补阳还五汤对脑缺血后12/15-脂氧合酶及相关炎性产物影响  

Effects of Buyang Huanwu Decoction(补阳还五汤)on 12/15-Lipoxygenase and Related Inflammatory Metoblites after Cerebral Ischemia-reperfusion in Rats

作　　者：高宇容 张亚星 巫芳华 戴嘉颖 刘微[1] GAO Yurong;ZHANG Yaxing;WU Fanghua;DAI Jiaying;LIU Wei(Guangzhou University of Chinese Medicine,Guangzhou 510006,Guangdong,China)

机构地区：[1]广州中医药大学,广东广州510006

出　　处：《辽宁中医药大学学报》2024年第5期13-18,共6页Journal of Liaoning University of Traditional Chinese Medicine

基　　金：国家自然科学基金项目(81673772);广东省中医药局科研项目(20221116)。

摘　　要：目的观察补阳还五汤对脑缺血再灌注后12/15-脂氧合酶(12/15-LOX)及下游相关代谢产物的影响。方法将雄性SD大鼠随机分为假手术组、模型组、补阳还五汤组和12/15-LOX抑制剂PD146176组。假手术组只分离组织和剥离血管,模型组建立大鼠大脑中动脉栓塞(middle cerebral artery occlusion,MCAO)模型,补阳还五汤组于造模术后给予补阳还五汤16 g·kg-1灌胃,2次/d,PD146176组于造模术前30 min给予PD14617610 mg·kg-1皮下注射(同补阳还五汤组做阳性对照)。缺血再灌注后6 h、24 h、48 h和72 h运用蛋白免疫印迹法(Western blot)检测脑组织12/15-LOX的表达水平;免疫荧光(IF)法观察大鼠脑组织中12/15-LOX蛋白与神经元标记物(NeuN)、小胶质细胞标记物(Iba1)及星形胶质细胞标记物(GFAP)的共定位;酶联免疫吸附(ELISA)法检测12/15-LOX抗炎代谢产物消退素D1(RvD1)的含量,以及促炎代谢产物12-羟基廿碳四烯酸(12-HETE)和15-羟基廿碳四烯酸(15-HETE)的含量;免疫荧光法检测髓过氧化物酶(MPO)的表达,反映炎症细胞浸润程度。蛋白免疫印迹法检测脑组织白细胞介素-6(IL-6)和白细胞介素-10(IL-10)表达水平。结果与假手术组比较,模型组在脑缺血再灌注不同时间点12/15-LOX表达均升高(P<0.01),与模型组比较,补阳还五汤组在脑缺血再灌注后6 h、24 h、48 h、72 h 12/15-LOX表达均降低(P<0.05或P>0.05)。12/15-LOX与NeuN和Iba1共定位重合较多,与GFAP共定位基本无重合。与假手术组比较,模型组RvD1含量降低(P<0.01),12-HETE和15-HETE含量升高(P<0.01),MPO和IL-6表达升高(P<0.01),IL-10表达降低(P<0.01);与模型组比较,PD146176组和补阳还五汤组RvD1含量升高(P<0.05,P<0.01),12-HETE和15-HETE含量降低(P<0.01),MPO表达减少(P<0.01),IL-6蛋白表达降低(P<0.05,P<0.01),补阳还五汤组IL-10蛋白表达升高(P<0.01)。结论补阳还五汤可能通过抑制12/15-LOX表达而增加抗炎产物RvD1,降低促炎产物12-HETE和15-HETE,以减�Objective To study the efffects of Buyang Huanwu Decoction(补阳还五汤,BYWHD)on 12/15-lipoxygenase(12/15-LOX)and its downstream metabolites after cerebral ischemia-reperfusion in rats.Methods Male SD rats were randomly divided into sham group,model group,PD146176 group and BYHWD group.In sham group,blood vessels was isolated;the middle cerebral artery occlusion(MCAO)was established for model group.BYHWD group was given intragastric treatment 16 g·kg-1 after MCAO surgery,once in the morning and once in the evening,and PD146176 group was given PD146176 subcutaneous injection 10 mg·kg-130 min before MCAO surgery.Western blot was used to detect the expression of 12/15-LOX in the brain at 6 h,24 h,48 h,72 h after ischemia-reperfusion.Immunofluorescence(IF)was used to observe colocalization of 12/15-LOX with NeuN,Iba1 and GFAP in rats brain tissue.Enzyme-linked immunosorbent assay(ELISA)was used to detect the anti-inflammatory metabolite resolvin D1(RvD1),the pro-inflammatory metabolite 12-hydroxyeicosatetraenoic acid(12-HETE)and 15-hydroxyeicosatetraenoic acid(15-HETE)of 12/15-LOX.Immunofluorescence was used to detect the expression of myeloperoxidase(MPO),which reflects the degree of inflammatory cell infiltration.Western blot was used to detect the expression of interleukin-6(IL-6)and interleukin-10(IL-10)in the brain after ischemia-reperfusion.Results Compared with sham group,the expression of 12/15-LOX was significantly increased in model group at different time points of cerebral ischemia-reperfusion(P<0.01).Compared with model group,the expression of 12/15-LOX in BYHWD group was decreased at 6 h,24 h,48 h and 72 h(P<0.05 or P>0.05)after cerebral ischemia-reperfusion.12/15-LOX mainly colocalized with NeuN and Iba1,almost no colocalization with GFAP;Compared with sham group, RvD1 was decreased(P <0.01),while 12-HETE and 15-HETE were increased(P <0.01),accompanied with increased MPO and IL-6(P <0.01),decreased IL-10 in the model group(P <0.01);Compared with model group,RvD1 was increased(P <0.05,P <0.01),whil

关 键 词：补阳还五汤 12/15-脂氧合酶 脑缺血再灌注 炎症 

分 类 号：R255.2[医药卫生—中医内科学]