The effect of growth factor signaling on keratocytes in vitro and its relationship to the phases of stromal wound repair.
OBJECTIVE
To determine the relationship between signaling by different growth factors and the phases of corneal stromal wound repair. The authors hypothesize that the process involves sequential signaling, resulting first in proliferation and then in extracellular matrix (ECM) synthesis.


METHODS
The effects of IGF-I, TGF-beta1, FGF-2, and PDGF on proliferation and ECM production by primary cultured bovine keratocytes were evaluated. DNA synthesis was determined by (3)H-thymidine incorporation, and maximal cell density was determined by measurement of DNA content. Relative levels of ECM components synthesized by keratocytes and secreted into the media were evaluated by (3)H-glycine incorporation into total ECM protein and collagen, by (3)H-glucosamine incorporation into chondroitin sulfate, keratan sulfate, and hyaluronan, and by Western blotting with antibodies specific to procollagen types Iota and IotaIotaIota.


RESULTS
FGF-2 stimulated the highest level of proliferation and the lowest level of glycosaminoglycan synthesis and inhibited the synthesis of collagen types Iota and IotaIotaIota. IGF-I, in contrast, stimulated the lowest level of proliferation and the highest levels of collagen synthesis. PDGF and TGF-beta1 had intermediate effects on proliferation and collagen synthesis. Although FGF-2 inhibited collagen production, it could be restored by subsequent treatment with IGF-I, TGF-beta1, and PDGF.


CONCLUSIONS
The results of this study showed that the level of proliferation induced by the growth factors was inversely related to the levels of collagen production. The authors suggest that FGF-2 initiates the hypercellular phase of corneal wound healing and that IGF-I and PDGF are involved in the restoration of a normal ECM.
Etheredge
Latia
L
Department of Pathology and Cell Biology, College of Medicine, University of South Florida, Tampa, Florida 33612-4799, USA. lethered@health.usf.edu
Kane
Bradley P
BP
Hassell
John R
JR
D000818:Animals; D015153:Blotting, Western; D002417:Cattle; D049109:Cell Proliferation / Q000187:drug effects*; D002478:Cells, Cultured; D002809:Chondroitin Sulfates / Q000378:metabolism; D024042:Collagen Type I / Q000378:metabolism; D024061:Collagen Type III / Q000378:metabolism; D003319:Corneal Stroma / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D004247:DNA / Q000096:biosynthesis; D004591:Electrophoresis, Polyacrylamide Gel; D005109:Extracellular Matrix / Q000378:metabolism; D016222:Fibroblast Growth Factor 2 / Q000494:pharmacology; D005347:Fibroblasts / Q000166:cytology* / Q000187:drug effects / Q000378:metabolism; D006820:Hyaluronic Acid / Q000378:metabolism; D007334:Insulin-Like Growth Factor I / Q000494:pharmacology; D036341:Intercellular Signaling Peptides and Proteins / Q000494:pharmacology*; D007632:Keratan Sulfate / Q000378:metabolism; D010982:Platelet-Derived Growth Factor / Q000494:pharmacology; D015398:Signal Transduction / Q000187:drug effects*; D053773:Transforming Growth Factor beta1 / Q000494:pharmacology; D014945:Wound Healing / Q000502:physiology*
D024042:Collagen Type I; D024061:Collagen Type III; D036341:Intercellular Signaling Peptides and Proteins; D010982:Platelet-Derived Growth Factor; D053773:Transforming Growth Factor beta1; D016222:Fibroblast Growth Factor 2; D007334:Insulin-Like Growth Factor I; D006820:Hyaluronic Acid; D002809:Chondroitin Sulfates; D004247:DNA; D007632:Keratan Sulfate