Analysis of DNA methylation of isolated glands in gastric cancers and the surrounding the intestinal glands and non -intestinal glands

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The crypt isolation technique was used to analysis methylation of tumor related genes of non-metaplastic g astric gland (NM), intestinal metaplasia gland (IM) and gastric cancer gland (GC). NM, IM and GC specimens were obtained from 35 patients with gastric cancer (intestinal type [n=22], diffuse type [n=13]). Three samples were obtained from cancer, antrum and body. After crypt isolation of each sample, the non-cancerous glands were classified into NM glands, incomplete intestinal metaplasia gland (IIM) and complete intestinal metaplasia gland (CIM) by double stain methods of alcian blue and MUC5AC immnostain. These samples were examined CpG island hypermethylation by Combine Bislufite Restriction Analysis (COBRA) for 8 genes (MLH1 , p16 , HPP1, RUNX3 , ZFP64 , RASSF2A, SFRP1, E-cadherin). IIM and CIM gland have significant higher methylation than NM gland. But, IIM and CIM had no significant differences in methylation status. The methylation GC gland and diffuse type GC gland had no significant differences. The methylation level of GC gland was significantly higher than corresponding NM gland, but no difference was observed in corresponding IM gland. The present data on NM gland, GIM gland and GC gland suggest that genetic alternations already occur within the surrounding noncancerous glands.status of intestinal type .