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为了探讨肿瘤坏死因子-α(tumor necrosis factor-α,TNFα)刺激的星形胶质细胞对神经元的作用,本研究将体外纯化培养的大鼠海马星形胶质细胞,用反相高效液相法测定TNFα刺激后细胞培养液内谷氨酸的含量;将TNFα刺激后的星形胶质细胞条件培养液(astrocytic conditioned medium,ACM)作用于培养的海马神经元,运用免疫细胞化学方法和图像分析技术研究神经元NF-κBp65和谷氨酸的表达。结果表明:(1)TNFα可明显促进星形胶质细胞释放谷氨酸,(2)ACM作用1 5 min即可诱导神经元NF-κBp65的核表达,30 min达高峰,180 min恢复至对照水平,(3)ACM作用60 min可使谷氨酸免疫反应阳性神经元平均光密度明显升高,持续至240 min。提示,TNFα刺激的星形胶质细胞可通过释放谷氨酸等可溶性物质使神经元快速激活、兴奋性升高。
In order to investigate the effects of tumor necrosis factor-α (TNFα) -stimulated astrocytes on neurons, in vitro cultured hippocampal astrocytes were cultured in vitro and purified by RP-HPLC Phase method to determine the content of glutamate in the cell culture solution stimulated by TNFα. The astrocytic conditioned medium (ACM) stimulated by TNFα was used to culture hippocampal neurons. Immunocytochemistry and immunocytochemistry Image analysis techniques were used to study the expression of NF-κBp65 and glutamate in neurons. The results showed that: (1) TNFα significantly promoted the release of glutamate from astrocytes; (2) The expression of NF-κBp65 in neurons was induced by ACM for 15 min, reaching the peak at 30 min and returned to the control at 180 min (3) 60 minutes after ACM treatment, the mean optical density of glutamate immunoreactive neurons increased significantly, reaching 240 min. Tip, TNFα-stimulated astrocytes through the release of soluble substances such as glutamate neurons rapid activation, increased excitability.