活化型肝星状细胞中miRNA-193下调α-平滑肌肌动蛋白的表达

miRNA-193 down-regulates the expression of a-smooth muscle actin in activative hepatic stellate cells

目的：研究活化型肝星状细胞（HSC）中miRNA-193（miR-193）下调肝纤维化相关基因的表达。方法：将大鼠miR-193的前体序列（pre-miR-193）克隆到真核表达载体pcDNA3．1（＋）中，经酶切和DNA测序鉴定，获得质粒pcDNA3．1-miR-193；通过脂质体转染法将质粒转染到大鼠的活化型肝星状细胞（HSC-T6）中，采用荧光素酶报告基因分析法和免疫印迹检测肝纤维相关基因的表达。结果：构建的真核表达载体pcDNA3．1-miR-193经酶切鉴定及DNA测序显示，目的片段的大小与预期结果一致；荧光素酶报告基因分析法和免疫印迹检测显示，重组质粒转染到活化型HSC-T6中后，肝纤维化相关基因的α-平滑肌肌动蛋白（a-SMA）的表达呈明显下降趋势，而胶原蛋白IαI和Iα2的下调作用不明显。结论：miR-193能抑制活化型肝星状细胞中肝纤维化相关基因α-SMA的表达。

Objective： To study how miRNA-193 down-regulates the expression of hepatic fibrosis related genes in activative hepatic steLLate ceils （MS（Z）. Methods： After searching for the precursor sequence of miR-193 （pre-miR-193） of rats in Genbank from NCBI, we constructed eukaryotic expression plasmid pcDNA3.1-miR-193 through cloning the pre-miR-193 into the eukaryotic expression vector, pcDNA3. 1 （＋）, and then identified it using the methods of restriction and DNA sequencing assays. The expressions of hepatic fibrosis-ralated genes （smooth muscle actin （a-SMA）, collagen IαI/a2） were evaluated in activative HSC line of rats （HSC-T6） through Western bolt assay when the plasmid pcDNA3.1-miR-193 was transfeted into HSC-T6. Results： The eukaryotic expression plasmid pcDNA3.1-miR-193, which was almost consistent with the result of anticipation, was constructed successfully through the methods of restriction and DNA sequencing assays. The expression of gene α-SMA presented dramatic decline through luciferase activity and Western blot assays after the plasmid pcDNA3.1-miR- 193 was transfected into HSC-T6, but the expressions were not obviously changed for genes of collagen [ α1/α2. Conclusion： miRNA-193 can down-regulate the expression of gene α-SMA in aetivative HSC.