肉苁蓉多糖对Aβ_(25-35)所致PC12细胞损伤BDNF基因表观遗传的调控作用  

CDPS reduces cell apoptosis induced by Aβ_(25-35) through brain-derived neurotrophic factor regulated by epigenetic modulation

作　　者：武燕 刘丹丹 马子兴 苗鑫 张晓菲 李刚 WU Yan;LIU Dan-dan;MA Zi-xing;MIAO Xin;ZHANG Xiao-fei;LI Gang(Inner Mongolia Medical University, Department of Pharmacy, HOHHOT 010110, China;State Key Laboratory of Bioactive Substance and Function of Natural Medicines, Institute of Materia Medica, Chinese Academy of Medical Sciences and Peking Union Medical College, Beijing 100050, China)

机构地区：[1]内蒙古医科大学药学院,呼和浩特010110 [2]中国医学科学院/北京协和医学院药物研究所天然药物活性物质与功能国家重点实验室,北京100050

出　　处：《中国新药杂志》2018年第12期1393-1400,共8页Chinese Journal of New Drugs

基　　金：国家自然科学基金资助项目(81260650);国家自然科学基金资助项目(81560685);天然药物活性物质与功能国家重点实验室开放课题项目(GTZK201710)

摘　　要：目的:探讨肉苁蓉多糖(CDPS)对Aβ_(25-35)诱导的大鼠肾上腺嗜络瘤细胞(PC12)损伤BDNF基因表观遗传的调控作用,分析组蛋白乙酰化对BDNF基因转录的影响。方法:采用Aβ_(25-35)诱导PC12细胞损伤建立阿尔茨海默病(AD)体外模型,在给予阳性药人参皂苷Rd(GSrd)、CDPS高、低浓度后,流式细胞术检测细胞凋亡情况;Hoechst33258染色观察细胞形态的变化。ELISA法检测组蛋白乙酰化酶(HATs)、组蛋白去乙酰化酶2(HDAC2)和BDNF的含量变化。Western Blotting检测P300和HDAC2的表达水平变化。染色质免疫共沉淀(CHIP)检测BDNF exonⅣ组蛋白H3乙酰化水平的变化和HDAC2含量变化。结果:20μmol·L^(-1)Aβ_(25-35)暴露PC12细胞48 h后成功建立AD细胞凋亡模型。GSrd组、CDPS高、低浓度作用48 h后,HATs表达增加(P<0.05),HDAC2表达下降(P<0.05),BDNF水平提高(P<0.05),并且有剂量依赖性。Ch IP分析证实Aβ_(25-35)诱导PC12细胞后BDNF基因exonⅣ组蛋白H3乙酰化水平明显下降,且降低呈HDAC2依赖性;给药48 h后,BDNF基因exonⅣ组蛋白H3乙酰化水平显著升高,HDAC2水平显著降低。结论:CDPS可能通过重建Ac-H3和HDAC2之间的平衡,抑制HDAC2的表达来活化BDNF基因转录从而起到保护PC12细胞、刺激PC12细胞再生的作用。Objective： To investigate the regulatory effect of Cistanche polysaccharides（ CDPS） on the epigenetic changes of BDNF gene induced by Aβ25-35 in rat adrenal sinophil tumor cells（ PC12）,and to analyze the effect of histone acetylation on BDNF gene transcription. Methods： The apoptosis of PC12 cells was induced by Aβ25-35 in vitro. Alzheimer’s disease（ AD） model was established in vitro. The apoptotic cells were detected by flow cytometry after the positive drug ginsenoside Rd（ GSrd） and CDPS were given. The changes of cell morphology were observed by Hoechst33258 staining. The levels of HATs,HDAC2 and BDNF were detected by ELISA method.The expression level of P300 and HDAC2 protein was detected by Western Blotting. The alterations of histone H3 acetylation and HDAC2 in BDNF exon Ⅳ were observed and analyzed by chromatin immunoprecipitation（ CHIP）.Results： After exposure of the PC12 cells 48 h by 20 μmol·L-1 Aβ25-35,the model of AD cell apoptosis was established successfully. Compared with model group,each group HATs expression increases（ P 〈 0. 05）,HDAC2 decrease（ P 〈 0. 05）,and BDNF levels increase（ P 〈 0. 05）,and there are dose dependent. Ch IP analysis confirmed that after Aβ25-35 induced P12 cells,BDNF exon Ⅳ histone H3 acetylation protein levels decreased significantly,and the reduction of histone acetylation is HDAC2-dependent. 48 h after CDPS administration,BDNF exon Ⅳ histone H3 acetylation protein levels increased significantly,and the level of HDAC2 decreased. Conclusion：CDPS had significant protective effects on PC12 cells exposed to Aβ25-35. The mechanism may be that the decrease of acetylation of HDAC2-dependent histone H3 induced the suppression of continuous transcription of BDNF gene.

关 键 词：肉苁蓉多糖 PC12细胞 组蛋白乙酰化酶2 组蛋白H3 

分 类 号：R742.5[医药卫生—神经病学与精神病学] R964[医药卫生—临床医学]