Molecular Pathways of Pain Generation in Osteoarthritis

描述(申请人提供)：疼痛是骨关节炎(OA)的主要症状，也是美国老年人口行动不便的主要原因之一。我们对慢性疼痛，特别是与骨性关节炎相关的慢性疼痛的机制缺乏了解，这是目前可用的治疗方案普遍无效的原因。缓解严重的骨性关节炎疼痛仍然是一个未得到满足的医疗需求，也是寻求手术干预的一个主要原因。这项建议的长期目标是确定骨性关节炎疼痛的起源和机制，从而能够识别新的靶点，并为骨性关节炎疼痛开发新的治疗方法和生物标志物。使用复杂的慢性进行性膝骨性关节炎小鼠模型，结合最先进的方法和有效的选择性抑制剂，该建议旨在系统地解决这一中心假说：膝骨性关节炎关节中的病理性基质转换慢性激活关节伤害性感受器，导致神经系统的可塑性变化，导致疼痛的开始和持续。目的1通过对实验性进行性骨性关节炎过程中支配周围神经系统(DRG)和背角(中枢神经系统)的变化进行纵向分析，明确趋化因子MCP-1和Fractalkine在慢性骨性关节炎疼痛中的时空作用。为了深入了解骨性关节炎周围神经系统(DRG)的痛觉调制并向中枢神经系统过渡，我们的目标是在进行性骨性关节炎的过程中，以时空的方式记录DRG和DH中细胞亚群激活的分子标志物。进一步，我们将测试干扰MCP-1/CCR2信号对疼痛行为和相关通路的影响。目的2探讨模式识别受体(PRR)及其配体(损伤相关分子模式，DAMPS)在关节伤害性感受器激活和骨关节炎相关疼痛中的作用。我们将使用新的Nav1.8-tdTomato报告小鼠，在DMM术后进行性膝骨性关节炎的过程中，可视化TLR2/4和RAGE在关节伤害性感受器上的表达。我们将以细胞内钙动员和单核细胞趋化蛋白-1释放为结果标记物，评估选定的DAMP激活感觉神经元的能力。为了在这个模型中评估DAMP信号对疼痛的贡献，我们将评估在痛感神经元中缺乏MyD88或TLR4的新型条件性基因敲除小鼠的疼痛相关行为。最后，目标3试图不同地针对骨关节炎关节的病理成分，以评估对疼痛和相关路径的影响。我们的目标是在进展性疾病的不同时间通过药物调节来不同地靶向软骨或软骨下骨，并监测与详细的关节组织病理学相关的疼痛/疼痛通路结果标记物。这将澄清关节中骨性关节炎疼痛的来源和调节。这些目标将导致相当大的收获，了解疼痛是如何在OA中产生和维持的，并为更有针对性和更安全的治疗方法铺平道路。

DESCRIPTION (provided by applicant): Pain is the major symptom in osteoarthritis (OA) and one of the leading causes of impaired mobility in the elderly population in the US. Our lack of understanding of the mechanisms underlying chronic pain in general, and chronic pain associated with OA in particular, accounts for the general ineffectiveness of currently available treatment options. Relief from severe OA pain remains an unmet medical need and a major reason for seeking surgical intervention. The long-term goal of this proposal is to define origins and mechanisms of pain in OA, thus enabling identification of new targets, and development of new therapies and biomarkers for OA pain. Using a sophisticated chronic progressive murine model of knee OA, in combination with state-of-the-art approaches and potent, selective inhibitors, this proposal aims to systematically address the central hypothesis: pathological matrix turnover in the OA joint chronically activates joint nociceptors and causes plasticity changes in the nervous system, resulting in the initiation and the persistence of pain. Aim 1 seeks to define a temporo-spatial role for the chemokines, MCP-1 and fractalkine, in chronic OA pain through longitudinal analysis of changes in the innervating DRG (peripheral nervous system) and dorsal horn (central nervous system) during the course of progressive experimental OA. In order to gain insight into OA pain modulation in the DRG (peripheral nervous system) and transition to the central nervous system, we aim to document molecular markers of activation of cellular subsets in DRG and DH in a temporo-spatial manner over the course of progressive OA. Further, we will test the effect of interfering with MCP-1/CCR2 signaling on pain behaviors and associated pathways. Aim 2 seeks to evaluate the contribution of Pattern Recognition Receptors (PRR) and their ligands (Damage Associated Molecular Patterns, DAMPs) to joint nociceptor activation and OA-associated pain. We will visualize TLR2/4 and RAGE expression on joint nociceptors in the course of progressive knee OA after DMM surgery, using novel Nav1.8-tdTomato reporter mice. We will evaluate the ability of selected DAMPs to activate sensory neurons, using intracellular Ca2+ mobilization and MCP-1 release as outcome markers. In order to assess the contribution of DAMP signaling to pain in this model, we will evaluate pain-related behaviors in novel conditional knock-out mice that lack Myd88 or TLR4 in pain-sensing neurons. Finally, Aim 3 seeks to differentially target pathological components of the OA joint in order to assess effects on pain and associated pathways. We aim to differentially target cartilage or subchondral bone through pharmacological modulation at different times during progressive disease, and monitor pain/pain pathway outcome markers in association with detailed joint histopathology. This will clarify sources and regulation of OA pain in the joint. These aims will lead to considerable gains in understanding how pain is in generated and maintained in OA, and pave the way for a more targeted and safer therapeutic approach.