银染PCR-SSCP检测粪便中结肠直肠癌抑癌基因突变的研究

Point Mutation Analysis on Tumor Suppressor Genes in Stool of Patients with Colorectal Carcinoma by Silver-Stain Polymerase Chain Reaction-Single Strand Conformation Polymorphism

目的：建立无创伤性结肠直肠癌筛查方法。方法：应用８％银染非变性聚丙烯酰胺凝胶（丙烯酰胺：甲叉丙烯酰胺＝２９：１）检测粪便中抑癌基因点突变。结果：４５例结肠直肠癌患者中，高分化癌１５例，中分化癌１８例，低分化癌 １２例； Ｄｕｋｅｓ分期中 Ａ期 １５例， Ｂ期 １６例， Ｃ期 ９例， Ｄ期 ５例；淋巴结转移 １４例，无淋巴结转移３１例。正常肠粘膜组织为对照组。粪便中 ＡＰＣ（ａｄｅｎｏｍａｔｏｕｓ ｐｏｌｙｐｏｓｉｓ ｃｏｌｉ）及／或 ＭＣＣ（ｍｕｔａｔｅｄ ｉｎ ｃｏｌｏｒｅｃｔａｌ ｃａｎｃｅｒ）基因突变２８例（６２．２％），占有ＡＰＣ及／或ＭＣＣ基因突变的结直肠癌组织的９３．３％（２８／３０例）；粪便ＡＰＣ及ＭＣＣ基因突变分别占 ＡＰＣ及 ＭＣＣ基因突变的结直肠癌组织的９１． ３％及 ８４ ６％。粪便中基因突变与肿瘤组织学类型、Ｄｕｋｅｓ分期、淋巴结转移、粪便潜血是否阳性及有无便血史、肿块大小、病灶部位及肿瘤形态统计学上无显著差异（Ｐ＞０．０５）。结论：该方法可望成为无创伤性结肠直肠癌普查、筛选、疗效观察及预后判断的方法。

Objective: The aim of this study was to seek a new approach for early detection and screening colorectal carcinoma. Methods: Point mutation analysis through 8% non-denaturant polyacrylamide gel electrophoresis (acrylamide: N, N, -methylene bisacrylamide = 29: 1 ) was applied to examine stool of patients with colorectal carcinoma. Results: Forty-five patients with different histological categories at all clinical stages and normal cases were examined. There were 14 individuals with lymph node metastasis while thirty-one without lymph node involvement. Of 45 samples investigated, twenty-eight patients with adenomatous polyposis colt(APC) or/and mutated in colorectal cancer(MCC) mutations in the stools were identified, which representing 62. 2% of all patients and 93. 3% of the patients in whom the tumor tissue with gene mutations. APC or MCC determined in the stool was 91. 3% or 84. 6% respectively corresponding to the carclnomas in which APC or MCC aberrants had been detectable. There were no significant relationships between genetic alterations in the stools and histological types, Dukes stages, lymph node involvements, sizes or locations of the tumor (P > 0. 05), and between genetic abnormalities in the stools and histomorphologies or blood occult tests of the patients (P > 0. 05). Conclusions: The method presented here may provides a useful noninvasive technique for screening, detection and prognosis of colorectal neoplasms.