p63和p73的表达与苯并(a)芘致H1299和16HBE细胞DNA损伤的关系

Associations between Expressions of p63,p73 and BaP-induced DNA Damage in H1299 and 16HBE Cells

背景与目的:研究p63与p73的mRNA表达与BaP致人肺腺癌细胞(H1299)和人支气管上皮细胞(16HBE)DNA损伤的关系。材料与方法:分别用不同浓度BaP(8、16、32、64和128μmol/L)处理H1299和16HBE两种细胞,在4 h和12 h时,使用相应的生化检测试剂盒分别测定细胞裂解液中MDA的水平和SOD、GSH-Px的活性,用qRT-PCR方法测定处理后细胞的p53、p63、p73、mdm2和mdm4的mRNA水平;用Comet实验评价细胞DNA损伤程度。结果:16、32和64μmol/L BaP处理4 h时,两种细胞MDA水平显著性升高,SOD和GSH-Px活性显著性下降(P<0.05)。用BaP处理H1299和16HBE细胞4 h和12 h时均观察到DNA损伤随浓度增加而加重,且呈剂量-效应关系(P<0.01),mdm2、mdm4mRNA表达水平升高(P<0.01)。不过仅在12 h时p53基因mRNA表达水平较对照组显著增加(P<0.01)。在4 h和12 h时点,仅在H1299细胞的p63和p73mRNA表达增加(P<0.05)。结论:在BaP致p53缺失的H1299细胞的DNA损伤中,BaP可能通过不依赖p53信号通路激活了p63和p73 mRNA的表达。

BACKGROUND AND AIM： The associations between the changes of p63 and p73 in mRNA levels and the BaP-induced DNA damage in H1299 cells and in 16HBE cells were investigated. MATERIALS AND METHODS： Both H1299 cells and 16HBE cells were treated with BaP at various concentrations （8, 16, 32, 64 and 128 μmol/L） for 4 h and 12 h.At the two time points, the levels of MDA,SOD and GSH-Px were measured using the test kits. The mRNA levels of p53,p63,p73,mdm2 and mdm4 genes were detected by RT-PCR assay. DNA damage in the cells were evaluated by Comet assay. RESULTS： At 4 h, enhanced MDA level was observed （P 〈 0.05） . However, levels of SOD and GSH-Px were significantly decreased（P 〈 0.05 for both） in both kinds of cells. After 16HBE and H1299 cells were treated for 4 h and 12 h, DNA damage and mRNA expression levels of mdm2 and mdm4 genes increased in a dose-dependent manner（P 〈 0.01 for all） . But enhanced mRNA expression level of p53 was observed only at 12 h（P 〈 0.01） In addition,significant up-regulation of p63 and p73 genes in mRNA levels at the two time points were observed in H1299 cells（ P 〈 0.05） . CONCLUSION： The BaP-induced DNA damage may be associated with enhanced mRNA levels of p63 and p73 genes in the p53-null H1299 cells.