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目的研究靶向自发性高血压大鼠热休克蛋白27(HSP27)基因的小干扰RNA(siRNA)对血管平滑肌细胞(VSMC)骨架及迁移的影响。方法以携带siRNA的慢病毒感染VSMC,以FITC-鬼笔环肽标记F-actin,在激光共聚焦显微镜下观察细胞骨架的形态变化;采用改良的Boyden微孔膜双槽法进行细胞迁移实验。结果正常培养未使用血管紧张素Ⅱ(AngⅡ)刺激的VSMC,细胞内无或仅存在少许无规则、散在分布的F-actin,经AngⅡ诱导后,VSMC内F-actin数量明显增加,纵向平行排列,且形成应力纤维丝。AngⅡ能够明显诱导VSMC细胞的迁移,pNL-HSP27-EGFP能明显抑制AngⅡ诱导的VSMC迁移,抑制率为40.0%(P<0.01)。结论靶向HSP27基因的siRNA能显著降低AngⅡ诱导的VSMC迁移,HSP27在细胞的迁移中起重要作用。
Objective To investigate the effects of heat shock protein 27 (HSP27) gene silencing RNA on the skeleton and migration of vascular smooth muscle cells (VSMCs) in spontaneously hypertensive rats. Methods VSMCs were infected with lentivirus carrying siRNA and F-actin was labeled with FITC-phalloidin. The morphological changes of cytoskeleton were observed under laser scanning confocal microscopy. Cell migration was performed using a modified Boyden microporous membrane dual-chamber method. Results The VSMCs stimulated with Ang Ⅱ were not cultured in normal conditions. There was no or only a small amount of F-actin in the cells. After induced by AngⅡ, the number of F-actin in VSMC significantly increased, and the longitudinal parallel arrangement , And forms a stress fiber filament. AngⅡ could obviously induce VSMC cell migration, pNL-HSP27-EGFP could significantly inhibit the migration of VSMC induced by AngⅡ, with the inhibition rate of 40.0% (P <0.01). Conclusion siRNA targeted to HSP27 can significantly reduce the migration of VSMC induced by AngⅡ, and HSP27 plays an important role in the migration of cells.