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目的: 研究胃癌患者手术前后外周血浆、对应原发肿瘤以及癌旁组织中p16基因启动子区异常甲基化的检出状况,探讨其在胃癌疗效评估中的应用。方法:采用甲基化特异性PCR方法,扩增84例胃癌患者的原发肿瘤、癌旁组织、手术前血浆中p16基因启动子区,以及上述患者中30例行胃癌根治性手术患者术后14~21天血浆中p16基因启动子区,以15例健康人血浆作为正常对照。采用凝胶电泳-溴乙锭显色以及高效液相色谱两种方法检测产物。结果: 84例患者样品中,原发肿瘤中存在p16 基因异常甲基化26例( 31. 0% ),癌旁组织中阳性2例(0. 02% ),术前血浆中阳性12例(14. 3% ), 15例健康人血浆检测均为阴性。患者血浆阳性者同时也存在对应肿瘤组织阳性。30例同时有手术前后血浆标本的胃癌根治手术患者中,原发肿瘤组织中p16异常甲基化14例,其中6例术前血浆阳性,术后5例转阴。在检测样本中,高效液相色谱的检测结果与凝胶电泳-溴乙锭显色结果一致。结论:胃癌患者外周血浆中可以检测到与原发肿瘤一致的p16异常甲基化,手术后血浆中p16甲基化状态的变化与手术治疗有关,高效液相色谱作为一种便捷的技术可以应用于PCR产物的检测。
OBJECTIVE: To study the detection of abnormal methylation of p16 gene promoter region in peripheral blood plasma of patients with gastric cancer before and after surgery, corresponding to the primary tumor and paracancerous tissues, and to explore its application in curative effect evaluation of gastric cancer. Methods: The methylation-specific PCR method was used to amplify the promoter region of p16 gene in 84 cases of gastric cancer and its preoperative plasma, and 30 cases of radical surgery of gastric cancer after operation The plasma p16 gene promoter region in 14-21 days, 15 healthy human plasma as a normal control. Gel electrophoresis - ethidium bromide and high performance liquid chromatography two methods to detect the product. Results: Among the 84 patients, there were 26 cases (31.0%) of abnormal methylation of p16 gene, 2 cases (0.2%) positive in adjacent tissues and 12 cases 14.3%), 15 cases of healthy plasma test were negative. Patients with positive plasma also exist corresponding to the positive tumor tissue. Thirty patients with gastric cancer underwent radical surgery before and after surgery had abnormal methylation of p16 in 14 cases, of which 6 cases were positive in preoperative plasma and 5 cases were negative after operation. In the test samples, the detection results of high performance liquid chromatography and gel electrophoresis - ethidium bromide color results. Conclusion: P16 abnormal methylation can be detected in the peripheral plasma of gastric cancer patients consistent with the primary tumor. The change of p16 methylation status after surgery is related to the surgical treatment. HPLC as a convenient technique can be applied Detection of PCR products.