GRANULOCYTE MACROPHAGE COLONY-STIMULATING FACTOR IS AN INTRINSIC KERATINOCYTE-DERIVED GROWTH-FACTOR FOR HUMAN MELANOCYTES IN UVA-INDUCED MELANOSIS/

Recently we demonstrated that endothelins secreted from human keratino cytes act as intrinsic mitogens and melanogens for human melanocytes i n UVB-induced melanosis. We show here that UVA-induced melanosis is as sociated with other keratinocyte-derived growth factors, secretion of which is specifically stimulated after exposure of human keratinocytes to UVA. Medium conditioned by UVA-exposed human keratinocytes elicite d a significant increase in DNA synthesis by cultured human melanocyte s in a UVA dose-dependent manner. Analysis of endothelin-1 and interle ukin (IL)-1 alpha in the conditioned medium by ELISA, both of which ar e major keratinocyte-derived cytokines involved in UVB-associated mela nocyte activation, revealed that UVA exposure did not cause human kera tinocytes to stimulate the secretion of the two cytokines. In contrast , the levels of several other cytokines such as IL-6, IL-8 and granulo cyte/macrophage colony-stimulating factor (GM-CSF) were significantly increased in the conditioned medium of human keratinocytes after expos ure to UVA at a dose of 1.0 J/cm(2). The gel chromatographic profile o f UVA-exposed keratinocyte-conditioned medium demonstrated that there were two factors (P-1 and P-2) with molecular masses of approx. 20 and 1 kDa respectively that stimulate DNA synthesis in human melanocytes, and the larger species (P-1) also increased melanization as assessed by [C-14]thiouracil incorporation. Quantitative analysis of cytokines in chromatographic fractions by ELISA revealed the P-1 fraction to be consistent with the molecular mass profile of GM-CSF. Furthermore the stimulatory effect of the P-1 fraction on DNA synthesis in human melan ocytes was neutralized by antibodies to GM-CSF, but not to basic fibro blast growth factor or stem cell factor. Binding and proliferation ass ays with recombinant GM-CSF demonstrated that human melanocytes posses s specific binding sites for GM-CSF(K-d 2.11 nM; binding sites, 2.5-3. 5 x 10(4) per cell), and recombinant GM-CSF at concentrations of more than 10 nM significantly stimulated DNA synthesis and melanization. Th ese findings suggest that GM-CSF secreted by keratinocytes plays an es sential role in the maintenance of melanocyte proliferation and UVA-in duced pigmentation in the epidermis.