Surfactant-induced cytotoxicity in cultures of human keratinocytes and a commercially available cell line (3T3)

Abstract

Biochemical parameters of cytotoxicity, such as the release of intracellular enzymes, appear to be useful for classification of irritant substances following in vivo chemical insult to the skin. Changes in activity of acid phosphatase (AP), a lysosomal enzyme, appear to parallel the development of the inflammatory response in laboratory animals after treatment with the known skin irritant sodium dodecyl sulphate (SDS). Neutral red (NR) uptake and AP were chosen as endpoints of cytotoxicity. NR and AP were measured in cultures of human epidermal keratinocytes, prepared from separate individuals, and in 3T3 cells following treatment with SDS. The NR 50 value (concentration producing a 50% reduction in NR uptake compared with controls) was similar in both cell types. AP in human keratinocyte cultures exhibited a peak activity, before declining at higher concentrations. This phenomenon was time dependent and was observed within 4 hr of treatment, but was not evident after a 24-hr exposure. The peak produced in 3T3 cells was negligible in comparison. The rate of NR uptake was also studied within the first 4 hr of exposure to SDS, which was comparable to the earlier time points at which AP was determined. The degree of inhibition of NR uptake was greater in human keratinocyte cultures than in 3T3 cells and a response was also elicited at lower dose levels in keratinocytes. AP may be a sensitive indicator of cytotoxicity in human keratinocytes and the data may be interpreted in relation to changes in lysosomal membrane function. This assay may be of value in assessment of skin irritation potential of aqueous soluble surfactants and chemicals that possess the ability to damage biological membranes.