Study on the molecular mechanism by which microglia transform into phagocytes

小胶质细胞转化为吞噬细胞的分子机制研究

• 批准号: 15500269
• 负责人: NAKAJIMA Kazuyuki
• 金额: $ 1.86万
• 依托单位: Soka University
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2003
• 资助国家: 日本
• 起止时间: 2003 至 2005
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/en/grant/KAKENHI-PROJECT-15500269/
• 关键词: Microgila; Macrophages; Phagocytes; Neuronal cell death; Neurobiology; 貪食; 細胞死; 顔面神経; 貧食細胞

Little has been known about the mechanism by which resting microglia change into active phagocytes. Therefore, I aimed to identify the specific molecule(s) associated with the induction of phagocytic activity in activated microglia.In the first year, we tried to establish an animal model of neuronal cell death induced by the injection of neurotoxic lectin RCA6O into facial nerve. However, the experiment was not carried out because RCA60 was not available in Japan. Instead, we used hydrogen peroxide or peroxinitrite to induce neuronal cell death. These radicals were found to be effective for injuring motoneurons in facial nucleus.In the second year, the molecules associated with the induction of phagocytic activity were explored by subtraction method in the hydrogen peroxide-injected facial nucleus. The subtraction method revealed that some molecules are induced in hydrogen peroxide-injected facial nucleus, but the most of the molecules belongs to proteases, other enzymes and cytoskelet … More ons. Despite the repeated experiments, we could not find out candidate molecules for phagocytic activity-related proteins. Apart from the in vivo model, we tried to prepare non-phagocytic microglia in vitro, and finally succeeded to do it by treating microglia with phorbol myristate acetate (PMA). Thus, it became possible to compare the proteins between phagocytic microglia and non-phagocytic microglia.In the last year, we continued to analyze the phagocytes-related proteins by subtraction method. However, the results were not significant. On the other hand, the comparison of proteins between phagocytic microglia and non-phagocytic microglia indicated that a molecule with molecular weight of approximately 90 kDa and pl 4.8 is present in phagocytic microglia. Therefore, the molecule (p90) was tried to be isolated from the cultured microglia. The microglial homogenate was applied to DEAE Sephadex, Hydroxyapatite, Sephacryl S200 column chromatographies, and subsequently SDS polyacrylamide gel electrophoresis/transblotting. Finally, p90 on PVDF membrane was analyzed for N-terminal amino acid sequence. The resultant sequence was Asp-Asp-Glu-Val-----(not opened). These results including the identification and the amino acid sequence of p90 remain to be confirmed. Less

关于静息小胶质细胞转变为活跃吞噬细胞的机制知之甚少。因此，我的目的是确定特定的分子（S）与诱导的吞噬活性在活化的小胶质细胞。在第一年，我们试图建立一个动物模型的神经细胞死亡的神经毒性凝集素RCA 6 O注射到面神经。然而，由于在日本没有RCA 60，因此没有进行实验。相反，我们使用过氧化氢或过氧亚硝酸盐诱导神经元细胞死亡。第二年，在注射过氧化氢的面神经核中，用减法探索与吞噬活性诱导相关的分子。差减法显示，注入过氧化氢后，面神经核内有部分分子被诱导，但大部分分子属于蛋白酶、其它酶和细胞色素 ...更多信息 一个。尽管反复实验，我们无法找到吞噬活性相关蛋白的候选分子。除了体内模型外，我们还尝试在体外制备非吞噬性小胶质细胞，并最终成功地用佛波醇肉豆蔻酸酯（PMA）处理小胶质细胞。因此，可以比较吞噬性小胶质细胞和非吞噬性小胶质细胞之间的蛋白质。在过去的一年中，我们继续通过减法分析吞噬细胞相关蛋白质。然而，结果并不显着。另一方面，吞噬性小胶质细胞和非吞噬性小胶质细胞之间的蛋白质比较表明，吞噬性小胶质细胞中存在分子量约为90 kDa且pI 4.8的分子。因此，试图从培养的小胶质细胞中分离分子（p90）。将小胶质细胞匀浆应用于DEAE Sephadex、羟基磷灰石、Sephacryl S200柱层析，随后进行SDS聚丙烯酰胺凝胶电泳/transblotting。最后，分析PVDF膜上的p90的N-末端氨基酸序列。所得序列为Asp-Asp-Glu-Val-（未打开）。这些结果包括p90的鉴定和氨基酸序列仍有待证实。少

项目成果

Nonparticipation of nuclear factor kappa B (NFκB) in the signaling cascade of c-Jun N-terminal kinase (JNK)-and p38 mitogen-activated protein kinase (p38MAPK)-dependent tumor necrosis factor alpha (TNFα) induction in lipopolysaccharide(LPS)-stimulated mic

核因子 kappa B (NFκB) 不参与脂多糖 (LPS) 中 c-Jun N 末端激酶 (JNK) 和 p38 丝裂原激活蛋白激酶 (p38MAPK) 依赖性肿瘤坏死因子 α (TNFα) 诱导的信号级联- 刺激麦克风

• 发表时间: 2006
• 期刊: Brain Res. 1073-1074
• 作者: Uesugi M;Uesugi M
• 通讯作者: Uesugi M

Enhancement of urokinase-type plasminogen activator (uPA) secretion, but not that of substrate plasminogen (PGn), by rat microglia stimulated with neuronal conditioned medium.

用神经元条件培养基刺激大鼠小胶质细胞，增强尿激酶型纤溶酶原激活剂 (uPA) 的分泌，但不增强底物纤溶酶原 (PGn) 的分泌。

• 发表时间: 2005
• 期刊: Neurosci Lett. 378
• 作者: Uesugi M;Uesugi M;Uesugi M;Nakajima K;Nakajima K;Nakajima K;Nakajima K;Nakajima K;Nakajima K
• 通讯作者: Nakajima K

Differential suppression of endotoxin-inducible inflammatory cytokines by nuclear factor kappa B (NFκB) inhibitor in rat microglia

• DOI: 10.1016/j.neulet.2006.03.014
• 发表时间: 2006-07-03
• 期刊: NEUROSCIENCE LETTERS
• 影响因子: 2.5
• 作者: Nakajima, K;Matsushita, Y;Kurihara, T
• 通讯作者: Kurihara, T

Nakajima K: "Suppression of lipopolysaccharide-dependent tumor necrosis factor α (TNFα) induction in rat microglia in which protein kinase Cα (PKCα) is downregulated"Neurosci.Lett.. 343・1. 33-36 (2003)

Nakajima K：“蛋白激酶 Cα (PKCα) 下调的大鼠小胶质细胞中脂多糖依赖性肿瘤坏死因子 α (TNFα) 诱导的抑制”Neurosci.Lett. 343・1 (2003)。

Nonparticipation of nuclear factor kappa B (NFκB) in the signaling cascade of c-Jun N-terminal kinase (JNK)- and p38 mitogen-activated protein kinase (p38MAPK)-dependent tumor necrosis factor alpha (TNFα) induction in lipopolysaccharide (LPS)-stimulated m

核因子 kappa B (NFκB) 不参与脂多糖 (LPS) 中 c-Jun N 末端激酶 (JNK) 和 p38 丝裂原激活蛋白激酶 (p38MAPK) 依赖性肿瘤坏死因子 α (TNFα) 诱导的信号级联-刺激m

• 发表时间: 2006
• 期刊: Brain Res. 1073-1074
• 作者: Uesugi M;Uesugi M;Uesugi M
• 通讯作者: Uesugi M