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目的通过检测氯乙烯染毒大鼠肝细胞MGMT基因启动子区甲基化的水平及m RNA的表达,探讨MGMT基因在氯乙烯致癌过程中的表观遗传机制。方法选取96只健康雄性SD大鼠,随机分为四组:阴性对照组和低剂量(5 mg/kg)、中剂量(25 mg/kg)、高剂量(125 mg/kg)三个氯乙烯染毒组。采用腹腔注射染毒,每周3次。每组分别于染毒第6、8、12周末随机处死8只,取其肝脏。采用甲基化特异性实时荧光定量(q MSP)检测大鼠肝细胞MGMT基因启动子区甲基化的水平,采取实时荧光定量(q PCR)测定MGMT m RNA表达量。结果染毒6周时,各氯乙烯染毒组MGMT基因启动子区甲基化水平均高于阴性对照组,随着染毒剂量的增加而下降,低剂量组和中剂量组与对照组相比,差异有统计学意义(P<0.05);MGMT m RNA表达量随着染毒剂量的增加而上升,且高剂量组与低剂量组和中剂量组相比,差异有统计学意义(P<0.05)。染毒8周时,MGMT基因启动子区甲基化水平和m RNA表达量在各组间差异无统计学意义(P>0.05)。染毒12周时,MGMT基因启动子区甲基化水平随着染毒剂量的增加而升高,且高剂量组与对照组相比,差异有统计学意义(P<0.05);但MGMT m RNA表达量在各组间差异无统计学意义(P>0.05)。结论氯乙烯可致大鼠肝细胞MGMT基因启动子区甲基化水平升高,可能参与氯乙烯致癌的表观遗传机制。
OBJECTIVE: To explore the epigenetic mechanism of MGMT gene in the carcinogenesis of vinyl chloride by detecting the methylation level of mGMT gene promoter and the expression of m RNA in rat hepatocytes exposed to vinyl chloride. Methods Ninety-six healthy male Sprague Dawley rats were randomly divided into four groups: negative control group and three low-dose (5 mg / kg), medium dose (25 mg / kg) and high dose (125 mg / Exposure group. Using intraperitoneal injection, 3 times a week. Each group were randomly sacrificed at the 6th, 8th and 12th week of exposure, respectively. The methylation level of MGMT gene promoter region in rat hepatocytes was detected by methylation-specific real-time fluorescence quantitative (q MSP), and the expression of MGMT m RNA was detected by real-time fluorescence quantitative PCR. Results At 6 weeks after exposure, the methylation level of MGMT promoter in each vinyl chloride exposure group was higher than that in the negative control group and decreased with the increase of exposure dose. Compared with the control group (P <0.05). The expression of MGMT m RNA increased with the increase of exposure dose, and there was significant difference between high dose group and low dose group and middle dose group (P <0.05). At 8 weeks after exposure, there was no significant difference in methylation level and m RNA expression between MGMT gene promoter regions (P> 0.05). At 12 weeks of exposure, the methylation level of MGMT gene promoter region increased with the increase of exposure dose, and there was significant difference between high dose group and control group (P <0.05). However, MGMT m There was no significant difference in RNA expression between groups (P> 0.05). Conclusion VCM can induce the elevation of methylation level of MGMT gene promoter in rat hepatocytes, which may be involved in the epigenetic mechanism of carcinogenesis of vinyl chloride.