PARG基因沉默在六价铬诱导细胞周期改变中的作用  

Effect of poly(ADP-ribose) glycohydrolase silencing on cell cycle alternation in response to six valence chromium

作　　者：黄海燕[1] 蔡剑锋[1] 刘建军[1] 夏菠[1] 杨淋清[1] 吴德生[1] 黄新凤[1] 杨细飞[1] 洪文旭[1] 庄志雄[1] 

机构地区：[1]深圳市疾病预防控制中心深圳现代毒理学重点实验室,广东深圳518055

出　　处：《毒理学杂志》2016年第3期200-204,共5页Journal of Toxicology

基　　金：国家自然科学基金(81370080;81001261);深圳市未来产业专项资金(ZDSYS20140509101335476)

摘　　要：目的深入了解PARG的功能及探讨PARG基因沉默在调节六价铬诱导细胞周期改变的作用。方法使用前期构建的PARG基因缺陷细胞(sh PARG细胞)和正常16HBE细胞作为研究对象,进行0、0.3、0.6、1.2、2.5和5.0μmol/L的六价铬[Cr(Ⅵ)]溶液染毒处理24 h,利用免疫荧光法检测PAR的表达、流式细胞仪观察细胞周期、RTPCR分析ATM和P53基因mRNA水平的表达。结果 Cr(Ⅵ)染毒处理后,正常16HBE细胞的S期明显延长,0、0.3、0.6、1.2、2.5和5.0μmol/L剂量组S期细胞比例分别为18.3%、21.6%、26.0%、30.9%、38.8%和43.2%,G2期明显缩短;sh PARG细胞的S期延长,但比例增幅远小于正常16HBE细胞,0、0.3、0.6、1.2、2.5和5.0μmol/L剂量组S期细胞比例分别为17.0%、19.0%、20.1%、21.2%、24.5%和31.3%。正常16HBE细胞内P53基因表达随Cr(Ⅵ)作用剂量的增加而逐渐升高,PARG缺陷细胞内P53基因表达改变不明显(与对照组相比,P>0.05);Cr(Ⅵ)染毒处理后,两种细胞内ATM基因的表达均增加,但正常16HBE细胞内增加更明显,如:5.0μmol/L的Cr(Ⅵ)作用时,正常16HBE细胞内ATM基因的表达升高可达6.67倍(与对照组相比,P<0.01),而sh PARG细胞内ATM基因表达仅升高2.27倍(与对照组相比,P<0.01)。结论 PARG基因沉默可对抗Cr(Ⅵ)诱导的细胞周期时相改变。Objective To explore the role of poly（ ADP-ribose） glycohydrolase（ PARG） and investigate the effect of PARG silencing on cell cycle alternation in response to six valence chromium. Methods The 16 HBE cells and sh PARG cells grown to 80% confluence were treated with different concentrations（ 0,0. 3,0. 6,1. 2,2. 5 and 5. 0 μmol / L） of Cr（ Ⅵ） for 24 h,then immunofluorescence assay was used to measure expression of PAR,flow cytometry analysis to detect cell cycle,quantitative real-time PCR analysis to detect the relative mRNA levels of P53 and ATM. Results Cr（ Ⅵ） enhanced S-phase arrest or delayed S-phase transition in 16 HBE cells and sh PARG cells. However,compared to 16 HBE cells,the proportion of S-phase of the cell cycle was lower in sh PARG cells（ P〈0. 05）. S-phase cells in the 16 HBE cells were 18. 3%,21. 6%,26. 0%,30. 9%,38. 8% and 43. 2% induced by 0,0. 3,0. 6,1. 2,2. 5 and 5. 0 μmol / L Cr（ Ⅵ）,which was17. 0%,19. 0%,20. 1%,21. 2%,24. 5% and 31. 3% in the sh PARG cells. Cr（ Ⅵ） induced concentration-dependent increase in expression of P53 gene in the 16 HBE cells. However,it was not detected in shRARG cells. Following the exposure to Cr（ Ⅵ）,the levels of the ataxia telangiectasia mutated（ ATM） were greater in the 16 HBE cells than the sh PARG cells. For example,the level of ATM was increased up to6. 67 times in 16 HBE cells induced by 5. 0 μmol / L Cr（ Ⅵ） for 24 h,but ATM expression increased only 2. 27 times in sh PARG cells.Conclusion PARG silencing could significantly improve cell cycle alternation induced by Cr（ Ⅵ）.

关 键 词：六价铬Cr(Ⅵ) 16HBE细胞 聚-ADP-核糖水解酶(PARG) 聚-ADP-核糖(PAR) 细胞周期 

分 类 号：R114[医药卫生—卫生毒理学]