文献类型：期刊文献

中文题名：黑逍遥散调控PINK1/Parkin信号通路干预线粒体自噬防治阿尔茨海默病的机制

英文题名：Mechanism of Hei Xiaoyaosan in Preventing and Treating Alzheimer's Disease via Regulation of PINK1/Parkin Signaling Pathway and Intervention in Mitophagy

作者：王虎平[1,2,3];裴文丽[1];韩玉梅[1];陈怡琴[1];杨娇[1]

第一作者：王虎平

机构：[1]甘肃中医药大学,兰州730000;[2]甘肃省中医方药挖掘与创新转化重点实验室,兰州730000;[3]甘肃省中药新产品创制工程实验室,兰州730000

第一机构：甘肃中医药大学

年份：2025

卷号：31

期号：19

起止页码：162

中文期刊名：中国实验方剂学杂志

外文期刊名：Chinese Journal of Experimental Traditional Medical Formulae

收录：;北大核心:【北大核心2023】；

基金：国家自然科学基金项目(82160862,81960828);甘肃省高校教师创新基金项目(2024A-083);第五批全国中医临床优秀人才研修项目(国中医药人教函[2022]239号);首批陇原青年英才项目(中共甘肃省委人才工作领导小组[2022]5号)。

语种：中文

中文关键词：阿尔茨海默病;黑逍遥散;PTEN诱导的激酶1(PINK1)/帕金蛋白(Parkin)信号通路;线粒体自噬

外文关键词：Alzheimer's disease;Hei Xiaoyaosan;PTEN-induced kinase 1(PINK1)/Parkin signaling pathway;mitophagy

摘要：目的:探究黑逍遥散调控PTEN诱导的激酶1(PINK1)/帕金蛋白(Parkin)信号通路干预线粒体自噬防治阿尔茨海默病(AD)的作用及其机制。方法:将50只4月龄雄性APP/PS1小鼠随机分为模型组,盐酸多奈哌齐组(0.65 mg·kg^(-1)),黑逍遥散高、中、低剂量组(22.10、11.05、5.53 g·kg^(-1)),另选取10只同月龄的雄性C57BL/6J小鼠作为空白组。给药90 d后进行Y迷宫试验测试学习记忆能力,苏木素-伊红(HE)染色观察小鼠海马组织形态学变化,荧光染料Fluoro-Jade B(FJB)染色观察小鼠神经元损伤及其数量变化情况,透射电镜观察小鼠线粒体损伤情况,免疫荧光观察小鼠海马区微管自噬相关蛋白1轻链3(LC3)及选择性自噬接头蛋白1(p62)等自噬指标的变化情况,蛋白免疫印迹法(Western blot)检测PINK1、Parkin、磷酸化(p)-Parkin、LC3、p62的蛋白表达水平,实时荧光定量聚合酶链式反应(Real-time PCR)检测PINK1、Parkin mRNA表达水平。结果:与空白组比较,模型组小鼠交替反应率下降(P<0.01);海马组织排列无序,细胞间隙较大,细胞核固缩较严重;神经元损伤程度加重,损伤细胞数量增加(P<0.01);线粒体肿胀破裂程度较重,PINK1、p-Parkin蛋白及mRNA表达水平下降(P<0.01);自噬蛋白p62免疫荧光强度增加(P<0.01),LC3荧光信号强度下降(P<0.01)。与模型组比较,盐酸多奈哌齐组,黑逍遥散高、中剂量组小鼠反应交替率上升(P<0.05,P<0.01);小鼠海马神经元损伤程度减轻,损伤细胞数量减少(P<0.01);细胞间隙缩小,细胞核固缩情况改善;线粒体肿胀破裂程度得到缓解;盐酸多奈哌齐组及黑逍遥散高剂量组p62荧光强度减少(P<0.05,P<0.01),LC3荧光强度增加(P<0.05,P<0.01),PINK1、p-Parkin蛋白表达上升(P<0.05,P<0.01),LC3表达增加(P<0.05,P<0.01),p62表达减少(P<0.05,P<0.01);盐酸多奈哌齐组及黑逍遥散高、中剂量组PINK1的mRNA水平上升(P<0.05,P<0.01),盐酸多奈哌齐组及黑逍遥散高剂量组Parkin的mRMA水平上升(P<0.05)。结论:黑逍遥散可改善APP/PS1小鼠的学习记忆能力及病理状态,其机制可能是通过调控PINK1/Parkin信号通路,增强线粒体自噬,修复受损神经细胞以防治AD。
Objective:This study aims to explore the effect and mechanism of Hei Xiaoyaosan in preventing Alzheimer's disease(AD)by regulating the PTEN-induced kinase 1(PINK1)/Parkin signaling pathway and intervening in mitophagy.Methods:Fifty 4-month-old male APP/PS1 mice were randomly divided into a model group,the donepezil hydrochloride group(0.65 mg·kg^(-1)),and the high-,medium-,and low-dose Hei Xiaoyaosan groups(22.10,11.05,and 5.53 g·kg^(-1)).Ten additional male C57BL/6J mice of the same age served as the blank control group.Following a 90-day treatment period,learning and memory functions were evaluated using the Y-maze test.Additionally,hippocampal morphology was examined through hematoxylin-eosin(HE)staining.Fluoro-Jade B(FJB)staining was employed to examine neuronal damage and the number of damaged cells.Transmission electron microscopy(TEM)was utilized to observe mitochondrial damage.Immunofluorescence staining was employed to assess the expression of autophagy-related proteins microtubule-associated protein 1 light chain 3(LC3)and selective autophagy adaptor protein 1(p62)in the hippocampus.Western blot analysis was performed to quantify the protein expression of PINK1,Parkin,phosphorylated Parkin(p-Parkin),LC3,and p62.Real-time quantitative polymerase chain reaction(Real-time PCR)was utilized to evaluate the mRNA expression levels of PINK1 and Parkin.Results:Compared with the blank group,the model group exhibited a decreased alternation rate in the Y-maze test(P<0.01),disordered hippocampal cell arrangement,enlarged intercellular space,and severe nuclear pyknosis.There was increased neuronal damage and number of damaged cells(P<0.01),severe mitochondrial swelling and rupture,reduced protein and mRNA expression levels of PINK1 and p-Parkin(P<0.01),increased immunofluorescence intensity of p62(P<0.01),and decreased fluorescence signal intensity of LC3(P<0.01).Compared with the model group,the donepezil hydrochloride group and the high-and medium-dose Hei Xiaoyaosan groups showed an increased alternation rate(P<0.05,P<0.01),alleviated hippocampal neuronal damage,and reduced numbers of damaged cells(P<0.01).There was a narrowed intercellular arrangement,improved nuclear morphology,and alleviated mitochondrial injury.The donepezil hydrochloride group and the high-dose Hei Xiaoyaosan group exhibited reduced p62 fluorescence intensity(P<0.05,P<0.01),increased LC3 fluorescence intensity(P<0.05,P<0.01),elevated protein expression of PINK1 and p-Parkin(P<0.05,P<0.01),increased LC3 expression(P<0.05,P<0.01),and decreased p62 expression(P<0.05,P<0.01).The donepezil hydrochloride group and the high-and medium-dose Hei Xiaoyaosan groups showed increased mRNA expression levels of PINK1(P<0.05,P<0.01),and the donepezil hydrochloride group and the high-dose Hei Xiaoyaosan group exhibited elevated mRNA levels of Parkin(P<0.05).Conclusion:Hei Xiaoyaosan can improve learning and memory abilities and ameliorate pathological damage in APP/PS1 mice.The mechanism may be related to the regulation of the PINK1/Parkin signaling pathway,enhancement of mitophagy,and repair of damaged neurons to prevent and treat AD.