巨噬细胞迁移抑制因子抑制剂ISO-1对脑缺血再灌注大鼠的影响

Effects of macrophage migration inhibitory factor inhibitor ISO-1 on brain ischemia-reperfusion in rats

目的:探讨巨噬细胞迁移抑制因子(MIF)对大鼠脑缺血再灌注损伤(I/R)后神经损伤及可能涉及的机制。方法:将42只雄性SD大鼠随机分为假手术组(sham)、缺血再灌注组(I/R)和MIF抑制剂ISO-1处理组(ISO-1)。利用大脑中动脉阻塞(MCAO)制备大鼠I/R模型。ISO-1组大鼠于再灌注同时腹腔注射ISO-1。采用Garcia评分和平衡木实验评定各组大鼠的神经功能损伤情况。通过2,3,5-三苯四唑氯化(TTC)染色检测大鼠脑梗死体积,Nissl染色观察大鼠神经元形态,TUNEL染色检测神经细胞凋亡情况,并采用免疫荧光染色观察大脑皮层中MIF在胶质细胞纤维酸蛋白(GFAP)、离子钙接头蛋白1(Iba-1)和神经元核抗原(Neu N)的表达变化,通过Western Blot检测MIF、腺苷酸活化蛋白激酶(AMPK)、乳酸脱氢酶A(LDHA)、丙酮酸激酶M2(PKM2)、葡萄糖转运蛋白-1(GLUT-1)在脑组织中表达情况。结果:MIF在MCAO大鼠脑组织梗死周边区明显增加,且主要位于神经元中。与I/R组相比,ISO-1组梗死体积明显减小,神经元凋亡数目显著减少,并能够抑制星形胶质细胞增殖活化,从而改善神经功能的损伤。AMPK、LDHA、PKM-2和GLUT-1蛋白表达上调,表明ISO-1可以促进糖代谢过程。结论:MIF抑制剂ISO-1对MCAO大鼠缺血再灌注后神经功能的恢复发挥重要的正向调控作用,可能与糖代谢相关的分子有关。

Objective:To investigate the effect of macrophage migration inhibitory factor(MIF)on nerve injury after cerebral ischemia-reperfusion injury(I/R)in rats and its potential mechanism.Methods:Forty-two male SD rats were randomly divided into sham group(sham),ischemia-reperfusion group(I/R)and MIF inhibitor ISO-1 treatment group(ISO-1).Middle cerebral artery occlusion(MCAO)was used to establish I/R model in rats.Rats in ISO-1 group were intraperitoneally injected with ISO-1 at the same time of reperfusion.Garcia score and balance beam walking test score were used to evaluate the neurologic impairment of rats in each group.2,3,5-triphenyltetrazolium chloride(TTC)staining was used to detect the area of cerebral infarction.Nissl staining was used to observe the morphology of neurons.TUNEL staining was used to detect the apoptosis of neurons.Immunofluorescence staining was used to observe the expression changes of MIF in glial fibrillary acidic protein(GFAP),ionocalcin-1(Iba-1),and neuronal nuclear antigen(Neu N)in cerebral cortex.Detecting the expression of MIF,AMP-activated protein kinase(AMPK),lactate dehydrogenase A(LDHA),pyruvate kinase M2(PKM2),and glucose transporter-1(GLUT-1)in brain tissue by Western Blot.Results:MIF was significantly increased in the peri-infarct region of I/R rats,and mainly located in neurons.Compared with I/R group,ISO-1 group improved neurological function by significantly reducing infarct size and apoptosis neurons number and inhibiting astrocyte proliferation and activation.The up-regulation of AMPK,LDHA,PKM2,and GLUT-1 protein expression indicates that ISO-1 may promote glucose metabolism.Conclusion:MIF inhibitor ISO-1 plays an important positive regulatory role in the recovery of neurological function after ischemia-reperfusion in MCAO rats,which may be related to the molecules involved in glucose metabolism.