Bromodomain and extra-terminal domain bromodomain inhibition prevents synovial inflammation via blocking IκB kinase-dependent NF-κB activation in rheumatoid fibroblast-like synoviocytes

Objective. To explore the roles of the bromodomain (Brd) and extra-terminal domain (BET) of chromatin adaptors in regulating synovial inflammation in RA. Methods. Fibroblast-like synoviocytes (FLSs) were isolated from synovial tissue from RA patients. A specific BET inhibitor, JQ1, and short hairpin RNA (shRNA) for Brd2 or Brd4 were used to evaluate the role of the BET Brd in inflammatory responses. Protein expression was measured by western blot or immunofluorescence staining. Nuclear factor kappa B (NF-kappa B) gene activity was detected by luciferase assay. The secretion and gene expression of cytokines and MMPs were evaluated by ELISA and real-time PCR, respectively. FLS proliferation was detected by BrdU incorporation. Results. Four Brd proteins, including Brd2, Brd3, Brd4 and Brdt, were expressed in FLSs from patients with RA and OA; however, the expression of Brd2 and Brd4 was increased in RA compared with that in OA. Treatment with JQ1, Brd2 shRNA or Brd4 shRNA decreased the production of pro-inflammatory cytokines (TNF alpha, IL-1 beta, IL-6 and IL-8), MMPs expression (MMP-1, MMP-3 and MMP-13) and proliferation by RA FLSs. BET inhibition downregulated TNF alpha-induced NF-kappa B-dependent transcription and expression of the NF-kappa B target genes. JQ1 suppressed the phosphorylation of I kappa B kinase beta and I kappa B alpha, and nuclear translocation of p65. Intraperitoneal injection of JQ1 in mice with collagen-induced arthritis reduced synovial inflammation, joint destruction and serum levels of the anti-CII antibodies TNF alpha and IL-6. Conclusion. This study implicates BET Brds as important regulators of I kappa B kinase/NF-kappa B-mediated synovial inflammation of RA and identifies BET proteins as novel therapeutic targets in inflammatory arthritis.