【摘要】 目的检测结晶型NiS诱发永生化人支气管上皮细胞系(16HBE)恶性转化过程中脆性组氨酸三联体基因(FHIT)和p16基因的变化,探讨镍化合物致癌的分子机制。方法采用RT-PCR、DNA序列分析、银染PCR-SSCP方法检测恶性转化细胞、成瘤细胞中FHIT基因和p16基因的变化。结果与对照组16HBE相比,转化细胞、成瘤细胞p16基因第2外显子、第2～3外显子未见突变,mRNA表达未见异常;FHIT基因第5,6,7,8外显子及第1～4外显子、第5～9外显子未见突变,但发现转化细胞、成瘤细胞FHIT基因的mRNA失表达或出现异常转录本;对其中FHIT基因第5～9外显子的一异常转录本测序分析显示,FHIT、第6,7,8外显子缺失,第5,9外显子异常拼接,且中间插入一个36 bp的小片段。结论在结晶型NiS诱发16HBE恶性转化过程中,p16基因在DNA和mRNA水平未见异常改变,提示p16基因在镍致癌过程中可能不发挥作用;FHIT基因在mRNA水平出现缺失或异常转录本,表明FHIT基因在镍致癌过程中可能发挥一定作用;镍诱导的FHIT基因异常,可能是将外源致癌物、染色体脆性部位不稳定性和细胞恶变相联系起来的一个分子事件,FHIT基因可能是镍等外源致癌物作用的主要靶基因之一。更多还原

【Abstract】 Objective To detect the alteration of fragile histidine triad (FHIT) gene and p!6 gene during malignant transformation of immortal human bronchial epithelial cell line (16HBE) induced by crystalline nickel sulfide, and study the molecular mechanism of nickel carcinogenesis. Methods Malignant transformed cells and tumorigenic cells were examined for the alteration of FHIT gene and p16 gene by RT-PCR,DNA sequencing and silver staining PCR-SSCP. Results Compared with those of control 16HBE , neither mutation of exon2 or exon2-3, abnormal expression in p16 gene nor mutation of FHIT exon5, 6, 7 and 8, exonl-4 or exon5-9 were observed in transformed cells and tumorigenic cells. But aberrant transcript or FHIT gene expression loss were observed in transformed cells and tumorigenic cells. One of the aberrant transcripts in FHIT gene, the deletion of exon6, exon? and exonS and an insertion of 36 bp sequence replacing exon6-8, was confirmed by sequencing. Conclusion FHIT gene, not p16 gene, could play a definite role in nickel carcinogenesis. Alterations of FHIT gene induced by crystalline NiS could be a molecular event associated with carcinogen , chromosome fragile site instability and cell malignant transformation, and FHIT gene could be one of the important target genes activated by exotic carcinogens.更多还原