The effect of a synthetic
peptide consisting of
Arg-Gly-Asp-Ser (RGDS), the amino acid sequence representing the fibroblast attachment site in
fibronectin (FN), was tested on migrating newt epidermal cells. In one approach, skin explants were placed on the bottom of
plastic dishes coated with human FN, human
fibrinogen (FGN), human
serum spreading factor (SF), or bovine
type I collagen. The explants were then incubated overnight in serum-free medium with or without RGDS. In these experiments exposure to 50 micrograms ml-1 of RGDS reduced migration over FN, FGN and SF to 2-7% of control levels. Two
peptides structurally dissimilar to RGDS (Val-Gly-Ser-Glu and
Thr-Pro-Arg-Lys), and two that are structurally similar (
Lys-Gly-Asp-Ser and
Arg-Gly-Glu-Ser), had no effect on explant migration even when used at concentrations higher than 50 micrograms ml-1. Upon removal of the
RGDS peptide, inhibited explants quickly recovered. In
collagen-coated dishes 50 micrograms ml-1 of RGDS was much less effective than in dishes coated with the other substrates. Raising the RGDS concentration in
collagen-coated dishes tenfold did not greatly increase the RGDS effect. When added to the medium bathing wounded limbs, 50 micrograms ml-1 of RGDS only moderately inhibited
wound closure. This concentration of
peptide, however, severely inhibited migration from skin explants in newt-plasma-coated-dishes and migration over pieces of newt-plasma-coated
plastic placed under one edge of a skin
wound. Increasing the RGDS concentration to 500 micrograms ml-1 resulted in almost total suppression of
wound closure.
Wounds exposed to this same concentration of
Lys-Gly-Asp-Ser closed normally. These results indicate that newt epidermal cells possess RGDS receptors and that these receptors are involved in epidermal
wound closure in vivo and in migration from skin explants onto
plastic coated with FN, FGN, SF and
collagen. The relative RGDS-insensitivity of
wound closure in vivo and in migration from explants onto
collagen may reflect in these instances the presence of a relatively high density of RGDS receptor binding sites on the substrate; the presence of RGDS receptor binding sites of relatively high affinity; or the participation of receptors other than those involved in migration over
plastic coated with FN, FGN or SF.