Src kinases in systemic sclerosis: Central roles in fibroblast activation and in skin fibrosis

Abstract

Objective. Src kinases are nonreceptor tyrosine kinases, which have been implicated in cytoskeletal organization and cell mobility. This study was undertaken to evaluate the potential of Src kinases as novel targets of antifibrotic therapies.Methods. Fibroblast cultures were obtained from 10 patients with systemic sclerosis (SSc) and 5 healthy subjects. Src signaling was inhibited using smallmolecule inhibitors and overexpression of a dominantnegative mutant of Src and of the endogenous inhibitor Csk. The expression of extracellular matrix proteins was analyzed by real-time polymerase chain reaction and by SirCol collagen assay. Toxic effects were excluded by MTT assay and staining for annexin V and propidium iodide. The mouse model of bleomycin-induced dermal fibrosis was used to assess the role of Src kinases in dermal fibrosis in vivo.Results. Stimulation with transforming growth factor ␤ and platelet-derived growth factor activated Src signaling in dermal fibroblasts from patients with SSc and healthy donors. Incubation with the Src kinase inhibitors or overexpressed mutant Src or Csk reduced the synthesis of messenger RNA for COL1A1, COL1A2, and fibronectin 1. A dose-dependent reduction in collagen release was also observed at the protein level. No inhibitory effects on proliferation and no increase in the number of apoptotic or necrotic fibroblasts were observed. Consistent with the in vitro data, inhibition of Src kinases prevented experimental dermal fibrosis. Dermal thickness, the amount of collagen protein, and the number of myofibroblasts were reduced in a dosedependent manner.Conclusion. These findings indicate that Src kinases play important roles in the activation of fibroblasts and in the development of experimental fibrosis. Thus, Src kinases might be interesting targets for novel antifibrotic therapies in SSc.

Therapeutic Targeting of Src Kinase in Myofibroblast Differentiation and Pulmonary Fibrosis

“…These profibrotic effects of TGF-b1 were inhibited by AZD0530 in human lung fibroblasts. Our results are consistent with previous findings that inhibition of Src kinase prevents collagen synthesis in cardiac and skin fibroblasts (Elkareh et al, 2007;Skhirtladze et al, 2008). a-SMA is critical for myofibroblast contractility (Hinz et al, 2007); therefore, reduced a-SMA expression leads to the reduced contractility in lung fibroblasts treated with AZD030.…”

The cannabinoid receptor CB2 exerts antifibrotic effects in experimental dermal fibrosis

“…Fivemicrometer sections were stained with hematoxylin and eosin. Dermal thickness was analyzed at 100-fold magnification by measuring the distance between the epidermal-dermal junction and the dermal-subcutaneous fat junction at 3 sites in lesional skin in each mouse (13). Collagen fibers were visualized by Masson's trichrome staining, according to the recommendations of the manufacturer (Sigma-Aldrich, Munich, Germany), and analyzed at 1,000-fold magnification.…”

Dissecting the enigma of scleroderma: possible involvement of the kynurenine pathway

“…TH17 lymphocytes promote the expression of the pro-fibrotic cytokines in scleroderma [53]. In addition, pp60 src released from the AHR on ligand binding plays a role in fibroblast activation [54].…”