Study of the molecular mechanism by which microglia are activated in the brain

由于小胶质细胞在体内的激活被认为在很大程度上影响神经元的退化和/或再生状态，我们分析了小胶质细胞在脑内激活的机制。(1)通过对大鼠面神经横断模型的观察，选择脑源性神经营养因子(BDNF)和ATP作为神经源性小胶质细胞激活因子的候选者。体外实验表明，脑源性神经营养因子可促进小胶质细胞分泌纤溶酶原(Pgn)和尿激活酶(UK)，并可引起细胞形态改变，促进Pgn和肿瘤坏死因子α(Tumor NectorForα)的分泌。因此，在体内，神经营养因子和/或ATP从损伤的神经元释放到细胞外间隙，激活小胶质细胞。(2)在抑制小胶质细胞功能的因素中，胶质细胞源性神经营养因子(GDNF)的活性最强。该因子不影响小胶质细胞的存活、形态和增殖活性，但抑制小胶质细胞分泌PGN和UK。GDNF被认为在包括面神经切断在内的脑损伤后期可观察到抑制小胶质细胞激活的作用。(3)采用脂多糖刺激系统建立体外模型，分析小胶质细胞激活诱导的信号转导机制。脂多糖可诱导小胶质细胞释放一氧化氮和肿瘤坏死因子α。这些分泌被特异性蛋白激酶C抑制剂抑制，提示蛋白激酶C与NO和肿瘤坏死因子α的释放有很强的相关性。尽管ERK、JNK和p38等MAP激酶均被内毒素激活，但p38的特异性抑制剂强烈抑制肿瘤坏死因子α的释放，提示p38与肿瘤坏死因子α的释放有关。这些细胞毒因子从激活的小胶质细胞释放出来可能受PKC信号通路和相关的MAP激酶活性的调节。

Since the activation of microglia in vivo has been believed to affect largely the state of neuronal degeneration and/or regeneration, we analyzed the mechanism by which microglia are activated in the brain.(1) Brain-derived neurotrophic factor (BDNF) and ATP were selected as candidates for neuron-derived microglial activation factors from the observation of rat facial nerve transection model. The in vitro study revealed that BDNF enhanced the secretion of plasminogen (PGn) and urokinase (UK), and ATP induced morphological change and enhanced the secretion of PGn and tumor necrosis factor α (TNFα) from microglia. Therefore, it is suggested that neurotrophins and/or ATP are released to extracellular spaces from injured neurons and activate microglia in vivo.(2) Among factors which can suppress microglial functions, glial cell line-derived neurotrophic factor (GDNF) was found to show the strongest activity. This factor did not affect survival, morphology and proliferative activity, but suppressed the secretion of PGn and UK from microglia. GDNF was considered to play a role on the suppression of microglial activation which was observed in later stage of brain injury including facial nerve transection.(3) To analyze the signal transduction mechanism in the induction of microglial activation, lipopolysaccharide (LPS)-stimulating system was used for in vitro model. LPS induced the release of nitric oxide (NO) and TNFα in microglia. These secretions were suppressed by the pretreatment with specific protein kinase C (PKC) inhibitor, suggesting the strong association of PKC with NO and TNFα releases. Although MAP kinases including ERK, JNK and p38 were all activated by the stimulation with LPS, specific inhibitor of p38 inhibited strongly the release of TNFα, suggesting the association of p38 with TNFα release. The release of these cytotoxic factors from activated microglia was suggested to be regulated by PKC signaling pathway and the associated MAP kinase activity.