Paeoniflorin-6'-O-benzene sulfonate down-regulates CXCR4-Gßγ-PI3K/AKT mediated migration in fibroblast-like synoviocytes of rheumatoid arthritis by inhibiting GRK2 translocation.

ABSTRACT

OBJECTIVE: This study aims to explore the effect of paeoniflorin-6'-O-benzene sulfonate (CP-25) on the migration of fibroblast-like synoviocytes (FLS) in rheumatoid arthritis (RA) and the mechanism focused on CXCR4-Gßγ-PI3K/AKT signaling. METHODS: Human synovial tissues were collected from RA and osteoarthritis (OA) patients. Immunohistochemistry (IHC) and Western blot were used to detect the protein expression of CXCR4, GRK2, Gßγ, PI3K, p-PI3K, AKT and p-AKT. Transwell was adopted to analyse the migration of fibroblast-like synoviocytes (FLS). Co-immunoprecipitation (Co-IP) and laser scanning confocal microscopy (LSCM) were used to detect the combination of GRK2 and Gßγ, the combination of PI3K and Gßγ. RESULTS: The expression level of CXCR4, GRK2, Gßγ, p-p85 and p-AKT were increased in RA synovial tissue according to the results of IHC and Western blot. In vitro, the migration of FLS was increased after stimulation of CXCL12, inhibition of Gßγ suppressed the migration and phosphorylation of p85 and AKT induced by CXCL12 in FLS, and CP-25 had the same effect as inhibition of Gßγ. The membrane expression of GRK2, Gßγ, PI3K and the combination of GRK2 and Gßγ, the combination of PI3K and Gßγ in FLS were increased after the stimulation of CXCL12, and CP-25 had an ability in reducing the membrane expression and the combination of these proteins. CONCLUSION: Excessive migration of FLS in RA was associated with over-activation of PI3K/AKT signaling, and the activity of Gßγ was involved in the over-activation of PI3K/AKT. CP-25 down-regulated CXCR4-Gßγ-PI3K/AKT signals by inhibiting GRK2-Gßγ complex membrane translocation.