Thrombin stimulates the production and release of a major surface-associated glycoprotein (fibronectin) in cultures of human fibroblasts

Abstract

Exposure of cultured human skin fibroblasts to thrombin in serum-free medium had several effects on fibronectin, a major cell surface-associated glycoprotein. Pericellular fibronectin fibrils, visualized by immunofluorescence, were lost after exposure for 4–20 h to thrombin (1–8 U/ml). Loss of fibronectin fibrils did not occur if thrombin was inhibited by phenylmethyl-sulfonyl fluoride (PMSF), N-α-tosyl-1-lysyl chloromethane (TosLysCH 2Cl), alpha-1-antithrombin, alpha-2-macroglobulin, or hirudin. Cell surface fibronectin, labeled by lactoperoxidase-catalysed iodination, and newly synthesized fibronectin, metabolically labeled with [ 3H]mannose, were lost after exposure for 20 h to thrombin. Within 60 min, increased concentrations of fibronectin were detected by radioimmunoassay in media of thrombin-treated cultures. Thrombin increased several-fold the total amount of fibronectin accumulating in cultures over a 20 h period by increasing the amount of fibronectin secreted or shed into the medium. Fetal calf serum, which contained inhibitors of thrombin and hence only low levels of thrombin activity (<0.05 U/ml), also stimulated fibronectin production but did not cause loss of pericellular fibronectin fibrils. Thrombin or serum, under the same experimental conditions, stimulated proliferation of human fibroblasts [46]. The effects of thrombin on fibronectin may be important in wound healing and tissue repair.