IFN-γ and TNF-α Synergize to Inhibit CTGF Expression in Human Lung Endothelial Cells

Connective tissue growth factor (CTGF/CCN2) is an angiogenetic and profibrotic factor, acting downstream of TGF-beta, involved in both airway-and vascular remodeling. While the T-helper 1 (Th1) cytokine interferon-gamma (IFN-gamma) is well characterized as immune-modulatory and anti-fibrotic cytokine, the role of IFN-gamma in lung endothelial cells (LEC) is less defined. Tumour necrosis factor alpha (TNF-alpha) is another mediator that drives vascular remodeling in inflammation by influencing CTGF expression. In the present study we investigated the influence of IFN-gamma and TNF-alpha on CTGF expression in human LEC (HPMEC-ST1.6R) and the effect of CTGF knock down on human LEC. IFN-gamma and TNF-alpha down-regulated CTGF in human LEC at the promoter-, transcriptional-and translational-level in a dose-and time-dependent manner. The inhibitory effect of IFN-gamma on CTGF-expression could be almost completely compensated by the Jak inhibitor AG-490, showing the involvement of the Jak-Stat signaling pathway. Besides the inhibitory effect of IFN-gamma and TNF-alpha alone on CTGF expression and LEC proliferation, these cytokines had an additive inhibitory effect on proliferation as well as on CTGF expression when administered together. To study the functional role of CTGF in LEC, endogenous CTGF expression was down-regulated by a lentiviral system. CTGF silencing in LEC by transduction of CTGF shRNA reduced cell proliferation, but did not influence the anti-proliferative effect of IFN-gamma and TNF-alpha. In conclusion, our data demonstrated that CTGF was negatively regulated by IFN-gamma in LEC in a Jak/Stat signaling pathway-dependent manner. In addition, an additive effect of IFN-gamma and TNF-alpha on inhibition of CTGF expression and cell proliferation could be found. The inverse correlation between IFN-gamma and CTGF expression in LEC could mean that screwing the Th2 response to a Th1 response with an additional IFN-gamma production might be beneficial to avoid airway remodeling in asthma.