LATENT TRANSFORMING GROWTH-FACTOR-BETA-1 ASSOCIATES TO FIBROBLAST EXTRACELLULAR-MATRIX VIA LATENT TGF-BETA BINDING-PROTEIN

The role of latent transforming growth factor-beta (TGF-beta) binding protein (LTBP) in the association of TGF-beta1 to the extracellular ma trix of cultured fibroblasts and HT-1080 fibrosarcoma cells was studie d by immunochemical methods. The matrices were isolated from the cells , and the levels of LTBP and TGF-beta1 were estimated by immunoblottin g and immunoprecipitation. LTBP, TGF-beta1, and its propeptide (latenc y-associated peptide, LAP) were found to associate to the extracellula r matrix. Immunoblotting analysis indicated that treatment of the cell s with plasmin resulted in a concomitant time and dose dependent relea se of both LTBP and TGF-beta1 from the extracellular matrix to the sup ernatant. Comparison of molecular weights suggested that plasmin treat ment resulted in the cleavage of LTBP from the high molecular weight f ibroblast form to a form resembling the low molecular weight LTBP foun d in platelets. Pulse-chase and immunoprecipitation analysis indicated that both the free form of LTBP and LTBP complexed to latent TGF-beta were efficiently incorporated in the extracellular matrix, from where both complexes were slowly released to the culture medium. Addition o f plasmin to the chase solution resulted, however, in a rapid release of LTBP from the matrix. Fibroblast derived LTBP was found to associat e to the matrix of HT-1080 cells in a plasmin sensitive manner as show n by immunoprecipitation analysis. These results suggest that the late nt form of TGF-beta1 associates with the extracellular matrix via LTBP , and that the release of latent TGF-beta1 from the matrix is a conseq uence of proteolytic cleavage(s) of LTBP.