巨噬样滑膜细胞焦亡激活膝骨关节炎痛敏及“温经活血”外治法干预效应的机制研究

Knee osteoarthritis is a common degenerative disease in orthopaedics which has high disability rate, It yet lacks specific treatment due to the unclear pathogenesis. How to relieve the pain is the most urgent problem needed to be solved. Our previous research preliminarily reveals the pathogenesis that how TRP ion channels mediate cold hyperalgesia and mechanical hyperalgesia of knee osteoarthritis, which can be intervened by external therapy like warming channels activating blood circulation.. The pyroptosis of macrophagocyte was a new caspase-1 dependent death pattern of cells, with release of inflammatory cytokine during its process. In previous studies, we find out that the therapy warming channels activating blood circulation external therapy not only inhibits the up-regulated expression of TRPs in synovial tissues of knee osteoarthritis to a certain degree, but also has a role on regulation of inflammatory cytokine which can activate the TRP ion channels. Through the pre-test, we have detected the markers of pyroptosis of macrophagocyte in synovial tissues of knee osteoarthritis, NLRP3, ASC, caspase-1 provides evidence supporting the hypothesis that the pyroptosis of macrophage-like synoviocytes maybe activate the hyperalgesia of knee osteoarthritis mediated by the activation of TRPs. And the therapy warming channels activating blood circulation may influence this part.. This project intends to unite mature technology like detection on pain-related behaviors of neurobiology, gene protein of molecular biology and electrophysiological property investigation of ion channels in cell membrane, will be applied to test the scientific hypothesis.

膝骨关节炎（KOA）是骨科常见退行性疾病，致残率高。其病理机制尚不明确，治疗方案长期存在争议。如何有效缓解疼痛是临床亟需解决的难题。我们前期研究初步揭示了TRPs通道介导KOA冷痛敏和机械痛敏的病理机制，以及“温经活血”中药外治对其存在干预。. 细胞焦亡是Caspase-1依赖性的细胞促炎死亡模式，其全程均有大量炎性因子释放。前期研究中我们发现“温经活血”中药外治除抑制KOA滑膜组织TRPs上调，对激活TRPs的炎性因子等前驱环节也有干预作用。我们预实验在KOA滑膜组织检测到巨噬细胞焦亡关键激活者：炎性小体的核心组分NLRP3、ASC、caspase-1，高度提示巨噬样滑膜细胞焦亡可激活TRPs介导的KOA痛敏，而温经活血中药外治对此环节存在影响。. 本课题拟应用神经生物学疼痛行为检测和分子生物学基因蛋白检测、细胞膜离子通道的电生理特性检测等成熟技术，验证这一科学假说。

疼痛是KOA最主要的临床症状和亟待解决的难题。本研究团队在上一个已结题的国家自然基金面上项目中阐明：TRPs离子通道家族是KOA疼痛的外周伤害感受器，以类似“开关”作用方式参与KOA疼痛，TRPs活化阈值降低是KOA外周痛敏的重要病理机制之一。冷刺激敏感通道TRPA1、TRPM8，机械刺激敏感通道TRPV1、TRPV4分别介导KOA冷痛敏、机械痛敏，表现为“遇寒则痛”、“遇劳则痛”的临床症状。“温经活血”外治法（代表方药：“易层”贴敷）不仅能降低TRPs的表达量，也对TRPs活化具有明确的阻断效应。然而，疼痛感知是“外周感知-信号形成-神经传导-中枢处理”的复杂过程，KOA疼痛在外周伤害感受器之外的病理环节有进一步研究价值，此外，“温经活血”外治法控制KOA疼痛的多元化药效靶点，亦存在深入的挖掘的必要。.在此基础上，本研究项目聚焦巨噬样滑膜细胞焦亡在背根神经节TRPs通道-冷痛敏/机械痛敏激活中的作用，以及“温经活血”外治法对上述途径的干预效应。研究表明，KOA病理过程中，滑膜组织巨噬细胞存在caspase1依赖的经典途径细胞焦亡，该过程由NLRP3炎症小体活化caspase1介导，释放IL-1β、IL-18等促炎因子形成炎症级联放大，不仅能够推动滑膜炎症、软骨破坏，还进一步活化DRG组织的TRPs通道，参与KOA外周痛敏。“温经活血”外治法能降低KOA滑膜巨噬细胞焦亡相关蛋白的表达水平，抑制细胞焦亡，进而阻断由DRG组织TRPs通道活化介导的KOA外周痛敏。.本研究项目顺利完成了所有既定的研究内容，达成了既定目标，并揭示了滑膜巨噬细胞焦亡相关蛋白、炎症级联风暴、TRPs通道均是“温经活血”外治法的有效干预靶点，阐明了“温经活血”外治法控制KOA滑膜炎症、阻断KOA外周痛敏、缓解KOA疼痛的病理机制，为“温经活血”外治法的临床推广和应用放大提供理论依据。