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Abstrato

Morphogenesis of Biologically Active Interfollicular Epidermis from Human Embryonic Stem Cell-derived Keratinocytes

Rasmussen CA, Schlosser SJ and Allen-Hoffmann BL

For patients suffering from extensive burns or chronic wounds, allogeneic cell-based therapies provide a means to restore viability and function to severely damaged cutaneous tissues. Pluripotent stem cells have been proposed as an allogeneic source for regenerative medicine applications, such as those required for burn and wound management. Ultimately the clinical utility of pluripotent stem cells relies on the ability to direct differentiation into the desired cell lineage, facilitate appropriate tissue formation, and confirm tissue-specific biological activity. We examined the capacity of human embryonic stem cell-derived keratinocytes (hES-DK) to undergo morphogenesis and form biologically active interfollicular epidermis using methods routinely employed to engineer skin substitutes for clinical applications. Throughout directed differentiation, the orderly sequence of epidermal gene expression mimicked the progression of fetal skin development. When introduced into three-dimensional organotypic culture, hES-DK cells formed a pluristratified tissue with architecture similar to that of the interfollicular epidermis. In hES-DK tissue the expression and localization of cell-cell adhesion proteins, markers of both early- and late-stage keratinocyte terminal differentiation, and host defense peptides were comparable to the patterns observed in keratinized stratified squamous epithelia generated from epidermal keratinocytes. Despite similar tissue morphology, functional analysis revealed that hES-DK tissues did not display robust cutaneous barrier function. However, hES-DK tissues were shown to possess antimicrobial activity, which represents the first demonstration of biological activity in a keratinized stratified squamous epithelium generated from a pluripotent stem cell-derived source. The successful generation of biologically active hES-derived stratified squamous epithelia represents a significant advance in the development of a hES-derived bioengineered human organ for clinical use.