关键词:

黄芪散, 阿尔茨海默病, 线粒体自噬, PTEN诱导激酶1/帕金蛋白信号通路, NLRP3炎症小体

Abstract:

This study aims to explore the impact of Huangqisan on synaptic plasticity in rat model of Alzheimer′s disease and to analyzed its potential mechanism.Fifty SPF grade male SD rats were randomly divided into normal group (Norm),model group (Mod),Huangqisan low dose group (HQS-L,1.2 g/kg),Huangqisan high dose group(HQS-H,4.8 g/kg) and positive group (Pos),with 10 rats in each group.Except for the normal group,the rats in other groups were injected with β-amyloid 25-35 into bilateral hippocampal to establish the model.Morris water maze test was used to detect the learning and memory function of the rats.Nissl staining was used to detect neuronal damage in hippocampus of rats.JC-1 probe was applied to detect the mitochondrial membrane potential in hippocampus by flow cytometry.Transmission electron microscope was used to detect the ultrastructural observation of neurons and synapses in rat hippocampus.ELISA was performed to measure the levels of pro-inflammatory cytokines interleukin-1β (IL-1β),interleukin-6 (IL-6),and tumor necrosis factor-α (TNF-α) in the rat hippocampus.Flow cytometry was used to detect the level of hippocampus reactive oxygen species (ROS).The expressions of PTEN-induced kinase 1 (PINK1),Parkin E3 ubiquitin protein ligase (Parkin),microtuble-associated protein light chain 3 (LC3B),ubiquitin-binding protein p62 (p62),dynamin-related protein (Drp1),mitofusion 1 (Mfn1),mitofusion 2 (Mfn2),NOD-like receptor protein 3 (NLRP3),growth-associated protein 43 (GAP43),N-methyl-D-aspartate receptor (NMDAR) subunit 2B (NR2B),postsynaptic dense protein 95 (PSD95),synaptosin (SYP) and brain-derived neurotrophic factor (BDNF) proteins in the hippocampus were detected by Western blot (WB).Compared with Norm group,the learning and memory function of rats in the Mod group was significantly reduced(P<0.01).Nissl bodies in neurons of hippocampus decreased or disappeared.The ultrastructure of neurons and synapses in hippocampus presented obvious pathological changes.Mitochondrial membrane potential decreased significantly and the structure was damaged,autolysosome has formed.The contents of IL-1β,IL-6 and TNF-α in hippocampus increased significantly (P<0.01),the level of ROS increased significantly(P<0.01),the protein expression of PINK1,Parkin,LC3B,Mfn1,Mfn2,GAP43,NR2B,PSD95,SYP and BDNF decreased significantly(P<0.05),and the protein expression of p62,Drp1,NLRP3 increased significantly(P<0.01).Compared with Mod group,the learning and memory function of AD rats improved significantly in the HQS-L group,HQS-H group and Pos group,which was mainly manifested by shortened the escape latency and elevated the number of crossing the platform (P<005,P<0.01).Nissl bodies in neurons of hippocampus increased,the ultrastructure of neurons and synapses in hippocampus was improved,the mitochondrial membrane potential was increased,and the mitophagy was increased.The contents of IL-1β,IL-6 and TNF-α in hippocampus decreased significantly (P<0.05,P<0.01),the level of ROS decreased significantly(P<0.01),the protein expression of PINK1,Parkin,LC3B,Mfn1,Mfn2,GAP43,NR2B,PSD95,SYP and BDNF increased significantly,and the protein expression of p62,Drp1,NLRP3 decreased significantly (P<0.05,P<0.01).The mechanism of Huangqisan improved synaptic plasticity in AD rats might be related to the activation of PINK1-Parkin pathway in hippocampus to promoted mitophagy,improved mitochondrial function,reduced ROS level and inhibited the activation of NLRP3 inflammasome.

Key words:

Huangqisan, Alzheimer′s disease, mitophagy, PTEN-induced putative kinase 1/Parkin signaling pathway, NLRP3 inflammatory corpuscles