TRPV1/SIRT1介导吴茱萸次碱抗AngⅡ诱导的血管平滑肌细胞衰老

目的探讨吴茱萸次碱(rutaecarpine,Rut)对长寿蛋白SIRT1表达及AngⅡ诱导的血管平滑肌细胞(vascular smooth muscle cells,VSMCs)衰老的影响。方法采用AngⅡ(1μmol·L^-1)孵育大鼠胸主动脉平滑肌细胞72 h,预先加入不同浓度的Rut(0.3、1、3μmol·L^-1),采用TRPV1拮抗剂CAPZ(10μmol·L^-1)和AMPK抑制剂Compound C(1μmol·L^-1)探讨TRPV1/AMPK是否介导Rut的保护效应。SA-β-Gal测定衰老细胞数目,DCFH-DA法测定细胞ROS水平。划痕愈合结合Transwell检测VSMCs迁移。Western blot检测VSMCs中长寿蛋白SIRT1和衰老相关蛋白p53、p21的表达以及p-AMPK水平。结果Rut明显地抑制AngⅡ诱导的VSMCs衰老和ROS生成,并抑制VSMCs迁移。预先给予TRPV1拮抗剂可取消Rut这一保护作用。AngⅡ可降低SIRT1的表达,给予Rut可剂量依赖性地恢复SIRT1的表达,且下调其下游衰老相关蛋白p53和p21的表达。AngⅡ可抑制p-AMPK,加入Rut能恢复p-AMPK水平。CAPZ和Compound C可消除Rut升高SIRT1表达的效应。结论Rut可上调SIRT1表达,抑制AngⅡ诱导的VSMCs衰老和迁移,其机制可能激活TRPV1/AMPK信号途径。

Rutaecarpine prevents AngⅡ-induced vascular smooth muscle cell senescence through TRPV1/SIRT1 pathway

Aim To evaluate the effects of Rutaecarpine(Rut)on the expression of SIRT1 and the senescence of vascular smooth muscle cells(VSMCs)induced by angiotensinⅡ.Methods VSMC senescence was induced by exposure to AngⅡ(1μmol·L^-1)for 72 h.VSMCs were treated with different concentrations of Rut(0.3,1,3μmol·L^-1).TRPV1 competitive antagonist CAPZ(10μmol·L^-1)and AMPK inhibitor Compound C(1μmol·L^-1)were used to explore whether TRPV1/AMPK mediated the protective effect of Rut.The quantity of senescent cells were determined by senescence-associated SA-β-Gal staining,and the intracellular ROS level was measured by(DCFH-DA)fluorescent probe.The migration ability of VSMCs was evaluated by Wound-healing assay combined with Transwell assay.The protein level of longevity protein SIRT1 and senescence-related proteins p53,p21 and AMPK phosphorylation level were detected by Western blot.Results Rut significantly inhibited AngⅡ-induced VSMC senescence and ROS production and prevented VSMCs migration.Preprocessing of TRPV1 antagonist CAPZ could abolish the protective effect of Rut.AngⅡinhibited the expression of longevity protein SIRT1.Rut recovered SIRT1 expression in a dose-dependent manner,while prevented the up-regulation of senescence-related proteins p53 and p21.AngⅡinhibited AMPK phosphorylation,pre-treatment with Rut restored AMPK phosphorylation level.CAPZ and Compound C eliminated the up-regulating function of Rut on SIRT1 expression.Conclusions Rut up-regulates the expression of SIRT1 and prevents the senescence and migration of VSMCs induced by AngiotensinⅡ,which is related to activation of the TRPV1/AMPK signaling pathway.