Cultured human melanocytes express functional Toll-like receptors 2–4, 7 and 9

Abstract

Toll-like receptors (TLRs) recognize pathogen-associated molecular patterns (PAMPs), which have been evolutionarily conserved in microbes. Human melanocytes are not simply pigment-producing cells but also have the phagocytic capacity and can produce pro-inflammatory mediators. However, the mechanisms of recognition of microbes by melanocytes have not yet been fully established. We investigated the TLRs 1-10 expression profile in human epidermal melanocytes and assessed their functions after triggering by their specific ligands. TLRs mRNA expression was determined by RT-PCR, and the TLR protein expression was measured by flow cytometry and immunofluorescence assays. After stimulation with various TLR ligands, the production of inflammatory cytokine IL-8 and IL-6 was measured by ELISA and the mRNA for chemokine CCL2, CCL3 and CCL5 was analyzed by real-time PCR. Phosphorylation of IkappaBalpha in TLR ligands-triggered melanocytes was determined by Western blot and the nucleus translocation of NF-kappaBp65 was analyzed by immunofluorescence. Human melanocytes constitutively expressed TLRs 1-4, 6, 7 and 9 mRNA. Ample amounts of TLRs 2-4, 7 and 9 were confirmed at protein level. Stimulation of melanocytes with TLRs ligands resulted in the release of cytokines (IL-8 and IL-6) and the mRNA accumulation of chemokines (CCL2, CCL3 and CCL5). Triggering of TLRs in melanocytes resulted in the up-regulation of phosphorylated IkappaBalpha and in the nucleus translocation of NF-kappaBp65. Present study indicates human melanocytes express a panel of functional TLRs. The ligation of TLRs can turn these cells into active players of the skin innate immunity.