Cao Qiong, Lu Jia, Kaur Charanjit, Sivakumar Viswanathan, Li Fan, Cheah Pike See, Dheen S Thameem, Ling Eng-Ang
Department of Anatomy, Yong Loo Lin School of Medicine, National University of Singapore, Singapore.
Glia. 2008 Aug 15;56(11):1224-37. doi: 10.1002/glia.20692.
Notch-1 receptor signaling pathway is involved in neuronal and glial differentiation. Its involvement in microglial functions, however, has remained elusive. This study reports the localization of Notch-1 receptor immunoreactivity in the amoeboid microglial cells (AMC) in the postnatal rat brain. By immunofluorescence, Notch-1 receptor was colocalized with its ligands, Jagged-1 and Delta-1, in the AMC in the corpus callosum and subventricular zone. Notch-1 immunopositive cells were confirmed to be microglia labeled by OX42 and lectin. Immunoexpression of Notch-1 receptor was progressively reduced with age. Western blot analysis showed that Notch-1 protein level in the corpus callosum in which the AMC were heavily populated was concomitantly decreased. In postnatal rats challenged with lipopolysaccharide (LPS), Notch-1 receptor immunofluorescence in AMC was noticeably enhanced. Furthermore, Notch-1 protein level in the corpus callosum was increased as revealed by Western blotting analysis. In primary microglial culture treated with LPS, mRNA expression of Notch-1 and its ligand Jagged-1 was upregulated but that of Delta-1 was reduced. The expression pattern of Notch-1 and its ligands was confirmed in murine BV-2 cells. Furthermore, Notch-1 neutralization with its antibody reduced its protein expression. More importantly, neutralization of Notch-1 concomitantly suppressed the mRNA expression of IL-6, IL-1, M-CSF, and iNOS; TNF-alpha, mRNA expression, however, was enhanced. Western blot confirmed the changes of protein level of the above except for IL-6, which remained relatively unaltered. It is concluded that Notch-1 signaling in the AMC and LPS-activated microglia/BV-2 cells modulates the expression of proinflammatory cytokines and nitric oxide.
Notch-1受体信号通路参与神经元和神经胶质细胞的分化。然而，其在小胶质细胞功能中的作用仍不明确。本研究报道了Notch-1受体免疫反应性在新生大鼠脑内阿米巴样小胶质细胞（AMC）中的定位。通过免疫荧光法，Notch-1受体与其配体Jagged-1和Delta-1在胼胝体和脑室下区的AMC中共定位。Notch-1免疫阳性细胞被证实为经OX42和凝集素标记的小胶质细胞。Notch-1受体的免疫表达随年龄增长而逐渐降低。蛋白质印迹分析表明，AMC大量聚集的胼胝体中Notch-1蛋白水平随之降低。在用脂多糖（LPS）攻击的新生大鼠中，AMC中Notch-1受体免疫荧光明显增强。此外，蛋白质印迹分析显示胼胝体中Notch-1蛋白水平升高。在用LPS处理的原代小胶质细胞培养物中，Notch-1及其配体Jagged-1的mRNA表达上调，但Delta-1的mRNA表达降低。在小鼠BV-2细胞中证实了Notch-1及其配体的表达模式。此外，用其抗体中和Notch-1可降低其蛋白表达。更重要的是，中和Notch-1同时抑制了IL-6、IL-1、M-CSF和iNOS的mRNA表达；然而，TNF-α的mRNA表达增强。蛋白质印迹证实了上述蛋白水平的变化，但IL-6相对未改变。结论是，AMC和LPS激活的小胶质细胞/BV-2细胞中的Notch-1信号调节促炎细胞因子和一氧化氮的表达。
