D-AP5对氟砷联合染毒原代培养海马神经元突触相关蛋白表达的影响  

Effect of D-AP5 on expression of synapse-associated proteins in hippocampal neurons treated by fluoride and arsenic co-exposure

作　　者：刘晓静[1] 杨洪蕊 刘颖[1] 曾洋[1] 张文丽 曹福源[3] 薛玲[1] 蒋守芳[1] LIU Xiao-jing YANG Hong-rui LIU Ying ZENG Yang ZHANG Wen-li CAO Fu-yuan XUE Ling JIANG Shou-fang(Hebei Province Coal Mine Health and Safety Laboratory, School of Public Health Medical Research Center Laboratory Animal Center, North China University of Science and Technology, Tangshan, Hebei 063210, China）.)

机构地区：[1]华北理工大学公共卫生学院河北省煤矿卫生与安全实验室,河北唐山063210 [2]华北理工大学医学实验研究中心,河北唐山063210 [3]华北理工大学实验动物中心,河北唐山063210

出　　处：《环境与职业医学》2017年第6期531-535,共5页Journal of Environmental and Occupational Medicine

基　　金：国家自然科学基金资助项目(编号:81102083);河北省自然科学基金(编号:H2015209088)

摘　　要：[目的]探讨谷氨酸NMDA受体拮抗剂D-2-氨基-5-磷酰基戊酸(D-AP5)对氟砷联合染毒后原代海马神经元突触相关蛋白表达的影响。[方法]将体外培养的海马神经元分为4组,空白对照组、氟砷联合染毒组(0.4μg/m L氟化钠+0.1μg/m L亚砷酸钠)、D-AP5组(25μmol/L D-AP5)和D-AP5+氟砷联合染毒组(0.4μg/m L氟化钠+0.1μg/m L亚砷酸钠+25μmol/L D-AP5)。各组海马神经元置于37℃、含有体积分数5%CO_2培养箱中培养24 h。把培养好的各组海马神经元用CCK8检测活性,用蛋白免疫印迹法检测海马神经元中NR2B、SYN及GAP43突触相关蛋白的表达水平。[结果]与空白对照组[(100.00±0.00)%]比较,氟砷联合染毒组[(75.81±6.27)%]、D-AP5+氟砷联合染毒组[(81.58±5.20)%]的细胞存活率均明显降低(P<0.05);与氟砷联合染毒组[(75.81±6.27)%]比较,D-AP5组[(98.78±3.53)%]、D-AP5+氟砷联合染毒组[(81.58±5.20)%]海马神经元存活率均明显升高(P<0.05)。氟砷联合染毒组(0.425±0.06)、D-AP5组(0.469±0.03)、D-AP5+氟砷联合染毒组(0.354±0.04)的NR2B相对表达量较空白对照组(0.572±0.05)均有所下降(P<0.05),D-AP5+氟砷联合染毒组(0.354±0.04)受体蛋白NR2B表达较氟砷联合染毒组(0.425±0.06)降低(P<0.05);氟砷联合染毒组与D-AP5+氟砷联合染毒组的SYN和GAP43相对表达量均较空白对照组低(P<0.05)。而D-AP5组与D-AP5+氟砷联合染毒组的SYN和GAP43表达较氟砷联合染毒组均升高(P<0.05)。[结论]氟砷联合染毒能够下调突触相关蛋白SYN及GAP-43表达水平,谷氨酸NMDA受体拮抗剂D-AP5可以抑制氟砷联合染毒对海马神经元突触相关蛋白表达的影响。[ Objective ] To test the effect of glutamate NMDA receptor antagonist D-2-amino-5-phosphonopentanoate （D-AP5） on the expression of synapse-associated proteins in primary cultured hippocampal neurons after fluoride and arsenic co-exposure. [ Methods ] In vitro cultured hippocampal neurons were divided into four groups, including control group, fluoride-arsenic co- exposure group （0.4 μg/mL sodium fluoride ＋0.1 μg/mL sodium arsenite）, D-AP5 group （25 μnol/L D-AP5）, and D-AP5＋ fluoride- arsenic co-exposure group （0.4 μg/mL sodium fluoride ＋0.1 I.tg/mL sodium arsenite＋25 p.mol/L D-AP5）. Hippocampal neurons in each group were cultured in 5%CO2 incubator at 37℃ for 24h. Cell viability was detected by CCK8, and the expression levels of GAP43, NR2B, and SYN proteins in hippocampal neurons were detected by Western blot.[ Results ] Compared with the control group [（100.00 ± 0.00）%], the cell viability rates of hippocampal neurons in the fluoride- arsenic co-exposure group [（75.81 ± 6.27）%] and the D-AP5＋ fluoride-arsenic co-exposure group [（81.58± 5.20）%] were significantly decreased （P〈0.05）. Compared with the fluoride-arsenic co-exposure group [（75.81 ± 6.27）%], the cell viability rates of hippocampal neurons in the D-AP5 group [（98.78 ± 3.53）%] and the D-AP5＋ fluoride-arsenic co-exposure group [（81.58 ± 5.20）%] were significantly increased （P 〈 0.05）. Compared with the control group （0.572 ± 0.05）, the protein expression levels of NR2B in the fluoride-arsenic co-exposure group （0.425 ± 0.06）, D-AP5 group （0.469 ± 0.03）, and D-AP5＋ fluoride-arsenic co-exposure group （0.354 ± 0.04） were significantly decreased （P 〈 0.05）. The protein expression level of NR2B in the D-AP5＋ fluoride-arsenic co-exposure group （0.354 ± 0.04） were significantly lower than that of the fluoride-arsenic co-exposure group （0.425 ± 0.06）. The protein expression levels of SYN and GAP43 in the fluoride-arsenic co-exposure group and the

关 键 词：氟 砷 D-2-氨基-5-磷酰基戊酸 海马神经元 突触素 突触相关蛋白43 

分 类 号：R994.6[医药卫生—毒理学]