梓醇-川芎嗪方调控STAT3改善阿尔茨海默病神经可塑性的机制研究  

Effect of Catalpol-Tetramethylpyrazine Prescription on Neuroplasticity in Alzheimer′s Disease

作　　者：陈慧泽 邓楚珺 孟泽宇 孟胜喜 CHEN Huize;DENG Chujun;MENG Zeyu;MENG Shengxi(Shanghai Sixth People′s Hospital Affiliated to Shanghai Jiao Tong University School of Medicine,Shanghai 200233,China;The Second Clinical Medical College,Heilongjiang University of Chinese Medicine,Harbin 150040,Heilongjiang,China)

机构地区：[1]上海交通大学医学院附属上海第六人民医院,上海200233 [2]黑龙江中医药大学第二临床医学院,哈尔滨150040

出　　处：《中西医结合心脑血管病杂志》2024年第21期3899-3906,共8页Chinese Journal of Integrative Medicine on Cardio-Cerebrovascular Disease

基　　金：上海市第六人民医院“脑科学与类脑研究”脑项目功能和针灸脑图谱光声成像技术应用研究(No.ynnkxyb202415);上海市第六人民医院医疗服务能级提升工程临床医疗技术骨干队伍培育项目(No.20220213);上海市第六人民医院院级课题(No.ynxg202218);上海交通大学医学院“大学生创新训练计划”项目(No.1723Z002、1824011);上海市进一步加快中医药传承创新发展三年行动计划项目(2021年—2023年)[No.ZY(2021-2023)-0205-04];华东片区及市级中医专科专病联盟建设项目[No.ZY(2021-2023)-0302]。

摘　　要：目的:探讨梓醇-川芎嗪(CT)方对阿尔茨海默病(AD)的作用及转录激活蛋白3(STAT3)的影响。方法:将野生型C57BL/6J小鼠作为对照组(Control),同时将粉样前体蛋白/早老素1双转染小鼠按照随机数字表法随机分为模型组(Model)、低剂量梓醇-川芎嗪方组(CT-L)、高剂量梓醇-川芎嗪方组(CT-H),每组6只。CT-L组和CT-H组分别给予梓醇-川芎嗪方药50、100 mg/kg灌胃处理。Control组和Model组均给予等体积的生理盐水灌胃。每日1次,连续灌胃8周。水迷宫实验探索梓醇-川芎嗪对AD小鼠学习记忆能力的影响,免疫组化检测海马组织中生长相关蛋白43(GAP-43)的表达。海马神经元HT-22细胞分为空白对照组(Control组)、模型组(Model组)、低剂量梓醇-川芎嗪方组(CT-L组)、高剂量梓醇-川芎嗪方组(CT-H组)、STAT3敲除+梓醇-川芎嗪方组(CT+siSTAT3组)。通过Aβ_(1-42)构建体外AD模型,通过转染siSTAT3构建STAT3沉默的神经元模型,并加入高、低剂量的梓醇-川芎嗪方培养24 h,细胞活性检测(CCK-8)检测细胞活力,蛋白免疫印迹法(Western Blot)检测Cyclin D1、Ki-67、B细胞淋巴瘤/白血病-2(Bcl-2)、Bcl-2相关X蛋白(Bax)、NF200、GAP-43、STAT3和磷酸化的STAT3(p-STAT3)蛋白表达水平,Brdu检测细胞增殖,流式细胞术和末端脱氧核苷酸转移酶介导的dUTP缺口末端标记测定法(TUNEL)染色检测细胞凋亡。结果:梓醇-川芎嗪方可以明显缩短APP/PS1双转染小鼠的逃避潜伏期,提高目标象限停留时间和目标象限停留次数,上调海马组织GAP-43的表达,其中高剂量的梓醇-川芎嗪方作用更显著(P<0.05或P<0.01)。梓醇-川芎嗪方可以恢复AD模型细胞的活力和增殖能力,上调Cyclin D1、Ki-67、NF200、GAP-43和STAT3蛋白的表达,下调凋亡相关蛋白Bax的表达并恢复了抗凋亡Bcl-2蛋白的水平,其中高剂量梓醇-川芎嗪方的作用更显著(P<0.05或P<0.01)。而沉默STAT3逆转了梓醇-川芎嗪方对AD神经元增殖和对NFObjective:To investigate the effect of catalpol-tetramethylpyrazine(CT)prescription on Alzheimer′s disease(AD).Methods:Wild-type C57BL/6J mice were used as Control group,and the mice double-transfused with powder precursor protein/presenilin 1 were randomly divided into Model group,low-dose CT prescription(CT-L)group and high-dose CT prescription(CT-H)group according to random number table method,with 6 mice in each group.The CT-L group and the CT-H group were given 50 and 100 mg/kg intragastric treatment of catalol-ligustrazine,respectively.Both Control group and Model group were given equal volume of normal saline intragastric administration.Once a day for 8 weeks.The effect of tetramethylpyrazine on the learning and memory ability of AD mice were explored by water maze experiment,and the expression of growth-related protein 43(GAP-43)in hippocampus was detected by immunohistochemistry.HT-22 cells of hippocampal neurons were divided into Control group,Model group,low-dose CT treatment(CT-L)group,high-dose CT treatment(CT-H)group,STAT3 knockout+CT treatment(CT+siSTAT3)group.The AD model in vitro was constructed by Aβ_(1-42),and the STAT3-silenced neuron model was constructed by transfection siSTAT3,and then cultured for 24 h with high and low doses of CT prescription.Cell activity assay(CCK-8)measured cell viability.The expression levels of Cyclin D1,Ki-67,B-cell lymphoma/leukemia-2(Bcl-2),Bcl-2 associated X protein(Bax),NF200,GAP-43,STAT3 and phosphorylated STAT3(p-STAT3)protein were detected by Western Blot.Cell proliferation was detected by Brdu.Apoptosis was detected by terminal deoxyribonucleotidyl transferase(TdT)-mediated dUTP nick end labeling(TUNEL).Results:CT prescription significantly shortened the escape latency of APP/PS1 double-transfected mice,increased the targed quadrant residence time and number of target quadrant stays,and up-regulated the expression of GAP-43 in hippocampus,and the effect of high dose of CT prescription was more significant(P<0.05 or P<0.01).CT prescription restored the ac

关 键 词：阿尔茨海默病 梓醇-川芎嗪方 转录激活蛋白3 轴突可塑性 实验研究 

分 类 号：R285.5[医药卫生—中药学]