CHARACTERIZATION AND PARTIAL-PURIFICATION OF A KERATINOCYTE-DERIVED GROWTH-FACTOR WITH WOUND-HEALING PROPERTIES

We have previously described the mit nic and wound-healing properties of keratinocyte-conditioned medium (KCM). In this study we investigate d the effect of KCM on the activation of second messenger systems and the expression of proto-onco ene in cultured human skin fibroblasts. W e also present a partial purification of the factor responsible for th e mitogenic and wound-healing effects of KCM. KCM was shown to increas e the expression of the proto-oncogenes c-fos, c-myc and c-jun. The ef fect of KCM on three second messenger systems was investigated. The ex tracellular release of choline metabolites was increased by 40 per cen t when cells were stimulated with KCM whereas the formation of cAMP an d hydrolysis of phosphatidyl inositol (PI) was unaffected. KCM was pur ified by ion exchange chromatography and filtration. The biologically active fraction was eluted from an SP column and retained its activity after filtration through a 3-kDa filter. The fraction was inactivated by heat and acid, indicative of a peptide origin. Furthermore, the ac tive fraction was shown to increase the extracellular release of choli ne metabolites and to stimulate re-epithelialization in wounds in huma n skin in vitro comparable to KCM. The study indicates that human kera tinocytes produce a <3 kDa peptide which may be partly responsible for the growth stimulatory and wound-healing properties of KCM. (C) 1997 John Wiley & Sons, Ltd.