Identification of leukemia inhibitory factor as a potent mast cell growth-enhancing factor produced by mouse keratinocyte cell line, KCMH-1.
Inoculation of KCMH-1 cells, a keratinocyte-derived cell line established from a chemically induced skin tumor, into the skin of mice results in accumulation of mast cells around the resulting tumors. The conditioned medium of KCMH-1 cells enhances the growth of mast cells in vitro when they are cultured in the presence of NIH/3T3 fibroblasts, suggesting an important role for keratinocytes in mast cell hyperplasia in the skin. The aim of this study was to identify this mast cell growth-enhancing factor (MCGEF) by screening a KCMH-1 cDNA library. We first established a polyclonal antibody raised against the partially purified factor obtained from KCMH-1-conditioned medium which neutralized the MCGEF activity in KCMH-1-conditioned medium. Expression cloning of 1 x 10(6) cDNAs from the KCMH-1 cDNA library led to 16 cDNAs. One of these cloned cDNAs was found to be leukemia inhibitory factor (LIF). Both LIF produced by COS cells and the recombinant protein obtained commercially showed MCGEF activity when added to mast cell/fibroblast cocultures. MCGEF activity in KCMH-1-conditioned medium was completely neutralized by an anti-LIF monoclonal antibody. These results suggest that MCGEF produced by KCMH-1 cells is identical to LIF.
Tanaka
T
T
Department of Dermatology, Hiroshima University Faculty of Medicine, Japan. toshikun@hiroshima-u.ac.jp
Morita
E
E
Mihara
S
S
Kanno
M
M
Yamamoto
S
S
D016475:3T3 Cells; D000818:Animals; D019556:COS Cells; D002460:Cell Line; D006131:Growth Inhibitors / Q000302:isolation & purification* / Q000494:pharmacology; D015850:Interleukin-6; D015603:Keratinocytes / Q000378:metabolism*; D053605:Leukemia Inhibitory Factor; D008222:Lymphokines / Q000302:isolation & purification* / Q000494:pharmacology; D008407:Mast Cells / Q000187:drug effects* / Q000502:physiology; D051379:Mice; D019089:Stem Cell Factor / Q000302:isolation & purification* / Q000494:pharmacology
D006131:Growth Inhibitors; D015850:Interleukin-6; D053605:Leukemia Inhibitory Factor; C507838:Lif protein, mouse; D008222:Lymphokines; D019089:Stem Cell Factor