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目的探讨胃癌组织中与细胞增殖、凋亡和转录有关的基因移码突变与微卫星不稳定性(MSI)的关系。方法采用酚-氯仿-异戊醇法从胃癌及切缘正常石蜡组织中提取DNA。组织切片中肿瘤成分不足50%时采用显微切割方法。以聚合酶链反应-单链构象多态性(PCR-SSCP)、直接DNA测序法检测MSI及转化生长因子βⅡ受体(TGFβRⅡ)、胰岛素生长因子Ⅱ受体(IGFⅡR)、凋亡诱导蛋白BAX和转录因子E2F4基因的移码突变。按照MSI的发生频率将胃癌患者分为3组:出现≥2个位点的MSI为高MSI,出现1个位点的MSI为低MSI,无MSI发生为微卫星稳定。结果61例胃癌患者中,12例(19.7%)为高MSI,11例(18.0%)为低MSI,38例(62.3%)为微卫星稳定。TGFβRⅡ、IGFⅡR、BAX和E2F4移码突变检出率分别为12例(19.7%)、3例(4.9%)、4例(6.6%)和10例(16.4%)。12例高MSI胃癌中,有10例TGFβRⅡ基因突变,3例IGFⅡR基因突变,4例BAX基因突变,10例E2F4基因突变;其移码突变率与高MSI发生密切相关。微卫星稳定组的肿瘤中未发现这些基因的突变。结论TGFβRⅡ、IGFⅡR、BAX和E2F4基因编码区重复序列是MSI发生的靶点,这些基因的移码突变在MSI胃癌的发生和发展中起了重要作用。
Objective To investigate the relationship between microsatellite instability (MSI) and the frameshift mutation of genes involved in cell proliferation, apoptosis and transcription in gastric cancer. Methods DNA was extracted from gastric cancer and normal paraffin tissues by phenol - chloroform - isoamyl alcohol method. Tissue sections of the tumor composition of less than 50% using microdissection method. PCR-SSCP and direct DNA sequencing were used to detect the expression of MSI and TGFβRⅡ, IGFⅡR, apoptosis-inducing protein BAX And frameshift mutations of the transcription factor E2F4 gene. Gastric cancer patients were divided into 3 groups according to the frequency of MSI: MSI with ≥2 sites showed high MSI, MSI with one site showed low MSI, and no MSI occurred with microsatellite stable. Results Of 61 patients with gastric cancer, 12 (19.7%) had high MSI, 11 (18.0%) had low MSI and 38 (62.3%) had microsatellite stable. The detection rates of frameshift mutation of TGFβRⅡ, IGFⅡR, BAX and E2F4 were 12 cases (19.7%), 3 cases (4.9%), 4 cases (6.6%) and 10 cases (16.4%). Ten cases of TGFβRⅡ gene mutation, three cases of IGFⅡR gene mutation, four cases of BAX gene mutation and ten cases of E2F4 gene mutation were found in 12 cases of high MSI gastric cancer. The frequency of the frameshift mutation was closely related to high MSI. No mutations in these genes were found in the microsatellite-stabilized tumors. Conclusion The repeats of TGFβRⅡ, IGFⅡR, BAX and E2F4 genes are the targets of MSI. The frameshift mutations of these genes play an important role in the occurrence and development of MSI gastric cancer.