六溴环十二烷(HBCD)对H4细胞和SK-N-AS细胞脱碘酶2和3表达的影响

六溴环十二烷(hexabromocyclododecane,HBCD)与多溴联苯醚(polybrominated diethyl ethers,PBDEs)复合污染体系,对人类健康尤其神经系统所造成的潜在危害及其机制一直是笔者课题组的研究方向。HBCD是广泛使用的溴化阻燃剂,与PBDEs一样,会通过干扰内分泌系统、影响甲状腺激素分泌及损伤神经系统,对生物体产生发育神经毒性。作为系列研究之一,本研究以H4人脑神经胶质瘤细胞和SK-N-AS人神经母细胞瘤细胞为体外生物模型,通过观察HBCD对H4细胞Ⅱ型脱碘酶(Dio2)和SK-N-AS细胞Ⅲ型脱碘酶(Dio3)表达的调控,初步探讨了HBCD对神经系统局部甲状腺激素水平的潜在影响。H4细胞和SK-N-AS细胞分别暴露于0、1、3和9μmol·L-1HBCD 24 h后,采用MTT法检测细胞活力,Western Blot和RT-PCR法分别分析Dio2和Dio3蛋白和基因的表达,酶联免疫吸附测定(ELISA)法检测H4细胞脑源性神经细胞营养因子(BDNF)的分泌。结果表明,HBCD以剂量依赖方式降低H4细胞和SK-N-AS细胞生存率,引起H4细胞Dio2蛋白和基因表达下调,而致SK-N-AS细胞Dio3蛋白和基因表达上调。此外,HBCD还降低H4细胞BDNF的分泌。这表明,HBCD很可能通过影响神经和胶质细胞脱碘酶的表达,影响脑局部甲状腺激素水平,从而引起神经系统损伤及发育神经毒性。

Effect of Hexabromocyclododecane (HBCD) on the Expressions of Deiodinases 2 and 3 in the Human Neuroglioma H4 Cells and Neuroblastoma SK-N-AS Cells

Being one of the chemicals in the complex of persistent organic pollutants, hexabromocyclododecane (HBCD), as a widely used brominated flame retardant, has been known to cause disturbance in the endocrine system, in particular, the thyroid hormone system, which may underlie its neurotoxicity and developmental neurotoxicity. This current study used the human H4 neuroglioma cells and human SK-N-AS neuroblastoma cells as the in vitro models, to investigate the effects of HBCD on the type 2 iodothyroninedeiodinase (Dio2) expression in the H4 cells and the type 3 iodothyroninedeiodinase (Dio3) expression in the SK-N-AS cells. Briefly, the H4 and SK-N-AS cells were separately exposed to HBCD for 24 h, and the cell viability was examined by the methyl-thiazolyl-tetrazolium (MTT) method; the levels of Dio2 and Dio3 protein and mRNA expression were quantitated by the Western Blot and the qRT-PCR, respectively; in addition, the secretion of brain-derived neurotrophic factor (BDNF) was examined by the ELISA in the H4 cells. The data showed that HBCD led to a loss of cell viability in both cell types in a dose-dependent manner; HBCD caused the reduction of both Dio2 protein and mRNA expressions in the H4 cells, and the elevation in the Dio3 protein and mRNA expressions in the SK-N-AS cells; moreover, HBCD also reduced the secretion of BDNF in the H4 cells as detected in the culture medium. Our data suggest that HBCD is likely to modulate the local level of thyroid hormones (in particular T3) in the brain by affecting the expression of deiodinated enzymes in the neurons and the glia, thereby resulting in the HBCD-mediated neurotoxicity and developmental neurotoxicity.