促红细胞生成素对发育期小鼠脑损伤后脑源性神经营养因子和酪氨酸受体激酶B表达的影响

Effects of erythropoietin on expression of brain-derived neurotrophic factor and tyrosine receptor kinase B after brain injury in mice of different developmental stages

目的探讨促红细胞生成素(EPO)对鹅膏蕈氨酸(Ibo)致不同发育期小鼠脑损伤后脑源性神经营养因子(BDNF)和酪氨酸受体激酶B(Trk B)mRNA及蛋白表达的影响。方法以120只昆明白小鼠作为实验动物,其中7日龄(P7)、21日龄(P21)和42日龄(P42)小鼠各40只;再将同日龄小鼠随机分为Ibo组(n=15)、Ibo+EPO组(n=15)和对照组(n=10)。Ibo组采用微量注射法向左侧海马注射Ibo 1μL;Ibo+EPO组注射Ibo 1μL后,腹腔注射5 000 U/(kg.d)EPO,连续5 d;对照组注射等体积生理盐水。各组小鼠在建模后第5天进行Y-电迷宫测试;Cresyl Violet染色观察海马神经元病理学变化;Real-Time PCR检测海马BDNF mRNA和TrkB mRNA的表达;ELISA法检测BDNF和TrkB蛋白的表达。结果术后Ibo组小鼠寻找安全区的正确率明显低于对照组(P<0.05),而EPO+Ibo组小鼠寻找安全区的正确率明显高于Ibo组(P<0.05)。光学显微镜观察结果显示:Ibo组小鼠海马组织神经元发生明显变性和细胞死亡。Ibo+EPO组和Ibo组海马神经元死亡率明显高于对照组(P<0.05);而Ibo+EPO组损伤较轻。Ibo组和Ibo+EPO组海马组织BDNF和TrkB的mRNA及蛋白表达量均显著高于对照组(P<0.05);其中,Ibo+EPO组BDNF和TrkB的mRNA及蛋白表达量均显著高于Ibo组(P<0.05)。结论 Ibo致脑损伤时,海马组织BDNF和TrkB mRNA及蛋白表达均明显增强,可能是一种保护性反应。EPO可减轻Ibo所致脑损伤,其机制可能与上调BDNF和TrkB的mRNA及蛋白的表达有关。

Objective To investigate the effects of erythropoietin（EPO） on expression of brain-derived neurotrophic factor（BDNF） and tyrosine receptor kinase B（TrkB）mRNA and protein after brain injury induced by ibotenic acid（Ibo） in mice of different developmental stages. Methods One hundred and twenty KM mice aged 7 d（n=40）,21 d（n=40） and 42 d（n=40） were selected,and those of the same age were randomly divided into Ibo group（n=15）,Ibo＋EPO group（n=15） and control group（n=10）,respectively.In Ibo group,1 μL Ibo was injected into left hippocampus.In Ibo＋EPO group,intraperitoneal injection of 5 000 U/（kg·d）EPO was performed for 5 consecutive days after injection of 1 μL Ibo into left hippocampus.Mice in control group were treated with same amount of normal saline.Y-maze tests were carried out 5 d after model establishment in each group,the pathological changes of neurons in hippocampus were observed with Cresyl Violet staining,the expression of BDNF mRNA and TrkB mRNA in hippocampus was detected by Real-Time PCR,and the expression of BDNF protein and TrkB protein was determined by ELISA. Results After operation,the percentage of mice entering safe arm in Ibo group was significantly lower than that in control group（P0.05）,while the percentage of mice entering safe arm in EPO＋Ibo group was significantly higher than that in Ibo group（P0.05）.Light microscopy revealed that degeneration and cell death of neurons in hippocampus occurred in Ibo group,and the death rates of neurons in hippocampus in Ibo＋EPO group and Ibo group were significantly higher than that in control group（P0.05）. However,the pathological changes of Ibo＋EPO group were more moderate.The expression of BDNF and TrkB mRNA and protein in Ibo group and Ibo＋EPO group was significantly higher than that in control group（P0.05）,and the expression of BDNF and TrkB mRNA and protein in Ibo＋EPO group was significantly higher than that in Ibo group（P0.05）. ConclusionThe expression of BDNF and TrkB mRNA and protein is significantly up-regulated in hippocampus of mice with Ibo-induced brain injury,which may be a protective reaction.EPO mitigates brain injury induced by Ibo,the mechanism of which may be related to the up-regulation of expression of BDNF and TrkB mRNA and protein.