~ Pergamon 0887-2333(94)E0005-E Toxic. in Vitro Vol. 8, No. 3, pp. 361-369, 1994 Copyright © 1994 Elsevier Science Ltd Printed in Great Britain. All rights reserved 0887-2333/94 $7.00 + 0.00 INTERLEUKIN-1 PRODUCTION AFTER TREATMENT WITH NON-IONIC SURFACTANTS IN A MURINE KERATINOCYTES CELL LINE E. CORSINI*, M. MARINOVICH*, L. MARABINIt, E. CHIESARA~" and C. L. GALLI* *Laboratory of Toxicology, Institute of Pharmacological Sciences, Via Balzaretti 9, 20133 Milan and *Department of Pharmacology "Emilio Trabucchi", Via Vanvitelli 32, 20129 Milan, Italy (Received 13 July 1993; revisions received 16 December 1993) Abstract--Detergents are well known irritating agents in human as well as in animal models. Using a murine keratinocyte cell line (HEL30) changes in the interleukin-l~t profile were characterized in response to three non-ionic detergents, all widely used in the cosmetics industry. The compounds used in this study were the most active (dodoxynol-9, A-9), moderate (polyglyceryl-4-1auryl ether, PEL) and mild (PEG-20- glyceryl ricinoleate + ricinoleamide DEA, PEG) in inducing cytotoxicity, measured as lactate dehydro- genase leakage and de novo protein synthesis, on the same cell line after 2 hr of treatment. All of the surfactants tested were able to induce IL-lct production both at a secretory and cell-associated level. However, in order to achieve a similar IL-l production different concentrations of surfactants were necessary. It was possible to calculate an ECs0 for IL-la release of 52.9/~g/ml for A-9, of 293.7/lg/ml for PEL and of greater than 9000#g/ml for PEG. At the concentration of 30~g/ml no release could be detected even after 24 hr of treatment with PEL or PEG. A time-course experiment also showed significant amounts of IL-l~t 20 min after treatment with A-9. These data confirmed A-9 as the most potent of the three non-ionic detergents tested in inducing IL-la release. The surfactants were also tested in vivo using the modified Draize test. Once again A-9 was the most active, followed by PEL and PEG. Considering the key role of IL-1 in the inflammatory response, the release of this cytokine by keratinocytes in vitro could be used as a more specific (in comparison with classical cytotoxic markers) and early marker to determine the irritant potential of water-soluble chemicals. INTRODUCTION Keratinocytes make up about 95% of the cell mass of the human epidermis, and are responsible for skin integrity. Over the past decade it has become clear that keratinocytes participate actively in skin inflam- mation (reviewed by Barker et al., 1991; MacKenzie and Sauder, 1990). Keratinocytes may act as trans- ducing elements converting exogenously applied xenobiotic stimuli into the production of endogen- ously derived immunoregulatory factors (Nickoloff et al., 1990). Among these immunoregulatory factors, it has been shown that many cytokines are produced by keratinocytes, including interleukin 1~ and fl (IL-1), IL-3, IL-6, IL-8, TNF-~ and GM-CSF (McKenzie and Sauder, 1990). Most of these proteins are involved in the initiation of inflammation, leuco- cytosis, and the acute-phase response. Among these cytokines, IL-I is constitutively produced by kerati- nocytes and retained in the cell (Kupper, 1988). Every time keratinocytes are damaged by chemicals, UVB light or infective agents IL-1 is released, this being a Abbreviations: A-9 = dodoxynol-9; ELISA = enzyme-linked immunosorbent assay; IL-l=interleukin-l; LDH= lactate dehydrogenase: PEG = PEG-20-glyceryl ricin- oleate-ricinoleamide DEA; PEL = polyglyceryl-4-1auryl ether. primary event in skin defence (Kupper, 1990a). IL-1 stimulates further release of IL-I and de novo pro- duction of other cytokines such as IL-6 and IL-8 from resting keratinocytes, thus amplifying the response (Kupper, 1989). Many environmental chemicals produce contact hypersensitivity or local inflammatory responses in the skin. Many compounds that cause T cell-medi- ated allergic contact dermatitis are also primary irritants (Kupper, 1990b). Therefore one of the early and common events during the inductive phase of skin sensitization or during an irritative response may be the production by keratinocytes of cytokines, which stimulate inflammatory and immunological responses (Kimber and Cumberbatch, 1992). Surfac- tants are well known irritating agents in human as well as in animal models. Since the initial damage caused by irritants is non-specific and may be simu- lated in vitro, several models have been proposed as alternatives to the in vivo test (Gay et al., 1992; Gueniche and Ponec, 1993). For detergents, we have previously found a good rank correlation between their relative toxicity in vitro in a murine keratinocyte cell line (HEL30) and the eye irritation produced in vivo in the Draize test (Marinovich et al., 1990). Comparative surfactant toxicity followed the general order of cationic > anionic > non-ionic > amphoteric. TIV 8/3--D 361