脂联素抑制甲基环糊精包被的胆固醇诱导的人血管平滑肌细胞泡沫化

Inhibitory effect of adiponectin on foaming process of human vascular smooth muscle cells induced by cholesterol:methyl-β-cyclodextrin

目的:体外研究脂联素(APN)对甲基环糊精包被的胆固醇(Chol:MβCD)诱导的人血管平滑肌细胞(VSMCs)泡沫化过程的影响,并探讨其可能的机制。方法:体外培养人主动脉平滑肌细胞,将细胞分为5组,即对照组(正常培养VSMCs)、泡沫细胞组(VSMCs与100 mg/L Chol:MβCD共孵育24 h)及低、中、高剂量APN组(在泡沫细胞组基础上分别加入2.5、5和10 mg/L APN干预24 h)。油红O染色及脂质含量计算分析各组细胞泡沫化情况,酶荧光法检测胞内胆固醇含量及细胞胆固醇流出率。RT-qPCR和Western blot法分别检测各组细胞内沉默信息调节因子1(SIRT1)和过氧化物酶增殖物激活受体γ辅激活因子1α(PGC-1α)mRNA和蛋白的表达。结果:与对照组相比,泡沫细胞组胞质内可见大量红色脂滴聚集,胞核突出,胞内总胆固醇(TC)、游离胆固醇(FC)和胆固醇酯(CE)的含量及CE/TC值均显著升高(P<0.05),细胞胆固醇流出率增加(P<0.05),符合泡沫细胞改变;SIRT1和PGC-1α的mRNA和蛋白表达水平均显著降低(P<0.05)。APN干预可呈浓度依赖性降低泡沫细胞内脂质沉积(P<0.05),减少胞内TC和CE含量及CE/TC值(P<0.05),促进细胞胆固醇流出(P<0.05);SIRT1和PGC-1α的mRNA和蛋白表达水平随着APN浓度的升高而逐步升高(P<0.05)。结论:APN在体外可呈浓度依赖性抑制Chol:MβCD诱导的VSMCs泡沫化,其机制可能与调控SIRT1/PGC-1α信号通路有关。

AIM:To explore the effect of adiponectin(APN)on the foaming process of human vascular smooth muscle cells(VSMCs)induced by cholesterol:methyl-β-cyclodextrin(Chol:MβCD)in vitro and related mecha-nisms.METHODS:Human VSMCs from aorta were cultured in vitro and divided into 5 groups,including control group(normal culture),foam cell group(incubated with 100 mg/L Chol:MβCD complexes for 24 h to establish VSMCs-derived foam cell model),and low-,middle-and high-dose APN groups(incubated with 2.5,5 and 10 mg/L APN for 24 h after modeling,respectively).Oil red O staining and calculation of lipid content were used to analyze cell foaming conditions.The cholesterol level and the cholesterol efflux rate were measured in each group.The expression of silent information regu-lator 1(SIRT1)and peroxisome proliferator-activated receptorγcoactivator-1α(PGC-1α)at mRNA and protein levels in each group was determined by RT-qPCR and Western blot,respectively.RESULTS:Compared with control group,marked accumulation of red lipid droplets and prominent nuclei in foam cell group were observed,the levels of total cholesterol(TC),free cholesterol(FC)and cholesterol ester(CE),the CE/TC ratio and the cholesterol efflux rate were signifi-cantly increased(P<0.05),while the expression of SIRT1 and PGC-1αat mRNA and protein levels was significantly de-creased(P<0.05).Treatment with APN reduced the intracellular red lipid droplets,the levles of TC and CE,and the ra-tio of CE/TC(P<0.05),and promoted efflux of intracellular cholesterol in a dose-dependent manner(P<0.05),while the expression of SIRT1 and PGC-1αat mRNA and protein levels was significantly elevated(P<0.05).CONCLUSION:APN inhibits the foaming of VSMCs induced by Chol:MβCD in a dose-dependent manner in vitro,which may be related to the regulation of SIRT1/PGC-1αsignaling pathway.