硫酸镍诱导16HBE细胞恶变过程中的K-ras、P15基因的改变及基因组不稳定性分析

Genomic Instability and Alterations of K-ras and P15 Genes in Transformed 16HBE Cells by Nickel Sulfate

目的 检测硫酸镍对K—ras基因和P15基因的改变及基因组不稳定性的影响，从而进一步探讨镍化合物致癌的分子机制。方法 采用聚合酶链反应-单链构象多态性分析方法探查硫酸镍在诱导16HBE细胞恶变过程中的K—ras基因Exon1和P15基因Exon2存在状况。采用随机扩增多态性技术对硫酸镍在诱导16HBE细胞恶变过程中的基因组不稳定性进行分析。结果 K—ras基因Exon1和P15基因Exon2未发生改变。本实验所选用的7条随机引物均能扩增出清晰、明显的条带，条带数在1～6条之间。7条引物中P1、P4、P7三条引物扩增的片段在实验组和对照组之间无差异。其余四条引物均有差异，对于同一随机引物他们都具有特异的带型。结论 P15基因第2外显子和K—ras琏因第一外显子可能不足硫酸镍作用的靶部位。在硫酸镍诱发细胞恶变转化过程中，基因组变得逐渐不稳定。

Objective To detect the alterations of K - ras gene and P15 gene and genomic instability in the malignant process of 16HBE cells induced by nickel sulfate. Methods The PCR single strand conformation polymorphism （PCR - SS- CP） was used to examine the K- ras gene Exon1 and P15 gene Exon2 alterations in the malignant process of 16HBE cells induced by nickel sulfate. The genomic instability was analyzed by random amplified polymorohic DNA（RAPD）. Results Compared with the negative control cells, alterations of P15 gene and K- ras gene were not observed and genomic instability was showed in the malignant process of 16HBE cells induced by nickel sulfate. The 7 random primers used in our experiments all induced 1 -6 clear and obvious bands. Among the 7 primers, the P1, P4, and P7 amplified segments were similar to the control. The other 4 primers induced different bands, however, and each showed its specific band pattern. Conclusions Nickel sulfate may not induce alterations of P15 gene and K- ras gene and it indicates that nickel sulfate has no effect on the P15 gene Exon2 and K - ras gene Exon1. But they could induce genomic instability, and the appearance of specific PCR bands suggests that the genome in the malignat transformed cells become more unstable.