Construction of an organotypic model from human embryonic stem cells

Introduction: The keratinized stratified epithelium which covers the exterior of the body and areas of the oral cavity forms a protective barrier. The development of an in-vitro model that can reproduce the appropriate mechanical and permeability characteristics of the normal tissue is critical to the formulation and delivery of therapeutic compounds. Current in-vitro organotypic models of keratinized stratified tissue are highly variable and do not reproduce the barrier properties of the parent tissue. Therefore, there is a need of a novel reproducible organotypic model (OM). OM differentiated from human embryonic stem cells (hESCs) holds great potential for future applications. Objective: To develop a novel OM from hESCs for the future research and clinical application. Materials and Methods: OM was generated on polycarbonate culture inserts. For the construction of OM, hESCs derived fibroblast (H9-ebF) were seeded with collagen I in the dermal compartment. hESCs derived keratinocyte (H9-Kert) were then seeded in the epidermal compartment. Cultures were kept in FAD Medium supplemented with 1.5 mmol/L calcium chloride and 50 ìg/mL ascorbic acid at the air-liquid interface for 10-15 days. Characterization of the OM was done by immunofluorescence staining. Results: Preliminary results showed the formation of stratified keratinized multilayer epithelium. Conclusion: This study brings a novel knowledge that hESCs derived reproducible invitro OM can be used for permeability testing with the minimum use of xenogenic and allogenic sources.