Integrating serum pharmacochemistry, network pharmacology, metabolomics and 16S rRNA sequencing to explore the mechanism of total flavonoids from Flemingia philippinensis in treating collagen induced arthritis rats.

BACKGROUD

Rheumatoid arthritis (RA) is a prevalent chronic autoimmune disease characterized by symmetric polyarthritis, resulting in pain and swelling in the synovial joints. Flemingia philippinensis, a traditional Chinese medicine, has been shown to be an effective treatment approach for anti-rheumatoid arthritis (RA), which still needs further research in its active ingredient and regulatory mechanisms.

PURPOSE

This study aimed at investigate the pharmacodynamic basis and intricate mechanism of action of Flemingia philippinensis (FPTF) in the treatment of RA based on integrated omics technologies.

METHODS

UPLC-Q-Orbitrap HRMS was first established to identify the active components of FPTF in blood and network pharmacology was then used to predict the key therapeutic targets and corresponding pathways of FPTF in treatment of RA. To substantiate the pharmacodynamic effects, a collagen-induced arthritis (CIA) animal model was employed to observe the anti-RA effects of FPTF through a series of indicators, including rat body weight, arthritis scoring, paw swelling, histopathological analysis of synovial tissue, and serum inflammatory factors. Subsequently, the potential mechanisms underlying the anti-RA efficacy of FPTF was elucidated by integrating metabolomics analysis with 16S rRNA gene sequencing. Specifically, the RT-qPCR experiment was further conducted to validate the pathways predicted by serum pharmacochemistry, network pharmacology, metabolomics and 16S rRNA gene sequencing.

RESULTS

A total of 10 compounds derived from FPTF were identified by serum sample analysis. Utilizing network pharmacology, we identified 117 common targets for FPTF in the treatment of RA. Notably, KEGG analysis highlighted the PI3K/AKT signaling pathway and the IL17 signaling pathway as key pathways associated with the anti-RA effects of FPTF. Pharmacodynamic studies showed that FPTF can significantly alleviate CIA-induced arthritis. Compared with the CIA model group, FPTF treatment significantly improved the expression of mRNA in the PI3K/AKT and IL-17 signaling pathways. Further investigation unveiled a total of 28 differential metabolites in serum samples, among which 21 metabolites were observed to be reversed following FPTF administration. Metabolomic profiling revealed pronounced perturbations in amino acid metabolism, fatty acid metabolism, and glycerophospholipid metabolism pathways in CIA rats, which were partially rectified by FPTF treatment. Additionally, 16S rRNA gene sequencing analysis indicated that FPTF could restore the gut microbiota balance disrupted by RA. RT-qPCR further confirmed that FPTF can modulate key enzymes in metabolic pathway analysis and gut microbiota metabolic pathways.

CONCLUSION

This study pioneeringly elucidates the potential pharmacodynamic material basis of FPTF for treatment of RA, detailing the regulated metabolic pathways and key gut microbiota genera involved. The findings provide a comprehensive understanding of mechanisms underlying the effects of FPTF in RA treatment.