滋肾活血方对2-VO模型大鼠海马CA1区线粒体自噬的调节及神经元再生的影响  

Effect of Zishen Huoxue Prescription on Regulation of Mitophagy and Neurogenesis in Hippocampal CA1 Region of Vascular Dementia Rats

作　　者：赵梓婷 伍大华[2] 张秀丽[3] 谢乐 刘桐赫 曾珊珊 施佳仪 姚婷[2] 谢瑶 李映辰 ZHAO Ziting;WU Dahua;ZHANG Xiuli;XIE Le;LIU Tonghe;ZENG Shanshan;SHI Jiayi;YAO Ting;XIE Yao;LI Yinchen(Hunan University of Chinese Medicine,Changsha 410208,China;Affiliated Hospital of Hunan Academy of Traditional Chinese Medicine,Changsha410006,China;Science and Technology Innovation Center,Hunan University of Chinese Medicine,Changsha 410208,China)

机构地区：[1]湖南中医药大学,长沙410208 [2]湖南省中医药研究院附属医院,长沙410006 [3]湖南中医药大学科技创新中心,长沙410208

出　　处：《中国实验方剂学杂志》2022年第20期45-52,共8页Chinese Journal of Experimental Traditional Medical Formulae

基　　金：国家自然科学基金面上项目(81874462);湖南省重点研发计划项目(2020SK2123);湖南省教育厅优秀青年基金项目(19B426)。

摘　　要：目的:探讨滋肾活血方通过调节线粒体自噬促进双侧颈总动脉永久性结扎(2-VO)模型大鼠海马CA1区神经元新生的作用机制。方法:2-VO法建立血管性痴呆大鼠模型,随机将60只SD大鼠分为假手术组、2-VO模型组、盐酸多奈哌齐组、滋肾活血方低、中、高剂量组(8.9、17.8、35.6 g·kg^(-1))。Morris水迷宫实验检测各组逃避潜伏期、跨越平台次数;透射电镜观察海马CA1区线粒体自噬情况;实时荧光定量聚合酶链式反应(Real-time PCR)检测海马CA1区PTEN诱导激酶1(Pink1)、帕金蛋白(Parkin)mRNA的表达;蛋白免疫印迹法(Western blot)检测海马CA1区线粒体自噬信号通路相关蛋白Parkin、抗增殖蛋白2(PHB2)、线粒体融合蛋白2(Mfn2)、动力学相关蛋白1(Drp1)蛋白的表达;Brdu法检测CA1区神经元再生情况。结果:与假手术组比较,2-VO模型组大鼠学习、空间记忆能力明显下降(P<0.05),海马CA1区见线粒体结构受损,自噬溶酶体形成,Parkin、Pink1 mRNA及Parkin、PHB2、Drp1蛋白明显上调(P<0.05),Mfn2蛋白表达量明显降低(P<0.05),海马CA1区神经元新生明显减少(P<0.05)。与模型组比较,滋肾活血方干预后可明显提高2-VO模型大鼠的学习、空间记忆能力(P<0.05),增加海马CA1区自噬小体数量并改善线粒体结构,进一步上调海马CA1区Parkin、Pink1 mRNA及Parkin、PHB2、Drp1蛋白表达(P<0.05,P<0.01),下调Mfn2蛋白表达(P<0.05,P<0.01),增加海马CA1区神经元新生数量(P<0.05,P<0.01)。结论:滋肾活血方对血管性痴呆大鼠海马CA1区神经元新生的促进作用与Pink1/Parkin信号通路介导的线粒体自噬有关。Objective:To investigate the effect of Zishen Huoxue prescription on promoting neurogenesis in hippocampal CA1 region of vascular dementia(VD)rats by regulating mitophagy.Method:The 2-VO method was used to establish the VD rat model and 60 SD rats were randomly divided into sham operation group,model group,donepezil hydrochloride group,and Zishen Huoxue prescription low-dose(8.9 g·kg^(-1)),medium-dose(17.8 g·kg^(-1))and high-dose(35.6 g·kg^(-1))groups.Morris water maze test was performed to detect the escape latency and the number of crossing platform in each group.The expression of phosphatase and tensin homology-induced kinase 1(PINK1)and Parkinson protein(Parkin)mRNA in hippocampal CA1 region was detected by Real-time fluorescent quantitative polymerase chain reaction(Realtime PCR).Western blot was used to determine the expression of mitochondrial autophagy signaling pathwayrelated proteins Parkin,prohibitin 2(PHB2),mitofusin 2(Mfn2)and dynamin-related protein 1(Drp1)in hippocampal CA1 region.The neurogenesis in hippocampal CA1 region was tested by Brdu method.Result:Compared with the conditions in the sham operation group,the learning and spatial memory abilities of the model group were decreased(P<0.05),with damaged mitochondrial structure and autolysosome formation in the hippocampal CA1 region.The expressions of Parkin,Pink1 mRNA and Parkin,PHB2,and Drp1 proteins were up-regulated(P<0.05),while the expression of Mfn2 protein and the neuronal regeneration in hippocampal CA1 region were reduced(P<0.05,P<0.05).Compared with the conditions in the model group,Zishen Huoxue prescription enhanced the learning and spatial memory abilities of VD rats(P<0.05),increased the number of autophagosomes in hippocampal CA1 region and improved the mitochondrial structure.The expression of Parkin,Pink1 mRNA and Parkin,PHB2,and Drp1 proteins in hippocampal CA1 region was up-regulated(P<0.05,P<0.01)while the expression of Mfn2 protein was down-regulated(P<0.05,P<0.01).The number of new neurons in hippocampal CA1 region was

关 键 词：血管性痴呆 CA1区 神经元新生 线粒体分裂蛋白 认知障碍 滋肾活血方 

分 类 号：R2-0[医药卫生—中医学] R22R285.5R289R33