论文部分内容阅读
目的 观察携带反义凝血酶受体 (ATR)和p2 1的双基因载体共表达后对血管平滑肌细胞增殖的抑制作用 ,为再狭窄基因治疗寻求新途径。 方法 以携带ATR或 和p2 1的单、双基因载体重组腺病毒伴随病毒(rAAV)感染培养的人主动脉平滑肌细胞 (hASMC) ,用半定量RT PCR检测各基因的整合与表达 ,MTT法测定病毒感染后不同时间点的细胞存活率。将携带AP双基因的rAAV导入WKY大鼠拉伤侧的颈总动脉 ,免疫组织化学法分别检测凝血酶受体 (TR)和p2 1两基因在动脉壁中的表达。 结果 RT PCR结果显示 ,TR单基因的mRNA表达降低 ,p2 1单基因表达升高 ,AP双基因得到了共表达 ;MTT法测定的生长曲线显示 ,双基因对hASMC增殖的抑制作用大于两个单基因 ;免疫组织化学证实 ,AP双基因导入后 ,TR基因的表达受到完全抑制 ,p2 1基因的表达明显增加 ,血管新生内膜与平滑肌细胞的增殖受到了抑制。 结论 ATR和p2 1双基因共表达在体外可明显抑制ASMC的增殖 ,在体内可明显抑制血管内膜新生和中膜的增生。
OBJECTIVE: To observe the inhibitory effect of co-expression of antisense thrombin receptor (ATR) and p2 1 double gene vectors on the proliferation of vascular smooth muscle cells (VSMCs), so as to seek new ways for the gene therapy of restenosis. Methods Human aortic smooth muscle cells (hASMC) were infected with the single and dual gene recombinant adenovirus associated virus (rAAV) carrying ATR or p21. The integration and expression of each gene were detected by semi-quantitative RT-PCR. MTT assay Cell viability at different time points after virus infection. The rAAV carrying AP double gene was transduced into the common carotid arteries of WKY rats. The expression of thrombin receptor (TR) and p21 gene in arterial wall were detected by immunohistochemistry. Results The results of RT-PCR showed that mRNA expression of TR gene was decreased, single gene of p2 1 was increased and AP double gene was co-expressed. The growth curve measured by MTT assay showed that double gene inhibited the proliferation of hASMC more than two single Immunohistochemistry confirmed that the expression of TR gene was completely inhibited after AP double gene introduction, the expression of p21 gene was significantly increased, and the proliferation of vascular neointimal and smooth muscle cells was inhibited. Conclusions Co-expression of ATR and p21 bi-genes can significantly inhibit the proliferation of ASMC in vitro and can significantly inhibit the proliferation of neointima and media in vivo.