In contrast to their murine counterparts, normal human keratinocytes and human epidermoid cell lines A431 and HaCaT fail to express IL-10 mRNA and protein

Abstract

SUMMARYIn mice, keratinocyte-derived IL-10 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 phenomenon can be detected in man. Freshly isolated or cultured normal human 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 variety of other stimuli. RNA was extracted at various time points post-stimulation and analysed by reverse transcriptase-polymerase chain reaction (RT-PCR) using four different IL-10-specific primer pairs and RNA from 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 and HaCaT cultures were negative for IL-10 protein, as tested by four different ELISAs and a bioassay. Murine keratinocytes, stimulated under comparable conditions and tested by the same techniques, displayed a strong expression of IL-10 mRNA and protein. Remarkably, an IL-10 mRNA signal could be detected in NHK after a second round of PCR amplification. Because NHK suspensions are contaminated with Langerhans cells, melanocytes and possibly fibroblasts, we tested pure populations of each individual cell type to determine the origin of this IL-10 mRNA. Our results clearly indicate that NHK, Langerhans cells and fibroblasts fail to express IL-10 and that melanocytes are the principal source of IL-10 mRNA in normal human epidermis.

cis-Urocanic Acid Initiates Gene Transcription in Primary Human Keratinocytes

“…In addition, unlike murine keratinocytes, it is debatable whether human keratinocytes produce IL-10 following UVR exposure (50). Some groups have demonstrated the production of IL-10 mRNA and its protein secretion in UV-and non-UV-irradiated keratinocytes (51,52), whereas others were unable to detect IL-10 mRNA or protein up to 24 h after UV irradiation (53)(54)(55). We were also unable to detect IL-10 mRNA or protein 24 h following UCA or SSR treatment.…”

Regulatory Activity of Autocrine IL-10 on Dendritic Cell Functions

“…Interestingly enough, primary DCs undergo spontaneous maturation in vitro on isolation from tissues such as the skin or the spleen (37,38), but the mechanisms responsible have not been defined yet. However, both human epidermal Langerhans cells and mouse spleen DCs do not express IL-10 mRNA (39,40), and this may explain, at least in part, their spontaneous maturation during shortterm culture. Also, DCs generated from CD1a ϩ progenitors (30) and Langerhans cells differentiated from monocytes or CD34 ϩ progenitors in the presence of TGF-␤ fail to synthesize IL-10 (41, 42), whereas peripheral blood CD83…”

Intradermal application of vitamin D3 increases migration of CD14+dermal dendritic cells and promotes the development of Foxp3+regulatory T cells

“…In line with in vitro data, we were able to detect elevated IL-10 levels in the cultures of skin biopsies injected with VitD, though this elevation may also result from cells other than DCs, but not KCs. 50 Nevertheless, KCs may be a major contributor in shaping immune responses following ID injection of VitD through the release of mediators that can influence DC functions. Although intradermally injected VitD rendered skin DCs tolerogenic, VitD concentrations applied in this model were 10 times higher than usually used in vitro on monocyte-derived or purified DCs and LCs.…”