PPARδ激动剂GW501516对高糖致体外神经-血管单元损伤的保护作用及机制研究  

The protective effect of PPARδagonist GW501516 on neurovascular unit injury induced by high glucose in vitro and its mechanism

作　　者：王赛 陈清杰[2] 张金玲 何业谱 陈辉[3] WANG Sai;CHEN Qing-jie;ZHANG Jin-ling;HE Ye-pu;CHEN Hui(Dept of Cardiovascular Medicine,Xianning Central Hospital,the First Affiliated Hospital of Hubei University of Science and Technology;Institute of Medicine,Faculty of Medicine,Hubei University of Science and Technology;Dept of Pharmacy,Xianning Central Hospital,the First Affiliated Hospital of Hubei University of Science and Technology,Xianning Hubei 437100,China)

机构地区：[1]咸宁市中心医院/湖北科技学院附属第一医院心血管内科 [2]湖北科技学院医学部医药研究院 [3]咸宁市中心医院/湖北科技学院附属第一医院药学部,湖北咸宁437100

出　　处：《中国药理学通报》2024年第10期1963-1970,共8页Chinese Pharmacological Bulletin

基　　金：国家自然科学基金面上资助项目(No 82270892)。

摘　　要：目的探讨PPARδ激动剂GW501516对高糖诱导的体外神经-血管单元(neuro-vascular unit,NVU)损伤的保护作用及其机制。方法体外分离、纯化和培养SD大鼠海马神经元(neurons,Neu)、星形胶质细胞和脑微血管内皮细胞,并建立NVU共培养体系。将NVU共培养体系细胞分为:对照组、高糖组(HG组)、HG+GW501516低、中、高浓度组(25、50、100 nmol·L^(-1))和HG+GW501516(100 nmol·L^(-1))+ANA12(TrkB抑制剂,5μmol·L^(-1))组。采用跨内皮电阻(transendothelial resistance,TEER)检测和渗漏实验检测评价NUV屏障功能;CCK-8实验检测共培养体系中Neu细胞增殖活性;ELISA法检测细胞上清液中炎症因子TNF-α、IL-6和IL-1β水平;化学法检测Neu细胞中氧化应激指标SOD、MDA和NO水平;流式细胞术检测Neu细胞凋亡水平;Western blot检测Neu细胞中PPARδ、Bax、Bcl-2、cleaved-caspase-3以及BDNF/TrkB通路相关蛋白BDNF、p-TrkB和TrkB表达水平。结果与对照组比较,HG组TEER值降低、渗漏值升高,Neu细胞增殖活性降低,上清液中TNF-α、IL-6、IL-1β及Neu细胞中MDA和NO水平升高,SOD水平降低,Neu细胞凋亡率及细胞中Bax、cleaved caspase-3蛋白表达水平升高,而PPARδ、Bcl-2、BDNF和p-TrkB/TrkB蛋白表达水平降低(P<0.05);与HG组比较,不同浓度GW501516处理后TEER值升高、渗漏值降低,Neu细胞增殖活性升高,上清液中TNF-α、IL-6、IL-1β及Neu细胞中MDA和NO水平降低,SOD水平升高,Neu细胞凋亡率及细胞中Bax、cleaved caspase-3蛋白表达水平降低,PPARδ、Bcl-2、BDNF和p-TrkB/TrkB蛋白表达水平升高(P<0.05),且具有浓度依赖性;然而,加入ANA12干预后会逆转GW501516对高糖条件下NVU损伤的改善作用。结论PPARδ激动剂GW501516通过激活BDNF/TrkB信号通路的转导改善高糖诱导的体外NVU损伤。Aim To explore the protective effect of PPARδagonist GW501516 on neuro-vascular unit(NVU)injury induced by high glucose in vitro and its mechanism.Methods SD rat hippocampal neurons(Neu),astrocytes(Ast)and brain microvascular endothelial cells(BMEC)were isolated,purified and cultured in vitro,and NVU co-culture system was established.NVU co-culture system cells were divided into the control group,high glucose group(HG group),HG+GW501516 low,medium and high concentration groups(25,50 and 100 nmol·L^(-1))and HG+GW501516(100 nmol·L^(-1))+ANA12(TrkB inhibitor,5μmol·L^(-1))group.NUV barrier function was evaluated by transendothelial resistance(TEER)test and leakage test;the proliferative activity of Neu cells in co-culture system was detected by CCK-8 assay;the levels of inflammatory cytokines TNF-α,IL-6 and IL-1βin cell supernatant were detected by ELISA;the levels of SOD,MDA and NO in Neu cells were detected by chemical method;the apoptosis level was detected by flow cytometry;the protein expression levels of PPARδ,Bax,Bcl-2,cleaved caspase-3,and BDNF/TrkB pathway-related proteins BDNF,p-TrkB,and TrkB in Neu cells were detected by Western blot.Results Compared with the control group,the TEER value decreased and leakage value increased in HG group;the proliferation activity of Neu cells decreased,the levels of TNF-α,IL-6,IL-1βin supernatant and MDA and NO in Neu cells increased,and the SOD level decreased;Neu cell apoptosis rate and expression levels of Bax and Cleaved caspase-3 increased,while the expression levels of PPARδ,Bcl-2,BDNF and p-TRKB/TrkB decreased(P<0.05).Compared with the HG group,after treatment with different concentrations of GW501516,TEER value increased,leakage value decreased,proliferation activity of Neu cells increased,levels of TNF-α,IL-6,IL-1β in supernatant and MDA and NO in Neu cells decreased,and SOD level increased,and apoptosis rate of Neu cells and expression levels of Bax and cleaved caspase-3 were decreased,and expression levels of PPARδ,Bcl-2,BDNF and p-TRKB/TrkB increa

关 键 词：PPARΔ GW501516 神经-血管单元 高糖 BDNF/TrkB信号通路 细胞凋亡 

分 类 号：R-332[医药卫生] R322.12R322.8R329.25R343R743.9R977.6