Bromodomain inhibitor JQ1 attenuates TGF-β-suppressed SOD2 expression and reduces proliferation in human primary pulmonary fibroblasts

Background: SOD2 converts intracellular superoxide to hydrogen peroxide and acts as an antioxidant enzyme. Low expression levels of SOD2 have been associated with smooth muscle cell activation and proliferation. We have identified low SOD2 expression in pulmonary fibroblasts from SSc and IPF fibrotic lung, in which fibro-proliferation is an early desease hallmark. A BRD4 inhibitor, JQ1 is being investigated for its inhibitory effects on cell proliferation in cancer. Here we investigate the effect of TGF-β on SOD2 expression, and the potential beneficial effects of JQ1 on SOD2 expression and proliferation in adult lung fibroblasts. Methods: Human primary adult lung fibroblasts where quiesced for 24 hrs in serum free DMEM and pre-treated with DMSO (0.1%), 300nM JQ1+, or its enantiomer JQ1-, for 2 hrs before stimulation with TGF-β1 at 2ng/ml for a further 24 hrs. SOD2 mRNA expression was determined in cell extracts by qRT-PCR. Proliferation assays were performed in 3% serum, with or without JQ1 for 24 hrs and 48 hrs, by assessing BrdU incorporation (Roche) during a 5 hr period before the time point. Cell toxicity was assessed by LDH assay. Results: Expression of SOD2 was reduced by approximately 70% (p<0.01) by TGF-β1 in the presence of the JQ1 enantiomer, which had no effect on expression. TGF-β suppressed SOD2 expression was enhanced 2 fold (p<0.01) by JQ1+ at 300 nM. This concentration of JQ1+ (but not of its enantiomer JQ1-) inhibited BrdU incorporation by 77% (p<0.01) and 43% (p<0.01), respectively. Conclusion: The data suggests bromodomain inhibitor JQ1 enhances SOD2 expression and inhibits proliferation in activated human pulmonary fibroblasts.