Sappanone A attenuates rheumatoid arthritis via inhibiting PI3K/AKT/ NF-κB and JAK2/STAT3 signaling pathways in vivo and in vitro

Objective: Sappanone A (SA), a bioactive compound in Caesalpinia sappan L., has anti-inflammation, antioxidant, and bone protection activities. But its effect on rheumatoid arthritis (RA) and the underlying mechanism are incompletely understood. Methods: Candidate targets of SA against RA were screened by network pharmacology and further validated by molecular docking. CIA rats and HFLS-RA were used to explore the effect and mechanism of SA on RA in vivo and in vitro, respectively. Macroscopic inspection (body weight, paw swelling, arthritis index), histological examination and micro-CT were used to evaluate the anti-RA effect of SA in vivo. ELISA and western blotting were used to explore the effects of SA on the levels of inflammatory cytokines in serum and the phosphorylation level of key proteins in tissue, respectively. Moreover, agonists and inhibitors of key proteins were used on HFLS-RA to explore the underlying mechanism of SA. Finally, immunofluorescence was utilized to explore the effects of SA on apoptosis in HFLS-RA. Results: SA significantly reduced arthritis index, alleviated paw swelling, and improved inflammatory cell infiltration and cartilage degradation in CIA rats. The levels of the pro-inflammatory cytokines including TNF-alpha, IL-1 beta, IL-6, and IL-17 were decreased while the level of the anti-inflammatory cytokine IL-10 was promoted by SA. The SA also down-regulated the protein phosphorylation levels of JAK2, STAT3, PI3K, AKT and p65 in vivo and in vitro. Furthermore, SA reversed the agonist-induced increase in phosphorylation levels of PI3K/AKT/NF kappa B and JAK2/STAT3 pathway-related proteins. In addition, SA acted on the phosphorylation levels of these proteins in the same trend as the pathway inhibitors and dose-dependently reduced the phosphorylation levels of PI3K/AKT/NF-kappa B pathway proteins. The immunofluorescence results suggested that SA could promote apoptosis in HFLS-RA. Conclusion: SA could inhibit inflammatory symptoms and bone destruction in CIA, and its mechanism may be related to the inhibition of PI3K/AKT/NF-kappa B and JAK2/STAT3 pathways. Hence, SA could be developed as a potential anti-RA therapeutic drug.