摘要
目的:检测Lactotransferrin(LTF)基因在胃癌细胞株BGC823内的甲基化和表达情况,探讨LTF的DNA启动子区甲基化异常与其在胃癌内表达的相关性。方法:用亚硫酸氢盐测序PCR方法(BSP)检测BGC823启动子区甲基化状态,使用real-time PCR和Western blot法分别检测LTF基因在BGC823内的mRNA和蛋白表达水平。同时使用去甲基化剂5-aza-CdR处理胃癌细胞株,观察给药前后的DNA甲基化状态和mRNA表达情况。结果:BGC823启动子甲基化率达53.6%,使用5-aza-CdR后,药物处理的两组(药物浓度2μmol/L组和药物浓度10μmol/L组)和对照组甲基化率差异和LTF的mRNA表达差异有统计学意义(P<0.05),并且随着BGC823启动子甲基化程度的下降,其mRNA和蛋白表达增加。而药物处理组间差异无统计学意义(P>0.05)。结论:LTF基因在胃癌细胞株BGC823内表现为较高甲基化状态,而且LTF的表达和其启动子区甲基化情况相关,使用去甲基化剂能逆转DNA的甲基化情况从而使其mRNA重新表达。
Objective： To detect the methylation status and mRNA expression of LTF （lactotransferrin） gene in BGC823 gastric cancer cell lines, to explore the relationship between the abnormal methylation of LTF gene’s promoter region and its expression in gastric cancer.Methods： Bisulifte genomic sequencing PCR （BSP） was adopted to detect the promotor methylation status of LTF. Real-time PCR and Western blot were respectively used to evaluate LTF expression in mRNA and protein level. Meanwhile, 5-aza-CdR was used to reverse the LTF genes’ methylation status, and after the treatment, methylation and mRNA expression status was recorded. Results： The promotor methylation rates of LTF in BGC823 was 53.6%. Compared to the control group, meth-ylation status and mRNA expression of LTF in drug treatment groups （the 5-aza-CdR concentration was 2 μmol/L and 10 μmol/L） were reversed by 5-aza-CdR, which showed statistical differences （P〈0.05）. But no diffenrence had shown between different treatment groups （P〉0.05）.Conclusion： The LTF methylation in promoter region is high in BGC823 which is related with the low expression of mRNA level. Demethylation agents can reverse the methylation status and re-express its mRNA.
出处
《温州医学院学报》
CAS
2014年第6期426-430,共5页
Journal of Wenzhou Medical College
