Hypoxia induces expression of connective tissue growth factor in scleroderma skin fibroblasts

Connective tissue growth factor (CTGF) plays a role in the fibrotic process of systemic sclerosis (SSc). Because hypoxia is associated with fibrosis in several profibrogenic conditions, we investigated whether CTGF expression in SSc fibroblasts is regulated by hypoxia. Dermal fibroblasts from patients with SSc and healthy controls were cultured in the presence of hypoxia or cobalt chloride (CoCl2), a chemical inducer of hypoxia-inducible factor (HIF)-1 alpha. Expression of CTGF was evaluated by Northern and Western blot analyses. Dermal fibroblasts exposed to hypoxia (1% O-2) or CoCl2 (1-100 mM) enhanced expression of CTGF mRNA. Skin fibroblasts transfected with HIF-1 alpha showed the increased levels of CTGF protein and mRNA, as well as nuclear staining of HIF-1 alpha, which was enhanced further by treatment of CoCl2. Simultaneous treatment of CoCl2 and transforming growth factor (TGF)-beta additively increased CTGF mRNA in dermal fibroblasts. Interferon-g inhibited the TGF-beta-induced CTGF mRNA expression dose-dependently in dermal fibroblasts, but they failed to hamper the CoCl2- induced CTGF mRNA expression. In addition, CoCl2 treatment increased nuclear factor (NF)-kappa B binding activity for CTGF mRNA, while decreasing I kappa B alpha expression in dermal fibroblasts. Our data suggest that hypoxia, caused possibly by microvascular alterations, up-regulates CTGF expression through the activation of HIF-1 alpha in dermal fibroblasts of SSc patients, and thereby contributes to the progression of skin fibrosis.