长期吸烟对大鼠肺组织p53、K-ras蛋白表达和基因突变的影响

THE STUDY OF INFLUENCE OF LONG-TERM SMOKING ON THE EXPRESSION AND MUTATION OF p53 AND K-ras GENE IN RAT LUNGS

目的观察长期吸烟对Wistar大鼠肺组织p53、K-ras表达的影响,研究吸烟与肺组织p53、K-ras基因突变的关系。方法建立Wistar大鼠被动吸烟模型。72只雄性Wistar大鼠分为实验组和对照组,实验组被动吸烟6个月,对照组正常呼吸条件下饲养,每个月末分别取6只大鼠肺组织,免疫组织化学观察p53和K-ras蛋白表达情况,聚合酶链式反应-单链构象多态性(PCR-SSCP)技术检测p53第5、6、7～8外显子和K-ras第1外显子的突变情况。结果免疫组织化学结果显示,p53蛋白表达于细胞核,K-ras蛋白表达于胞浆,吸烟组p53、K-ras蛋白表达强度与正常组比较有显著性差异。随着吸烟时间的增加,p53、K-ras蛋白阳性表达率均随吸烟时间的延长而升高。PCR-SSCP显示,p53基因随吸烟时间的延长突变率增加;K-ras基因突变率随吸烟时间延长增加不显著。结论吸烟可以导致大鼠肺组织p53、K-ras蛋白表达增强和基因突变率增加,随吸烟时间的延长呈上升趋势,为吸烟对肺的组织基因突变的判定及吸烟者肺癌的早期诊断提供理论依据,具有重要的理论和应用价值。

Objective To observe the influence of long-term smoking on the expression of p53 and K-ras in rat lung tissues, and to study the relationship of smoking to the mutation of p53 and K-ras gene. Methods The model of Wistar rat smoking was built up. Seventy-two healthy male Wistar rats were divided into the experimental group and control group at random. The rats of the experimental group were compelled to smoke, and the rats of the control group were given the same condition as the experimental group was, without smoking. The rats of the experimental group were smoked for 6 months. At the end of each month, 6 rats were chosen from the two groups respectively, their lung tissues were sampled and immunohistochemistry was applied to observe the expression of p53 and K-ras in lung tissues. Finally, the mutation which might happen in the exon 5, 6, 7-8 of p53 and the exon 1 of K-ras was examined by PCR-SSCP. Results The p53 protein was expressed in cell nucleus and K-ras in cytoplasm. The positive ratio of protein expression was increased with the extension of smoking time. The mutation of p53 was increased as the smoking time extended. But the effect of smoking time was not that significant on the mutation of K-ras. Conclusion Smoking can strengthen the expression of p53 and K-ras protein and can also result in gene mutation. As the time of smoking extended, those phenomenons were tending to rise. That provided the theoretical evidence which can be used to judge the lesion of lung tissues caused by smoking and help the early diagnosis of smoking-related lung carcinomas. It is of great theoretical and application values.