NADH对二乙基亚硝胺所致L02人肝细胞株p53基因突变和c-erbB2基因表达的影响

Effect of NADH on Mutations of p53 Gene and Expression of c-erbB2 Gene in L02 Cells Induced by Diethylnitrosamine

背景与目的:研究NADH对二乙基亚硝胺(DENA)所致L02人肝细胞株p53基因突变和c-erbB2基因表达的影响。材料与方法:采用聚合酶链反应-单链构象多态性(PCR-SSCP)及聚合酶链反应-限制性酶切片段长度多态性(PCR-RFLP)方法检测DENA所致的L02细胞p53基因突变;Southern blot分析DENA对c-erbB2基因表达的影响;研究辅酶NADH的抗突变作用。结果:PCR-SSCP分析显示NADH保护的DL02-Ⅲ细胞和DL02-B细胞p53exon7突变率均较DENA致突变组降低,差异有统计学意义(P<0.01);PCR-RFLP分析结果显示NADH降低DENA所诱发的L02细胞p53基因第7外显子249位密码子点突变率(P<0.01)。Southern blot检测结果也显示NADH可抑制DENA所致的L02细胞c-erbB2基因的表达上调。结论:还原型辅酶NADH具有抗突变作用,可降低DENA所致的L02细胞p53基因的突变,并抑制c-erbB2基因的表达。

BACKGROUND AND AIM：To investigate effects of NADH on mutations of p53 gene and c-erbB2 gene in L02 cells induced by diethylnitrosamine（DENA）.MATERIALS AND METHODS：The anti-mutagenesis effects of NADH on DENA-induced mutations of p53 gene in L02 cells were investigated by PCR-SSCP and PCR-RFLP analysis.The effect of NADH on c-erbB2 gene amplification was assessed by Southern blotting analysis.RESULTS：PCR-SSCP analysis showed that the mutation rate of p53 exon7 in DL02-Ⅲ and DL02-B cells in NADH protection groups decreased to 33.3% （4/12） and 50% （6/12）, respectively. The difference was significant（P 〈 0.01）compared with DENA mutagenesis group. PCR-RFLP analysis showed that NADH decreased the mutation frequency of the 249th codon of p53 in L02 cells induced by DENA. Southern blotting showed that NADH down-regulated the amplification of c-erbB2 gene in L02 cells induced by DENA. CONCLUSION： Reduced coenzyme NADH had anti-mutagenesis effects and could decrease mutation of p53 and inhibit the expression of c-erbB2 in L02 ceils induced by DENA.