黄芪多糖通过调节肠道菌群纠正Treg/Th17细胞失衡治疗类风湿关节炎的实验研究

Rheumatoid arthritis (RA) is a clinical refractory disease with high morbidity and disability. Previous studies revealed that gut microbiota could affect the development of RA. Based on our prophase studies of small sample size we found that: 1)Transplantation of specific intestinal strains can relieve RA; 2)Oral administration of Astragalus Polysaccharide (APS, 20kD) can regulate Treg/Th17 imbalance and improve the prognosis of collagen-induced arthritis (CIA) mice, but its mechanism needs further clarification. Considering that APS can not be absorbed into blood by oral administration directly, we hypothesize that APS corrects Treg/Th17 imbalance by regulating intestinal microbiota to treat RA. Firstly, the key bacteria affected by APS in the treatment of RA and their relationship with Treg/Th17 changes were investigated in the CIA model mice; Then, we colonize the key bacteria to the SKG spontaneously arthritis mice, CIA model mice and sterile mice to observe the effects and finally clarify the mechanism of APS to treat RA. We believe that the results of this study can lay the foundation for the treatment of rheumatoid arthritis from intestinal microbiota, and provides theoretical basis for the modernization of traditional Chinese medicine and the screening and development of similar traditional Chinese medicine.

类风湿关节炎(RA)是一种发病率高，致残率高的临床难治疾病。既往研究揭示肠道菌群可通过调节Treg/Th17平衡影响RA的进展。课题组前期基于小样本量的研究发现：1)移植特定肠道菌株可治疗RA；2)口服给予黄芪多糖（APS，20kD）能调节Treg/Th17失衡，改善胶原诱导性关节炎模型(CIA)小鼠的预后，然其机制亟待进一步明确。结合口服APS不直接吸收入血，本研究提出APS通过“调节肠道菌群，纠正Treg/Th17细胞失衡”治疗RA的科学假设。项目拟通过在大样本量CIA模型鼠中，首先明确APS治疗RA的关键菌及其与Treg/Th17变化的关系，其次定植关键菌于SKG自发性关节炎模型鼠、CIA模型鼠和无菌小鼠，反复验证该菌在RA治疗中的作用，最终明确APS治疗RA的可能机制。项目的实施可为从肠道菌群调节出发治疗类风湿关节炎治疗奠定基础，并为类似中药筛选和开发及中药现代化提供理论依据。

类风湿性关节炎（Rheumatoid arthritis, RA）是一种发病率高，致残率高的临床难治自身免疫性疾病。课题组前期研究发现，从中药黄芪中提取分离得到其主要活性成分黄芪多糖，能改善关节炎模型小鼠的预后，然其机制尚不明确。本项目以胶原诱导建立的实验性关节炎小鼠模型，探讨黄芪多糖抗实验性关节炎的作用机制。课题组从预防性和治疗性给药两个方面评价了黄芪多糖对RA防治作用，研究发现预防性和治疗性均显著降低类风湿性关节炎小鼠发病率、严重程度等级评分以及小鼠骨关节骨损失严重程度。进一步采用流式细胞术、ELISA等技术手段研究发现，黄芪多糖可以有效降低小鼠血液中炎症因子IL-17和TNF-a 含量，增加IL-10含量，显著升高小鼠脾脏中Treg/Th17比例。通过16S rDNA测序技术分析发现，经过黄芪多糖处理后，小鼠肠道菌群门、目、科、属、种5类ASVs地位丰度明显降低，模型组中毛螺菌科（Lachnospiraceae）和厌氧支原属（Anaeroplasma）、Blautia 以及梭菌属Clostridiales的丰度显著低于正常组，拟杆菌科Bacteroides和分节丝状菌Candidatus_Arthromitus在模型组中显著增高，而经黄芪多糖处理后，部分菌可逐渐回调至正常状态。最后，利用菌群移植技术研究发现，移植预防组小鼠粪便后，小鼠的发病率、严重程度均明显减轻，小鼠血液中TNF-a以及IL-17含量显著降低，而IL-10含量升高，且显小鼠脾脏中Treg/Th17比例明显升高。本研究初步明确黄芪多糖治疗类风湿性关节炎的可能机制，为黄芪多糖临床上防治类风湿关节炎治疗奠定实验基础，同时为类似中药开发提供了一种新的理论依据。