二苯乙烯苷经GSK-3β途径对干预tau蛋白磷酸化机制研究  

Effect of TSG on tau phosphorylation via GSK-3βpathway

作　　者：蒙婉莹 刘超宇 夏小燕 李彦炳 李振中 朱晓莹[4] 廖艳花 黄忠仕 MENG Wan-ying;LIU Chao-yu;XIA Xiao-yan;LI Yan-bing;LI Zhen-zhong;ZHU Xiao-ying;LIAO Yan-hua;HUANG Zhong-shi(College of Pharmacy,Guangxi University of Chinese Medicine,Nanning 530200,China;College of Basic Medicine,Youjiang Medical University for Nationalities,Baise 533000,China;College of Pharmacy,Youjiang Medical University for Nationalities,Baise 533000,China;College of Clinical Medicine,Youjiang Medical University for Nationalities,Baise 533000,China)

机构地区：[1]广西中医药大学药学院,广西南宁530200 [2]右江民族医学院基础医学院,广西百色533000 [3]右江民族医学院药学院,广西百色533000 [4]右江民族医学院临床医学院,广西百色533000

出　　处：《海南医学院学报》2022年第14期1059-1067,共9页Journal of Hainan Medical University

基　　金：广西自然科学基金资助项目(2018GXNSFAA294153)。

摘　　要：目的:观察二苯乙烯苷(TSG)如何经GSK-3β途径干预Aβ_(25-35)致拟痴呆大鼠模型的tau蛋白磷酸化过程。方法:选用24月龄雄性SD大鼠36只,随机分为正常组、假手术组、模型组及TSG低、中、高剂量组(TSG分别为0.033、0.1、0.3 g/kg),每组各6只。模型组、TSG各剂量组按大鼠脑立体定位图谱选择海马区域注射Aβ_(25-35)溶液致痴呆模型(假手术组注射等体积生理盐水)。经Aβ_(25-35)海马区造模筛选后开始灌胃给药,每组每日灌胃给药1次,连续4周。各组灌胃4周相应药物后,采用HE染色观察大鼠海马和皮层区神经细胞的结构;IHC检测各组大鼠脑组织PKC、PKA蛋白的表达;实时荧光定量PCR检测GSK‑3β、PKA、PI3K、PKB、PKC mRNA表达情况;蛋白免疫印迹法(Westernblot)检测GSK-3β、PI3K、PKB、p-tau蛋白表达情况。结果:与正常组比较,模型组大鼠神经细胞的数量较少,排列较稀疏无序;GSK‑3β、PKA mRNA表达水平呈上调趋势(P<0.05);PI3K、PKB、PKC mRNA表达水平明显降低(P<0.05);PI3K蛋白表达水平降低(P<0.05)及PKB蛋白表达水平降低(P<0.01);PKC蛋白在海马及皮层的表达水平均显著降低(P<0.01),PKA蛋白在海马及皮层的表达水平均呈上调趋势(P<0.05);p-tau蛋白相对表达量升高(P<0.05)。与模型组比较,各给药组大鼠神经细胞数量有一定回升;GSK‑3β、PKA mRNA表达水平有下降的趋势(P<0.05);PI3K、PKB、PKC蛋白及mRNA表达水平明显上调(P<0.05),p-tau相对表达量呈下降趋势(P<0.05)。结论:TSG对Aβ_(25-35)致拟痴呆大鼠模型海马组织内的GSK-3β、PI3K、PKC、PKB、PKA具有调控作用,通过调节这些因子的表达,抑制tau蛋白过度磷酸化,改善tau蛋白过度磷酸化的现象。Objective:To observe how TSG interferes with tau phosphorylation in Aβ25-35 induced dementia model via GSK-3βpathway.Methods:Thirty-six male SD rats aged 24 months were randomly divided into control group,sham group,model group,low-dose,medium-dose and high-dose of TSG groups(TSG,0.033,0.1,0.3g·kg-1),with 6 rats in each group.Model group and TSG dose groups were injected Aβ25-35 into hippocampus by brain stereoscopic locator to induce dementia model(sham group was injected with equal volume of normal saline).Six SD rats of the same age were selected as normal group.Fourteen days after modeling by Aβ25-35,the rats was given TSG by gavage once a day in each group for 4 weeks.After 4 weeks of treatment with TSG,HE staining was used to observe the structural changes of nerve cells in brain tissue,IHC to observe the expres-sion of PKC and PKA protein in brain tissue,real-time PCR to observe the mRNA expressions of GSK‑3β,PKA,PI3K,PKB and PKC,and Western blot was used to observe the expression of GSK-3β,p-tau,PI3K and PKB protein in hippocampus.Results:Compared with the normal group,the number of nerve cells in the model group was less,and the arrangement was sparse and disordered.The expression of PI3K,PKB mRNA were significantly decreased(P<0.01),the expression of GSK‑3β,PKA mRNA increased(P<0.05),the expression of PKC mRNA was decreased(P<0.05),the protein expression level of PKC(P<0.01),PI3K(P<0.05)and PKB(P<0.01)were decreased,the expression levels of PKC protein in hippocampus and cortex were significantly decreased(P<0.01),the expression levels of PKA protein in hippocampus and cortex were increased(P<0.05).Compared with model group,the number of nerve cells in each administration group increased to a certain extent,the expression levels of P-tau,GSK-3βand PKA protein and mRNA were significantly decrease(P<0.05),the expression levels of PI3K,PKB and PKC protein and mRNA were significantly up-regulated(P<0.05).Conclusion:TSG can regulate GSK-3β,PI3K,PKC,PKB and PKA in the hippocampus of Aβ25-35 ind

关 键 词：二苯乙烯苷 阿尔茨海默症 糖原合成酶激酶-3 磷脂酰肌醇激酶 蛋白激酶C 蛋白激酶B 蛋白激酶A 

分 类 号：R965[医药卫生—药理学]