MT1-MMP和PCSK9的联合抑制上调LDLR表达的研究

Study on combined inhibition of MT1-MMP and PCSK9 up-regulating LDLR expression

目的:探究Ⅰ型膜基质金属蛋白酶(MT1-MMP)和γ-分泌酶或前蛋白转化酶枯草溶菌素9(PCSK9)对低密度脂蛋白受体(LDLR)表达的影响。方法:应用DsiRNA分别沉默人肝细胞中MT1-MMP和PCSK9的表达并对细胞进行培养,选择DAPT抑制细胞中的γ-分泌酶,实时荧光定量PCR和免疫印迹法分别检测mRNA和蛋白质的表达水平。结果:MT1-MMP与γ-分泌酶或PCSK9的联合抑制可较单独抑制组进一步提高原代肝细胞中LDLR的表达水平;PCSK9^(-/-)小鼠再感染靶向小鼠MT1-MMP的shRNA腺相关病毒(AAV),会增加肝脏LDLR水平,降低血浆中的胆固醇浓度。结论:MT1-MMP和γ-分泌酶或PCSK9协同调节LDLR的表达。联合抑制MT1-MMP和γ-分泌酶或PCSK9可提高LDLR表达,并降低血浆中LDL胆固醇的水平,从而降低因现有手段无法有效控制血浆胆固醇水平的患者发生心血管病的风险。

Objective:To investigate the effects of membrane type-1 matrix metalloproteinase(MT1-MMP)andγ-secretase or proprotein convertase subtilisin/kexin type 9(PCSK9)on the expression of low-density lipoprotein receptor(LDLR).Methods:DsiRNA was used to silence MT1-MMP and PCSK9 expression in cultured human hepatocytes,and the cells were cultured.DAPT was selected to inhibitγ-secretase in cells,and real-time fluorescence quantitative PCR and immunoblotting were used to measure mRNA and protein expression levels,respectively.Results:The combined inhibition of MT1-MMP andγ-secretase or PCSK9 further increased the expression level of LDLR in primary hepatocytes compared with the single inhibition group.The reinfection of PCSK9^(-/-)mice with shRNA adeno-associated virus(AAV)targeting MT1-MMP in mice increased the level of LDLR in liver and reduced the concentration of cholesterol in plasma.Conclusion:MT1-MMP andγ-secretase or PCSK9 synergistically regulate LDLR expression.Combined inhibition of MT1-MMP andγ-secretase or PCSK9 has the potential to further increase the expression of LDLR and reduce the level of plasma LDL cholesterol,thereby reducing the risk of cardiovascular disease in patients whose plasma cholesterol levels can not be successfully controlled by currently available strategies.