AIBP and regulation of neuropathic pain

项目摘要 持续性疼痛状态，由炎性疾病引起，如关节炎、糖尿病、HIV和 化疗等对生活质量有非常负面的影响。其中一个共同特点是 启动事件是释放损伤相关分子模式（DAMP）分子， Toll样受体-4（TLR 4）。我们以前的研究表明，TLR 4在介导从 急性至持续性疼痛。TLR 4以及其他炎症受体定位于细胞膜上的脂筏微区。 质膜富含胆固醇和鞘磷脂的脂筏促进配体介导的受体 二聚化和下游信号传导。从质膜上去除胆固醇减少脂筏 并且经常导致受体功能的抑制。我们发现，分泌型apoA-I结合蛋白（AIBP） 加速胆固醇清除，破坏脂筏，阻止TLR 4二聚化，抑制小胶质细胞 LPS的炎症反应。此外，由于AIBP与TLR 4结合，当TLR 4 被激动剂激活，我们建议AIBP将胆固醇清除靶向于含有激活的 TLR4。在我们最近的工作中，我们还发现这一机制与神经病理性疼痛的调节有关 states.鞘内注射重组AIBP可预防LPS诱导的触觉异常性疼痛， 逆转了顺铂诱导的异常性疼痛。基于这些发现，我们建议有针对性的AIBP- 介导的脂筏破坏及其对TLR 4信号传导的影响可能是一种潜在的治疗策略， 治疗神经性疼痛状态。本文的具体目的是：（1）检验AIBP 靶向携带活化TLR 4的脂筏;（2）检验AIBP减少胶质细胞活化的假设， 神经病理性疼痛的小鼠模型中的神经炎症;和（3）鉴定内源性神经炎症的起源和功能。 脊髓中的AIBP。为了验证这些假设，我们提出了实验，在体内和分离的神经胶质细胞， 阐明AIBP-TLR 4结合和脂筏机制，并表征胶质细胞活化和 神经炎症状态。我们还将制备具有突变的apoA-I或TLR 4结合界面的AIBP变体， 验证拟议的机制。使用我们独特的AIBP敲除和AIBP flox/flox小鼠，我们将鉴定 内源性AIBP在脂筏调节和神经炎症中的作用。总之，我们提出的实验 将阐明AIBP减少神经炎症和减轻神经病理性疼痛的机制。我们 研究还表明，提高中枢神经系统中的AIBP水平可能是一种新的治疗方法， 持续性疼痛状态。

Project Summary Persistent pain states, arising from inflammatory conditions, such as in arthritis, diabetes, HIV, and chemotherapy among others, have an extraordinary negative impact on quality of life. A common feature of these initiating events is the release of damage-associated molecular pattern (DAMP) molecules, which can activate Toll-like receptor-4 (TLR4). Our previous studies suggest that TLR4 is critical in mediating the transition from acute to persistent pain. TLR4 as well as other inflammatory receptors localize to lipid raft microdomains on the plasma membrane. Lipid rafts, enriched with cholesterol and sphingomyelin, facilitate ligand-mediated receptor dimerization and downstream signaling. Removal of cholesterol from the plasma membrane reduces lipid rafts and often results in inhibition of receptor function. We have found that the secreted apoA-I binding protein (AIBP) accelerates cholesterol removal, disrupts lipid rafts, prevents TLR4 dimerization and inhibits microglia inflammatory responses to LPS. Furthermore, because AIBP binds to TLR4 and its affinity increases when TLR4 is activated by an agonist, we propose that AIBP targets cholesterol removal to lipids rafts harboring activated TLR4. In our recent work, we have also found that this mechanism is relevant to regulation of neuropathic pain states. Intrathecal injections of recombinant AIBP prevented LPS-induced tactile allodynia and, remarkably, reversed established cisplatin-induced allodynia. Based on these findings, we propose that targeted, AIBP- mediated disruption of lipid rafts and its effects upon TLR4 signaling can be a potential therapeutic strategy in treating neuropathic pain states. The Specific Aims of this proposal are: (1) to test the hypothesis that AIBP targets lipid rafts harboring activated TLR4; (2) to test the hypothesis that AIBP reduces glial activation and neuroinflammation in mouse models of neuropathic pain; and (3) to identify the origin and function of endogenous AIBP in the spinal cord. To test these hypothesis, we propose experiments, in vivo and in isolated glial cells, to elucidate the AIBP-TLR4 binding and lipid raft mechanism, and to characterize glial activation and neuroinflammation states. We will also make AIBP variants with mutated apoA-I or TLR4 binding interfaces to validate the proposed mechanism. Using our unique AIBP knockout and AIBP flox/flox mice, we will identify the role of endogenous AIBP in lipid raft regulation and neuroinflammation. In summary, our proposed experiments will elucidate the mechanisms by which AIBP reduces neuroinflammation and alleviates neuropathic pain. Our studies may also suggest that raising AIBP levels in the CNS may be a novel therapeutic approach to treat persistent pain states.