摘要
目的观察大鼠肾小球系膜细胞中内脏脂肪素(Visfatin)对肾素血管紧张素系统(RAS)mRNA表达的影响及探讨其机制。方法分别构建Visfatin表达载体质粒及Visfatin RNAi表达载体质粒,将细胞分为8组:正常糖对照组、正常糖+NF-κB特异性抑制剂吡咯烷二硫基甲酸酯(PDTC)组、过表达Visfatin组、过表达Visfatin+PDTC组、空白过表达载体组、空白过表达载体+PDTC组、Visfatin RNAi组及空白沉默载体组,用Realtime-PCR方法检测血管紧张素原(AGT)、血管紧张素转化酶(ACE)、血管紧张素Ⅱ1型受体(AT1R)、血管紧张素Ⅱ2型受体(AT2R)等RAS相关基因表达,使用电泳迁移率变动分析法(EMSA)检测NF-κB活性,比较各组间基因表达及NF-κB活性的差异;以及通过观察PDTC处理对照组、过表达Visfatin组、空白过表达载体组前后上述指标的变化。结果细胞过表达Visfatin后,NF-κB活性、Visfatin、AGT、AT1R mRNA表达量显著升高,分别为正常糖对照组的1.52、11.42、2.85、1.25倍(P均<0.01);转染RNAi质粒pSIREN-Visfatin/sh RNA后,NF-κB活性、Visfatin、AGT、AT1R mRNA表达量显著降低,分别为正常糖对照组的10.90%、26.83%、30.00%、73.47%(P均<0.01)。经PDTC处理的各组NF-κB活性、AGT mRNA表达量与相应的未经PDTC处理组相比显著降低(P<0.01),正常糖+PDTC干预组NF-κB活性、AGT mRNA表达量是正常糖对照组的21.60%、34.59%;过表达Visfatin+PDTC干预组的NF-κB活性、AGT mRNA表达量是过表达Visfatin组的37.96%、19.72%;空白过表达载体+PDTC干预组NF-κB活性、AGT mRNA表达量是空白表达载体组的52.53%、33.08%,差异均具有统计学意义(P<0.01)。结论在大鼠肾小球系膜细胞中,Visfatin可通过NF-κB引起RAS相关基因表达的变化。
Objective To investigate whether visfatin induced the mRNA expression of renin-angiotensin system（RAS） by NF-κB pathway in rat mesangial cell.Methods The rat mesangial cells were transfected with the plasmids by using Lipofectamine method to overexpress or silence visfatin,or treated with pyrrolidinedithiocarbamate（PDTC）.The mRNA expression levels of visfatin,angiotensinogen（AGT）,angiotensin converting enzyme（ACE）,angiotensinⅡtype-1 receptor（AT1R）,angiotensinⅡtype-2 receptor（AT2R） was measured by realtime PCR.The NF-κB activity was detected by electrophoretic mobility shift assay.Results The NF-κB activity,and the mRNA levels of visfatin,AGT and AT1R in the overexpression of visfatin group,were significantly higher than those in the control group.The opposite effect was observed in the visfatin RNAi group.When the control group,overexpression of visfatin group,or the group transfected with blank expression plasmids were treated with PDTC,the NF-κB activity,the mRNA levels of visfatin and AGT were significantly reduced compared with the control group.Conclusion Visfatin upregulated the expression of RAS-related genes probably by NF-κB pathway.
出处
《热带医学杂志》
CAS
2012年第5期553-557,共5页
Journal of Tropical Medicine
基金
广州市科技计划项目(2011J4100114)
