin, as well as that both LmcMF and SmcMF expressed type Ⅰ collagen at a lower level than primary IMFs. Finally, we investigated the expression level of lipopolysaccharide (LPS) receptor-related proteins, as well as the response of the cells to LPS treatment. We found that th
维普期刊数据库