六溴环十二烷对视黄醛类X受体α、孕烷X受体、过氧化物酶体增殖物活化受体γ的影响及其相互作用  

Effects of hexabromocyclododecanes on retinoid X receptor α,pregnane X receptor and peroxisome proliferator-activated receptor γ

作　　者：万克艳 蒋友胜[2] 张建清[2] 王晓辉[2] 雷毅雄[1] 李云秀[3] 肖悦[3] 梅树江[2] 周健[2] 

机构地区：[1]广州医科大学公共卫生学院,广州511436 [2]深圳市疾病预防控制中心 [3]四川大学公共卫生学院

出　　处：《卫生研究》2017年第3期396-403,共8页Journal of Hygiene Research

基　　金：国家自然科学基金(No.21677103);广东省科技厅产业技术研究与开发资金(No.2013B030800001)

摘　　要：目的探讨六溴环十二烷(HBCDs)对小鼠神经母细胞瘤细胞N2a增殖的影响以及对3个重要细胞核受体:视黄醛类X受体α(RXRα)、过氧化物酶体增殖物活化受体γ(PPARγ)、孕烷X受体(PXR)表达及其相互作用的影响。方法用不同浓度的3种非对映异构体(±)α-HBCD,(±)β-HBCD,(±)γ-HBCD处理N2a细胞,Cell counting kit-8(CCK-8)法检测HBCD对N2a的细胞毒性作用;流式细胞术检测HBCD对N2a细胞周期的影响;实时荧光定量(RT-PCR)和免疫蛋白印迹(Western blot,WB)法分别用于检测3个细胞核受体RXRα、PPARγ、PXR和下游靶基因细胞色素P450亚酶CYP3A11的mRNA及蛋白表达水平的变化;免疫共沉淀技术分析RXRα、PXR、PPARγ受体间的相互作用。结果β-HBCD对N2a的细胞毒性明显大于α-HBCD,γ-HBCD没有明显的细胞毒性。α-HBCD、β-HBCD对N2a细胞增殖的抑制作用呈时间-剂量效应关系(P<0.05),其半数抑制浓度(IC_(50))分别为60.07和10.52μmol/L,γ-HBCD的细胞毒性较小,镜下可见黑色絮状物,CCK-8法未能测定出其IC_(50);α-、β-HBCD会使细胞周期阻滞在G2/M期;染毒24 h后,RXRα、PPARγ、PXR及CYP3A11的mRNA及蛋白表达水平均呈现上升趋势(P<0.05);在N2a细胞内,α-HBCD染毒前后,RXRα与PPARγ、PXR之间始终存在交互作用关系。结论α-HBCD、β-HBCD对N2a细胞具有增殖抑制作用,细胞周期主要阻滞在G2/M期。α-HBCD、β-HBCD均可诱导3种细胞核受体RXRα、PPARγ和PXR的表达升高,PXR受体下游表达基因CYP3A11的表达也明显升高(P<0.05)。RXRα与PPARγ、PXR三个细胞核受体之间始终存在交互作用,但是受体间相互作用的分子机制有待深入研究。Objective To investigate the effect of hexabromocyclododecanes（ HBCDs） on cell proliferation and the expression of the three important cell nuclear receptor of retinoic X receptor α（ RXRα）,peroxisome proliferator-activated receptor-γ（ PPARγ）,pregnane X receptor（ PXR） and their interaction in Neuro-2a（N2a）.Methods Neuro-2a cells were treated with different concentrations of diastereoisomers,of HBCDs which were α-HBCD,β-HBCD,γ-HBCD,respectively,and cell toxicity was analyzed using the cell counting kit-8（ CCK-8） assay. The impact of HBCDs on cell cycles of Neuro-2a were analyzed by flow cytometry analysis,and the expression levels in mRNA and protein for the three nuclearreceptors（ RXRα,PPARγ,PXR andits target genes CYP3A11） were determined by RT-PCR and Western blot, respectively. The interaction between the receptors of RXRα, PXR, PPARγ was explored by immunoprecipitation. Results Cytotoxicity of β-HBCD was the greatest among the three diastereoisomers,it was significantly greater than α-HBCD,however cytotoxicity of γ-HBCD for the Neuro-2a cells couldn ＇t be determined. Moreover α-HBCD,β-HBCD induced significant cytotoxicity in a time-dose-response relationship to Neuro-2a cells（ P〈0. 05）,IC（50） of α-HBCD,β-HBCD to Neuro-2a cells were 60. 07 and 10. 52 μmol/L,respectively. α-,β-HBCD blocked the cell cycle at G2/M phase. The expression levels in mRNA and protein of RXRα,PPARγ,PXR,CYP3A11 were significantly increased after cells exposure to α-HBCD and β-HBCD 24 h. An interaction between RXRα,PPARγ and PXR in Neuro-2a cells existed no matter before and after exposure to HBCD. Conclusion α-HBCD,β-HBCD inhibit proliferation of Neuro-2a cells,cell cycle mainly was arrested at G2/M phase. α-HBCD,β-HBCD could up-regulated the expression levels of RXRα,PPARγ,PXR. Meanwhile,the expression of CYP3A11 which is downstream gene of PXR also significantly increased（ P〈0. 05）. Interaction between RXRα,PPARγ and PXR exist whether or not exposure to α

关 键 词：六溴环十二烷 视黄醛类X受体α 过氧化物酶体增殖物活化受体Γ 孕烷X受体 细胞周期 

分 类 号：R730.264[医药卫生—肿瘤] R994.6[医药卫生—临床医学]