论文部分内容阅读
目的 观察腺病毒介导反义AT1基因转染对培养的大鼠动脉平滑肌细胞 (VSMCs)增殖的影响。方法 用定向克隆和同源重组方法构建携带人反义AT1基因的复制 -缺陷型重组腺病毒 (Ad/CMV·ahAT1) ,转染体外培养的VSMCs,用RT PCR半定量法和免疫组化法检测AT1R的表达 ,用3 H TdR掺入实验和流式细胞仪检测VSMCs的DNA合成和增殖指数。结果 与对照组相比 ,转染Ad/CMV·ahAT1后 4 8h的VSMCs,AT1R的mRNA表达低 5 0 % ,蛋白表达也显著低于对照组 (P <0 0 1)。给予AngⅡ刺激的VSMCs的3 H TdR掺入量和增殖指数明显高于空白对照组 (分别为P <0 0 5和P <0 0 1) ;转染Ad/CMV·ahAT1组3 H TdR掺入量和增殖指数则显著降低 (与DMEM组比P <0 0 5 ,与AngⅡ对照组比P <0 0 1)。 结论 腺病毒介导的反义AT1R转染 ,通过抑制AT1R的表达 ,明显抑制VSMCs的增殖和AngⅡ刺激的VSMCs增殖
Objective To observe the effect of adenovirus-mediated antisense AT1 gene transfection on the proliferation of cultured rat arterial smooth muscle cells (VSMCs). Methods Recombinant adeno-associated virus (Ad / CMV-ahAT1) carrying human antisense AT1 gene was constructed by directional cloning and homologous recombination. VSMCs cultured in vitro were transfected by semi-quantitative RT-PCR and immunohistochemistry The expression of AT1R was detected. DNA synthesis and proliferation index of VSMCs were detected by 3 H TdR incorporation assay and flow cytometry. Results Compared with the control group, the mRNA expression of AT1R was 50% lower and the protein expression was significantly lower in VSMCs transfected with Ad / CMV · ahAT1 48 h (P <0.01). The amount of 3 H TdR incorporation and proliferation index of VSMCs stimulated with AngⅡwere significantly higher than that of the blank control group (P <0.05 and P <0.01, respectively); 3 H TdR incorporation of Ad / CMV · ahAT1 group Volume and proliferation index were significantly lower (compared with DMEM group P <0 05, compared with Ang Ⅱ control group P <0 01). Conclusions Adenovirus-mediated antisense AT1R transfection can significantly inhibit the proliferation of VSMCs and the proliferation of VSMCs stimulated by AngⅡ by inhibiting the expression of AT1R