PURPOSE: Thrombospondin-1 (TSP-1) mediates chemotaxis, cell proliferation, angiogenesis, and protease regulation in healing. TSP-1 also binds platelet-derived growth factor (PDGF) and transforming growth factor beta (TGF-beta). This study confirms the role of TSP-1 and defines the relationship between TSP-1 and PDGF in proliferative tissue repair. METHODS: Purified TSP-1 was analyzed for bound PDGF. Cultured fibroblast growth response to TSP-1 and recombinant PDGF was studied and the effects of antibodies against TSP-1, PDGF, and TGF-beta on this response were evaluated. Levels of TSP-1 and PDGF and relative proteolytic activity in fluid collected from 10 skin graft donor sites were then assessed by ELISA and a protease assay kit. The effect of proteolysis on TSP-bound PDGF and free recombinant PDGF was studied by adding trypsin and measuring the remaining PDGF by ELISA. RESULTS: TSP-1 promoted dose-dependent fibroblast growth. While antibody to TGF-beta had no effect on promotion, antibody to both TSP-1 and PDGF eliminated this. Since a strong correlation of TSP-1 with PDGF levels was found and strong proteolysis was seen in all samples, we proposed that TSP-1 protected PDGF from proteolysis. Consistent with this, we found PDGF bound to TSP-1 was 33% less degraded than free PDGF upon trypsinization. CONCLUSIONS: These results suggest that TSP-1 stabilizes PDGF, enhancing the biological effects of PDGF in proliferative tissue repair. This effect of TSP-1 along with its matrix-modulating activities may have important clinical utility regarding topical growth factor therapy in wound healing, since high proteolytic activity is believed to be partially responsible for limiting the efficacy of this treatment.
PURPOSE:Thrombospondin-1 (TSP-1) mediates chemotaxis, cell proliferation, angiogenesis, and protease regulation in healing. TSP-1 also binds platelet-derived growth factor (PDGF) and transforming growth factor beta (TGF-beta). This study confirms the role of TSP-1 and defines the relationship between TSP-1 and PDGF in proliferative tissue repair. METHODS: Purified TSP-1 was analyzed for bound PDGF. Cultured fibroblast growth response to TSP-1 and recombinant PDGF was studied and the effects of antibodies against TSP-1, PDGF, and TGF-beta on this response were evaluated. Levels of TSP-1 and PDGF and relative proteolytic activity in fluid collected from 10 skin graft donor sites were then assessed by ELISA and a protease assay kit. The effect of proteolysis on TSP-bound PDGF and free recombinant PDGF was studied by adding trypsin and measuring the remaining PDGF by ELISA. RESULTS:TSP-1 promoted dose-dependent fibroblast growth. While antibody to TGF-beta had no effect on promotion, antibody to both TSP-1 and PDGF eliminated this. Since a strong correlation of TSP-1 with PDGF levels was found and strong proteolysis was seen in all samples, we proposed that TSP-1 protected PDGF from proteolysis. Consistent with this, we found PDGF bound to TSP-1 was 33% less degraded than free PDGF upon trypsinization. CONCLUSIONS: These results suggest that TSP-1 stabilizes PDGF, enhancing the biological effects of PDGF in proliferative tissue repair. This effect of TSP-1 along with its matrix-modulating activities may have important clinical utility regarding topical growth factor therapy in wound healing, since high proteolytic activity is believed to be partially responsible for limiting the efficacy of this treatment.
Authors: William Foulsham; Thomas H Dohlman; Sharad K Mittal; Yukako Taketani; Rohan Bir Singh; Sharmila Masli; Reza Dana Journal: Ocul Surf Date: 2019-06-05 Impact factor: 5.033