Increased Alleviation of Bone Destruction in Individuals with Rheumatoid Arthritis via the Coinhibition of the METTL3 and YTHDF1 Axis by the Combination of Triptolide and Medicarpin

Rheumatoid arthritis (RA) is a progressive autoimmune disease characterized by bone destruction that is primarily caused by the overactivation of osteoclasts (OCs), which are critical therapeutic targets. Triptolide (TP) has strong anti-RA effects but is limited by its narrow therapeutic window and associated toxicity, necessitating combination therapy to increase its efficacy and reduce side effects . Medicarpin (Med), a flavonoid with anti-inflammatory and anti-bone destruction properties, has shown potential in reducing osteoclastogenesis . However, the mechanisms underlying the synergistic effects of TP and Med on RA treatment remain unclear. We addressed this issue by evaluating the effects of TP, Med, and their combination on a collagen-induced arthritis (CIA) rat model, with a focus on bone erosion as the primary research endpoint. We subsequently performed experimental validation in an in vitro OC differentiation model to assess the impacts of these treatments on OC formation and function. Based on polymerase chain reaction (PCR) microarray data from RA patients, further investigations focused on N 6 -methyladenosine (m 6 A) methylation and its regulatory factors, methyltransferase-like 3 (METTL3) and YT521-B homology domain family protein 1 (YTHDF1), which have been identified as potential targets of TP and Med. Key findings revealed that the TP and Med combination significantly alleviated bone destruction and inhibited OC differentiation, exerting stronger effects at lower doses than either drug alone. Mechanistically, TP and Med synergistically modulated METTL3 and YTHDF1 to suppress osteoclastogenesis through distinct m 6 A methylation pathways, contributing to the mitigation of RA-associated bone destruction. Overall, our data highlight the potential of the m 6 A modification as a therapeutic mechanism for the combined use of TP and Med for RA treatment, providing a theoretical basis for the clinical application of herbal active ingredient combinations.