补阳还五汤促进自噬抑制H_(2)O_(2)诱导的人脐静脉内皮细胞衰老机制研究  

Mechanism Research of Buyang Huanwu Decoction(补阳还五汤)on Promoting Autophagy and Inhibiting Aging of Human Umbilical Vein Endothelial Cells Induced by H_(2)O_(2)

作　　者：曾乙凡 闵冬雨[1] 刘勇明[1] 袁常斌 ZENG Yifan;MIN Dongyu;LIU Yongming;YUAN Changbin(Affiliated Hospital of Liaoning University of Traditional Chinese Medicine,Shenyang 110032,Liaoning,China;Liaoning University of Traditional Chinese Medicine,Shenyang 110847,Liaoning,China)

机构地区：[1]辽宁中医药大学附属医院,辽宁沈阳110032 [2]辽宁中医药大学,辽宁沈阳110847

出　　处：《辽宁中医药大学学报》2024年第10期10-14,共5页Journal of Liaoning University of Traditional Chinese Medicine

基　　金：国家自然科学基金面上项目(82174114);沈阳市科学技术局公共卫生研发专项(医工结合协同创新研究项目)(22-321-32-14);辽宁中医药大学附属医院育苗工程项目(YM202133)。

摘　　要：目的观察补阳还五汤(BYHW)对过氧化氢(H_(2)O_(2))诱导的衰老内皮细胞沉默信息调节因子1(silent information regulator 1,SIRT1)-自噬通路的作用。方法以人脐静脉内皮细胞(HUVEC)作为研究对象,H_(2)O_(2)诱导的内皮细胞作为模型,体外培养HUVECs,并将细胞分为正常组、模型组(H_(2)O_(2)400μmol/L)、BYHW组(H_(2)O_(2)400μmol/L+BYHW 2500μg/mL)。衰老相关β半乳糖苷酶(SA-β-Gal)染色检测细胞衰老,蛋白质免疫印迹法检测SIRT1、纤溶酶原激活物抑制因子1(PAI-1)、泛素结合蛋白p62及微管相关蛋白1轻链3β(LC3)蛋白的表达,ELISA法检测细胞上清液中超氧化物歧化酶(superoxide dismutase,SOD)、丙二醛(malondialdehyde,MDA)含量,免疫荧光观察SIRT1、LC3的表达。结果400μmol/L的H_(2)O_(2)处理可成功诱导HUVECs衰老。与正常组相比,衰老模型组中SA-β-Gal阳染率增加,SIRT1、LC3蛋白表达降低,SOD含量降低,PAI-1、p62蛋白表达升高,MDA含量升高,细胞免疫荧光显示SIRT1、LC3表达降低。与模型组比较,BYHW组细胞蓝染比率降低,PAI-1、p62蛋白表达降低,SOD水平升高,SIRT1、LC3蛋白表达升高,细胞免疫荧光BYHW组SIRT1、LC3表达明显升高。结论在过氧化氢诱导的HUVECs衰老模型中,补阳还五汤能够上调SIRT1,激活自噬,从而对衰老的内皮细胞起到保护作用。Objective To observe the effect of the Buyang Huanwu Decoction(BYHW)on silent information regulator 1(SIRT1)-autophagy pathway induced by hydrogen peroxide(H_(2)O_(2)).Methods Human umbilical vein endothelial cells were used as the research subjects,and hydrogen peroxide cell senescence was induced as a model.HUVEC was cultured in vitro,and the cells were divided into normal group(H_(2)O_(2)400μmol/L)and BYHW group(H_(2)O_(2)400μmol/L+BYHW 2500μg/mL).Cell senescence was detected by senescence-relatedβgalacsidase staining,expression of SIRT1,PAI-1,p62 and LC3 protein was detected by protein immunoblotting,superoxide dismutase(SOD)and malondialdehyde(MDA)content in cell supernatant was detected by ELISA,and the expression of SIRT1 and LC3 was observed by immunofluorescence.Results H_(2)O_(2) treatment with 400μmol/L successfully induced HUVECs senescence.Compared with control group,positive staining rate of SA-β-Gal increased,expression of SIRT1 and LC3 decreased,content of SOD decreased,expression of PAI-1 and p62 increased,content of MDA increased in model group,and expression of SIRT1 and LC3 decreased in the cellular immunofluorescence.Compared with the model group,the blue staining ratio in BYHW group was decreased,PAI-1 and p62 protein expression decreased,SOD level increased,SIRT1 and LC3 protein expression increased,and expression of SIRT1 and LC3 was increased significantly in cell immunofluorescence.Conclusion In a hydrogen oxide-induced HUVECs senescence model,the rehydration system can upregulate SIRT1 and activate autophagy,thus protecting against senescent endothelial cells.

关 键 词：衰老 内皮细胞 补阳还五汤 SIRT1 自噬 

分 类 号：R285[医药卫生—中药学]