Keratinocytes play an important role in skin irritation. In an attempt to investigate mechanistic bases of human skin irritation response, we recently identified the upregulation by skin irritants of adipose differentiation related protein (ADRP) in reconstituted human epidermis. ADRP is a lipid-storage-droplet-associated protein, governing deposition and release of lipids from droplets. The purpose of this study was to characterize, in a human keratinocyte cell line (NCTC 2544), sodium-dodecyl-sulfate-induced ADRP expression, to identify the biochemical events that lead to ADRP expression, and to understand its function in sodium dodecyl sulfate cytotoxicity. Sodium dodecyl sulfate induced a concentration- and time-related production of ADRP that was associated with lipid droplet accumulation. Lipid accumulation following sodium dodecyl sulfate treatment was due to intracellular redistribution rather than lipid neosynthesis, as indicated by equivalent 14C-oleate and 14C-acetate incorporations. Other skin irritants, namely benzalkonium chloride, tributyltin, and 12-O-tetradecanoylphorbol 13-acetate, also induce lipid droplet accumulation. Sodium-dodecyl-sulfate-induced ADRP expression and lipid droplet accumulation were modulated by the calcium chelator BAPTA, indicating a role of calcium in ADRP induction. Decrease of sodium-dodecyl-sulfate-induced ADRP expression by specific ADRP antisense oligonucleotide resulted in increased cytotoxicity, indicating a protective role of ADRP and lipid accumulation in the process of cell damage induced by skin irritants. ADRP expression was also induced in vivo following treatment with sodium dodecyl sulfate in an experimental model of skin irritation, indicating that the in vitro model represents irritation.

Skin irritation is a complex phenomenon, and keratinocytes play an important role in it. We have recently characterized the expression and protective role of adipose differentiation related protein (ADRP) in skin irritation. In particular, ADRP expression is induced to recover functional cell membrane following the cell damage caused by skin irritants.The purpose of this study was to characterize in a human keratinocyte cells line (NCTC 2544) the biochemical events that lead to ADRP expression following SDS treatment, and in particular, to investigate the role of transcription factor SP-1. Analysis of ADRP promoter region revealed the presence of a potential binding site for the transcription factor SP-1 close to the start site. Evaluated by measuring the DNA binding activity, we found that SDS induced a dose and time related SP-1 activation, which was correlated with SDS-induced ADRP mRNA expression. Furthermore, SDS-induced SP-1 activation, ADRP mRNA expression and lipid droplets accumulation could be modulated by mithramycin A, an antibiotic that selectively binds to the GC box preventing SP-1 binding and gene expression. This demonstrated that SDS-induced ADRP expression was mediated in part through the transcription factor SP-1. In addition, SDS-induced SP-1 activation and ADRP expression could be modulated by the calcium chelator BAPTA, indicating a role of calcium in ADRP-induction.Thus, every time an irritant perturbs the membrane barrier, it renders the membrane leaky and allows extracellular calcium to enter the cells, an event that provides the upstream mechanisms initiating the signaling cascade that triggers the activation of SP-1 and culminates in the enhancement of ADRP expression, which helps to restore the normal homeostasis and ultimately repairs the to membrane.