RELEASE OF TRANSFORMING GROWTH FACTOR-BETA-1 FROM THE PERICELLULAR MATRIX OF CULTURED FIBROBLASTS AND FIBROSARCOMA CELLS BY PLASMIN AND THROMBIN

A sensitive immunoblotting assay was developed for the detection of transforming growth factor (TGF)-beta1 from cell extracts and culture medium. HT-1080 human fibrosarcoma cells and human fibroblasts were used as models for the secretion and proteolytic release of pericellular matrix-associated TGF-beta1. Analysis of the pericellular matrices of the cells indicated that the majority of cell-layer associated TGF-beta1 was associated with the pericellular matrix. Treatment of the cells with plasmin or thrombin released the matrix-associated TGF-beta1 to the culture medium. Assays for the biological activity of plasmin-released TGF-beta1 by Mv1Lu cell growth inhibition assays indicated that the majority was in the latent form. Northern hybridization analyses indicated that the mRNA levels of TGF-beta1 were not elevated during the proteinase treatment. Experiments using radiolabeled TGF-beta1 indicated that exogenous active TGF-beta1 associates mainly with the presumed TGF-beta1 receptors that were not retained in the extracellular matrix preparations. These results indicate that a major fraction of latent TGF-beta1 that is produced by the cells is deposited to and remains associated with the pericellular matrices of cultured fibroblasts and fibrosarcoma cells, and that matrix-associated TGF-beta1 is very susceptible to release by various proteolytic enzymes.