7-Hydroxycoumarin mitigates the severity of collagen-induced arthritis in rats by inhibiting proliferation and inducing apoptosis of fibroblast-like synoviocytes via suppression of Wnt/β-catenin signaling pathway

Abstract

Background7-Hydroxycoumarin (7-HC) as a coumarin compound is widely found in Chinese herbs and exhibits diverse biological activities. Promoting cell apoptosis of fibroblast-like synoviocytes (FLS) is a meaningful strategy for rheumatoid arthritis (RA). Though the protective effect of 7-HC on RA experimental models has been reported, the specific mechanisms, especially the possible relationships of this effect to regulating FLS proliferation and apoptosis, still need clarification. PurposeThis study clarified the therapeutic effects of 7-HC on collagen-induced arthritis (CIA) in rats and explored the underlying mechanisms. MethodsIn vivo, 7-HC (15, 30 or 60 mg/kg) was intraperitoneally given to CIA rats, and its therapeutic effect and anti-inflammatory activity were evaluated. Ki67 immunohistochemistry, TUNEL assay and synovial proteins detection were conducted. In vitro, after treating with 7-HC (20, 40 or 80 μM) in TNF-α-stimulated RA FLS (MH7A cell line), cell proliferation and apoptosis were examined. The involvement of Wnt/β-catenin pathway was checked in vivo and in vitro. Results7-HC attenuated the severity of rat CIA, evidenced by the reduction of paw swelling, arthritis index, joint damage, collagen type II antibody serum level, and IL-1β, IL-6, TNF-α production in serum and synovium. Particularly, 7-HC in vivo had anti-proliferative and pro-apoptotic effects on CIA rat synovial cells, indicated by reduced synovial Ki67 expression, raised synovial apoptosis index, decreased Bcl-2 protein level and increased level of Bax and cleaved caspase 3 protein. Further, 7-HC in vitro suppressed proliferation and promoted apoptosis of TNF-α-stimulated MH7A cells by regulating the mitochondrial pathway. Mechanistically, 7-HC treatment inhibited Wnt/β-catenin pathway, suggested by the reduction of pathway-related proteins (e.g. Wnt1, LRP6, p-GSK-3β (Ser9), β-catenin, cyclin D1 and c-Myc), the recovery of GSK-3β activity and the inhibition of β-catenin nuclear translocation. As expected, combined use of lithium chloride, an activator of Wnt/β-catenin signaling, reversed the anti-proliferative and pro-apoptotic effects of 7-HC in vitro. Conclusion7-HC relieved the severity of rat CIA by inhibiting cell proliferation and inducing apoptosis of rheumatoid FLS via inhibition of Wnt/β-catenin pathway.