RUNX3基因在胃癌中表达及其表达下调机制的研究

【作者】 曾超；

【导师】 贺修胜；

【作者基本信息】 南华大学 ， 病理学与病理生理学， 2006， 硕士

【摘要】 (一)胃癌组织中RUNX3基因的表达及其突变、杂合性缺失、甲基化检测 目的 为阐明RUNX3基因异常与胃癌发生发展的关系，本文从mRNA及蛋白质水平，检测胃癌组织中RUNX3基因的表达；并从基因的结构上，分析胃癌组织中RUNX3基因的突变、杂合性缺失及甲基化状况，评价RUNX3基因异常在胃癌发生及演进过程中作用与地位。 方法 采用RT-PCR技术，分析胃癌组织RUNX3基因mRNA的表达状况；应用WesternBlot、免疫组织化学技术，检测胃癌发生发展过程中RUNX3蛋白表达水平的改变；施用MSP、PCR-SSCP和微卫星结合PCR-变性聚丙烯酰胺凝胶电泳技术，从基因结构(DNA)水平，分析胃癌组织中RUNX3突变、杂合性缺失、甲基化。 结果 免疫组织化学法结果显示：RUNX3蛋白表达定位于正常胃粘膜上皮细胞的胞浆及胞核；45例正常胃粘膜组织中，RUNX3蛋白均为阳性表达，表达率为100％；45例胃癌组织中呈阳性反应者仅为23例，阳性率为51.13％；结果表明胃癌组织中存在RUNX3蛋白的表达下调，其阳性表达率明显低于正常胃粘膜组织(P＜0.05)。RUNX3蛋白在不同分化程度胃癌中的阳性表达率分别为：高分化组80.00％(12／15)，中分化组53.84％(7／13)，低分化组23.52％(4／17)，表明RUNX3蛋白表达与胃癌分化程度有关，差异有显著性意义(P＜0.05)。有淋巴结转移胃癌组RUNX3蛋白阳性表达率仅为33.33％(7／21)，无淋巴结转移胃癌组RUNX3蛋白阳性表达率为66.67％(16／24)，差异有显著性意义(P＜0.05)，结果提示，胃癌中RUNX3蛋白表达的降低可能与淋巴结转移有关。 Western Blot结果显示：25例胃癌组织中RUNX3蛋白表达强度为0.495±0.161，而在相对应的正常胃粘膜组织中其表达强度为1.274±0.108，RUNX3蛋白表达较正常胃粘膜组织明显降低。RUNX3在高、中、低分化胃癌组中蛋白表达强度分别为更多还原

【Abstract】 Part I Mutatioin, deletion, methylation and expression of RUNX3 gene in gastric carcinomaObjective we research the relation between abnormal expression of RUNX3 gene and incidence of gastric carcinoma. This article examine expression of RUNX3 from level of mRNA and protein and detect the mutation, LOH and methylation of RUNX3 gene from level of DNA in gastric carcinoma. We evaluate the influence of abnormal expression of RUNX3 gene for incidence and progression of gastric carcinoma.Methods we analyse the expression state of mRNA of RUNX3 gene by RT-PCR. Usingwestern blot and S-P immunohistochemical method, we dectect the change of the level of RUNX3 protein in gastric carcinoma.Using MSP, PCR-SSCP and denaturing PAGE gel electrophoresis, we interpret the mutation, deletion and methylation of RUNX3 gene in the level of DNA in gastric carcinoma.Results The levels of RUNX3 protein were examined in gastric carcinoma and adjacent normal gastric tissue by imrnunohistochemistry. The RUNX3 protein staining was located on the cell nucleus and cytoplasm. The positive rates of RUNX3 protein expression were 51.13% in gastric carcinoma and 100% in normal gastric mucosa respectively. The positive rate of RUNX3 protein expression in gastric carcinoma was significantly lower than that in normal gastric mucosa. The positive rate of RUNX3 protein expression was 80.00% (12/15) in well differentiated, 53.84% (7/13) in moderate differentiated and 23.52% (4/17) in poor differentiated gastric carcinoma respectively, which indicated the expression of RUNX3 protein was related to the degree of differentiation of gastric carcinoma. The positive rate of RUNX3 expression in node-positive cases was 33.33% (7/21), which was significantly lower than that in node-negative cases 66.67% (16/24) .The expression of RUNX3 was related with lymph node metastasis.The levels of RUNX3 protein were examined in gastric carcinoma and adjacent normal gastric lesions by western blot. The levels of RUNX3 protein were 1.274±0.108 in normal gastric lesions, while in gastric carcinoma they were 0.495±0.161. The level of RUNX3更多还原