Toddalia asiatica extract attenuates adjuvant-induced arthritis by modulating colon Th17/Treg balance and colony homeostasis

Ethnopharmacological relevance: Given the adverse effects of the current principal treatments, there is still a great need for effective cures for rheumatoid arthritis (RA), an immune-mediated disease. Toddalia asiatica (L.) Lam is a traditional medicinal herb that can be used for RA treatment because of its anti-inflammatory and analgesic properties. Aim of the study: To investigate the possible effects of Toddalia asiatica extract (TAE) on intestinal immunity and the intestinal bacterial flora in a rat model of RA. Materials and methods: The anti-arthritis effect of TAE was evaluated in arthritis rats induced by complete Freund's adjuvant-induced arthritis (AIA). Arthritis index (AI) scores, systemic inflammation scores, histopathologic changes in the colon and ankle were detected by hematoxylin and eosin staining. Western blot analysis was performed to assess the protein expression of IL-17A, RORC, IL-18, IL-6, FOXP3, IL-10 in the colon. RT-PCR was performed to assess the expression of the colon's mRNA. Finally, changes to the gut microbiome by sequencing 16S rDNA. Microbial function prediction was performed using PICRUSt with the KEGG databases and correlation analysis was carried out by computing Spearman's rank correlations. Results: demonstrated that TAE administration at a dose of 3 g/kg dramatically decreased AI scores, systemic inflammation scores, and histopathologic lesions of the ankle and colon in AIA rats. TAE was found to significantly reduce the expression levels of Th17-related proteins and mRNAs (IL-17A, RORC, IL-18 and IL-6) in the colon, while increasing the expression levels of Treg-related proteins and mRNA (IL-10 and FOXP3), which helped restore the balance of Th17/Treg immune cells in the colon. Meanwhile, TAE was also found to be capable of remodeling the gut microbiota in AIA rats. Depleting RA-associated genera and thereby increasing alpha-diversity enriched the gut microbiota's diversity and shifted the community composition dramatically, leading to the increase of Firmicutes_unclassified, Ruminococcaceae_unclassified, Muribaculum, Subdoligranulum, Lachnospira, Marvinbryantia, and the reduction of RA-related bacteria Ligilactobacillus, Streptococcus and Eubacterium-eligensgroup. Furthermore, PICRUSt analysis revealed that metabolic pathways were associated with TAE treatment, with metabolic pathways dominating. Among them, metabolic pathways were predominant. Correlation studies showed that a total of 9 microorganisms, including Ligilactobacillus, Eubacterium-eligens-group and Subdoligranulum, were significantly associated with Th17/Treg expression. Conclusions: This study demonstrates that TAE is a low-toxicity poly alkaline drug that can rapidly and effectively improve joint symptoms in RA rats and increases beneficial intestinal bacteria and decreases harmful ones, which is associated with modulating Th17/Treg interactions in intestinal T cells and reversing microbial disorders.