General description

Human skin contains both dermal and epidermal layers of cells connected by basement membrane ECM proteins. Dermal layers contain nerves, hair follicles, blood vessels, and fibroblasts, etc. Epidermal layers contain the squamous epithelium which consists of the basal, spinous, granular and cornified layers and serves a barrier to the outside enviromenment including pathogens and infection. In normal skin, keratinocytes replicate in the basal/parabasal layer and differentiate into the above layers. In the granular layer, keratinocytes undergo senescence and cornify generating the cornified layer. Organotypic 3D skin models can be generated by differentiating primary human keratinocytes (PHKs) using transwell culture inserts (Millicell). Briefly, PHKs are seeded onto a dermal equivalent consisting of rat tail collagen and human fibroblasts. The whole assembly is cultured until the PHK monolayer becomes confluent and then raised to the medium-air interface for 10 or more days. PHKs then stratify and differentiate into terminally differentiated squamous epithelium. Recetly, these organotypic 3D skin models have been used by the cosmetic industry as an alternative to animal testing.