芒柄花素保护BV2小胶质细胞糖氧剥夺再灌注损伤及其机制研究  

Protective effects of formononetin against oxygen-glucose deprivation/reperfusion injury in BV2 microglia and associated mechanism

作　　者：韦丁玲 王湄 王文秀 曹丽平 何前松 Wei Dingling;Wang Mei;Wang Wenxiu;Cao Liping;He Qiansong(The First Clinical Medical College of Guizhou University of Traditional Chinese Medicine;Department of Neurology,The First Affiliated Hospital of Guizhou University of Traditional Chinese Medicine)

机构地区：[1]贵州中医药大学第一临床医学院,贵阳550001 [2]贵州中医药大学第一附属医院神经内科,贵阳550001

出　　处：《重庆医科大学学报》2025年第2期244-249,共6页Journal of Chongqing Medical University

基　　金：国家自然科学基金资助项目(编号:82160850);贵州省科技基金资助项目(编号:黔科合基础[2020]1Z071)。

摘　　要：目的:探讨芒柄花素(formononetin,FN)对糖氧剥夺再灌注(glucose and oxygen deprivation/reperfusion,OGD/R)刺激的BV2小胶质细胞多聚ADP核糖聚合酶1[poly(ADP-ribose)polymerase 1,PARP1]/聚(ADP-核糖)糖水解酶[poly(ADP-Ribose)glycohydrolase,PARG]信号通路及神经炎症的影响。方法:建立OGD/R BV2细胞模型,分为空白对照组、OGD/R组、10μm FN组、OGD/R+10μm FN处理组、OGD/R+抑制剂PJ34(10μm)处理组、OGD/R+抑制剂Ethacridine lactate(7.5μm)处理组。采用免疫荧光法(immunofluorescence method,IF)检测BV2细胞核因子-κB p65(Nuclear factor-kappa B p65,NF-κB p65)蛋白核转移及p53、凋亡诱导因子(apoptosis-inducing factor,AIF)、Toll样受体4(Toll-like receptor 4,TLR4)蛋白表达;免疫印迹法检测(Western blot,WB)PARP1/PARG通路相关蛋白表达;酶联免疫吸附试验(enzyme-linked immunosorbent assay,ELISA)检测血清γ干扰素(interferon-γ,IFN-γ)、白细胞介素-1β(interleukin-1β,IL-1β)以及肿瘤坏死因子-α(tumor necrosis factor-α,TNF-α)含量。结果:WB结果显示,与对照组相比,进行糖氧剥夺6 h后再进行复糖复氧培养1 h,细胞中PARP1、PARG表达明显升高(P=0.000);与OGD/R组相比,在OGD/R后加入FN处理,BV2细胞中PARP1(P=0.000)和PARG(P=0.000)蛋白表达量明显下降,Iduna蛋白表达量明显上升(P=0.000);在OGD/R后加入PARP抑制剂PJ34处理后,PARP1蛋白表达量明显下降(P=0.000),但PARG和Iduna蛋白表达量无明显变化(P=0.061);在OGD/R后加入PARG抑制剂Ethacridine lactate处理后,BV2细胞中PARG蛋白表达量明显下降(P=0.000),但PARP1和Iduna蛋白表达量无明显变化(P=0.072)。IF结果显示,与OGD/R组相比,OGD/R后加入FN,细胞中p53、AIF、TLR4、NF-κB p65表达均明显下降,且NF-κB p65核转移明显减少;在OGD/R后加入PARP抑制剂PJ34或PARG抑制剂Ethacridine lactate,p53、AIF、TLR4表达同样下降,NF-κB p65核转移也明显减少。ELISA结果显示,与OGD/R组BV2细胞相比,在OGD/R后加入FN处理,细胞中IL-1β(P=Objective:To investigate the effects of formononetin on the PARP1/PARG signaling pathway and neuroinflammation in BV2 microglia activated by oxygen-glucose deprivation/reperfusion(OGD/R).Methods:An OGD/R model was established with BV2 cells.These microglia were divided into blank control group,OGD/R group,formononetin(10μm)group,OGD/R+formononetin(10μm)group,OGD/R+PJ34(PARP1 inhibitor,10μm)group,and OGD/R+ethacridine lactate(PARG inhibitor,7.5μm)group.We measured the nuclear translocation of nuclear factor-kappa B(NF-κB)p65 and the expression of p53,apoptosis-inducing factor(AIF),and Toll-like receptor 4(TLR4)proteins in BV2 cells by immunofluorescence assay;measured the expression of PARP1/PARG pathway-related proteins by Western blot;and determined the content of interferon(INF)-γ,interleukin(IL)-1β,and tumor necrosis factor-α(TNF-α)by enzyme-linked immunosorbent assay(ELISA).Results:Western blot results showed that compared with the control group,the expression of PARP1 and PARG in cells was significantly increased after 6-hour oxygen-glucose deprivation followed by 1-hour reperfusion(P<0.05);compared with the OGD/R group,the OGD/R+formononetin group had significantly decreased expression of PARP1 and PARG proteins(P<0.05)and significantly increased expression of Iduna protein(P<0.05);the OGD/R+PJ34 group had significantly decreased expression of PARP1 protein(P<0.05),but the expression of PARG and Iduna proteins had no significant changes(P>0.05);the OGD/R+ethacridine lactate group had significantly decreased expression of PARG protein(P<0.05),but had no significant changes in the expression of PARP1 and Iduna proteins(P>0.05).Immunofluorescence assay results showed that compared with the OGD/R group,formononetin significantly reduced the expression of p53,AIF,TLR4,and NF-κB p65 and the nuclear translocation of NF-κB p65 in OGD/R cells;and both PJ34 and ethacridine lactate significantly reduced the expression of p53,AIF,and TLR4 and the nuclear translocation of NF-κB p65 in OGD/R cells.ELISA resul

关 键 词：芒柄花素 BV2小胶质细胞 糖氧剥夺再灌注 多聚ADP核糖聚合酶1抑制剂 聚(ADP-核糖)糖水解酶抑制剂 

分 类 号：R926[医药卫生—药学]