摘要
该研究旨在探讨三七皂苷R1能否通过调控miRNA-34a/SIRT1/p53通路发挥延缓H2O2诱导的血管内皮细胞衰老的作用。以人脐静脉内皮细胞（HUVEC）作为研究对象,建立H2O2诱导的衰老模型,以白藜芦醇作为阳性对照药,实验分为青年组、H2O2模型组、三七皂苷R1组、白藜芦醇组。药物组提前加入三七皂苷R1（30μmo L·L^-1）或白藜芦醇（10μmo L·L^-1）干预24 h,然后用H2O2（100μmo L·L^-1）造模4 h,最后换成完全培养基培养24 h。采用衰老特异性β-半乳糖苷酶（SA-β-gal）染色法鉴定细胞衰老程度,CCK-8法检测细胞增殖能力,流式细胞仪分析细胞周期,WST-1法检测细胞内超氧化物歧化酶（SOD）活力,QRT-PCR检测miRNA-34a,SIRT1和p53的核酸表达,Western-blot技术检测SIRT1,p53和衰老相关蛋白p21,p16的蛋白表达。结果发现,与模型组相比,三七皂苷R1组和白藜芦醇组可有效减低衰老内皮细胞的β-半乳糖苷酶染色阳性率,增强细胞增殖能力和SOD活力,减少G0/G1期细胞比例,增加S期细胞比例,抑制了miRNA-34a和p53的核酸表达,促进了SIRT1的核酸表达,此外抑制了p53,p21和p16的蛋白表达,促进了SIRT1的蛋白表达,差异均有统计意义（P〈0.05）。该研究表明,H2O2诱导的衰老的内皮细胞可以作为衰老研究的模型,三七皂苷R1延缓血管内皮细胞衰老的效果明显,其机制可能是通过调控miRNA-34a/SIRT1/p53通路延缓了血管内皮细胞衰老的进程。
This study aimed to investigate the effect of notoginsenoside R1 in delaying H2O2-induced vascular endothelial cell senescence through microRNA-34 a/SIRT1/p53 signal pathway. In this study,human umbilical vein endothelial cells（ HUVECs） were selected as the study object; the aging model induced by hydrogen peroxide（ H2O2） was established,with resveratrol as the positive drug. HUVECs were randomly divided into four groups,youth group,senescence model group,notoginsenoside R1 group and resveratrol group. Notoginsenoside R1 group and resveratrol group were modeled with 100 μmo L·L^-1 H2O2 for 4 h after 24 h treatment with notoginsenoside R1（ 30 μmo L·L^-1） and resveratrol（ 10 μmo L·L^-1） respectively. At the end,each group was cultured with complete medium for 24 h. The degree of cellular senescence was detected by senescence-associated β-galactosidase（ SA-β-Gal） staining kit,the cell viability was detected by cell counting kit-8,the cell cycle distribution was analyzed by flow cytometry,and the cellular SOD activity was detected by WST-1 method in each group. The expressions of SIRT1,p53,p21 and p16 proteins in HUVECs were detected by Western blot. In addition,the mRNA expressions of miRNA-34 a,SIRT1 and p53 in HUVECs were assayed by Real-time PCR.These results indicated that notoginsenoside R1 significantly reduced the positive staining rate of senescent cells,enhanced the cell proliferation capacity and intracellular SOD activity,decreased the proportion of cells in G0/G1 phase,and increased the percentage of cells in S phase simultaneously compared with the senescence model group. Moreover,notoginsenoside R1 decreased the mRNA expressions of miRNA-34 a and p53 and the protein expression of p53,p21 and p16. At the same time,notoginsenoside R1 increased the protein and mRNA expressions of SIRT1. The differences in these results between the senescence model group and the notoginsenoside R1 group were statistically significant（ P〈0. 05）. However,there was not statistically significant difference in these results between the notoginsenoside R1 group and the resveratrol group. In conclusion,the senescence of endothelial cells induced by H2O2 can be used as a model for studying aging. Notoginsenoside R1 has an obvious anti-aging effect on vascular endothelial cells in this study. The possible mechanism is that notoginsenoside R1 can delay the senescence process of vascular endothelial cells induced by H2O2 by regulating microRNA-34 a/SIRT1/p53 signal pathway.
作者
赖小华
雷燕
杨静
修成奎
LAI Xiao-hua;LEI Yan;YANG Jing;XIU Cheng-kui(Gttangdong Metabolic Disease Research Center of Integrated Chinese and Western Medicine, Institu, te of Chinese Medicinal Sciences, Guangdong Pharmacetttieal University, Guangzhou 510006, China;Beijing Key Laboratory of Traditional Chinese Medicine Basic Research on Prevention and Treatment of Major Disease Experimental Research Center, China Academy of Chinese Medical Sciences, Beijing 100700, China)
出处
《中国中药杂志》
CAS
CSCD
北大核心
2018年第3期577-584,共8页
China Journal of Chinese Materia Medica
基金
国家自然科学基金项目(81503448,81673822)
中国中医科学院自主选题项目(ZZ2015011)
