Study On The Mechanism Of Daphnetin Improving Rheumatoid Arthritis By Regulating Intestinal Microbiota

Background:Rheumatoid arthritis(RA)is a chronic autoimmune disease characterized by swelling of joints,inflammatory cell infiltration and damage to articular cartilage.The pathogenesis of rheumatoid arthritis is still unclear,but accumulated studies have shown that in addition to mobilizing the systemic immune response of patients,the occurrence and development of rheumatoid arthritis is closely related to the composition of intestinal microbiota.Daphnetin is a natural coumarin compound,which has a wide range of pharmacological activities,such as antiinflammatory,antibacterial,anti-tumor.However,the therapeutic effect of daphnetin on rheumatoid arthritis remains to be further confirmed,and the mechanisms involved remain unclear.Therefore,it is of great significance to study the role of daphnetin in the treatment of rheumatoid arthritis and its immune regulation mechanism.Objective:To study the pharmacological action of daphnetin in the treatment of rheumatoid arthritis and elucidate its immune regulation mechanism,and to provide theoretical basis and new thought for the treatment of rheumatoid arthritis.Methods:The adjuvant-induced rheumatoid arthritis(AIA)model was established in this study.The mice were divided into normal control group,arthritis group,daphnetin treatment group(low dose)and daphnetin treatment group(high dose).The model severity was reflected by observing and recording the disease progression of mice.The Treg cells frequency in inguinal lymph nodes of mice were detected by flow cytometry.H&E staining was used to observe the pathological changes of joints.Inflammatory factors IL-1β,IL-6 and TNF-α in serum were detected by ELISA.The 16S rRNA gene sequencing of mice feces reflected the changes of intestinal microbiota.Fecal Microbiota Transplantation(FMT)experiment confirmed that the effect of daphnetin on arthritis was closely related to its regulation of intestinal microbiota.Metabolites related to the tricarboxylic acid cycle in feces and serum were detected by HPLC and LC-MS respectively.Dimethyl fumarate stimulated naive T cells of lymph nodes to detect the differentiation of Treg cells frenquncy in vitro.Results:1.Daphnetin releases the progression of the disease.2.Daphnetin restored the proportion of Treg cells and the levels of related cytokines in arthritic mice.3.Daphnetin modifies gut microbiota composition in arthritic mice 4.The protective effect of daphnetin could be transmitted through the transfer of mice fecal microbiota.5.Daphnetin increases the levels of fumarate in serum and feces of arthritic mice.6.Fumarate promotes differentiation of Naive T cells into Treg cells in vitro.Conclusion:Daphnetin regulates the composition and abundance of intestinal microbiota in rheumatoid arthritis mice,leading to the up-regulation of fumarate level in mice,further promoting the differentiation of Treg cells,and improving the symptoms of rheumatoid arthritis.