葛根素对氧糖剥夺细胞模型CaM、CaMKⅡ、MECP2、BDNF及Akt表达的影响  

Effects of puerarin on the expressions of CaM,CaMKⅡ,MECP2,BDNF and Akt in oxygen and glucose deprivation cell models

作　　者：高震[1] 陈婉莹 张萌 陈梦燚 王虎清[1] 吴海琴[1] GAO Zhen;CHEN Wan-ying;ZHANG Meng;CHEN Meng-yi;WANG Hu-qing;WU Hai-qin(Department of Neurology,The Second Affiliated Hospital of Xi'an Jiaotong University,Xi'an 710004;Department of Neurology,Taikang Tongji (Wuhan) Hospital,Wuhan 430000,China)

机构地区：[1]西安交通大学第二附属医院神经内科,陕西西安710004 [2]泰康同济(武汉)医院神经内科,湖北武汉430000

出　　处：《西安交通大学学报（医学版）》2019年第1期153-157,共5页Journal of Xi’an Jiaotong University（Medical Sciences）

基　　金：国家自然科学基金面上资助项目(No.81270415);陕西省重点研发计划项目(No.2017SF-181)~~

摘　　要：目的研究葛根素对氧糖剥夺(oxygen and glucose deprivation,OGD)血管性痴呆细胞模型细胞黏附分子(CaM)、钙调蛋白激酶Ⅱ(CaMKⅡ)、脑源性神经营养因子(BDNF)及Akt表达的影响。方法选取生长良好的PC12细胞传代、分化,行OGD准备血管性痴呆细胞模型,随机分为对照组、模型组及低、中、高剂量葛根素组。MTT法测定细胞存活率并确定合适的葛根素干预浓度及OGD处理时间;检测乳酸脱氢酶(LDH)释放量评定细胞损伤程度,鉴定细胞模型;Western blot检测CaM、CaMKⅡ、MECP2、BDNF及Akt蛋白的表达水平。结果 PC12细胞存活率随OGD时间延长而逐渐降低,呈时间依赖性;PC12细胞存活率随葛根素浓度增加而逐渐升高,呈浓度依赖性。葛根素有效干预浓度为0.1～10μmol/L;OGD最佳处理时间为6h。与对照组相比,模型组LDH释放量明显增高(P<0.05);葛根素干预组LDH释放量随葛根素浓度增加而减少(P<0.05)。模型组CaM蛋白表达明显升高,BDNF表达量明显减少(P<0.05),MECP2表达及CaMKⅡ、Akt蛋白磷酸化水平均未见明显变化(P>0.05)。葛根素干预可下调CaM蛋白水平,提高MECP2、BDNF的表达及CaMKⅡ磷酸化水平,中、高剂量葛根素组亦能升高Akt蛋白磷酸化水平(P<0.05)。结论葛根素可能通过提高Ca2+-CaM复合物介导CaMKⅡ自身磷酸化水平,诱导MECP2磷酸化,上调BDNF的表达,激活下游PI3K-Akt通路,抑制凋亡基因及蛋白表达,发挥神经保护作用。Objective To observe the effects of puerarin on the expressions of CaM,CaMKⅡ,BDNF and Akt in vascular dementia cell models induced by oxygen and glucose deprivation(OGD).Methods The passaged well-differentiated PC12 cells were randomly divided into control group,model group,and low-dose,medium-dose and high-dose intervention groups.Vascular dementia cell model was established by OGD.Suitable OGD time and concentration of puererin were obtained from the cell viability measured by MTT assay.The release of LDH was measured to assess the extent of cell damage and identify cell models.The expressions of CaM,CaMKⅡ,MECP2,BDNF and Akt were detected by Western blot.Results PC12 cells with OGD prolonged viability decreased in a time-dependent manner,with increased concentrations of puerarin increased in a concentration-dependent manner.Effective intervention of puerarin was 0.1-10μmol/L and optimal time of OGD was 6 h.Compared with control group,the release of LDH in model group was significantly increased(P<0.05),while the release in puerarin group was decreased with the increase of puerarin concentration(P<0.05).In model group,the expression of CaM was significantly increased and that of BDNF was significantly decreased(P<0.05).However,the expression of MECP2 and the phosphorylation of CaMKⅡand Akt did not differ(P>0.05).Puerarin could down-regulate the level of CaM protein,increase the expressions of MECP2 and BDNF and the phosphorylation of CaMKⅡ,and also increase the phosphorylation of Akt in addition to the low-dose group(P<0.05).Conclusion The neuroprotective effect of puerarin may be related to the increase of the autophosphorylation of CaMKⅡmediated by Ca^2+-CaM complex,induce the phosphorylation of MECP2,up-regulate the expression of BDNF and activate the PI3K-Akt pathway to inhibit the expression of apoptotic genes and proteins.

关 键 词：葛根素 氧糖剥夺 PC12细胞 CAM CaMKⅡ MECP2 BDNF Akt 

分 类 号：R543.[医药卫生—心血管疾病]