摘要
目的研究活血胶囊对H_(2)O_(2)诱导的小鼠主动脉平滑肌细胞(mASMC)衰老的抑制作用及分子机制。方法①实验分为对照组、活血胶囊150μg/mL组、活血胶囊300μg/mL组、活血胶囊750μg/mL组,WST-1法检测各组细胞活力。②实验分为对照组、H_(2)O_(2)组、活血胶囊+H_(2)O_(2)组,采用SA-β-gal染色法观察细胞衰老情况,采用qRT-PCR法检测细胞衰老相关基因p21和衰老相关分泌因子细胞间黏附分子-1(ICAM-1)、白细胞介素-8(IL-8)、白细胞介素-6(IL-6)及沉默信息调节蛋白1(SIRT1)mRNA表达量,ELISA法检测细胞培养上清中ICAM-1、IL-8、IL-6水平,流式细胞仪分析细胞周期变化。结果活血胶囊150μg/mL组、活血胶囊300μg/mL组细胞活力与对照组比较差异均无统计学意义(P均>0.05),活血胶囊750μg/mL组明显高于对照组(P<0.05)。SA-β-gal染色显示,H_(2)O_(2)组衰老细胞明显增多,活血胶囊+H_(2)O_(2)组衰老细胞明显较H_(2)O_(2)组少。H_(2)O_(2)组p21、ICAM-1、IL-8、IL-6 mRNA相对表达量均明显高于对照组(P均<0.05),且活血胶囊+H_(2)O_(2)组p21、ICAM-1、IL-6 mRNA相对表达量均明显低于H_(2)O_(2)组(P均<0.05)。H_(2)O_(2)5 h组SIRT1 mRNA相对表达量明显低于对照组(P<0.05),活血胶囊+H_(2)O_(2)组明显高于H_(2)O_(2)5 h组(P<0.05)。H_(2)O_(2)组细胞周期停滞在G1期,G1期细胞比例明显高于对照组(P<0.05),S期细胞比例明显低于对照组(P<0.05);活血胶囊+H_(2)O_(2)组G1期细胞比例明显低于H_(2)O_(2)组(P<0.05),S期细胞比例明显高于H_(2)O_(2)组(P<0.05)。结论活血胶囊可抑制H_(2)O_(2)诱导的mASMC衰老,其作用与促进细胞中SIRT1表达、抑制ICAM-1和IL-6表达有关。
Objective It is to study the inhibitory effect of Huoxue capsule on H_(2)O_(2)induced senescence of mouse aortic smooth muscle cells(mASMC)and its molecular mechanism.Methods①The experiment was divided into control group,Huoxue capsule 150μg/mL group,Huoxue capsule 300μg/mL group and Huoxue capsule 750μg/mL group,the cell viability of each group was detected by WST-1 method.②The experiment was divided into control group,H_(2)O_(2)group,and Huoxue capsule+H_(2)O_(2)group,the cell senescence was observed by SA-β-gal staining,the expression of cell senescence related genes p21,senescence related secretory factor intercellular adhesion molecule-1(ICAM-1),interleukin-8(IL-8),interleukin-6(IL-6),and silent information regulatory protein 1(SIRT1)mRNA were detected by qRT PCR.The levels of ICAM-1,IL-8,and IL-6 in cell culture supernatant were detected by ELISA,the cell cycle changes were analyzed by flow cytometry.Results There was no significant difference in cell viability among the Huoxue capsule 150μg/mL group,Huoxue capsule 300μg/mL group and the control group(all P>0.05),the cell viability in the Huoxue capsule 750μg/mL group was significantly higher than that in the control group(P<0.05).SA-β-gal staining showed that the number of senescent cells in H_(2)O_(2)group was significantly increased,and the number of senescent cells in Huoxue capsule+H_(2)O_(2)group was significantly less than that in H_(2)O_(2)group.The relative expression of p21,ICAM-1,IL-8,IL-6 mRNA in H2O2 group was significantly higher than that in control group(all P<0.05),and the relative expression of p21,ICAM-1,IL-6 mRNA in Huoxue capsule+H_(2)O_(2)group was significantly lower than that in H_(2)O_(2)group(all P<0.05).The relative expression of SIRT1 mRNA in H_(2)O_(2)5 h group was significantly lower than that in the control group(P<0.05),and Huoxue capsule+H_(2)O_(2)group was significantly higher than H_(2)O_(2)5h group(P<0.05).The cell cycle of H_(2)O_(2)group stagnated in G1 phase,the proportion of cells in G1 phase was significantly higher than that of the control group(P<0.05),and the proportion of cells in S phase was significantly lower than that of the control group(P<0.05);The proportion of cells in G1 phase in Huoxue capsule+H_(2)O_(2)group was significantly lower than that in H_(2)O_(2)group(P<0.05),and the proportion of cells in S phase was significantly higher than that in H_(2)O_(2)group(P<0.05).Conclusion Huoxue capsule could inhibit the senescence of mASMC cells induced by H_(2)O_(2),and its effect is related to promoting the expression of SIRT1 and inhibiting the expression of ICAM-1 and IL-6 in cells.
作者
霍雪萍
曹情雯
王海芳
赵向绒
王晶
董静
马运峰
HUO Xueping;CAO Qingwen;WANG Haifang;ZHAO Xiangrong;WANG Jing;DONG Jing;MA Yunfeng(School of Basic Medical Science of Xi’an Jiaotong University,Xi’an 710061,Shaanxi,China;Shaanxi Provincial Hospital of Traditional Chinese Medicine,Xi’an 710003,Shaanxi,China;Shaanxi Key Laboratory of Integrated Traditional and Western Medicine for Prevention and Treatment of Cardiovascular Diseases,Shaanxi University of Traditional Chinese Medicine,Xi’an 712046,Shaanxi,China;Shaanxi Provincial People’s Hospital,Xi’an 710068,Shaanxi,China;The Second Affiliated Hospital of Shaanxi University of Traditional Chinese Medicine,Xi’an 712000,Shaanxi,China)
出处
《现代中西医结合杂志》
CAS
2022年第20期2818-2823,共6页
Modern Journal of Integrated Traditional Chinese and Western Medicine
基金
陕西省中医管理局项目(2019-ZZ-JC025)
国家自然科学基金项目(81573823)
陕西省中医管理局平台项目(JCPT028)
陕西中医药大学第二附属医院学科创新团队项目(2020XKTD-B02)。
