艾灸调控NLRP3炎症小体及相关信号通路治疗克罗恩病的作用机制研究

As a kind of inflammatory bowel disease, Crohn’s disease (CD) has been universally considered an intractable disease and its incidence has been increasing in China. It’s been confirmed that moxibustion is effective in treating mild-to-moderate CD. NLRP3 inflammasome has been revealed closely related to the pathogenesis of CD. Based on the previous work, targeting NLRP3 inflammasome and its upstream and downstream signaling pathways, this study was to discuss the relationship between the NLRP3/ASC/caspase-1 signaling pathway and the development of CD, as well as the intervention effect of moxibustion; also to observe the regulatory effect of moxibustion on the positive control elements P2X7R, ROS, and cathepsin B, and explore the change of mitophagy/NLRP3 inflammasome in intestinal mucosal epithelial cells and the intervention of moxibustion, for proving that the NLRP3/ASC/caspase-1 signaling pathway is closely related to the development of CD, and moxibustion reduces intestinal inflammation and restores intestinal homeostasis via regulating the upstream positive control molecules P2X7R, ROS, and cathepsin B of NLRP3/ASC/caspase-1 and the negative control of mitophagy in colonic mucosa, inhibiting the hyperactivation of NLRP3 inflammasome, and down-regulating the hyper-expression of IL-1β and IL-18 in intestinal mucosa.

克罗恩病(CD)属炎症性肠病,是世界公认的疑难病症,国内发病率日趋上升,艾灸治疗轻中型CD疗效确切。研究证实NLRP3炎症小体与CD发病关系密切。基于前期研究成果,本项目以NLRP3炎症小体及其上下游信号通路为切入点,探讨NLRP3/ASC/caspase-1通路与CD发生发展的关系及艾灸的干预作用;并观察艾灸对ATP/P2X7R、ROS、cathepsin B炎症小体正调控途径的调节作用,探讨线粒体自噬/NLRP3炎症小体在肠黏膜上皮细胞的变化与艾灸的干预效应,拟证实NLRP3/ASC/caspase-1通路与CD发生发展密切相关,艾灸通过调节CD肠黏膜NLRP3/ASC/caspase-1上游正调控途径(P2X7R、ROS、cathepsin B)和线粒体自噬负调控途径,抑制NLRP3炎症小体过度活化,进而减少肠黏膜IL-1β和IL-18过度表达,达到减轻肠道炎症、恢复肠道稳态之目的。

克罗恩病(CD)是世界公认的疑难病症,艾灸治疗轻中型CD疗效确切。NLRP3炎症小体与CD发病关系密切,本项目以NLRP3炎症小体及其上下游信号通路为切入点探讨艾灸治疗CD的抗炎免疫机制。结果发现：①隔药灸能改善CD大鼠结肠病理损伤,缓解炎症反应,下调结肠NLRP3炎症小体(NLRP3、ASC、caspase-1)及其下游炎症因子IL-1β、IL-18的表达；提示抑制结肠NLRP3炎症小体的过度活化可能是隔药灸治疗CD的作用机制之一。②隔药灸能降低CD大鼠结肠ATP含量,下调P2X7R、Pannexin-1、NF-κB p65蛋白及mRNA表达；降低CD大鼠结肠ROS含量,下调TXNIP、TRX蛋白及mRNA表达；提示隔药灸可能通过下调CD大鼠结肠ATP/P2X7-Pannexin-1和ROS-TXNIP信号通路,进而抑制下游NLRP3炎症小体的异常活化发挥治疗作用。③隔药灸能调节CD大鼠结肠自噬相关蛋白LC3II、Beclin-1的表达,抑制自噬活性；隔药灸能上调CD大鼠结肠mTOR磷酸化蛋白及mRNA表达,抑制ULK1、ATG13蛋白及mRNA表达；提示隔药灸可能通过调控mTOR/ULK1/ATG13信号通路抑制结肠自噬活性,进而下调NLRP3、IL-1β的表达发挥抗炎作用。④隔药灸能上调CD大鼠结肠cAMP、PKA蛋白表达；激活cAMP/PKA信号通路,促进泛素蛋白Ub表达以及NLRP3与Ub共表达,进而降低NLRP3、IL-1β的表达,可能是隔药灸治疗CD的作用机制之一。研究结果从NLRP3炎症小体及其上下游信号通路调控角度揭示了艾灸治疗CD的信号转导机制和关键信息分子,为艾灸治疗CD疗效机制研究提供实验依据与理论基础,有助于推动艾灸疗法在CD等肠腑病症中的进一步应用。