Effect of Granulocyte-macrophage Colony-stimulating Factor and Ascorbic Acid Co-supplementation on the Fibroblast Proliferation and Collagen Synthesis

Seo, Sung Ig; Han, Seung Kyu; Kim, Hyung Jun; Kim, Woo Kyung

Abstract

Granulocyte-macrophage colony-stimulating factor (GM-CSF) is naturally generating protein that has the healing effect in normal wounds as well as infected nonhealing wounds and ulcers. Ascorbic acid has also been known to stimulate fibroblast stimulation and collagen synthesis. However, there are no reports about the effect of GM-CSF and ascorbic acid co-supplementation in wound healing. Therefore, in this report, we examined the effect of GM-CSF and ascorbic acid co-supplementation on the proliferation of human dermal fibroblasts and collagen synthesis which play a crucial role in wound healing process in vitro. To determine an optimal GM-CSF and ascorbic acid concentration for human dermal fibroblast proliferation and collagen synthesis, the cells were incubated with various concentrations of GM-CSF and ascorbic acid. The activity of fibroblast proliferation was determined by MTT assay. To measure the amount of collagen production, the collagen type I carboxy-terminal propeptide enzyme immunoassay was used. The best fibroblast proliferation was observed at the co-supplementation of 0.5 microgram/ml GM-CSF with 25 microgram/ml ascorbic acid and 1 microgram/ml GM-CSF with 12.5 microgram/ml ascorbic acid. Maximal stimulation of collagen synthesis was observed at the co-supplementation of 1 microgram/ml GM-CSF with 12.5 microgram/ml ascorbic acid. The collagen synthesis per cell was also maximal at concentration of 1 microgram/ml GM-CSF with 12.5 microgram/ml ascorbic acid. This results demonstrates that GM-CSF and ascorbic acid co-supplementation increases human dermal fibroblast proliferation and collagen synthesis and the concentration is the critical factor in vitro.