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目的:观察促红细胞生成素(EPO)对谷氨酸(Glu)诱导的大脑皮质神经元损伤的保护作用和核因子-κB(NF-κB)介导的细胞内信号转导机制。方法:采用1日龄Wistar大鼠皮层神经细胞体外原代培养技术,建立Glu兴奋性毒性神经细胞损伤模型。实验分为正常对照组、Glu组、EPO组和吡咯烷二硫氨基甲酸(PDTC,NF-κB活化阻断剂)组。倒置显微镜下观察细胞的形态学改变;MTT法测定细胞存活率;流式细胞术测定细胞凋亡率评价细胞损伤程度。结果:Glu处理后神经细胞形态明显受损,EPO组神经细胞形态趋于正常,PDTC组细胞形态与Glu组相似;与正常对照组相比,Glu组神经细胞存活率明显下降、凋亡率明显升高,差异均有统计学意义(P<0.01);而与Glu组相比,EPO组神经细胞存活率明显增高下降、凋亡率明显下降,差异均有统计学意义(P<0.01)。与EPO组相比,PDTC组细胞存活率明显下降、凋亡率明显升高,差异均有统计学意义(P<0.01),这些变化与Glu组相似。结论:EPO对Glu诱导的大脑皮质神经元损伤的具有保护作用,其胞内信号转导机制涉及到NF-κB的活化。
Objective: To observe the protective effect of erythropoietin (EPO) on glutamate (Glu) -induced cerebral cortical neuron injury and the mechanism of nuclear factor-κB (NF-κB) -mediated intracellular signal transduction. Methods: The primary culture of cortical neurons in Wistar rats was used to establish the model of Glu excitotoxic neuronal injury. The experiment was divided into normal control group, Glu group, EPO group and pyrrolidine dithiocarbamate (PDTC, NF-κB activation blocker) group. The morphological changes of the cells were observed under an inverted microscope. The cell viability was measured by MTT assay. The apoptosis rate was evaluated by flow cytometry to evaluate the cell damage. Results: The morphological changes of nerve cells were obviously impaired after treatment with Glu. The morphology of neurons in EPO group tended to be normal. The morphology of PDTC group was similar to Glu group. Compared with normal control group, the survival rate of neuronal cells in Glu group was significantly decreased and the apoptosis rate was obvious (P <0.01). Compared with Glu group, the survival rate of neurons in EPO group was significantly increased and decreased, the apoptosis rate was significantly decreased, the difference was statistically significant (P <0.01). Compared with the EPO group, the cell viability was significantly decreased and the apoptosis rate was significantly increased in the PDTC group (P <0.01). These changes were similar to those in the Glu group. Conclusion: EPO has a protective effect on Glu-induced neuronal damage in the cerebral cortex. Its intracellular signal transduction mechanism involves the activation of NF-κB.