发酵红参总皂苷对高糖培养大鼠心肌间质成纤维细胞的保护作用及其机制  

Protective effect of fermented red ginseng total saponins on rat myocardial interstitial fibroblasts cultured with high glucose and its mechanism

作　　者：曲萌[1] 翁诗雅 郑鸿 李焱 高润泽 王胜告[3] 于春艳[2] 陈博学 董志恒[2] QU Meng;WENG Shiya;ZHENG Hong;LI Yan;GAO Runze;WANG Shenggao;YU Chunyan;CHEN Boxue;DONG Zhiheng(Department of Biochemistry and Molecular Biology,College of Basic Medical Sciences,Beihua University,Jilin 132013,China;Department of Pathology,College of Basic Medical Sciences,Beihua University,Jilin 132013,China;Department of Anatomy,College of Basic Medical Sciences,Beihua University,Jilin 132013,China)

机构地区：[1]北华大学基础医学院生物化学与分子生物学教研室,吉林吉林132013 [2]北华大学基础医学院病理学教研室,吉林吉林132013 [3]北华大学基础医学院解剖学教研室,吉林吉林132013

出　　处：《吉林大学学报（医学版）》2021年第5期1201-1208,共8页Journal of Jilin University：Medicine Edition

基　　金：吉林省卫健委卫生与健康技术创新项目(2020J010);吉林省中医药管理局科技项目(2018109);国家级大学生创新训练项目(202010201002,202110201028);吉林省大学生创新训练项目(201910201098);北华大学研究生创新计划项目(2019070);北华大学科研平台项目(20140301);北华大学大学生创新训练项目(202010201157)。

摘　　要：目的:观察发酵红参总皂苷(FRGTS)对高糖培养的大鼠心肌间质成纤维细胞(cFb)增殖和胶原合成的作用,并阐明其作用机制。方法:采用胰酶消化和差速贴壁法分离提取1~3 d龄SD大鼠cFb细胞,培养、传代后,取2~3代细胞用于实验;将细胞分成正常糖对照组(5.5 mmol·L^(-1) D-葡萄糖)、高糖组(25 mmol·L^(-1) D-葡萄糖)、高糖+15 mg·L^(-1) FRGTS组(F15组)和高糖+30 mg·L^(-1) FRGTS组(F30组);采用MTT法检测各组cFb细胞增殖活性,酶联免疫吸附测定(ELISA)法检测各组cFb细胞培养上清液中Ⅰ型和Ⅲ型胶原蛋白水平,实时荧光定量PCR法(RT-qPCR)检测各组cFb细胞中Smad3和Smad7 mRNA表达水平,Western blotting法检测各组cFb细胞中尾加压素Ⅱ(UⅡ)、转化生长因子β1(TGF-β1)、Smad3和Smad7蛋白表达水平。结果:与正常糖对照组比较,高糖组cFb细胞增殖活性明显升高(P<0.01),Ⅰ型和Ⅲ型胶原蛋白水平明显升高(P<0.01);与高糖组比较,F15组和F30组cFb细胞增殖活性降低(P<0.05),其中F30组抑制作用强于F15组。发酵红参总皂苷干预24 h后,F15组和F30组细胞培养上清液中Ⅰ型及Ⅲ型胶原蛋白水平明显低于高糖组(P<0.05),其中F30组作用较F15组更为明显。与正常糖对照组比较,高糖组cFb细胞中UⅡ和TGF-β1蛋白表达水平明显升高(P<0.01),Smad3 mRNA和蛋白的表达水平明显升高(P<0.01),而Smad7 mRNA和蛋白的表达水平则明显降低(P<0.05);与高糖组比较,发酵红参总皂苷干预24 h后,F15组和F30组cFb细胞中UⅡ和TGF-β1蛋白表达水平明显降低(P<0.01),Smad3 mRNA和蛋白表达水平明显降低(P<0.05或P<0.01),而Smad7 mRNA和蛋白表达水平则明显升高(P<0.05或P<0.01),其中F30组作用优于F15组。结论:FRGTS能有效抑制高糖所致的纤维化效应,保护糖尿病大鼠心肌,其作用机制与阻抑cFb细胞中UⅡ活化和调节TGF-β1/Smads传导通路有关。Objective:To observe the effect of fermented red ginseng total saponins(FRGTS)on the proliferation and collagen synthesis of the rat myocardial interstitial fibroblasts(cFb)cultured in high glucose,and to clarify their mechanisms.Methods:The cFb were isolated and extracted from 1-3 old SD rats by trypsin digestion and differential adhesion method.After culture and passage,the 2-3 generations of cFb were taken for experimental study.The cells were divided into normal glucose control group(5.5 mmol·L^(-1)D-glucose),high glucose group(25 mmol·L^(-1)D-glucose),high glucose+15 mg·L^(-1)FRGTS group(F15 group),and high glucose+30 mg·L^(-1)FRGTS group(F30 group).MTT assay was used to detect the proliferation activities of cFb in various groups,ELISA method was used to detect the levels of collagenⅠand collagenⅢproteins in the cFb supernatant in various groups,Real-time fluorescence quantitative PCR(RT-qPCR)was used to detect the expression levels of Smad3 and Smad7 mRNA,and Western blotting method was used to detect the expression levels of urotensinⅡ(UⅡ),transforming growth factor beta 1(TGF-β1),Smad3 and Smad7 proteins in the cFb in various groups.Results:Compared with normal glucose control group,the proliferation activities of cFb in high glucose group was significantly increased(P<0.01),and the levels of collagenⅠand collagenⅢproteins were significantly increased(P<0.01).compared with high glucose group,the proliferation abilities of cFb in F15 and F30 groups were decreased(P<0.05),and the inhibitory effect of F30 group was stronger than that of F15 group.After 24 h of intervention with FRGTS,the levels of collagenⅠand collagenⅢproteins in the cFb supernatant were significantly lower than those in high glucose group(P<0.05),and the effect of F30 group was more significant than that of F15 group.Compared with normal glucose control group,the expression levels of UⅡand TGF-β1 proteins in the cFb in high glucose group were significantly increased(P<0.01),the mRNA and protein expression levels of

关 键 词：发酵红参总皂苷 高糖 细胞增殖 尾加压素Ⅱ 转化生长因子β1 SMADS信号通路 

分 类 号：R256.2[医药卫生—中医内科学] R587.1[医药卫生—中医学]