BRD3 regulates the inflammatory and stress response in rheu-matoid arthritis synovial fibroblasts

Individual functions of members of the bromodomain (BRD) and extra-terminal (BET) protein family underlying the anti-inflammatory effects of BET inhibitors in rheumatoid arthritis (RA) are incompletely understood. Here we aimed to analyze regulatory functions of BRD3, an under-studied member of the BET protein family, in RA synovial fibroblasts (FLS). BRD3 was silenced in FLS prior to stimulation with TNF. Alternatively, FLS were treated with I-BET. Transcriptomes were analyzed by RNA sequencing (RNAseq), followed by pathway enrichment analysis. We confirmed results for selective target genes by Real-time PCR, ELISA and Western blotting. BRD3 regulates the expression of several cytokines and chemokines in FLS, and positively correlates with inflammatory scores in the RA synovium. In addition, RNAseq pointed to a profound role of BRD3 in regulating FLS proliferation, metabolic adaption, and response to stress, including oxidative stress, and autophagy. BRD3 acts as an upstream regu-latory factor that integrates the response to inflammatory stimuli and stress conditions in FLS and executes many functions of BET proteins that have previously identified using pan-BET inhibitors. Overall design: To study the function of BRD3 in RA synovial fibroblasts, we silenced BRD3 expression in synovial fibroblasts using lentiviral particles. Cells were stimulated with TNF or left untreated and harvested 24h after stimulation.