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目的研究腺相关病毒载体介导人血管内皮生长因子基因(AAV-VEGF121)的体外促血管内皮细胞生长作用。方法 RT-PCR法获取人VEGF121基因,克隆入pAAV-MCS载体中,构建成pAAV-VEGF121(处理组),以空白载体克隆入pAAV-MCS病毒(对照组);两组分别在AAV-293细胞中包装成pAAV-VEGF121病毒颗粒和空白病毒颗粒。相同条件下,两组分别感染人脐静脉内皮细胞(HUVEC),半定量RT-PCR检测细胞内VEGF121基因mRNA的表达;CCK-8法测取OD450nm值,绘制细胞增殖曲线;电子显微镜观察细胞超微结构的改变。结果成功构建AAV-VEGF121;半定量RT-PCR显示处理组细胞内VEGF121的mRNA表达水平高于对照组(P<0.01);细胞增殖曲线显示处理组细胞增殖能力明显高于对照组;透射电镜显示处理组细胞内与蛋白质合成有关的细胞器增生明显活跃。结论 AAV-VEGF121基因对血管内皮细胞的生成具有促进作用。
Adeno-associated virus vector-mediated human vascular endothelial growth factor gene (AAV-VEGF121) in vitro vascular endothelial cell growth. Methods The human VEGF121 gene was obtained by RT-PCR and cloned into pAAV-MCS vector to construct pAAV-VEGF121 (control group). The recombinant plasmid was cloned into pAAV-MCS vector (control group) Packaged as pAAV-VEGF121 virus particles and blank virus particles. Under the same conditions, the two groups were infected with human umbilical vein endothelial cells (HUVECs) respectively, and the expression of VEGF121 mRNA was detected by semi-quantitative RT-PCR. The OD450nm was measured by CCK-8 and the cell proliferation curve was drawn. Microstructure changes. Results AAV-VEGF121 was successfully constructed. The mRNA expression of VEGF121 in the treated group was higher than that in the control group (P <0.01) by semi-quantitative RT-PCR. The cell proliferation curve showed that the proliferative capacity of the treated group was significantly higher than that of the control group The proliferation of organelles involved in protein synthesis in the treated group was markedly active. Conclusion AAV-VEGF121 gene can promote the formation of vascular endothelial cells.