Chromones from Saposhnikovia divaricata modulate m6A RNA methylation-mediated macrophage polarization by targeting CBLL1 to ameliorate RA.

BACKGROUND

Over-activated M1 macrophages, which destroyed the balance of macrophage phenotype in the synovial microenvironment, play a pivotal role in rheumatoid arthritis (RA) progression. Increasing evidence indicates that m6A RNA methylation serves as a crucial epigenetic regulatory mechanism in modulating the transition from M0 to M1 macrophages. Saposhnikovia divaricata (Turcz.) Schischk treats RA through immune regulation. In our previous study, chromones (CHR) from Saposhnikovia divaricata could alleviate RA by inhibiting the pathological manifestations of synovial inflammation, but the mechanism of action remains to be elucidated.

PURPOSE: To elucidate the pharmacology and mechanism of CHR on m6A RNA methylation mediated M1 macrophages for RA treatment.

METHODS

In CIA mice, joint inflammation was assessed clinically and histologically, while bone changes were quantified by micro-CT and H&E staining. In vitro, the effects of CHR on macrophage polarization were analyzed by examining cell surface proteins, effector cytokine secretion, and master transcription factor expression. Target determination was interrogated through dot blot combined with the GEO database, corroborated by the sgRNA of CBLL1. Downstream mechanistic pathway validation mainly included quantification of expression via RIP-qPCR, CETSA, and DARTS. A co-culture system of MH7A and polarized THP-1 cells was utilized to assess synovial inflammation.

RESULTS

CHR relieved paw swelling and joint damage and regulated macrophage polarization in CIA mice. Moreover, polarized macrophages were significantly repressed. CHR also modulated the expression of M1 macrophage-associated proteins to influence abnormal characteristics of synovial inflammation. Furthermore, CHR could directly bind to CBLL1 and inhibit its expression and function in macrophages. By knocking down CBLL1, CHR might be involved in regulating macrophage polarization by modulating CBLL1-mediated STAT1/NF-κB expression.

CONCLUSION

CHR could inhibit M1 macrophage polarization by regulating CBLL1-mediated m6A RNA methylation to treat RA.