食管反流大鼠肿瘤诱发过程中p53基因突变的研究

MUTATION OF p53 IN CARCINOGENESIS OF ESOPHAGEAL TUMOR CONSEQUENT TO REFLUX OF DIFFERENT GASTRO INTESTINAL SECRETIONS

通过动物实验研究不同反流状态下食管肿瘤诱发过程中 p5 3基因的突变情况。选用SD大鼠进行不同手术 ,制作单纯胃食管反流 (G组 )、单纯十二指肠食管反流 (D组 )、胃十二指肠混合液食管反流 (DG组 )及无反流对照 (C组 )模型 ,于术后 4周开始注射食管致癌剂甲基戊基亚硝胺 (MANA) ,共 15周 ,于术后 2 0、2 6、40周分批取得食管组织 ,进行基因组DNA抽提 ,合成 p5 3基因的第 5、6、7、8外显子的 4对引物 ,进行PCR扩增。扩增产物变性后进行非变性聚丙烯酰胺凝胶电泳 (PCR SSCP) ,电泳后凝胶以硝酸银染色。结果显示PCR扩增未见 p5 3基因缺失。 2 0周时D组及DG组各有 1只 (10 % )标本检测出p5 3突变。此后随时间延长各组突变增加 ,40周时D组 (31 6 % )、DG组 (33 3 % )的突变率显著高于C组 (15 4% )和G组 (11 7% ) ,P均 <0 0 5 ,后两者差异无显著性意义 (P >0 0 5 )。提示十二指肠内容物食管反流可导致食管粘膜上皮p5 3基因的突变率增加 ,这可能是它促进食管肿瘤发生的机制之一。而单纯胃液反流不增加食管的 p5

The aim of the study is to evaluate the presence of mutations of p53 gene in RE and esophageal tumor induced by reflux of different gastro intestinal secretions. Gastroesophageal reflux(G), duodenoesophageal reflux(D) and duodeno gastroesophageal reflux(DG) model and control group(C) were reproduced by operations. All rats were given carcinogen (methyl n amyl nitrosamine) and their esophagi were harvested at 20, 26, 40 weeks after the operation. DNA was extracted and PCR amplification for 5, 6, 7, 8 exon of p53 followed by SSCP and AgNO 3 staining were used to detect mutation of p53 gene. It was showed that p53 gene mutations occurred in D and DG group at 20 weeks, and then more mutations were found at 26 and 40 weeks. The total rate of p53 mutation was 15 4% in groupc, 11 7% in G, 31 6% in D,and 33 3% in DG.The changes in the last two groups were significantly more compared with the former two groups.It is suggested that reflux of duodenal contents might promote mutations in p53 gene in esophageal mucosa, and reflux of gastric juice alone has lesser impact on p53 mutation.