豨桐丸对痛风促炎信号轴NLRP3-IL-1β-NF-κB活化的调控机制研究

In recent years, with the improvement of standard of living and change of diet, the incidence of gout increases rapidly and gout has become a common and frequently-occurring disease in the clinic. There are some defects of anti-gout drugs in current clinical use such as limited efficacy and terrible side effects. Therefore, it is important to explore safe and effective traditional Chinese medicine with the function of anti-gout. XiTong Wan has been empirically used to treat gout, however, its underlying mechanisms remain unclear. In this research, considering the core position of IL-1β in gout inflammation, and the key role of NLRP3 inflammasome on IL-1β production, as well as the crucial effect of nuclear transcriptional factor NF-κB on biological activity of IL-1β, we hypothesize that XiTong Wan inhibits gout inflammation through regulation of the pro-inflammatory signal axis “NLRP3-IL-1β-NF-κB”. Hence, we will: (i) investigate the effect of XiTong Wan on the monosodium urate-induced gouty arthritis in rats. (ii) investigate the effects of XiTong Wan on the monosodium urate-induced NLRP3 inflammasome activation and IL-1β production, as well as IL-1β-induced NF-κB activation in macrophages. (iii) investigate the effect of XiTong Wan on the monosodium urate-stimulated macrophages whose IL-1 receptor gene is silenced by RNA interference method. Depending on all above experiments, we can illuminate the important role of pro-inflammatory signal axis “NLRP3-IL-1β-NF-κB” on the alleviated effect of XiTong Wan for gout. Meanwhile, we can discover the new mechanisms of XiTong Wan in gout treatment and it may contribute to made up prescriptions for gout in the clinic of traditional Chinese medicine.

随着人们生活水平的提高、饮食结构的改变，近年来痛风的发病率不断升高。当前治疗痛风药物存在疗效有限、副作用大等缺陷，故从中医药中探寻安全、有效的治疗痛风方剂意义重大。豨桐丸是治疗痛风的常用方剂，但其药理机制至今不甚清楚。本课题立足于IL-1β在痛风炎症中的核心地位，着眼于NLRP3炎性体在IL-1β成熟表达及NF-κB在IL-1β生物活性中的关键作用，提出“豨桐丸通过调控NLRP3-IL-1β-NF-κB促炎信号轴而拮抗痛风炎症发展”的假说。为此我们拟观察：①豨桐丸对尿酸钠诱导的大鼠痛风性关节炎发展的影响；②豨桐丸对巨噬细胞NLRP3炎性体活化、IL-1β成熟表达以及NF-κB活化的影响。通过以上研究以阐明“NLRP3-IL-1β-NF-κB”促炎信号轴在豨桐丸治疗痛风中的重要作用，从全新的角度揭示豨桐丸治疗痛风的现代药理机制，为中医药临床治疗痛风的遣方用药提供重要参考。

随着人们生活水平的提高、饮食结构的改变，近年来痛风的发病率不断升高。当前治疗痛风药物存在疗效有限、副作用大等缺陷，故从中医药中探寻安全、有效的治疗痛风方剂意义重大。豨桐丸是治疗痛风的常用方剂，但其药理机制至今不甚清楚。本课题立足于IL-1β在痛风炎症中的核心地位，着眼于NLRP3炎性体在IL-1β成熟表达及NF-κB在IL-1β生物活性中的关键作用，提出“豨桐丸通过调控NLRP3-IL-1β-NF-κB促炎信号轴而拮抗痛风炎症发展”的假说。通过研究，我们发现：（1）豨桐丸使MSU诱导的关节炎大鼠足肿胀显著减轻，步态评分和组织学评分均显著减少，同时关节组织中的破骨细胞数量及TNF-α，IL-1β，IL-6和NLRP3炎性体表达也都显著降低；（2）豨桐丸使MSU刺激的RAW264.7细胞上清中IL-1β含量明显降低；同时豨桐丸使MSU刺激的RAW264.7细胞内pro-IL-1β、pro-caspase-1和caspase-1的表达显著减少；（3）豨桐丸使MSU刺激的RAW264.7细胞内IKKβ活性和IκBα含量明显降低，豨桐丸也使MSU刺激的RAW264.7细胞核中p65水平和NF-κB与DNA的结合活性均明显降低，同时豨桐丸显著抑制了MSU诱导的RAW264.7细胞内MAPKs信号转导通路激活。预先应用RNAi 技术阻断巨噬细胞IL-1RI受体表达，我们发现豨桐丸明显抑制了MSU刺激的巨噬细胞中P65与DNA结合活性，但并没有完全阻断。上述结果表明，豨桐丸通过下调MSU激活的“NLRP3-IL-1β-NF-κB 促炎信号轴”的活化而抑制痛风炎症发生、发展。本课题从全新的角度揭示豨桐丸治疗痛风的现代药理机制，为中医药临床治疗痛风的遣方用药提供重要参考。