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One of the primary points of regulation of transforming growth factor-␤ (TGF-␤) activity is control of its conversion from the latent precursor to the biologically active form. We have identified thrombospondin-1 as a major physiological regulator of latent TGF-␤ activation. Activation is dependent on the interaction of a specific sequence in thrombospondin-1 (K 412 RFK415 ) with the latent TGF-␤ complex. Platelet thrombospondin-1 has TGF-␤ activity and immunoreactive mature TGF-␤ associated with it. We now report that the latency-associated peptide (LAP) of the latent TGF-␤ complex also interacts with thrombospondin-1 as part of a biologically active complex. Thrombospondin⅐LAP complex formation involves the activation sequence of thrombospondin-1 (KRFK) and a sequence (LSKL) near the amino terminus of LAP that is conserved in TGF-␤ 1-5 . The interactions of LAP with thrombospondin-1 through the LSKL and KRFK sequences are important for thrombospondin-mediated activation of latent TGF-␤ since LSKL peptides can competitively inhibit latent TGF-␤ activation by thrombospondin or KRFKcontaining peptides. In addition, the association of LAP with thrombospondin-1 may function to prevent the reformation of an inactive LAP⅐TGF-␤ complex since thrombospondin-bound LAP no longer confers latency on active TGF-␤. The mechanism of TGF-␤ activation by thrombospondin-1 appears to be conserved among TGF-␤ isoforms as latent TGF-␤ 2 can also be activated by thrombospondin-1 or KRFK peptides in a manner that is sensitive to inhibition by LSKL peptides.

Glucose is a key factor in the development of diabetic complications, including diabetic nephropathy. The development of diabetic glomerulosclerosis is dependent on the fibrogenic growth factor, transforming growth factor-beta (TGF-beta). Previously we showed that thrombospondin-1 (TSP-1) activates latent TGF-beta both in vitro and in vivo. Activation occurs as the result of specific interactions of latent TGF-beta with TSP-1, which potentially alter the conformation of latent TGF-beta. As glucose also up-regulates TSP-1 expression, we hypothesized that the increased TGF-beta bioactivity observed in rat and human mesangial cells cultured with high glucose concentrations is the result of latent TGF-beta activation by autocrine TSP-1. Glucose-induced bioactivity of TGF-beta in mesangial cell cultures was reduced to basal levels by peptides from two different sequences that antagonize activation of latent TGF-beta by TSP, but not by the plasmin inhibitor, aprotinin. Furthermore, glucose-dependent stimulation of matrix protein synthesis was inhibited by these antagonist peptides. These studies demonstrate that glucose stimulation of TGF-beta activity and the resultant matrix protein synthesis are dependent on the action of autocrine TSP-1 to convert latent TGF-beta to its biologically active form. These data suggest that antagonists of TSP-dependent TGF-beta activation may be the basis of novel therapeutic approaches for ameliorating diabetic renal fibrosis.

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