miRNA-33对Hcy干预RAW264.7巨噬细胞衍生的泡沫细胞表达ABCA1/ABCG1的影响  

Effect of miRNA-33 on ABCA1/ABCG1 expression in RAW264.7 macrophage-derived foam cells after Hcy treatment

作　　者：代佩 高奋[1] 高宏伟 王远 冯高洁 张秦风 白瑞[1] 秦卫伟[1] 李虹[1] 宋晓苏[1] DAI Pei;GAO Fen;GAO Hong-wei;WANG Yuan;FENG Gao-jie;ZHANG Qin-feng;BAI Rui;QIN Wei-wei;LI Hong;SONG Xiao-su(Department of Cardiology,Second Hospital of Shanxi Medical University,Taiyuan 030001,China)

机构地区：[1]山西医科大学第二医院心内科,山西太原030001

出　　处：《中国病理生理杂志》2019年第2期212-217,共6页Chinese Journal of Pathophysiology

基　　金：山西省自然科学基金资助项目(No.2014011040-3)

摘　　要：目的:研究同型半胱氨酸(Hcy)是否通过微小RNA-33 (miRNA-33)抑制三磷酸腺苷结合盒转运体A1(ABCA1)和三磷酸腺苷结合盒转运体G1(ABCG1)的表达,从而降低胆固醇逆转运(RCT)效率。方法:复制RAW264.7巨噬细胞经氧化型低密度脂蛋白(ox-LDL)诱导为泡沫细胞的模型,油红O染色确定模型是否复制成功,用LipofectamineTM2000将miRNA-33 mimics和miRNA-33 inhibitor分别转染入细胞内,再给予5 mmol/L的Hcy干预24 h后进行实验;用油红O染色观察细胞内脂滴情况;real-time PCR和Western blot检测ABCA1和ABCG1的mRNA和蛋白表达水平;液体闪烁计数法检测细胞内胆固醇流出率;高效液相色谱分析细胞内胆固醇含量。结果:(1)与空白对照组相比,miRNA-33 mimics组细胞内脂滴含量增加,ABCA1和ABCG1的mRNA和蛋白表达量降低(P <0.05),细胞内胆固醇含量逐渐增加(P <0.05),细胞内胆固醇流出率逐渐减少(P <0.05);(2)与空白对照组相比,miRNA-33 inhibitor组检测结果与miRNA-33 mimics组的结果相反,差异具有统计学意义(P <0.05);(3)空白对照组、miRNA-33 mimics-NC组和miRNA-33 inhibitor-NC组3组间相比,所有检测结果的差异均无统计学显著性。结论:Hcy可间接通过诱导miRNA-33表达而抑制ABCA1和ABCG1的蛋白表达,降低RCT效率。AIM:To study whether homocysteine(Hcy)inhibits the expression of ATP-binding cassette transporter A1(ABCA1)and ATP-binding cassette transporter G1(ABCG1)by microRNA-33(miRNA-33)signaling,and reduces the efficiency of reverse cholesterol transport(RCT).METHODS:RAW264.7 macrophages were induced by oxidized low-density lipoprotein(ox-LDL)to establish foam cell model.Oil red O staining was used to determine whether the model was established successfully.miRNA-33 mimics and miRNA-33 inhibitor were transfected into the cells by Lipofectamine 2000,and the cells were exposed to Hcy at concentration of 5 mmol/L for 24 h.The intracellular lipid droplets were observed by Oil red O staining.The expression of ABCA1 and ABCG1 at mRNA and protein levels was determined by real-time PCR and Western blot.The cellular cholesterol content was analyzed by HPLC,and effluent rate of cholesterol was detected by the method of liquid scintillation counting.RESULTS:Compared with blank control group,the lipid content in miRNA-33 mimics group was increased,and the expression of ABCA1 and ABCG1 at mRNA and protein levels was decreased(P<0.05).The intracellular cholesterol content was increased gradually(P<0.05),and the cellular cholesterol efflux rate was gradually decreased(P<0.05)in miRNA-33 mimics group.Compared with blank control group,the testing results in miRNA-33 inhibitor group were the opposition of those in miRNA-33 mimics group(P<0.05).No diffe-rence of the above indexes among blank control group,miRNA-33 mimics-NC group and miRNA-33 inhibitor-NC group was observed.CONCLUSION:Hcy inhibits the mRNA and protein expression of ABCA1 and ABCG1 through miRNA-33 signaling,and reduces the efficiency of RCT in RAW264.7 macrophage-derived foam cells.

关 键 词：微小RNA-33 同型半胱氨酸 胆固醇逆转运 三磷酸腺苷结合盒转运体A1 三磷酸腺苷结合盒转运体G1 

分 类 号：R543.5[医药卫生—心血管疾病] R363.2[医药卫生—内科学]