mGluR5 inhibits microglial activation and neuronal cell death after TBI

描述(申请人提供)：创伤性脑损伤(TBI)导致神经细胞死亡，合并星形胶质细胞增殖和炎症与小胶质细胞激活相关，这有助于不可逆转的组织损伤。这种继发性伤害在侮辱后几秒钟到几分钟内开始，可能会持续几天、几周，甚至可能持续几个月到几年。我们实验室的最新证据显示，中枢神经系统的慢性炎症在受伤后至少持续6个月。炎症可能会导致创伤后的慢性神经退化，以及阿尔茨海默病(AD)或帕金森综合症等疾病。我们实验室的初步研究表明，炎症可能通过激活小胶质细胞中的mGluR5受体来抑制。此外，我们的数据表明，mGluR5的作用可能是通过对NADPH氧化酶的作用来介导的，该酶作用于小胶质细胞产生活性氧物种(ROS)，并可能在小胶质细胞损伤后的持续激活中发挥重要作用。建议的研究旨在解决下列假设：(1)抑制NADPH氧化酶可减少脑损伤后小胶质细胞的激活、促炎因子的产生和相关的神经功能障碍；(2)mGluR5刺激可减弱小胶质细胞的激活和继发性神经细胞死亡，并改善脑损伤后的功能结果；(3)mGluR5而不是mGluR1调节小胶质细胞的激活和神经毒性，部分是通过抑制NADPH氧化酶来实现的；(4)G1q蛋白信号转导通路是mGluR5信号转导事件的关键组成部分，导致抑制小胶质细胞NADPH氧化酶和小胶质细胞的激活。其具体目的是证明：(1)NADPH氧化酶在脑损伤后小胶质细胞激活及相关促炎因子产生中的重要性；(2)与mGluR5整体敲除动物相比，mGluR5刺激可减弱脑损伤后小胶质细胞的激活，减少神经细胞死亡，并改善功能结局；mGluR5全局敲除动物显示出更大的小胶质细胞激活和神经细胞死亡。我们还将利用条件/诱导基因敲除小鼠来区分小胶质细胞和神经元mGluR5的相对作用；(3)在多种小胶质细胞刺激的细胞培养模型中，mGluR5而不是mGluR1调控小胶质细胞激活和神经毒性；(4)利用体外小胶质细胞模型，G1q-蛋白质信号转导通路由mGluR5刺激启动，并且通过防止NADPH氧化酶复合体的激活而对于减弱小胶质细胞活性至关重要。公共卫生相关性全国每年有100万起创伤性脑损伤(TBI)事件。脑外伤患者有证据表明，损伤后小胶质细胞激活时间延长，至少持续3周。在这项拟议的工作中，我们将研究小胶质细胞和神经元mGluR5的神经保护作用，这将为脑外伤的新治疗方法提供初步研究。

DESCRIPTION (provided by applicant): Traumatic brain injury (TBI) causes neuronal cell death, combined with astroglial proliferation and inflammation associated with the activation of microglia, which contributes to irreversible tissue damage. Such secondary injury begins within seconds to minutes after the insult and may continue for days, weeks and potentially even months to years. Recent evidence from our laboratory shows chronic inflammation in the CNS lasting at least 6 months after injury. Inflammation may contribute to chronic neurodegeneration after trauma as well as to disorders such as Alzheimer's disease (AD) or Parkinson's syndrome. Pilot studies in our laboratory have indicated that inflammation may be inhibited by activation of mGluR5 receptors in microglia. Furthermore, our data suggest that mGluR5 effects may be mediated by actions on the NADPH oxidase enzyme, which functions to produce reactive oxygen species (ROS) by microglia and may play a significant role in persistent activation of microglia following injury. The proposed studies are intended to address the following hypotheses: (1) NADPH oxidase inhibition reduces microglial activation, production of pro-inflammatory factors and associated neurological dysfunction after TBI; (2) mGluR5 stimulation attenuates microglia activation and secondary neuronal cell death, and improves functional outcomes after TBI; (3) mGluR5, but not mGluR1, modulates microglial activation and neurotoxicity in primary microglia cultures, a microglial cell line, and microglia/neuronal co-cultures, in part through inhibition of NADPH oxidase; and (4) the G1q-protein signal transduction pathway is the critical component of the mGluR5 signal transduction events leading to inhibition of microglial NADPH oxidase and suppression of microglial activation. Specific aims are to demonstrate: (1) the importance of NADPH oxidase in microglial activation and correlated production of pro-inflammatory factors in the chronic neuronal cell loss and associated neurological dysfunction after TBI; (2) that mGluR5 stimulation attenuates microglial activation, decreases neuronal cell death and improves functional outcomes after TBI in contrast to mGluR5 global knockout animals that show greater microglial activation and neuronal cell death. We will also distinguish the relative roles of microglial versus neuronal mGluR5 using conditional/inducible knockout mice; (3) that mGluR5, but not mGluR1, modulates microglial activation and neurotoxicity in multiple cell culture models of microglial stimulation; and (4) that the G1q-protein signal transduction pathway is initiated by mGluR5 stimulation and is critical for attenuation of microglial activity by preventing activation of the NADPH oxidase enzyme complex, using in vitro microglial cell models. PUBLIC HEALTH RELEVANCE There are 1 million incidences of traumatic brain injury (TBI) per year nationally. Patients with TBI demonstrate evidence of prolonged microglial activation that lasts at least 3 weeks post-injury. The proposed work, in which we will investigate the neuroprotective effects of microglial and neuronal mGluR5, will provide initial investigation to a novel therapeutic approach for TBI.