RELEASE OF STEM-CELL FACTOR FROM A HUMAN KERATINOCYTE LINE, HACAT, ISINCREASED IN DIFFERENTIATING VERSUS PROLIFERATING CELLS

Stem cell factor, a recently discovered growth factor for hematopoieti c stem cells, mast cells, and melanocytes, was initially reported to b e produced by fibroblasts. In this study, we investigated the secretio n of this factor from human HaCaT cells during in vitro culture and co mpared it to synthesis by cells in the skin. Release of stem cell fact or from freshly cultured keratinocytes was comparable to that of HaCaT cells and was nearly half that produced by fibroblasts and umbilical vein endothelial cells. No stem cell factor was detectable in culture supernatants of melanocytes. HaCaT cells underwent spontaneous differe ntiation after a period of proliferation until confluency. Depending o n duration of culture, they released increasing amounts of stem cell f actor similar to 150 pg/10(6) cells on day 3 (proliferating cells) vs similar to 450 pg/10(6) cells on day 14 (differentiating cells) measur ed by enzyme-linked immunosorbent assay. Stimulation for 24 h with the calcium ionophore A 23187 (10(-6) to 10(-8) M) further enhanced relea se. Western blot analysis of HaCaT cell lysates with a stem cell facto r antibody revealed two proteins with the known molecular weights of m embrane-bound and soluble stem cell factor. By semiquantitative revers e transcriptase polymerase chain reaction, full-length as well as spli ced type stem cell factor mRNA was found to be increased in differenti ating versus proliferating HaCaT cells. Keratinocytes are thus potenti ally important sources of ste, cell factor in human skin, and HaCaT ce lls provide a useful model for further studies of stem cell factor fro m keratinocytes.