let-7a对血管平滑肌细胞迁移、增殖功能调控作用的体外研究

目的:观察let-7a对体外培养血管平滑肌细胞(VSMC)迁移和增殖能力的影响。方法:分离大鼠胸腹主动脉VSMC,并传代培养。将培养的VSMC通过慢病毒表达载体分别转染let-7a模拟物(实验组)及其阴性对照物(阴性对照组),或不做转染处理(空白对照组)。通过绿色荧光蛋白的表达估计转染效率;用real-time PCR检测各组细胞let-7a的表达水平;用细胞划痕试验和CCK-8法分别检测各组细胞的迁移能力及增殖情况。结果:荧光显微镜观察显示,96 h后细胞的转染效率达85%以上;real-time PCR结果显示,实验组VSMC的let-7a表达水平明显高于阴性对照组和空白对照组(均P<0.01);细胞划痕试验与增殖检测结果显示,实验组的细胞迁移数及增殖率明显低于阴性对照组和空白对照组(均P<0.01)。阴性对照组与空白对照组间上述项目无明显差异(均P>0.05)。结论:let-7a对体外培养的VSMC迁移与增殖有抑制作用。

Regulatory effect of let-7a on migration and proliferation of vascular smooth muscle cells: an in vitro study

Objective: To investigate the effects of let-7a on the migration and proliferation ability of vascular smooth muscle cells (VSMCs) cultured in vitro. Methods: VSMCs were isolated from the thoracoabdominal aorta of rats and then, were cultured and subcultured. Subsequently, the cultured VSMCs were assigned either to let-7a mimics (experimental group) or its negative control (negative control group) transfection using lentiviral expression vector, or no transfection treatment (blank control group). Transfection efficiency was estimated by expression of green fluorescence protein. In the cells of each group, the let-7a expression was detected by real-time PCR method, and the cell migration ability and proliferation status were determined by scratch test and CCK-8 assay, respectively. Results: Fluorescence microscopic observation revealed that the cell transfection efficiency reached more than 85%. Real-time PCR results showed that the let-7a expression was significantly higher in VSMCs of experimental group than that of either negative control group or blank control group (both P<0.01). The results of cell scratch test and proliferation measurement demonstated that both the migration and proliferative ability of the cells of experimental group were significantly decreased compared with those of negative control group and blank control group (all P<0.01), while all of the above parameters showed no obvious differences between negative control group and blank control group (all P>0.05) Conclusion: Let-7a has inhibitory effect on migration and proliferation of VSMCs cultured in vitro.