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目的探讨转移抑制基因TIMP3(金属组织蛋白酶抑制因子-3)在胃癌和胃癌细胞系中启动子甲基化及其蛋白和mRNA表达的特点,并分析两者之间的关联。方法应用MSP法检测TIMP3基因启动子区的甲基化状态;RT-PCR和免疫组织/细胞化学法检测TIMP3基因mRNA和蛋白的表达;观察DNA去甲基化剂(5-aza-2’-deoxycitydine)对人胃癌细胞系(MGC803和HGC-27)细胞系TIMP3基因表达的影响。结果TIMP3基因于胃癌组织中的甲基化频率为17/33(51.5%);四个胃癌细胞系均存在TIMP3基因的甲基化,而MGC803、BGC823和HGC-27仍有该基因的表达或低水平表达;在去甲基化剂的作用下TIMP3基因的表达水平明显上调。结论TIMP3的异常甲基化是其表达下调或沉默的主要原因并存在部分甲基化的倾向。该指标可作为胃癌的敏感性表观遗传学标记物在胃癌的筛查和诊断中应用。
Objective To investigate the promoter methylation and protein and mRNA expression of the metastasis suppressor gene TIMP3 (TIMP3) in gastric cancer and gastric cancer cell lines and to analyze the relationship between the two. Methods The methylation status of TIMP3 gene promoter region was detected by MSP method. The mRNA and protein expression of TIMP3 gene was detected by RT-PCR and immunohistochemistry. The effects of DNA demethylating agent (5-aza-2’- deoxycitydine on human gastric cancer cell lines (MGC803 and HGC-27) cell line TIMP3 gene expression. Results The methylation frequency of TIMP3 gene in gastric cancer tissue was 17/33 (51.5%). The methylation of TIMP3 gene was found in all four gastric cancer cell lines, while the expression of TIMP3 gene still existed in MGC803, BGC823 and HGC-27 cells. Low level of expression; in the role of demethylating agent TIMP3 gene expression was significantly increased. Conclusions Aberrant methylation of TIMP3 is the main reason for the down-regulation or silencing of TIMP3, and the tendency of partial methylation exists. This index can be used as a sensitive epigenetic marker for gastric cancer in gastric cancer screening and diagnosis.