摘要
目的探讨利血平与大鼠主动脉血管平滑肌细胞(RA-VSMC)自噬的关系。方法用含利血平或者厄尔平衡盐缓冲液(EBSS)培养基体外培养RA-VSMC,MTT法检测对照组、5μmol/L利血平组、10μmol/L利血平组、20μmol/L利血平组、50μmol/L利血平组RA-VSMC的细胞活力;mRFP-GFP-LC3双荧光体系示踪RA-VSMC的自噬小体;实时荧光定量PCR(qPCR)以及蛋白免疫印迹(WB)实验检测对照组、10μmol/L利血平组、50μmol/L利血平组和EBSS组RA-VSMC自噬相关基因LC3和Beclin1的表达。结果MTT结果显示,与对照组比较,5μmol/L利血平组、10μmol/L利血平组、20μmol/L利血平组和50μmol/L利血平组细胞活力均显著降低(P<0.05或P<0.01);mRFP-GFP-LC3双荧光体系结果显示,对照组和10μmol/L利血平组细胞中无明显自噬小体,50μmol/L利血平组和EBSS组细胞形成明显自噬小体;qPCR结果显示,与对照组比较,10μmol/L利血平组LC3 mRNA表达水平差异无统计学意义(P>0.05),50μmol/L利血平组和EBSS组LC3 mRNA表达水平显著升高(P<0.05或P<0.01);10μmol/L利血平组、50μmol/L利血平组和EBSS组Beclin1 mRNA表达水平显著升高(P<0.01);WB结果显示,与对照组比较,10μmol/L利血平组、50μmol/L利血平组和EBSS组LC3-Ⅱ/LC3-Ⅰ比率显著升高(P<0.01),10μmol/L利血平组、50μmol/L利血平组和EBSS组Beclin1蛋白表达水平显著升高(P<0.01)。结论利血平可能通过自噬机制调控VSMC的增殖。
Objective To explore the relationship of Reserpine and the autophagy of rat aortic vascular smooth muscle cells(RA-VSMC).Methods RA-VSMC was cultured in medium with Reserpine or Earles balanced salt solution(EBSS)in vitro.MTT assay was used to detect the effect of Reserpine on the proliferation of RA-VSMC with 5μmol/L Reserpine,10μmol/L Reserpine,20μmol/L Reserpine and 50μmol/L Reserpine.The dual-fluorescence mRFP-GFP-LC3 was used to trace the autophagosomes of RA-VSMC.Real-time fluorescence quantitative PCR(qPCR)and Western blot(WB)were used to detect the expression of LC3 and Beclin1.Results The MTT results showed that compared with the control group,the cell viability in the 5μmol/L Reserpine group,10μmol/L Reserpine group,20μmol/L Reserpine group and 50μmol/L Reserpine group were significantly reduced(P<0.05 or P<0.01).The results of the dual-fluorescence mRFP-GFP-LC3 showed that there were no significant autophagosomes in the cells of the control group and the 10μmol/L Reserpine group,and the 50μmol/L Reserpine group and EBSS group formed significant autophagosomes.qPCR results showed that compared with the control group,there was no significant difference in LC3 mRNA expression level in the 10μmol/L Reserpine group and the control group(P>0.05).LC3 mRNA expression levels in 50μmol/L Reserpine group and EBSS group increased significantly(P<0.05 or P<0.01).Beclin1 mRNA expression levels was significantly increased in 10μmol/L Reserpine group,50μmol/L Reserpine group and EBSS group(P<0.01).WB results showed that,compared with the control group,the 10μmol/L Reserpine group,50μmol/L Reserpine group and EBSS group LC3-Ⅱ/LC3-Ⅰratio increased significantly(P<0.01),Beclin1 protein expression was significantly increased in 10μmol/L Reserpine group,50μmol/L Reserpine group and EBSS group(P<0.01).Conclusion Reserpine may regulate the proliferation of vascular smooth muscle cells through autophagy.
作者
王顺民
杨晓丹
曾勇
黄露
雍苏南
WANG Shunmin;YANG Xiaodan;ZENG Yong;HUANG Lu;YONG Su′nan(Department of Cardiology,the First Hospital of Hu′nan University of Chinese Medicine,Hu′nan Province,Changsha 410007,China;Department of Cardiology,the First Clinical College of Traditional Chinese Medicine,Hu′nan University of Chinese Medicine,Hu′nan Province,Changsha 410208,China;Department of Emergency,the First Hospital of Hu′nan University of Chinese Medicine,Hu′nan Province,Changsha 410007,China)
出处
《中国医药导报》
CAS
2020年第16期4-7,I0004,共5页
China Medical Herald
基金
国家自然科学基金面上项目(81473616)
国家自然科学基金资助项目(81804077)
湖南省中医药科研计划重点项目(201753)
湖南省自然科学基金(青年基金)项目(2019JJ50468)。
关键词
利血平
血管平滑肌细胞
增殖
自噬
Reserpine
Vascular smooth muscle cells
Proliferation
Autophagy
