Use of Human Skin Recombinants as an in vitro Model for Testing the Irritation Potential of Cutaneous Irritants

Abstract

Two human skin recombinants, the epidermis reconstructed on the deepidermized dermis (RE-DED) or on fibroblast-populated collagen matrix (Living Skin Equivalent, LSE^TM), were used to study the irritating effect of sodium lauryl sulfate (SLS). The extent of cytotoxicity induced after a 24-hour exposure period to increasing concentrations of SLS (0-5%) was evaluated on the basis of (1) morphological perturbations, (2) changes in the expression of differentiation-specific protein markers (keratin 1, 10, 6, 16, involucrin and transglutaminase), (3) cell membrane integrity (LDH leakage) and (4) release of proinflammatory mediators (PGE₂, IL-1, IL-6 and IL-8). SLS induced significant changes in epidermal morphology and changes in the expression and localization of differentiation-specific protein markers when applied topically in concentrations higher than 1 % on RE-DED and higher than 0.1% on LSE. The exposure of both human skin recombinants to SLS resulted in a dose-dependent release of LDH, PGE₂and IL-1Α and in an increase in keratinocyte intracellular IL-1 levels. Upon application of 5% SLS on RE-DED the total (intra- and extracellular) IL-1 levels remained high but due to cell damage the intracellular IL-1 level was markedly decreased and the extracellular IL-1 level increased. Similar observations have been made with LSE after application of 0.5% SLS. However, with LSE the extracellular IL-1Α levels were found to be about 100 times lower than those measured with RE-DED. Exposure of LSE to SLS induced a marked increase of IL-6 production in fibroblasts incorporated in the collagen matrix. Contrary to LSE, both intra- and extracellular levels of IL-6 were low in unexposed controls and were only marginally modulated by the exposure of the RE-DED to SLS. In addition, a dose-dependent increase in IL-8 release was observed upon application of SLS on RE-DED. The results of the present study indicate that concentrations of SLS required to induce epidermal irritancy in vitro approximate those inducing irritation in human skin in vivo. All parameters used in the present study for evaluation of toxicity can serve as useful endpoints for screening of contact skin irritancy in vitro. Compared to RE-DED, the LSE seems to be more susceptible to SLS. The differences in sensitivity between LSE and RE-DED can be ascribed to reported differences in their stratum corneum barrier function.

Characterization of Reconstructed Skin Models

“…However, for application of skin models in large-scale studies, quality control is of crucial importance. Since these models have mainly been used for relatively small-scale and in-house toxicity studies by research laboratories [8][9][10][11], their suitability for standardized screening and prediction of skin irritation and penetration in humans has not yet been fully accepted by regulatory authorities.…”

Comparative Study of Molecular Mechanisms of Skin Irritation After Acute Exposure to m-XYLENE in Rats and Guinea Pigs

“…These cytokines are considered as a critical signal in the cascade of events leading to skin irritation and inflammation (Barker, et al, 1991;Ponec and Kempenaar, 1995). Among the number of cytokines, IL-l a is a major interleukin present in keratinocytes (Kupper, et al, 1986) and is associated with skin irritation (Corsini, et al, 1996a).…”

Irritant Contact Dermatitis: Mechanisms to Repair

“…Different mechanisms have been associated with ICD and these are largely dependent on the nature of the irritant. Experimental and animal models have demonstrated that solvents such as acetone extract lipids from the stratum corneum [13], and anionic surfactants like sodium lauryl sulphate (SLS), cocbetaine and sodium dodecan sulphonate (SDS) damage keratin, involucrin, profilaggrin, and other protein structures responsible for preventing oversaturation of the stratum corneum and disorganization of the lipid bilayers [14][15][16].…”