IN CONTRAST TO THEIR MURINE COUNTERPARTS, NORMAL HUMAN KERATINOCYTES AND HUMAN EPIDERMOID CELL-LINES A431 AND HACAT FAIL TO EXPRESS IL-10 MESSENGER-RNA AND PROTEIN

In mice, keratinocyte-derived IL-IO is up-regulated by ultraviolet-B ( UVB) radiation and plays a major role in UVB-induced immunosuppression . The present study was designed to examine whether a comparable pheno menon can be detected in man. Freshly isolated or cultured normal huma n keratinocytes (NHK) and keratinocyte cell lines A431 and HaCaT were stimulated with graded doses of UVB (up to 200 J/m(2)) or with a varie ty of other stimuli. RNA was extracted at various time points post-sti mulation and analysed by reverse transcriptase-polymerase chain reacti on (RT-PCR) using four different IL-10-specific primer pairs and RNA f rom monocytes or T cells as positive controls. We failed to detect IL- 10 mRNA in NHK from 40 different donors (breast, abdomen, leg, scalp, foreskin) and in A431 and HaCaT cells, irrespective of the stimulation used and despite successful stimulation. Supernatants of NHK, A431 an d HaCaT cultures were negative for IL-IO protein, as tested by four di fferent ELISAs and a bioassay. Murine keratinocytes, stimulated under comparable conditions and tested by the same techniques, displayed a s trong expression of IL-10 mRNA and protein. Remarkably, an IL-10 mRNA signal could be detected in NHK after a second round of PCR amplificat ion. Because NHK suspensions are contaminated with Langerhans cells, m elanocytes and possibly fibroblasts, we tested purl populations of eac h individual cell type to determine the origin of this IL-IO mRNA. Our results clearly indicate that NHK, Langerhans cells and fibroblasts f ail to express IL-10 and that melanocytes are the principal source of IL-IO mRNA in normal human epidermis.