摘要
目的:研究前蛋白转换酶枯草溶菌素9(PCSK9)siRNA对THP-1源性巨噬细胞CD36、SR-A1及SR-B1表达的影响。方法:以THP-1源性巨噬细胞为研究对象,应用Lipofectamine 2000转染不同浓度PCSK9 siRNA进THP-1源性巨噬细胞中,RT-PCR及Western blot筛选出最有效的siRNA,再转染入THP-1源性巨噬细胞,24 h后加入ox-LDL处理24 h,采用油红O染色检测细胞内脂质蓄积情况,RT-PCR分析细胞CD36、SR-A1及SR-B1表达。结果:浓度为80 nmol/L的PCSK9 siRNA基因沉默效应最佳;油红O染色结果表明ox-LDL组细胞内脂质蓄积情况最为明显,PCSK9 siRNA转染组次之;PCSK9 siRNA转染组CD36 mRNA表达水平相对于ox-LDL组降低(P<0.05),而ox-LDL组和PCSK9 siRNA转染组SR-A1及SR-B1 mRNA表达无明显差异。结论:PCSK9可能通过影响摄取脂质的细胞膜表面受体CD36表达,参与动脉粥样硬化的发生发展。
Objective：To investigate the effects of PCSK9 siRNA on CD36,SR-A1 and SR-B1 expressions in THP-1 derived macrophages.Methods：The siRNA for PCSK9 gene were transfected into THP-1 derived macrophages using positive ion liposome Lipofectamine 2000.Transfection efficiency was assessed by fluorescence microscope assay.The expression of PCSK9 in THP-1 derived macrophages at 24 h after transfection was detected by RT-PCR and Western blot.The most efficient siRNA was selected for transfection.At 24 h after transfection,macrophages were treated with ox-LDL for another 24 h.Then the intracellular lipid accumulation was observed by oil red O staining,and expressions of CD36 mRNA,SR-A1 mRNA and SR-B1 mRNA were detected by RT-PCR.Results：siRNA of 80 nmol/L showed the strongest inhibition for the gene expression of PCSK9,and ox-LDL-induced accumulation of cholesterol and upregulation of CD36 mRNA expression in THP-1 derived macrophages decreased by application of PCSK9 siRNA（P 〈 0.05）.However,PCSK9 siRNA caused no effect on ox-LDL-induced upregulation of SR-A1 mRNA and SR-B1 mRNA expression in macrophages.Conclusion：PCSK9 may be involved in atherosclerosis development by inhibiting the expression of CD36.
出处
《南京医科大学学报（自然科学版）》
CAS
CSCD
北大核心
2011年第5期673-678,共6页
Journal of Nanjing Medical University(Natural Sciences)
基金
湖南省应用基础研究计划重点项目(2008FJ2006)
湖南省科技厅计划项目(2009TP4057-2,2010TP4008-2)
湖南省教育厅重点科研项目(10A105)
湖南省高校科技创新团队支持计划资助
