抑制PPAR-α表达对ET-1诱导的心肌肥大和PI3K/Akt/GSK3β-NFATc4通路的影响  

Effects of PPAR-α silence on ET-1 induced myocardial hypertrophy and PI3K/Akt/GSK3β-NFATc4 signal pathway

作　　者：李瑞芳[1] 乐康[2] 高洁[2] 杨国庆[2] 鲍颖霞[2] 刘培庆[2] 

机构地区：[1]河南科技大学医学院药理教研室,河南洛阳471003 [2]中山大学药学院药理毒理实验室,广东广州510080

出　　处：《中国病理生理杂志》2009年第12期2289-2294,共6页Chinese Journal of Pathophysiology

基　　金：国家自然科学基金资助项目(No.30772576);广东省自然科学基金重点资助项目(No.7117380)

摘　　要：目的:研究过氧化物酶体增殖物激活受体-α(PPAR-α)在病理性心肌肥大中的作用及其信号机制。方法:应用Invitrogen's Stealth RNAi抑制心肌细胞PPAR-α的表达;采用[3H]-亮氨酸掺入法和RT-PCR检查心肌细胞蛋白质合成和心房利钠因子(ANF)mRNA的表达;采用Western blotting法检测Akt/GSK3β的磷酸化表达;应用免疫荧光技术检测NFATc4的胞核移位。结果:(1)RSS304168是最有效的PPAR-αRNAi,特异性地抑制了PPAR-α的表达。(2)非诺贝特预处理抑制了内皮素-1(ET-1)诱导的心肌细胞肥大(蛋白质合成和ANF mRNA的表达);RSS304168加强了ET-1的诱导效应,而且逆转了非诺贝特对心肌肥大的抑制效应。(3)非诺贝特降低了ET-1诱导的Akt/GSK3β的磷酸化表达,RSS304168增强了ET-1的诱导效应,ET-1和RSS304168的上述作用可被PI3K阻断剂LY294002所阻断;RSS304168逆转了非诺贝特对Akt/GSK3β的磷酸化表达的负性调控作用。(4)非诺贝特抑制了ET-1诱导的NFATc4的胞核移位;而RSS304168加强了ET-1的诱导作用,逆转了非诺贝特对NFATc4胞核移位的抑制作用。结论:PPAR-α激活可以通过PI3K/Akt/GSK3β-NFATc4通路抑制ET-1诱导的心肌肥大反应。AIM ： To investigate the role and signal mechanism of PPAR - α in the pathogenesis of cardiac hypertrophy. METHODS： Small interfering RNA （siRNA） was applied to efficiently silence the gene expression of PPAR -α in cardiac myocytes. [ 3H ] leucine incorporation assay was performed to measure protein synthesis. Reverse transcription - polymerase chain reaction （RT -PCR） was used to analyze the mRNA level of atrial natriuretic factor （ANF） and PPAR - α. Western blotting analysis was performed to investigate the levels of phosphorylation of protein kinase B （PKB/Akt） and glycogen synthase kinase 3β （GSK3β）. Immunofluorescence analysis was used to examine the cellular localization of NFATc4. RESULTS ： （ 1 ） RSS304168 was the most efficient stealth RNAi duplex to specifically inhibit PPAR - α expression. （2）RSS304168 significantly potentiated the ET - 1 - induced cardiomyocyte bypertrophy and enhanced ET - 1 - induced protein synthesis and ANF mRNA expression in cardiomyocytes. Moreover, RSS304168 completely reversed the inhibitory effects of fenofibrate on ET- 1 -induced protein synthesis and ANF mRNA expression. （3）RSS304168 enhanced ET - 1 - induced phosphorylation of Akt at Ser473 and GSK3β at Set9. The effects of ET - 1 or ET - 1 combined with RSS304168 on phosphorylation of Akt/GSK3β were completely blocked by LY294002, a PI3K specific inhibitor. Fenofibrate markedly inhibited ET - 1 -induced phosphorylation of Akt/GSK3β while RSS304168 abolished these effects of fenofibrate. （4）Fenofibrate prevented the nuclear translocation of NFATc4 induced by ET- 1 while RSS304168 abolished this effect of fenofibrate. CONCLUSION： Activation of PPAR - αinhibits ET - 1 - induced cardiomyocyte hypertrophy through blocking Akt/GSK3β -NFATc4 signaling pathways.

关 键 词：RNA干扰 心肌肥大 过氧化物酶体增殖物活化受体α 糖原合成酶激酶3Β 活化T细胞核因子 

分 类 号：R541[医药卫生—心血管疾病]