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目的探讨缺氧诱导活化的小胶质细胞在SD大鼠海马神经元缺氧损害中的作用机制。方法建立共培养体系,应用原位缺口末端标记(TUNEL)法、化学发光法探讨不同组别神经元生长状况以及Caspase-3活性;采用免疫荧光法、格里斯试剂法(Griess Reagent)、还原WST-1法、酶联免疫吸咐测定(ELISA)等方法检测各组培养液中NO、O2-以及TNF-α的表达水平。结果缺氧12h,N9细胞培养液可抑制常规培养的神经元生长增殖活力,同时可加重由缺氧抑制的共培养体系中神经元活力;既可诱导常规培养的神经元凋亡,又可促进共缺氧培养的神经元凋亡;较之于单纯神经元培养系和常规共培养系,共缺氧培养系的培养液中NO、O2-、TNF-α3类应激性神经毒性因子产量最高。结论小胶质细胞活化在缺氧诱发的神经元损害中发挥了重要的作用,其活化后产生的神经毒性分子是效应分子。
Objective To investigate the mechanism of hypoxia-induced activated microglial cells in hypoxic damage of hippocampal neurons in SD rats. Methods The co-culture system was established. The neuronal growth status and Caspase-3 activity in different groups were investigated by TUNEL and chemiluminescence. The immunofluorescence and Griess Reagent -1 method and enzyme linked immunosorbent assay (ELISA) were used to detect the expression of NO, O2- and TNF-α in each group. Results After hypoxia for 12 hours, N9 cells could inhibit the growth and proliferation of neurons in routine culture and increase the activity of neurons in the co-culture system inhibited by hypoxia. The N9 cells could induce the apoptosis of neurons in routine culture, Compared with pure neuronal and conventional coculture lines, the total production of NO, O2- and TNF-α3-type stress neurotoxic factors in the culture medium of total anoxic culture was the highest . Conclusion Microglial activation plays an important role in hypoxia-induced neuronal damage. The activated neurotoxic molecules are effector molecules.