脊髓Treg介导的BDNF/TrkB-ASIC1a信号通路参与大鼠带状疱疹后遗神经痛的分子机制研究

Post herpetic neuralgia (PHN) is one of the most common causes in elderly individuals with chronic neuropathic pain. The pathogenesis of PHN remains largely unknown and treatment options are very limited. In the central nervous system (CNS), regulatory T cell (Treg) could reduce the inflammation of microglia . However, there were few reports about the mechanism of Treg-mediated immunity in CNS. Our previous results show that Brain-derived neurotrophic factor/ Tyrosine kinase receptor B (BDNF/TrkB) is involved in the development of choronic pain. Recent studies in our laboratory indicate that the protein expression of acid-sensing ion channels 1a (ASIC1a) in spinal cord was greatly enhanced in PHN rats. Therefore, our hypothesis is that immune dysfunction of Treg in post herpetic neuralgia leads to the glia cells though BDNF upregulation , then BDNF released within the spinal cord enhances TrkB receptor and ASIC1a signaling on the spinal cord neurons. The present project will combine in vivo and vitro experiments by using flow cytometry,immunohistochemistry,pach-clamp, behavioral test and molecular biology methods to explore the immune mechanisms of spinal Treg via BDNF/trkB-ASIC1a signaling in post herpetic neuralgia in rats. These will provide novel strategy for developing therapeutic drugs targeting the neuro-immune-inflammation network.

带状疱疹后遗神经痛(PHN)是造成中老年人神经病理性疼痛的常见原因之一，其发病机制尚未完全阐明，缺乏有效的治疗手段。已有研究表明中枢神经系统中的调节性T细胞(Treg)可降低胶质细胞炎症反应，然而其具体的分子机制却鲜有报道。我们前期结果表明BDNF/TrkB参与慢性痛的发生与维持，预实验结果发现PHN动物模型的脊髓中枢ASICla蛋白表达显著升高。因此，我们假设Treg功能失调导致胶质细胞激活分泌大量BDNF，释放的BDNF作用于脊髓背角神经元的TrkB受体并激活ASIC1a信号从而导致PHN。本项目将结合在体和离体实验，采用流式细胞技术、组织免疫荧光、电生理膜片钳、行为学以及分子生物学等方法，着重探究脊髓Treg调控BDNF/TrkB-ASIC1a信号参与大鼠带状疱疹后遗神经痛的分子机制，从而为开发以神经-免疫-炎症网络为靶点的镇痛药物提供新思路。

Tropomyosin受体激酶B（TrkB）是脑源性神经营养因子（BDNF）的内源性高亲和力受体，并在初级感觉神经元中大量表达。 有证据表明，BDNF参与由水痘-带状疱疹病毒感染的再激活所引起的疱疹后神经痛（PHN），但是其详细机制尚未明确。此外，酸敏感离子通道3（ASIC3）是参与慢性神经性疼痛的主要因素，主要在感觉神经元中大量表达。本项目旨在研究BDNF / TrkB通过调节背根神经节神经元ASIC3信号参与PHN的分子机制。本研究使用树脂毒素（RTX）建立大鼠PHN模型。我们发现RTX诱发机械性异常性疼痛，上调TrkB，ASIC3，TRAF6，nNOS和c-Fos蛋白表达，并增加DRGs的神经元兴奋性。 ASIC3抑制剂逆转了RTX注射诱导的这些变化，但并未改变BDNF和TrkB蛋白的表达。同时，TrkB抑制剂逆转了RTX注射诱导的所有这些变化。我们进一步发现RTX可以增加BDNF的表达，并上调BDNF与DRGs中神经元或卫星胶质细胞的共表达。此外，外源性BDNF通过PC12细胞中的TrkB受体上调了ASIC3信号传导，增加了NO水平，并增强了IL-6，IL-1β，TNF-α的水平，而shRNA-ASIC3转染逆转了外源性BDNF引起的作用。这些发现表明，抑制BDNF / TrkB可以通过调节ASIC3信号通路降低RTX诱导的机械异常性疼痛，降低炎症因子水平并逆转DRG中的神经元兴奋性，这可能为治疗带状疱疹后神经痛提供潜在的新型治疗靶点。