MAPK/ERK信号通路在大鼠视神经损伤后小胶质细胞活化中的作用及机制研究

目的 探讨丝裂原激活蛋白激酶(MAPK)/细胞外信号调节蛋白激酶(ERK)信号通路在大鼠视神经损伤后小胶质细胞活化中的作用及机制。方法 将100只成年雄性大鼠随机分为实验组50只和对照组50只,实验组采用荧光金双侧上丘逆行标记视网膜神经节细胞并在培养7d后制作视神经损伤模型,免疫组织化学方法检测2组视网膜铺片小胶质细胞计数情况,应用蛋白免疫印记法检测2组MAPK通路的ERK蛋白表达水平及磷酸化蛋白水平,并应用免疫组化法检测2组视网膜白介素10(IL-10)和肿瘤坏死因子(TNF-α)等表达水平; 向实验组视网膜铺片添加ERK1/2通路阻断剂PD98059,再次检测其IL-10和TNF-α等表达水平及小胶质细胞计数情况。结果 与对照组比较,实验组ERK蛋白表达水平及磷酸化蛋白水平降低,视网膜铺片小胶质细胞计数降低,IL-10阳性率和TNF-α阳性率升高(P<0.05)。与阻断前比较,实验组阻断后6 h的IL-10和TNF-α等表达水平升高,小胶质细胞计数降低(P<0.05)。结论 MAPK/ERK信号通路在大鼠视神经损伤后小胶质细胞活化中具有保护作用,其机制可能通过抑制炎症因子IL-10和TNF-α等表达而减少其视神经损伤并促进小胶质细胞活化。

The role and mechanism of MAPK/ERK signaling pathway in the activation of microglia after optic nerve injury in rats

ObjectiveTo investigate the role of mitogen activated protein kinase(MAPK)/ extracellular signal regulated protein kinase(ERK)signaling pathway in the activation of microglia after optic nerve injury in rats.Methods 100 adult male rats were randomly divided into experimental group(50 rats)and control group(50 rats), the experimental group had Fluorogold retrograde labeling bilateral superior retinal ganglion cells and were made into optic nerve injury model after cultured for 7d. Immunohistochemical method was applied in detecting retinal microglial cell count of two groups. Western blot was applied in detecting MAPK pathway ERK protein and its phosphorylation protein expression level of two groups, and immunohistochemistry was applied in detecting retinal interleukin 10(IL-10)and tumor necrosis factorα(TNF-α)expression level in two groups. The ERK1/2 blocking agent PD98059 was added to the retina of the experimental group, and the IL-10 and TNF-αexpression levels and retinal microglial cell count were detected again.Results Compared with the control group, the ERK protein and its phosphorylation protein expression level in the experimental group were decreased, the retinal microglial cell count of the experimental group was decreased, and the IL-10 positive rate and the TNF-α positive rate of the experimental group were increased(P<0.05). Compared with before obstruction, the IL-10 positive rate and the TNF-α positive rate in the experimental group at 6^th h after obstruction was increased, while retinal microglial cell count of the experimental group in the experimental group at 6^th h after obstruction was decreased(P<0.05).Conclusion MAPK/ERK signaling pathway played a protective role in activated microglia after nerve injury in the rat optic, its mechanism might be through inhibition of inflammatory cytokines IL-10 and TNF-α expression to reduce the optic nerve injury and promote the activation of microglia.