| 慢病毒载体CV072介导的Mfn2过表达抑制肝星状细胞活化 |
| |
| 引用本文： | 朱含章,葛珂,刘凌,沈伟敏,贾长库. 慢病毒载体CV072介导的Mfn2过表达抑制肝星状细胞活化[J]. 中国病理生理杂志, 2019, 0(6): 1112-1117 |
| |
| 作者姓名： | 朱含章 葛珂 刘凌 沈伟敏 贾长库 |
| |
| 作者单位： | 浙江大学医学院附属杭州市第一人民医院肝胆胰外科 |
| |
| 基金项目： | 浙江省自然科学基金资助项目(No.LY17H030002) |
| |
| 摘 要： | 目的:构建携带线粒体融合蛋白2(Mfn2)基因的慢病毒载体,并探讨Mfn2过表达对大鼠肝星状细胞活化的抑制作用及其减少肝纤维化相关因子生成的机制。方法:构建含Mfn2的慢病毒过表达载体CV072-pCMV-Mfn2-EGFP,转染肝星状细胞;荧光显微镜下观察细胞中绿色荧光蛋白的表达及转染效率;RT-qPCR和Western blot法检测转染后细胞中Mfn2的mRNA和蛋白表达水平;Western blot检测Bax、Bcl-2、cleaved caspase-3、α-SMA、TGF-β1、Smad2和Smad3的水平;ELISA检测Ⅰ、Ⅲ和Ⅳ型胶原蛋白水平。结果:与各对照组相比,慢病毒过表达载体CV072-pCMV-Mfn2-EGFP转染后,细胞促凋亡蛋白表达增多,TGF-β1/Smad通路蛋白TGF-β1、p-Smad和p-Smad3的蛋白水平均降低,纤维化相关蛋白α-SMA、I型胶原蛋白、Ⅲ型胶原蛋白和IV型胶原蛋白的水平降低(P<0.01)。结论:转染慢病毒过表达载体CV072-pCMV-Mfn2-EGFP在体外可有效抑制肝星状细胞活化,并可能通过抑制TGF-β1/Smad通路减少肝纤维化相关因子的生成。
|
| 关 键 词： | 线粒体融合蛋白2 肝纤维化 肝星状细胞 细胞凋亡 TGF-Β1/SMAD信号通路 |
Lentiviral vector CV072-mediated Mfn2 over-expression inhibits hepatic stellate cell activation |
| |
| ZHU Han-zhang,GE Ke,LIU Ling,SHEN Wei-min,JIA Chang-ku. Lentiviral vector CV072-mediated Mfn2 over-expression inhibits hepatic stellate cell activation[J]. Chinese Journal of Pathophysiology, 2019, 0(6): 1112-1117 |
| |
| Authors: | ZHU Han-zhang GE Ke LIU Ling SHEN Wei-min JIA Chang-ku |
| |
| Affiliation: | (Department of Hepatopancreatobiliary Surgery, Hangzhou First People’s Hospital, TheAffiliated Hospital of Medical School of Zhejiang University, Hangzhou 310006 , China) |
| |
| Abstract: | AIM: To construct a lentiviral vector carrying mitofusin 2(Mfn2), and to investigate the inhibitory effect of Mfn2 on the activation of rat hepatic stellate cells and its mechanism of reducing the formation of hepatic fibrosis-related factors. METHODS: The lentiviral over-expression vector CV072-pCMV-Mfn2-EGFP containing Mfn2 was constructed and transfected into the hepatic stellate cells. The expression of green fluorescent protein was observed under fluorescence microscope, and the transfection efficiency was evaluated. The protein levels of Bax, Bcl-2, cleaved caspase-3,α-SMA, TGF-β1, Smad2 and Smad3 were detected by Western blot. The levels of type I collagen, type III collagen and type IV collagen in the cell culture supernatants were determined by ELISA. RESULTS: Compared with control group, the apoptosis of the hepatic stellate cells transfected with lentivirus over-expression vector CV072-pCMV-Mfn2-EGFP was increased, and the protein levels of proapoptotic molecules Bax and cleaved caspase-3 were increased(P<0.01). TGF-β1/Smad pathway-related proteins TGF-β1, p-Smad2 and p-Smad3 were decreased, and the levels of fibrosis-related proteins α-SMA, type I collagen, type III collagen and type IV collagen were decreased(P<0.01). CONCLUSION: Transfection of lentiviral over-expression vector CV072-pCMV-Mfn2-EGFP effectively inhibits hepatic stellate cell activation in vitro and may reduce the production of hepatic fibrosis-related factors by inhibiting TGF-β1/Smad pathway. |
| |
| Keywords: | Mitofusin 2 Liver fibrosis Hepatic stellate cells Apoptosis TGF-β1/Smad signaling pathway |
| 本文献已被 CNKI 维普 等数据库收录！ |
|
