Effect of Epidermal Growth Factor on Bone Formation inVitro*

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Abstract
Epidermal growth factor (EGF), a peptide isolated from mouse submaxillary glands which stimulates cell proliferation, was examined for its effects on bone formation in vitro. To study the effect of EGF on bone formation, we examined its effects on the incorporation of [3H]thymidine into DNA, [3H]uridine into RNA, and [3H]proline into collagenase-digestible (CDP) and noncollagen protein as well as on DNA content in cultured 21-day-old fetal rat calvaria. EGF caused a dose-dependent stimulation of the incorporation of [3H]thymidine into DNA at concentrations of 0.1–10 ng/ml and increased the total DNA content. The effect on [3H]thymidine incorporation first appeared at 3 h, was maximal at 18 h, and was sustained for up to 96 h. The effect of EGF was opposed by cortisol. EGF had a small stimulatory effect on [3H]uridine incorporation, related to an increase in uptake of the label. EGF (1–100 ng/ml) caused a dose-related inhibition of the incorporation of [3H]proline into CDP without affecting noncollagen protein. The effect on CDP appeared at 12 h and was sustained for 96 h. Cortisol did not alter the inhibitory effect of EGF on CDP at 24 h, while labeled CDP was greater with EGF and cortisol than with cortisol alone at 96 h. EGF decreased the labeling of CDP in the presence of insulin and serum. Histological sections showed an increase in mitotic index after colcemid arrest and in periosteal fibroblastic proliferation in EGF-treated bones. The effect of EGF on [3H]thymidine incorporation, but not on CDP labeling, was opposed by theophylline and isobutylmethylxanthine. Our studies indicate that EGF stimulates DNA synthesis in bone and inhibits bone collagen synthesis in vitro.