14,15-EET对氧糖剥夺/再灌注模型中星形胶质细胞分泌BDNF及共培养体系中神经元凋亡的影响

Effect of 14,15-EET on Release of BDNF from Astrocytes and Neuronal Apoptosis in a Co-culture System after Oxygen-glucose Deprivation and Reperfusion

目的通过外源性应用14,15-环氧二十碳三烯酸(14,15-EET)干预星形胶质细胞,探讨14,15-EET对氧糖剥夺/再灌注(OGD/R)模型中星形胶质细胞分泌脑源性神经营养因子(BDNF)及共培养体系中神经元凋亡的影响。方法采用氧糖剥夺法建立体外神经元与星形胶质细胞缺血损伤模型,于再灌注期使用外源性14,15-EET孵育星形胶质细胞,采用酶联免疫吸附实验(ELISA)检测不同时间点星形胶质细胞分泌BDNF的变化,随后应用Transwell小室和神经元共培养,采用Tunnel染色检测神经元凋亡。结果与对照试剂组相比,14,15-EET可显著刺激OGD/R损伤后星形胶质细胞分泌BDNF,其作用时间可持续至撤药后至少48 h。此外,14,15-EET孵育的OGD星形胶质细胞可减少共培养体系中OGD神经元的凋亡,这种神经保护作用可被BDNF TrkB受体抑制剂k252a部分逆转。结论在OGD损伤的情况下,外源性应用14,15-EET可刺激星形胶质细胞释放BDNF,后者通过与共培养体系中的神经元TrkB受体相结合,激活下游信号通路,发挥神经元保护作用。

Objective To investigate whether post-treatment of cultured astrocytes with 14,15-epoxyeicosatrienoic acids(14,15-EET)would increase the brain derived neurotrophic factor(BDNF)secretion after oxygen-glucose deprivation and reperfusion(OGD/R),and if this effect would subsequently protect neurons during reperfusion after OGD in the co-cultured system.Methods Astrocytes and neurons were subjected to OGD/R.Exogenous 14,15-EET were applied to astrocytes in the reperfusion period and ELISA was then performed to measure BDNF secretion from astrocytes at different time points following OGD/R.After that,the OGD neurons were co-cultured with the astrocytes that were previously incubated with DMSO or 14,15-EET.Tunnel staining was used to detect neuronal apoptosis.Results BDNF secretion was significantly promoted by application of 14,15-EET on astrocytes in the reperfusion stage after OGD.Exposure of OGD neurons to astrocyte media previously conditioned with 14,15-EET reduced the neuronal apoptosis,but the pro-survival effect could be partly reversed by TrkB inhibitor k252 a.Conclusion Exogenous administration of 14,15-EET augments BDNF secretion from astrocytes,which increases TrkB receptor occupancy on neurons and promotes neuronal survival after OGD/R.