Burnett School of Biomedical Sciences, College of Medicine, University of Central Florida, 4000 Central Florida Blvd, Orlando, FL 32816, USA.
Stroke. 2011 Sep;42(9):2589-94. doi: 10.1161/STROKEAHA.111.614834. Epub 2011 Jul 7.
Activation of Notch worsens ischemic brain damage as antisense knockdown or pharmacological inhibition of the Notch pathway reduces the infarct size and improves the functional outcome in a mouse model of stroke. We sought to determine whether Notch activation contributes to postischemic inflammation by directly modulating the microglial innate response.
The microglial response and the attendant inflammatory reaction were evaluated in Notch1 antisense transgenic (Tg) and in nontransgenic (non-Tg) mice subjected to middle cerebral artery occlusion with or without treatment with a γ-secretase inhibitor (GSI). To investigate the impact of Notch on microglial effector functions, primary mouse microglia and murine BV-2 microglial cell line were exposed to oxygen glucose deprivation or lipopolysaccharide in the presence or absence of GSI. Immunofluorescence labeling, Western blotting, and reverse-transcription polymerase chain reaction were performed to measure microglial activation and production of inflammatory cytokines. The nuclear translocation of nuclear factor-κB in microglia was assessed by immunohistochemistry. The neurotoxic potential of microglia was determined in cocultures.
Notch1 antisense mice exhibit significantly lower numbers of activated microglia and reduced proinflammatory cytokine expression in the ipsilateral ischemic cortices compared to non-Tg mice. Microglial activation also was attenuated in Notch1 antisense cultures and in non-Tg cultures treated with GSI. GSI significantly reduced nuclear factor-κB activation and expression of proinflammatory mediators and markedly attenuated the neurotoxic activity of microglia in cocultures.
These findings establish a role for Notch signaling in modulating the microglia innate response and suggest that inhibition of Notch might represent a complementary therapeutic approach to prevent reactive gliosis in stroke and neuroinflammation-related degenerative disorders.
Notch 的激活会加重缺血性脑损伤，而反义寡核苷酸敲低或 Notch 通路的药理学抑制可减少梗死面积并改善中风小鼠模型的功能预后。我们试图通过直接调节小胶质细胞固有反应来确定 Notch 激活是否会导致缺血后炎症。
Notch1 反义转基因（Tg）和非转基因（非-Tg）小鼠接受大脑中动脉闭塞（MCAO），并用γ-分泌酶抑制剂（GSI）治疗或不治疗，评估小胶质细胞反应和伴随的炎症反应。为了研究 Notch 对小胶质细胞效应功能的影响，将原代小鼠小胶质细胞和小鼠 BV-2 小胶质细胞系暴露于氧葡萄糖剥夺或脂多糖存在或不存在 GSI 的情况下。通过免疫荧光标记、Western blot 和逆转录聚合酶链反应测量小胶质细胞激活和炎症细胞因子的产生。通过免疫组织化学评估小胶质细胞核因子-κB 的核易位。在共培养物中测定小胶质细胞的神经毒性潜能。
Notch1 反义小鼠的同侧缺血皮质中活化的小胶质细胞数量明显低于非-Tg 小鼠，促炎细胞因子的表达也减少。 Notch1 反义培养物和用 GSI 处理的非-Tg 培养物中的小胶质细胞激活也减弱。GSI 显著降低了核因子-κB 的激活和促炎介质的表达，并显著减弱了共培养物中小胶质细胞的神经毒性活性。
这些发现确立了 Notch 信号在调节小胶质细胞固有反应中的作用，并表明抑制 Notch 可能代表预防中风和神经炎症相关退行性疾病中反应性神经胶质增生的一种互补治疗方法。
