Insulin-transferrin-selenium as an alternative to foetal serum for epidermal equivalents.

Abstract

Organotypic skin models are powerful tools for research in development, ageing and diseases. They have become more and more complex with the use of multiple cell types. This requires a culture medium adapted to optimize the development of such in vitro skin. Foetal bovine serum (FBS) is the most complete supplement in existence at the moment, providing at once growth factors, vitamins, hormones and other circulating compounds. However, this cocktail suffers from batch variability and its animal origin is ethically questionable. More importantly, its biological activities may interfere with the study of certain signalling pathways. Here, we present a strategy for constructing an epidermal equivalent using a defined culture medium without serum. An epidermal equivalent was constructed with primary human keratinocytes cultured using an insulin-transferrin-selenium (ITS) medium. Determination of steady-state gene expression levels and the immunohistological characterization of keratinocyte markers were performed to compare the ITS medium condition with a reference model, where keratinocytes were co-cultured with fibroblasts in the presence of FBS. The data show that the ITS medium promoted the expression of keratinocyte proliferation and differentiation markers at the protein and transcript levels in a similar way to that of the reference model. We show that culture using the ITS medium appears as a viable replacement for FBS in the construction of epidermal equivalents, opening the way to signal transduction studies.