羟基红花黄色素A通过激活自噬抑制ANGⅡ诱导的VAFs迁移

Hydroxysafflor yellow A represses AngⅡ-induced migration through activation of autophagy in VAFs

目的探讨羟基红花黄色素A(hydroxysafflor yellow A,HSYA)对血管紧张素Ⅱ(angiotensin Ⅱ,AngⅡ)诱导的血管外膜成纤维细胞(vascular adventitial fibroblasts,VAFs)迁移的作用。方法组织法培养SD大鼠VAFs,细胞分为正常组、AngⅡ组、HSYA组、HSYA与自噬抑制剂Bafilomycin A1共处理组、Bafilomycin A1组;采用免疫荧光实验鉴定VAFs;CCK-8法分别建立AngⅡ和HSYA的量效时效曲线;划痕实验检测细胞迁移;采用MDC法检测各组细胞自噬小体数量;Western blot实验检测自噬相关蛋白Beclin1、LC3、p62表达。结果培养的细胞鉴定为VAFs;AngⅡ呈剂量、时间依赖性增强VAFs细胞活力,HSYA呈剂量、时间依赖性抑制VAFs细胞活力;AngⅡ明显促进VAFs迁移,HSYA以及HSYA和Baf共处理均可抑制AngⅡ诱导的VAFs迁移,而Baf处理则促进VAFs迁移;HSYA明显增加了自噬斑点数量,并促进自噬相关蛋白LC3Ⅱ、Beclin1的表达,抑制p62蛋白表达。结论HSYA通过激活自噬抑制AngⅡ诱导的VAFs迁移,改善血管重构。

Aim To explore if hydroxysafflor yellow A(HSYA)inhibits angiotensinⅡ(AngⅡ)-induced cell migration by activating autophagy in vascular adventitia fibroblasts(VAFs).Methods Primary rat VAFs were isolated and verified by immunofluorescence staining.The cultured cells were randomly divided into normal group,AngⅡgroup,HSYA group,Co-treatment group of HSYA and autophagy inhibitor Bafilomycin A1,and Bafilomycin A1 group.CCK-8 method was used to establish dose-effect and time-effect curve of AngⅡand HSYA on VAFs.Wound healing assay was applied to detect cell migration.MDC method was employed to detect the number of autophagosomes.Western blot was utilized to detect the level of Beclin1,LC3 and p62.Results The cultured cells proved to be VAFs.AngⅡincreased VAFs viability and HSYA decreased VAFs cell viability in a dose-and time-dependent manner.Compared with normal group,AngⅡsignificantly promoted VAFs migration,HSYA and co-treatment of HSYA and Baf both inhibited the migration of VAFs induced by AngⅡ.While Baf promoted VAFS migration.Compared with AngⅡgroup,HSYA significantly promoted autophagy,increased the number of autophagy spots,the level of Beclin 1,LC3Ⅱ/Ⅰ,and decreased the level of p62.Conclusion HSYA inhibits AngⅡ-induced cell migration by activating autophagy in VAFs,which plays a role in improving vascular remodeling.