Fn14, non-canonical NF-kappaB and downstream signaling in neuropathic pain

背景：正在进行的阿片类药物流行在一定程度上是由合法使用为 神经性疼痛。使用非成瘾药物控制疼痛的替代途径是主要目标 神经病理性疼痛研究。周围神经损伤后细胞因子和趋化因子表达上调 在中枢，它们在神经病理性疼痛的发病机制中起到了机械作用。反应性星形胶质细胞 背角表现出一种慢性激活的促炎分泌(CAPS)表型，其特征是 分泌多种因子，包括白细胞介素6(IL-6)、趋化因子C-C基序配体2(CCL2)、趋化因子 C-X-C基序配体1(CXCL1)和神经生长因子(NGF)。PNI后星形细胞帽的表型 通过神经元趋化因子受体促进炎症和神经元过度激活，导致 神经性疼痛。已知规范的NF-κB信号在其中扮演着重要的角色，但令人惊讶的是，尽管其 记录了包括大脑在内的大多数器官在多种炎症条件下的重要性，非 规范的NF-κB信号通过p52：RelB异源二聚体在PNI中尚未被发现。新作品 来自我们实验室的新发现表明，非典范的NF-κB信号通过肿瘤坏死 因子样弱诱导因子/成纤维细胞生长因子诱导14(Fn14)轴是突出的 PNI后，尤其是在背角星形胶质细胞中，可能与Sur1-1的转录表达有关 Trpm4在这些细胞中的表达。此外，我们的初步数据表明，胶质细胞Sur1-Trpm4在 调节IL-6、CCL2和CXCL1的表达。我们的中心假设是，在背角星形胶质细胞中 PNI后，TWEAM诱导的非规范的NF-κB信号是Sur1-Trpm4表达的上游调节因子， 而Sur1-Trpm4反过来调节下游效应物IL-6、CCL2和CXCL1的表达， 可促进慢性神经炎、神经元过度激活和神经病理性疼痛。 描述：这个项目有三个机械目标。在目标1中，Genetic(TWINE-/-，Fn14-/-，P100-/-， AbcC8-/-，全局的或GFAP特异的或GFAP和位点特异的)和药理的(抗TWINE抗体， 优降糖)将进行实验，以确定非典型性NF-κB和Sur1在 两种神经病理性疼痛模型：坐骨神经套叠和坐骨神经痛模型中坐骨神经与背角胶质细胞GFAP的表达 N.暴露于HIV/gp120蛋白。在目标2中，我们将扩展来自染色质的体内和体外数据 免疫沉淀法研究非典型性核因子-κB在功能性Sur1-Trpm4表达中的作用 频道。在目标3中，我们将证实和扩展体内和体外数据，以确定Sur1- Trpm4调节钙/钙调神经磷酸酶依赖的IL-6、CCL2和CXCL1转录这个项目是第一个 研究神经病理性疼痛中背角和坐骨神经胶质细胞中非典型的核因子-κB，将有助于识别新的 使用非成瘾性药物缓解神经病理性疼痛的可用药靶点。

BACKGROUND: The ongoing opioid epidemic is driven in part by legitimate use of opioids prescribed for neuropathic pain. Targeting alternative pathways for pain control using non-addicting drugs is a major goal of neuropathic pain research. After peripheral nerve injury (PNI), cytokines and chemokines are upregulated centrally, where they contribute mechanistically to the pathogenesis of neuropathic pain. Reactive astrocytes in the dorsal horn exhibit a chronically activated, pro-inflammatory secretory (CAPS) phenotype characterized by secretion of numerous factors, including interleukin-6 (IL-6), chemokine C-C motif ligand 2 (CCL2), chemokine C-X-C motif ligand 1 (CXCL1) and nerve growth factor (NGF). The post-PNI astrocytic CAPS phenotype contributes to both inflammation and neuronal hyperactivation via neuronal chemokine receptors, leading to neuropathic pain. Canonical NF-κB signaling is known to play a crucial role, but surprisingly, despite its documented importance in numerous inflammatory conditions involving most organs including the brain, non- canonical NF-κB signaling via the p52:RelB heterodimer has not been identified previously in PNI. New work from our laboratory demonstrates the novel finding that non-canonical NF-κB signaling by the tumor necrosis factor-like weak inducer of apoptosis (TWEAK) / fibroblast growth factor-inducible 14 (Fn14) axis is prominent after PNI, especially in dorsal horn astrocytes, and may be responsible for transcriptional expression of Sur1- Trpm4 in these cells. Moreover, our preliminary data suggest that glial Sur1-Trpm4 plays a crucial role in regulating the expression of IL-6, CCL2 and CXCL1. Our central hypothesis is that, in dorsal horn astrocytes post-PNI, TWEAK-induced non-canonical NF-κB signaling is an upstream regulator of Sur1-Trpm4 expression, and that Sur1-Trpm4, in turn, regulates the expression of the downstream effectors, IL-6, CCL2 and CXCL1, which promote chronic neuroinflammation, neuronal hyperactivation and neuropathic pain. DESCRIPTION: This project has three mechanistic aims. In Aim 1, genetic (Tweak–/–, Fn14–/–, p100–/–, Abcc8–/–, either global or GFAP-specific or GFAP-&-site-specific) and pharmacological (anti-TWEAK antibody, glibenclamide) experiments will be carried out to determine the role of non-canonical NF-κB and Sur1 in GFAP-expressing glia in sciatic n. vs. dorsal horn in two neuropathic pain models: sciatic n. cuffing and sciatic n. exposure to HIV/gp120 protein. In Aim 2, we will expand upon in vivo and in vitro data from chromatin immunoprecipitation to establish the role of non-canonical NF-κB in the expression of functional Sur1-Trpm4 channels. In Aim 3, we will corroborate and expand upon in vivo and in vitro data to establish the role of Sur1- Trpm4 in regulating Ca2+/calcineurin-dependent transcription of IL-6, CCL2 and CXCL1. This project is the first to study non-canonical NF-κB in dorsal horn vs. sciatic n. glia in neuropathic pain, and will help identify novel druggable targets to ameliorate neuropathic pain using non-addictive drugs.