PROLIFERATIVE AND MATRIX SYNTHESIS RESPONSE OF CANINE ANTERIOR CRUCIATE LIGAMENT FIBROBLASTS SUBMITTED TO COMBINED GROWTH-FACTORS

We investigated the effects of growth factors on the proliferation and matrix synthesis of anterior cruciate ligament fibroblasts. Fibroblas ts from the anterior cruciate ligaments of dogs were transferred at th e second passage in a defined medium. Epidermal growth factor, platele t-derived growth factor-AB, transforming growth factor-beta(1), insuli n-like growth factor-1, and insulin, combined two by two following a 5 x 5 logarithmic concentration matrix, were added. Tridimensional curv es showing cell proliferation at 24 hours against the concentration of two effecters were obtained for each combination. Collagen and proteo glycan productions were quantified using [C-14]glycine and Na-2[S-35]O -4. Ratios of type I:III collagen and hydrodynamic size distributions of proteoglycans were assayed, respectively, by sodium dodecyl sulfate -polyacrylamide gel electrophoresis and gel filtration chromatography. Epidermal growth factor had an effect nearly equivalent to that of pl atelet-derived growth factor-AB on cell proliferation. Both had a grea ter effect than insulin-like growth factor-1, which in turn had a grea ter effect than both the effect of insulin or the nearly equivalent ef fect of transforming growth factor-beta(1). Neither platelet-derived g rowth factor-AB nor insulin has a significant effect by itself on coll agen production. Epidermal growth factor slightly decreases collagen p roduction as well as the type I:III collagen ratio; both transforming growth factor-beta(1) and insulin-like growth factor-1 increase the sa me parameters. Epidermal growth factor inhibits the stimulation induce d by transforming growth factor-beta(1). Similarly, insulin decreases the response to insulin-like growth factor-1. Proteoglycan production was significantly increased by all growth factors in this study, with transforming growth factor-beta(1) having the strongest effect. Small hydrodynamic size of proteoglycan was correlated to a high level of pr oteoglycan biosynthesis. The results map be readily applied to tissue engineering or provide a basis for in vivo investigations.