Use of XTT-assay to assess the cytotoxicity of different surfactants and metal salts in human keratinocytes (HaCaT). A feasible method for in vitro testing of skin irritants

Abstract

Because of the increasing need of reliable skin irritation tests and in order to reduce the number of animal experiments, in vitro alternatives have to be developed. We studied four surfactants and five metal salts for their cytotoxic potency in HaCaT cells, a spontaneously immortalized human keratinocyte line. The endpoint used to assess cellular viability was metabolization of the tetrazolium salt XTT (2,3-bis(2-methoxy-4-nitro-5-sulfophenyl)-5-[(phenylamino)carbonyl]-2H -tetrazolium hydroxide). The tested substances revealed a significant rank order of their cytotoxicity at an exposure time of 24 h. It was 1) benzalkonium chloride, 2) sodium laurylsulphate, and 3) Tween 20 (polyoxyethylene sorbitanmonolaurate) and Tween 80 (polyoxyethylene sorbitanmonooleate), for the surfactants; and 1) potassium bichromate, 2) copper sulphate, 3) cobalt chloride and palladium chloride, and 4) nickel sulphate, for the metal salts. There is an excelient correlation to the rank order of their known irritative potency in vivo. Being practicable and effective, the presented XTT-assay on HaCaT cells would be well suitable for an initial orientating screening of substances, subsequently followed by irritation tests directly in humans.