Enhanced expression of ephrins and thrombospondins in the dermis of patients with early diffuse systemic sclerosis: potential contribution to perturbed angiogenesis and fibrosis

Objective. To determine the skin and fibroblast expression of ephrins (EphB4 and EphrinB2) and thrombospondins (TSPs: TSP1 and TSP2) in patients with SSc. Methods. All experiments were performed in skin sections and dermal fibroblasts issued from control and clinically involved/non-involved SSc skin biopsies. Dermal fibroblasts were stimulated with hypoxia or TGF-β, or treated with TGF-β-neutralizing antibodies. Ephrin and TSP mRNA levels were assessed in skin tissue and dermal fibroblasts by in situ hybridization and quantitative RT-PCR, respectively, and protein levels were assessed by immunohistochemistry and western blots, respectively. Results. Enhanced ephrin and TSP mRNA and protein levels were observed in clinically involved SSc skin. EphrinB2, TSP1 and TSP2 mRNA and protein levels were also up-regulated in non-involved SSc skin. Similar mRNA and protein levels of ephrinB2 and EphB4 were detected in unstimulated and stimulated control and SSc dermal fibroblasts. TSP1 and TSP2 mRNA and protein levels were significantly increased in fibroblasts issued from involved and non-involved SSc skin. This up-regulation was not modified by hypoxic exposure, but was markedly reduced by the addition of TGF-β-neutralizing antibodies. Stimulation of healthy fibroblasts with TGF-β significantly increased TSP1 and TSP2 mRNA and protein levels. Conclusion. EphB4 and EphrinB2 are up-regulated in clinically involved skin of SSc patients, suggesting their participation in SSc-perturbed angiogenesis. TSP1 and TSP2 are up-regulated in both clinically involved and non-involved SSc skin and are constitutively overexpressed in a TGF-β-dependent and hypoxia-independent manner in SSc dermal fibroblasts, suggesting their potential early contribution in SSc pathogenesis.