Braut-Boucher, F., J. Pichon, J. Wantyghem, M.-P. Muriel, M. Giner, J. Gont and M. Aubery. Human Keratinocyte Models: Assessment of Cell Adhesion and Dermotoxicity using Fluorescent Probes. Toxic. in Vitro 1997. 11: 601-611. [Reprinted with permission from Elsevier Science]

sodium dodecyl sulfate - 00151-21-3; 2,4-dinitrophenol - 00051-28-5

To assess the molecular and cellular events that occur in the skin during biological and pharmaco-toxicological processes, we developed different in vitro models. Two major systems are described: (1) relatively simple ones such as normal human keratinocytes (NHK) grown in monolayer or continuous culture of spontaneously immortalized keratinocyte cells, the HaCaT cell line. This cell line forms a monolayer and displays the same phenotypic morphology, pattern of differentiation markers as NHK. (2). More complex models such as NHK multilayers differentiated on a synthetic porous membrane. Indeed, NHK grown at the air-liquid interface of culture inserts may undergo epidermal differentiation in 21 days (Noel-Hudson et al., 1995a). Under the same culture conditions, no stratification of the HaCaT cell line was obtained. NHK and/or HaCaT monolayers were used to study the cell surface molecules involved in heterologous cell interactions, and to estimate the cytotoxic effects of different compounds through a sensitive fluorimetric microtitration assay. When cell adhesion was measured with calcein-AM labelled lymphocytes, it appeared that lymphocytes display the same behaviour toward NHK or HaCaT cells. The importance of the activation status of each cell and the involvement of a ₂ and a _{3b 1} integrins in lymphocyte-keratinocyte interactions were demonstrated. Likewise cytotoxicity of SDS and DNP was easily and rapidly detected with calcein-AM and Alamar blue probes. Skin models in combination with fluorescent probes offer promising alternatives for assessing cell interactions as well as cytotoxic effects.