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目的:探讨高蛋氨酸饮食诱导高同型半胱氨酸血症(Hhcy)对大鼠主动脉组织Bcl-2基因启动子区甲基化水平的影响。方法:健康6周龄雄性Wistar大鼠24只,体重(160±10)g,适应性喂养1周后随机分为对照组和Hhcy组,每组12只。对照组给予AIN-93G配方饲料,Hhcy组饲以含1.7%蛋氨酸的AIN-93G配方饲料。喂养18周后,全自动生化仪测定血浆同型半胱氨酸浓度。HE染色观察大鼠主动脉组织形态学变化,免疫组织化学检测大鼠主动脉组织Bcl-2表达;巢式降落式PCR联合甲基化特异性PCR检测Bcl-2基因启动子区甲基化水平,荧光RT-PCR检测主动脉组织Bcl-2基因mRNA表达。结果:18周蛋氨酸饮食可以诱导大鼠Hhcy;HE染色主动脉组织呈现动脉粥样硬化早期形态学变化,免疫组织化学显示Hhcy组主动脉组织Bcl-2表达增加;Hhcy组主动脉组织Bcl-2基因启动子区甲基化水平低于对照组(P<0.05),Bcl-2基因mRNA表达高于对照组(P<0.05)。结论:Hhcy通过降低Bcl-2甲基化水平,使Bcl-2表达增加,平滑肌细胞凋亡减弱,可能是其参与动脉粥样硬化形成的分子机制之一。
AIM: To investigate the effect of hyperhomocysteinemia-induced hyperhomocysteinemia on the methylation of Bcl-2 promoter in rat aortic tissue. Methods: Twenty-four healthy male Wistar rats aged 6 weeks, weighing 160 ± 10 g, were randomly divided into control group and Hhcy group, 12 rats in each group. The control group was given AIN-93G formula feed, and Hhcy group was fed AIN-93G formula diet containing 1.7% methionine. After 18 weeks of feeding, plasma homocysteine concentration was determined by automatic biochemical analyzer. The morphological changes of the aorta were observed by HE staining. The expression of Bcl-2 was detected by immunohistochemical staining. The methylation level of Bcl-2 promoter was detected by nested-PCR and methylation-specific PCR Fluorescence RT-PCR was used to detect the mRNA expression of Bcl-2 in aorta. Results: Methionine diet for 18 weeks could induce Hhcy in rats. HE staining of aorta tissue showed early morphological changes of atherosclerosis. Immunohistochemistry showed that the expression of Bcl-2 in aortic tissue of Hhcy group was increased. Bcl-2 The methylation level of gene promoter was lower than that of the control group (P <0.05), and the mRNA expression of Bcl-2 gene was higher than that of the control group (P <0.05). Conclusion: Hhcy may be one of the molecular mechanisms involved in the development of atherosclerosis by decreasing the level of Bcl-2 methylation, increasing the expression of Bcl-2 and decreasing the apoptosis of smooth muscle cells.