DNMT1在SD大鼠心肌成纤维细胞增殖中的作用  

Role of DNMT1 onproliferation of cardiac fibroblasts in SD rats

作　　者：丁季飞 陶辉[1,2] 石开虎[1,2] 徐盛松[1,2] DING Ji-fei;TAO Hui;SHI Kai-hu;XU Sheng-song(Dept of Cardiothoracic Surgery, the Second Affiliated Hospital , Cardiovascular Research Center, Anhui Medical University, Hefei 230601, China)

机构地区：[1]安徽医科大学第二附属医院心胸外科,安徽合肥230601 [2]安徽医科大学第二附属医院心血管病研究中心,安徽合肥230601

出　　处：《中国药理学通报》2020年第8期1129-1133,共5页Chinese Pharmacological Bulletin

基　　金：安徽省自然科学基金资助项目(No 1808085MH231);国家自然科学基金资助项目(No 81570295)。

摘　　要：目的探讨DNA甲基化转移酶1(DNA methyltransferase 1,DNMT1)在SD大鼠心肌成纤维细胞(cardiac fibroblasts,CFs)增殖中的作用。方法取1~3 d的SD大鼠的心脏,剪碎消化后提取CFs,在显微镜下观察鉴定形态。细胞贴壁生长后分别向各组细胞转染siDNMT1(小干扰DNMT1)及阴性对照组(NC)24~48 h,运用qRT-PCR方法检测DNMT1、α-平滑肌肌动蛋白(α-SMA)和Ⅰ型胶原前胶原A1(Col1A1)的mRNA表达量;CCK-8法检测细胞增殖活性;Western blot检测DNMT1、α-SMA和Col1A1的蛋白表达量;细胞周期实验检测细胞周期分布情况。结果CFs中siDNMT1组中DNMT1表达下调。Western blot及qRT-PCR结果显示,α-SMA和Col1A1在siDNMT1组中的表达明显下降。CFs转染siDNMT1后培养48 h,CFs活力明显下降。细胞周期实验表明CFs转染siDNMT148 h后,与NC组相比,G 0和G 1期细胞比例升高明显,细胞周期被阻滞明显。结论DNMT1表达下调能抑制CFs的增殖,提示DNMT1可能是心肌纤维化发生发展中的关键调节点。Aim To investigate the role of DNA methyltransferase 1(DNMT1)in the proliferation of cardiac fibroblasts(CFs)in SD rats.Methods The hearts of SD rats were taken from 1 to 3 days.After cutting and digesting,CFs were extracted by differential adherence centrifugation and observed under microscope.After cells were adherently grown to appropriate density,the cells were transiently transfected with siDNMT1(small interfering DNMT1)and negative control group(NC)for 24 to 48 h.qRT-PCR was used to detect mRNA expression of DNMT1,α-smooth muscle actin(α-SMA)and type I collagen procollagen A1(Col1A1);cell proliferation activity by CCK-8 assay;protein expression of DNMT1,α-SMA and Col1A1 by Western blot;cell cycle distribution by cell cycle assay.Results The expression of DNMT1 was down-regulated in the small interfering DNMT1 group in CFs.The results of Western blot and qRT-PCR showed that the expression ofα-SMA and Col1A1 was significantly reduced in the small interference group.After CFs were transiently transfected with siDNMT1 for 48 h,the activity of CFs decreased significantly.Cell cycle assay showed that CFs were transiently transfected with siDNMT1 for 48 h,and the proportion of cells in G0 and G1 phase was significantly higher than that in NC group,and the cell cycle was arrested during this phase.Conclusions Down-regulation of DNMT1 can inhibit the proliferation of CFs,suggesting that DNMT1 may be a key regulatory point in the development of myocardial fibrosis.

关 键 词：DNMT1 心肌纤维化 心肌成纤维细胞 增殖 Α-SMA COL1A1 

分 类 号：R-332[医药卫生] R322.11