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目的 探讨p16~(INK4a)和p14~(ARF)基因变异与胃癌发生的关系。方法 应用聚合酶链反应(PCR)、PCR-单链构象多态性(SSCP)、PCR甲基化分析法和RT-PCR分别检测48例胃癌及癌旁组织中p16~(INK4a)和p14~(ARF)基因纯合性缺失、突变、CpG岛甲基化及mRNA表达状况。结果(1)胃癌组织p16~(INK4a)和p14~(ARF)基因总缺失率为35.4％(17/48),癌旁组织均未见纯合性缺失。(2)31例无纯合性缺失的胃癌及癌旁组织均未见p16~(INK4a)和p14~(ARF)基因点突变。(3)胃癌组织p16~(INK4a)和p14~(ARF)基因总甲基化率为47.90％(23/48),癌旁组织仅2例甲基化,两者差异有显著性(P<0.01)。(4)胃癌组织p16~(INK4a)mRNA缺失率为47.9％(23/48),其中E1α和E2共甲基化者p16~(INK4a)mRNA缺失率(100％,6/6)明显高于其他类型甲基化者(11.8％,2/17)(P<0.01);胃癌组织p14~(ARF)mRNA缺失率为45.8％(22/48),其中E1β和E2共甲基化者p14~(ARF)mRNA缺失率(100％,3/3)明显高于其他类型甲基化者(15％,3/20)(P<0.05)。(5)低未分化癌组p16~(INK4a)和p14~(ARF)mRNA共同缺失率(36.7％,11/30)明显高于高中分化癌组(5.6％,1/18)(P<0.05)。结论 胃癌中p16~(INK4a)和p14~(ARF)基因失活多由纯合性缺失和5’CpG岛甲基化所致,其表达缺失与胃癌的发生密切相关。
Objective To investigate the relationship between p16 INK4a and p14 ARF gene mutations and gastric carcinogenesis. Methods The expressions of p16 (INK4a) and p14 ~ (p14) in gastric cancer tissues and paracancerous tissues were detected by polymerase chain reaction (PCR), PCR-single strand conformation polymorphism (SSCP), PCR methylation analysis and RT- (ARF) gene homozygous deletion, mutation, CpG island methylation and mRNA expression status. Results (1) The total deletion rate of p16 INK4a and p14 ARF in gastric cancer tissues was 35.4% (17/48). There was no homozygous deletion in adjacent tissues. (2) There was no point mutations in p16 (INK4a) and p14 (ARF) genes in 31 cases of gastric cancer and paracancerous tissue without homozygous deletion. (3) The total methylation rate of p16 INK4a and p14 ARF gene was 47.90% (23/48) in gastric cancer tissues and only 2 cases were methylated in paracancer tissues, the difference was significant (P < 0.01). (4) The loss rate of p16 INK4a mRNA in gastric cancer tissue was 47.9% (23/48), in which the deletion rate of p16 INK4a mRNA (100%, 6/6) was significantly higher than that of E1α and E2 co-methylation (11.8%, 2/17) (P <0.01). The deletion rate of p14 ARF mRNA in gastric cancer tissue was 45.8% (22/48) (100%, 3/3) were significantly higher than those in other types of methylation (15%, 3/20) (P <0.05). (5) The rates of co-deletion of p16 INK4a and p14 ARF mRNA in poorly differentiated carcinoma were significantly higher than those in high differentiated cancer group (5.6%, 1/18) (36.7%, 11/30) ). Conclusion Inactivation of p16 INK4a and p14 ARF genes in gastric cancer is mostly caused by homozygous deletion and 5’CpG island methylation. The lack of expression is closely related to the occurrence of gastric cancer.