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目的探讨胞内S-腺苷同型半胱氨酸(S-adenosylhomocysteine,SAH)聚集损伤血管内皮细胞的分子基础,为阐明膳食高蛋氨酸摄入促进心血管疾病的病理机制提供实验依据。方法以永生化的人脐静脉内皮细胞(HUVEC)为研究对象,经不同浓度的S-腺苷同型半胱氨酸水解酶(SAHH)强效抑制剂3-deazaadenosine(DZA)处理24,48或72 h后,利用酶联免疫吸附测定法(ELISA)测定HUVEC内炎性分子单核细胞趋化蛋白-1(MCP-1)的含量,荧光定量-PCR法(qRT-PCR)检测MCP-1 mRNA的相对表达量,甲基化特异PCR法(MSP)分析MCP-1基因启动子甲基化状态。结果HUVEC经DZA处理后,与正常对照相比,胞内MCP-1的含量升高,mRNA的相对表达量上调,但其启动子甲基化方式并不发生改变。结论内皮细胞SAH升高上调MCP-1的表达可能是膳食高蛋氨酸摄入促进心血管疾病的分子机制,但这种作用与MCP-1基因启动子的甲基化状态无关。
Objective To investigate the molecular basis of intracellular accumulation of S-adenosylhomocysteine (SAH) in vascular endothelial cells and provide experimental evidence for elucidating the pathogenic mechanism of dietary high methionine intake for cardiovascular diseases. Methods Immortalized human umbilical vein endothelial cells (HUVECs) were treated with various concentrations of S-adenosylhomocysteine hydrolase (SAHH) potent inhibitor 3-deazaadenosine (DZA) for 24, 48 or After 72 h, the level of MCP-1 in HUVEC was determined by enzyme-linked immunosorbent assay (ELISA) and the level of MCP-1 was detected by qRT-PCR mRNA relative expression, methylation specific PCR (MSP) analysis MCP-1 gene promoter methylation status. Results After treated with DZA, the intracellular MCP-1 level increased and the relative mRNA level increased, but the methylation pattern of HUVEC did not change. Conclusion The up-regulation of MCP-1 expression by elevated SAH in endothelial cells may be the molecular mechanism of dietary high methionine intake for cardiovascular disease, but this effect is not related to the methylation status of MCP-1 promoter.