DNA聚合酶β对苯并[a]芘致细胞遗传毒性及基因组遗传不稳定性的影响

目的 研究DNA聚合酶β(polβ)表达水平对苯并[a]芘(BaP)致细胞遗传毒性及基因组遗传不稳定性的影响,为BaP的致癌分子机制提供实验依据.方法 采用3种具有相同遗传背景的小鼠胚胎成纤维细胞[polβ野生型(polβ+/+)、polβ缺陷型(polβ-/-)和polβ高表达型(polβoe)]作为模型,检测BaP对细胞的氧化损伤作用,细胞遗传毒性和基因组遗传不稳定性.结果 随着BaP浓度的增加,3种细胞的存活率和克隆形成能力均下降.5.00和20.00 μmol/L BaP染毒时,polβ-/-细胞产生的ROS荧光强度均大于polβ+/+和polβoe细胞,差异均有统计学意义(P＜0.05).在5.00和20.00μmol/L时,polβ-/-卢细胞SOD活力为(76.56±2.84)和(62.78±4.28) U/mg pro,低于对照组[(84.85±3.59) U/mg pro]和polβ+/+细胞[(85.21 ±3.20)和(76.90±3.38) U/mg pro],20.00 μmol/L BaP染毒组polβoe细胞SOD活力[(82.59±4.64) U/mg pro]低于对照组[(88.58±6.77) U/mg pro]但高于相同浓度染毒的polβ +/+细胞,差异均有统计学意义(P＜0.05).1.25、5.00和20.00 μmol/L BaP染毒组的polβ-/-细胞的微核形成率明显高于polβ +/+细胞,5.00和20.00 μmol/LBaP染毒组的polβ oe细胞的微核形成率明显低于pol β+/+细胞,差异均有统计学意义(P＜0.05).5.00和20.00 μmol/L BaP染毒组polβ-/-细胞的HPRT基因突变频率为26.16×10-6和37.51×10-6,polβ oe细胞为27.68×10-6和38.63×10-6,均明显高于polβ +/+细胞(19.76×10-6和24.78×10-6),差异均有统计学意义(P＜0.05).结论 Polβ在保护细胞免受BaP造成的细胞毒性和遗传毒性方面具有积极的作用,其正常表达对于维持细胞基因组遗传稳定性具有重要的意义.

Effects of DNA polymerase beta on the genotoxicity and genetic instability induced by benzo(a)pyrene

Objective To explore the effects of DNA polymerase β expression level on the genotoxicity and genetic instability induced by benzo (a) pyrene (BaP),and provide experimental the basis for further study on the carcinogenic molecular mechanism of BaP.Methods Three kinds of cell lines with the identical genetic background,polβ wild-type cells (polβ +/+),polβ null cells (polβ -/-) and polβ overexpression cells (polβ oe) were applied as cellular models.The oxidative damage,genotoxicity and genetic instability induced by BaP were analyzed by using different methods respectively.Results Cell viability and colony forming ability of 3 kinds of cell lines exposed to BaP decreased with BaP.After treated with 5 and 20 μmol/L concentration of BaP,fluorescence intensity of polβ-/- cell line was significantly higher than that of other two cell lines (P＜0.05).When treated with 5.00 μmol/L and 20.00 μ mol/L concentration of BaP,the SOD activities (76.56±2.84 and 62.78±4.28 U/mg pro) of polβ-/- cell line were significantly lower than that (84.85±3.59) of control group and those (85.21±3.20 and 76.90±3.38 U/mg pro) of polβ +/+ cell line.In 20.00 μmol/L BaP group.SOD activity (82.59±4.64 U/mg pro) of polβoe cell line was lower than that (88.58t6.77 U/mg pro) of control but higher than that of polβ +/+ cell line (P＜0.05).In 1.25,5.00 and 20.00 μ mol/L concentration BaP groups,the micronucleus rates of polβ-/- cell line were much higher than those of polβ+/+ cell line (P＜0.05).In 5.00 and 20.00 μmol/L concentration BaP groups,the micronucleus rates of polβ oe cell line were significantly lower than those of polβ+/+ line (P＜0.05).In 5.00 and 20.00 μmol/L concentration BaP groups,HPRT gene mutation frequencies (26.16×10-6 and 37.51×10-6; 27.68×10-6 and 38.63×10-6) in polβ-/- cells and polβ oe cells were significantly higher than those (19.76×10-6 and 24.78×10-6) of polβ +/+ cells (P＜0.05).Conclusion Polβ could play a role in protecting the cells from the genotoxicity and genetic instability induced by BaP,and the normal expression level of polβ was indispensable for maintaining genome stability.