溃疡性结肠炎及伴发癌的p53改变和微卫星不稳定性

目的:探讨溃疡性结肠炎和不典型增生、癌变组织的p53基因改变和微卫星不稳定性状况.方法:采用SP法检测21例70个手术切除的溃疡性结肠炎标本和25例正常人的P53蛋白表达;人工微切割-PCR-SSCP/HA-克隆测序技术检测溃疡性结肠炎标本p53基因第5-8外显子的点突变;SSLP-测序技术检测10个微卫星位点的改变情况.结果:P53蛋白免疫组化发现25例正常人全部阴性;溃疡性结肠炎患者4例阳性,其中炎症性、低度不典型增生(LGD)、高度不典型增生(HGD)与癌变阳性标本分别为0/5例、1/7例、2/7例和1/2例.SSCP/HA、SSLP结果各有2例阳性,其中1例癌变病例癌组织 p53基因第6外显子及Bat26微卫星位点均阳性,其LGD区Bat26也阳性;1例LGD病例Bat26位点阳性;另1例HGD病例p53基因第8外显子阳性,后者经测序证实发生了移码突变.21例标本在TGFβRII(A)10、IGFIIR(G)8、IGFIIR(CT)5、TGFβRII(GT)3、BAX(G)8、hMSH3(A)8、hMSH6(C)8、TCF4(A)9和DPC4(CA)179个微卫星位点都阴性.结论:溃疡性结肠炎和癌变黏膜的早期就存在P53蛋白的过表达和微卫星不稳定性,前者部分是基因突变的结果.

Alterations of p53 gene and microsatellite instability in ulcerative colitis and ulcerative colitis-associated colorectal cancer

OBJECTIVE: To study the expression of p53 protein, incidence of p53 gene mutation and microsatellite instability in ulcerative colitis, dysplasia of colonic mucosa and ulcerative colitis-associated colorectal cancer. METHODS: P53 protein expression was detected by immunohistochemistry in 70 specimens from 21 cases of ulcerative colitis and 25 colonic mucosa specimens from normal subjects. The specimens of ulcerative colitis were examined for the mutation in exon 5, 6, 7, 8 of p53 gene with microdissection-PCR-SSCP/HA-clone-sequencing technique and the alterations in 10 microsatellite loci with microdissection-PCR-SSLP-clone- sequencing technique. RESULT: None of 25 normal specimens was p53-positive immunohistochemically, while 4/21 of ulcerative colitis specimens were p53-positive. P53- positive rate in inflammatory mucosa of ulcerative colitis specimens was 0/5, while that was 1/7, 2/7 and 1/2 in low-grade displasia (LGD), high-grade displasia (HGD) and carcinoma, respectively. The abnormal exons were detected by SSCP and confirmed by sequencing in 2 out of 21 cases: one was exon 6 in a case with carcinoma and the other was exon 8 in a HGD case; both had positive P53 expression. Two cases showed positive in Bat26 locus by SSLP: one was a LGD case, the other was a case of carcinoma, who also had abnormal exon 6 of p53 gene. Other 9 microsatellite loci, (TGF beta RII(A)(10), IGFIIR(G)(8), IGFIIR(CT)(5), TGF beta RII(GT)(3), BAX(G)(8), hMSH3(A)8, hMSH6(C)(8), TCF4(A)(9) and DPC4 (CA)(17)) were showed negative in all cases. CONCLUSION: P53 gene mutations and microsatellite instability may be one of the mechanisms for higher risk of carcinogenesis in ulcerative colitis.