siRNA干扰热休克蛋白27基因对自发性高血压大鼠血管平滑肌细胞骨架及迁移的影响

The Effects of Heat Shock Protein 27 siRNA Interference on Cytoskeleton and Migration of Vascular Smooth Muscle Cells from Spontaneously Hypertensive Rats

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 study the effect of lentiviral RNA interference（RNAi） sequence targeting heat shock protein 27 （HSP27）gene of spontaneously hypertensive rats（SHR）on the cell cytoskeleton and migration of vascular smooth muscle cells （VSMC） from SHR.Methods VSMC were infected by the described lentivirus.The F-actin polymerization was detected by FITC-Phalloidine staining and examined by confocal microscopy.Modified Boyden chamber technique was used for the assessment of VSMC migration.Results Ang Ⅱ resulted in a substantial increase in the number of stress fibers and the rearrangement of these structures into ordered parallel arrays.The reorganization of actin cytoskeleton stimulated by Ang Ⅱ could be inhibited by pNL-HSP27-EGFP.The migration of VSMC induced by Ang Ⅱ were significantly inhibited by treating with pNL-HSP27-EGFP with a inhibition rate of 40.0%（P〈0.01）.Conclusion Migration of VSMC in response to Ang Ⅱ was significantly inhibited by the RNAi targeting HSP27 gene.HSP27 may be a potential target for the interventional treatment of pathological process related to cell migration.