ABSTRACT
Demonstrating temporal variation in the expression of messenger RNA (mRNA) for growth factors may give some indication as to whether growth factor synthesis is regulated in wound healing. The aim of this study was to evaluate the expression of insulin-like growth factors (IGF) I and II in the wound. Two wound models, an incisional model and a subcutaneous sponge implant model, were used in this study. The RNA was extracted and reverse transcribed and mRNA was amplified using polymerase chain reaction (PCR). Semiquantitation of PCR products was accomplished using [3H]dGTP incorporation. Levels of expression for both IGF-I and -II were found to be low in unwounded skin and at 12 hr postwounding. However, in both wound models expression increased substantially from 1 to 21 days postwounding. Both factors also were found to be expressed by fibroblasts and polymorphonuclear leukocytes (PMN). Additionally, two transcripts were found for IGF-II, the larger of which appeared to be specific for PMN and possibly cells involved in angiogenesis. Levels of message expressed in healing wounds for IGF-I and -II appear to be regulated with the highest levels of message found at time points coinciding with fibroblast predominance in the wound. Since fibroblasts are known to both secrete and respond to IGF-I, it is possible that IGF-I and IGF-II are acting to influence fibroblast differentiation and function in the later stages of wound healing.
Subject(s)
Insulin-Like Growth Factor II/metabolism , Insulin-Like Growth Factor I/metabolism , Wound Healing/physiology , Animals , Base Sequence , Electrophoresis , Male , Molecular Probes/genetics , Molecular Sequence Data , Polymerase Chain Reaction , Polymorphism, Restriction Fragment Length , Rats , Rats, Inbred F344ABSTRACT
Growth factors and amino acids (AA) are required for cell proliferation. A comparison of the AA composition of wound fluid (WF) to that of Eagle's medium reveals that AA in WF may be limiting to cell replication. Yet WF supports fibroblast replication and stimulates AA uptake. Epidermal growth factor (EGF) stimulates fibroblast replication and stimulates human wound healing when applied topically. We evaluated the interactions between EGF and AA concentrations found in WF. Wound fibroblasts were cultured in media prepared to mimic the AA concentrations found in WF on days 1, 5, and 10 and in the presence of varying concentrations of EGF. Fibroblasts cultured in all three experimental media showed a dose response to EGF for both tritiated-thymidine uptake (proliferation) and AA uptake. The fibroblast proliferation in response to EGF was augmented by the AA composition of day-5 WF. These data show a dose-dependent effect of EGF on fibroblast replication and AA uptake in the absence of serum that is augmented by the particular AA combination found in day-5 WF and suggests that an optimal physiologic AA profile may aid in EGF stimulation of wound fibroblast replication.
