Li Jian-Hua, Shi Xian-Zhe, Liu Min, Wang Yan, Yu Zhi-Hong, Xu Guo-Wang, Lu Shen
Laboratory of Molecular Biology, Experimental Center, The Second Affiliated Hospital, Dalian Medical University, Dalian, Liaoning, 116027, P.R.China.
Ai Zheng. 2005 Mar;24(3):273-7.
BACKGROUND & OBJECTIVE: Mismatch repair (MMR) deficiency, characterized by microsatellite instability (MSI), is a major cause of hereditary nonpolyposis colorectal carcinoma (HNPCC). Our previous study showed that loss of MMR protein expression can also be seen in gastric carcinoma, but MMR gene mutation seldom occur. Promoter methylation, a main cause of suppressor gene inactivation, might also lead to defect of MMR gene function. This study was designed to investigate the relationship of hMLH1 gene promoter methylation status to MSI in gastric carcinoma, and analyze its action as a risk factor of carcinogenesis.
DNA was extracted from 52 samples of gastric carcinoma and their adjacent non-cancerous gastric mucosa. Polymerase chain reaction (PCR) was used to amplify microsatellite loci BAT-26, D17S261, D3S1283, D2S123, and D3S1611. MSI was studied by capillary electrophoresis. Methylation of hMLH1 gene promoter was detected by restrictive endonuclease digestion, expression of hMLH1 protein was detected by immunohistochemistry.
Of the 52 specimens of gastric carcinoma, 13 with high MSI (MSI-H), 2 with low MSI (MSI-l), and 37 with microsatellite stability (MSS). Frequency of hMLH1 promoter methylation was significantly higher in the 13 specimens of gastric carcinoma with MSI-H than in the 39 specimens of gastric carcinoma with MSI-L or MSS (100% vs. 2.6%, P<0.01). Furthermore, frequency of hMLH1 promoter methylation was significantly higher in the 13 specimens of adjacent non-cancerous gastric mucosa of gastric carcinoma with MSI-H than in the 39 specimens of adjacent non-cancerous gastric mucosa of gastric carcinoma with MSI-L or MSS (46.2% vs. 2.6%, P<0.01). The methylation status was accordant with loss of hMLH1 protein expression. Methylation of hMLH1 gene promoter had no relation with differentiation and clinical stage of gastric carcinoma.
Methylation of hMLH1 gene promoter exists in gastric carcinoma tissues with MSI-H, and their adjacent non-cancerous gastric mucosa, it may be a risk factor of carcinogenesis of gastric carcinoma.
错配修复（MMR）缺陷以微卫星不稳定（MSI）为特征，是遗传性非息肉病性结直肠癌（HNPCC）的主要病因。我们先前的研究表明，MMR蛋白表达缺失在胃癌中也可见，但MMR基因突变很少发生。启动子甲基化作为抑癌基因失活的主要原因，也可能导致MMR基因功能缺陷。本研究旨在探讨hMLH1基因启动子甲基化状态与胃癌中MSI的关系，并分析其作为致癌危险因素的作用。
从52例胃癌及其癌旁非癌胃黏膜组织中提取DNA。采用聚合酶链反应（PCR）扩增微卫星位点BAT-26、D17S261、D3S1283、D2S123和D3S1611。通过毛细管电泳研究MSI。采用限制性内切酶消化检测hMLH1基因启动子的甲基化，采用免疫组织化学检测hMLH1蛋白的表达。
52例胃癌标本中，13例为高度微卫星不稳定（MSI-H），2例为低度微卫星不稳定（MSI-L），37例为微卫星稳定（MSS）。hMLH1启动子甲基化频率在13例MSI-H胃癌标本中显著高于39例MSI-L或MSS胃癌标本（100%对2.6%，P<0.01）。此外，hMLH1启动子甲基化频率在13例MSI-H胃癌的癌旁非癌胃黏膜标本中显著高于39例MSI-L或MSS胃癌的癌旁非癌胃黏膜标本（46.2%对2.6%，P<0.01）。甲基化状态与hMLH1蛋白表达缺失一致。hMLH1基因启动子甲基化与胃癌的分化程度和临床分期无关。
hMLH1基因启动子甲基化存在于MSI-H的胃癌组织及其癌旁非癌胃黏膜中，可能是胃癌发生的危险因素。
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