同型半胱氨酸对人主动脉平滑肌细胞ATP结合盒转运蛋白A1启动子甲基化水平及表达的影响

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摘要目的探讨同型半胱氨酸（Hcy）对人主动脉平滑肌细胞ATP结合盒转运蛋白A1（ABCA1）启动子甲基化水平及表达的影响。方法培养人主动脉平滑肌细胞，将其分为两组：对照组和实验组。对照组不加Hcy，实验组加入不同浓度Hcy（50、100、200、500 μmol/L）培养48 h。闪烁计数法检测胆固醇流出率；实时定量PCR检测ABCA1 mRNA表达水平；Western-blotting检测ABCA1蛋白质表达；巢式甲基化PCR检测ABCA1启动子甲基化水平。结果对照组各时间点胆固醇流出率比较差异无统计学意义（P > 0.05）；各Hcy浓度组不同时间比较显示，24、48 h胆固醇流出率均明显低于0、12 h，差异均有统计学意义（P < 0.05）。与对照组比较，0、12 h各Hcy浓度组胆固醇流出率差异无统计学意义（P ＞ 0.05），24、48 h各Hcy浓度组胆固醇流出率显著降低，差异均有统计学意义（P < 0.05）。与对照组比较，实验组ABCA1 mRNA相对表达水平下降（均P < 0.05），ABCA1蛋白质相对表达水平下降（P < 0.05），ABCA1启动子甲基化水平显著升高（P < 0.05）；不同Hcy浓度之间比较，ABCA1mRNA表达、ABCA1蛋白质表达和ABCA1启动子甲基化水平差异均无统计学意义（P ＞ 0.05）。结论 Hcy使人主动脉平滑肌细胞ABCA1启动子甲基化水平升高及表达下降，引起胆固醇流出率降低，可能是其引起动脉粥样硬化的病理机制之一。

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	王慧媛1 张紫荆1 袁封艳1 郑文倩1 马旭1 翟晓娟2 高胜利2
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关键词 ：同型半胱氨酸, ATP结合盒转运蛋白A1, 甲基化, 血管平滑肌细胞

Abstract：Objective To investigate the effect of homocysteine (Hcy) on the promoter methylation and expression of ATP binding cassette transporter A1 (ABCA1) in human aortic smooth muscle cells. Methods The human aortic smooth muscle cells were divided into two groups: the control group and the experimental group. The cells in the control group were cultured without Hcy, however cells in the experimental group were cultured with different concentrations of Hcy (50, 100, 200, 500 μmol/L). The cholesterol outflow rate was detected by scintillation counting technique, the expression of ABCA1 mRNA was detected by real-time quantitative PCR, the expression of ABCA1 protein was detected by Western-blotting, the ABCA1 promoter methylation was detected by nested detection of PCR. Results There was no significant difference in cholesterol efflux rate at each time point in the control group (P > 0.05). Compared with Hcy treatment for 0 h and 12 h, after treatment of 24 h and 48 h the cholesterol efflux rate of the experimental were significantly decreased (P < 0.05). After treated with Hcy for 0 h and 12 h, compared with the control group, there was no significant difference in cholesterol efflux rate (P > 0.05). After treated with Hcy for 24 h and 48 h, compared with the control group, cholesterol efflux rate of the experimental group was significantly decreased (P < 0.05). Compared with control group, ABCA1 mRNA relative expression level decreased (P < 0.05), the relative expression level of ABCA1 protein was decreased (P < 0.05), ABCA1 promoter methylation level was significantly increased (P < 0.05). There was no significant difference in ABCA1mRNA expression, ABCA1 protein expression and ABCA1 promoter methylation level between different Hcy concentrations groups (P > 0.05). Conclusion Hcy can increase the methylation level of ABCA1 promoter and decrease the expression of ABCA1 in aortic smooth muscle cells that lead to lower cholesterol efflux rate, which may be one of the pathological mechanisms of atherosclerosis.

Key words： Homocysteine ATP binding cassette transporter A1 Methylation Vascular smooth muscle cells

基金资助:山西省高等学校大学生创新创业训练计划项目（2017534）。

通讯作者: 高胜利（1975.2-），男，硕士，副教授；研究方向：心血管疾病的表观遗传机制。

引用本文:

王慧媛1 张紫荆1 袁封艳1 郑文倩1 马旭1 翟晓娟2 高胜利2,3. 同型半胱氨酸对人主动脉平滑肌细胞ATP结合盒转运蛋白A1启动子甲基化水平及表达的影响[J]. 中国医药导报, 2018, 15(2): 19-22.
WANG Huiyuan1 ZHANG Zijing1 YUAN Fengyan1 ZHEN Wenqian1 MA Xu1 ZHAI Xiaojuan2 GAO Shengli2,3. Effects of homocysteine on promoter methylation level and expression of ATP binding cassette transporter A1 in human aortic smooth muscle cells. 中国医药导报, 2018, 15(2): 19-22.

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