抑肝散提取物对环境内分泌干扰物神经毒性的抑制作用  

The inhibitive effect of Yigansan extract on neurotoxicity of environmental endocrine disruptors

作　　者：穆轶[1] 朱彧 MU Yi;ZHU Yu(Department of Pharmacy,The Second Hospital,Tianjin Medical University,Tianjin 300211,China;Department of Clinical Laboratory,Tianjin Haihe Hospital,Tianjin 300350,China)

机构地区：[1]天津医科大学第二医院药学部,天津300211 [2]天津市海河医院检验科,天津300350

出　　处：《天津医科大学学报》2022年第2期151-155,共5页Journal of Tianjin Medical University

基　　金：天津市中医药管理局中医中西医结合科研课题(2019125)。

摘　　要：目的:考察抑肝散提取物抑制环境内分泌干扰物(EEDs)双酚A(BPA)和邻苯二甲酸(2-乙基己基)酯(DEHP)神经毒性的作用并分析其机制。方法:小鼠神经母细胞瘤细胞Neuro-2a细胞经BPA(0.1 mmol/L)+DEHP(0.1 mmol/L)与3 mg/mL(低剂量组)和6 mg/mL(高剂量组)抑肝散乙醇/氯仿提取物共培养72 h,观察细胞凋亡和氧化损伤(活性氧簇,ROS)的变化。按照暴露剂量为25μg/(kg·d)BPA和50μg/(kg·d)DEHP给予初断乳雄性Sprague-Dawley(SD)大鼠BPA连续灌胃暴露90 d,同时将抑肝散乙醇/氯仿提取物按300 mg/kg(低剂量组)和600 mg/kg(高剂量组)给大鼠灌胃给药,通过Y迷宫实验观察大鼠短期记忆能力(自发交替正确率)的变化。取大鼠海马组织后,检测海马组织总抗氧化能力(T-AOC)、丙二醛(MDA)、谷胱甘肽(GSH)、游离脂肪酸(FFAs)葡萄糖含量和线粒体膜电位(MMP)的变化,分析星形胶质细胞胶质纤维状酸性蛋白(GFAP)和巢蛋白(nestin)、caspase-3以及哺乳动物雷帕霉素靶蛋白(mTOR)信号通路活性的变化。结果:Neuro-2a细胞经BPA+DEHP暴露后,24、48、72 h细胞增殖均受到抑制(t=3.951、8.823、7.060,均P<0.05),早期和晚期凋亡率均上调(t=14.072、6.590,均P<0.05),抑肝散提取物可促进细胞增殖(与暴露组相比,低剂量组t=2.963、5.814、2.101,均P<0.05,高剂量组t=3.796、10.370、5.311,均P<0.05),抑制早期和晚期细胞凋亡(与暴露组相比,低剂量组t=0.770、1.475,均P<0.05,高剂量组t=6.187、4.368,均P<0.05)。大鼠经BPA+DEHP暴露后,短期记忆能力显著下降(t=2.123,P<0.05),其海马组织氧化应激、GFAP表达和caspase-3活性上调(t=3.634、5.043、8.914,均P<0.05),nestin表达、线粒体膜电位、MDA、GSH、T-AOC和mTOR磷酸化水平下调(t=8.398、9.979、3.754、9.621、7.193、25.982,均P<0.05),抑肝散提取物可缓解BPA+DEHP暴露后大鼠短期记忆能力损伤(与暴露组相比,低剂量组t=1.339,P>0.05,高剂量组t=2.293,P<0.05)以及海马组织的上述表型改变(�Objective:To investigate the neurotoxicity of Yigansan extract in the treatment of environmental endocrine disruptors bisphenol A(BPA)and di-(2-ethylhcxyl)phthalate(DEHP)and analyze its mechanism.Methods:Mouse neuroblastoma cell line Neuro-2a cells were co-cultured with BPA(0.1 mmol/L)+DEHP(0.1 mmol/L)and 3 mg/mL(low dose group)and 6 mg/mL(high dose group)Yigansan ethanol/chloroform extract for 72 h,and apoptosis and oxidative damage(ROS)were measured.According to the exposure dose of 25μg/(kg·d)BPA and 50μg/(kg·d)DEHP,the first weaned male Sprange-Dawley(SD)rats were given BPA for continuous gastric exposure for 90 days.At the same time,the ethanol/chloroform extract of Yigansan was administered at 300 mg/kg(low-dose group)and 600 mg/kg(high-dose group)were intragastrically administered to rats,and the changes in short-term memory ability(spontaneous alternation accuracy)of rats were observed by Y maze experiment.After taking the rat hippocampus tissue,the changes of total antioxidant capacity(T-AOC),malondialdehyde(MDA),glutathione(Glutathione,GSH),free fatty acid(FFAs)glucose content and mitochondrial membrane potential(MMP)of hippocampus were measured.The astrocyte glial fibrillary acidic protein(GFAP)and nestin,caspase-3 and mammalian rapamycin target protein mTOR signal pathway activity changes were analyzed.Results:After Neuro-2a cells are exposed to BPA+DEHP,cell proliferation in 24,48 and 72 h was inhibited(t=3.951,8.823,7.060,all P<0.05),and the early and late apoptosis rate was increased(t=14.072,6.590,all P<0.05).Yigansan extract could promote cell proliferation(compared with exposure group,t=2.963,5.814,2.101,all P<0.05 in low-dose group,t=3.796,10.370,5.311,all P<0.05 in high-dose group)and inhibit cell early and late apoptosis(compared with exposure group,t=0.770 and 1.475,all P>0.05 in low-dose group,t=6.187 and 4.368,all P<group).After the rats were exposed to BPA+DEHP,the short-term memory ability was significantly decreased(t=2.123,P<0.05),the hippocampal oxidative stress,GFAP expression an

关 键 词：抑肝散 环境内分泌干扰物 神经毒性 

分 类 号：R9[医药卫生—药学]