Long-Term Expansion of Mouse Primary Epidermal Keratinocytes Using a TGF-β Signaling Inhibitor

Abstract

Mouse models have been used to study the physiology and pathogenesis of the skin. However, propagation of mouse primary epidermal keratinocytes remains challenging. In this chapter, we introduce a newly developed protocol that enables long-term expansion of p63⁺ mouse epidermal keratinocytes in low-Ca²⁺ media without the need of progenitor cell purification steps or support by a feeder cell layer. Pharmacological inhibition of TGF-β signaling in crude preparations of mouse epidermis robustly increases proliferative capacity of p63⁺ epidermal progenitor cells while preserving their ability to differentiate. Suppression of TGF-β signaling also permits p63⁺ epidermal keratinocytes to form macroscopically large clones in 3T3-J2 feeder cell co-culture. This simple and efficient approach will facilitate the use of mouse models by providing p63⁺ primary epidermal keratinocytes in quantity.

Keywords: Mouse models; Primary epidermal keratinocytes; Small-molecule inhibitors; TGF-signaling; Transcription factor p63.