miR-425-5p对脂多糖诱导的肠道L细胞GLP-1分泌的影响及机制研究  

Effect of miR-425-5p on GLP-1 secretion in intestinal L cells induced by lipopolysaccharide and its mechanism

作　　者：王娇[1] 韦丽瑞 黄凤姣 赵雪楠 郭丰[1] 吴丽娜[1] 刘彦玲 秦贵军[1] Wang Jiao;Wei Lirui;Huang Fengjiao;Zhao Xuenan;Guo Feng;Wu Lina;Liu Yanling;Qin Guijun(Department of Endocrinology,the First Affiliated Hospital of Zhengzhou University,Zhengzhou 450052,China)

机构地区：[1]郑州大学第一附属医院内分泌与代谢病科,450052

出　　处：《中华内分泌代谢杂志》2021年第7期646-652,共7页Chinese Journal of Endocrinology and Metabolism

基　　金：国家自然科学基金(81500646);河南省科技项目(192102310064)。

摘　　要：目的探讨miR-425-5p对脂多糖诱导的肠道L细胞胰升糖素样肽1(GLP-1)分泌的影响及其机制。方法采用脂多糖孵育肠道L细胞系GLUTag细胞,检测miR-425-5p和GLP-1的表达和分泌;采用MTT法测定细胞活力,流式细胞术检测细胞凋亡。采用实时定量PCR和Western印迹法检测miR-425-5p、磷酸酶和张力蛋白同源基因(PTEN)、胰升糖素原mRNA和蛋白的表达。通过检测TOP/FOP比率测定Wnt/β-连环蛋白(β-catenin)信号通路的活性。采用双荧光素酶报告基因实验和染色质免疫共沉淀(ChIP)明确miR-425-5p与PTEN、β-catenin之间的相互作用。结果在GLUTag细胞中,随着脂多糖浓度的升高,miR-425-5p表达增加,活性GLP-1水平降低,细胞凋亡增加,细胞活力下降;而且miR-425-5p参与脂多糖对GLP-1表达和肠道L细胞活力的调节作用。抑制miR-425-5p可降低胰升糖素原mRNA表达和TOP/FOP比率,提高PTEN蛋白水平,抑制细胞活力。在脂多糖诱导下,miR-425-5p通过靶向PTEN上调β-catenin的表达水平,而β-catenin作为顺式作用元件诱导胰升糖素原的转录,进而促进GLP-1的表达。结论在脂多糖诱导的肠道L细胞中,miR-425-5p通过靶向PTEN调控β-catenin水平进而促进GLP-1的分泌。Objective To investigate the effect of miR-425-5p on glucagon-like peptide-1(GLP-1)secretion in intestinal L cells induced by lipopolysaccharide(LPS),and to explore its mechanism.Methods GLUTag cells of intestinal L cell line were incubated with LPS to determine the levels of miR-425-5p and GLP-1.Cell viability was determined by MTT assay,and cell apoptosis was detected by flow cytometry.Quantitative real time-PCR and western blot were performed to determine the expressions of miR-425-5p,phosphatase and tensin homology(PTEN),proglucagon,and GLP-1.Activity of Wnt/β-catenin signaling pathway was determined by detecting TOP/FOP ratio.Interaction among miR-425-5p,PTEN,andβ-catenin was analyzed using luciferase activity assay and chromatin immunoprecipitation(ChIP)assay.Results In GLUTag cells,with the elevation of LPS concentration,the expression of miR-425-5p and the apoptosis rate were increased,while the level of active GLP-1 and the cell viability were decreased.MiR-425-5p was involved in the regulation of LPS on GLP-1 secretion and intestinal L cell viability.Inhibition of miR-425-5p reduced the mRNA expression of proglucagon and the TOP/FOP ratio,increased PTEN protein level,and inhibited cell viability.In LPS-treated GLUTag cells,miR-425-5p increased the level ofβ-catenin by targeting PTEN,andβ-catenin acted as a cis-acting element to induce the transcription of proglucagon and promote the secretion of GLP-1.Conclusion In LPS-induced intestinal L cells,miR-425-5p promotes the expression of GLP-1 by targeting PTEN to modulateβ-catenin.

关 键 词：miR-425-5p 脂多糖 磷酸酶和张力蛋白同源基因 Β-连环蛋白 肠道L细胞 

分 类 号：R587.1[医药卫生—内分泌]