Spatially Guided Construction of Multilayered Epidermal Models Recapturing Structural Hierarchy and Cell-Cell Junctions.

A current challenge in three-dimensional (3D) bioprinting of skin equivalents is to recreate the distinct basal and suprabasal layers and to promote their direct interactions. Such a structural arrangement is essential to establish 3D stratified epidermis disease models, such as for the autoimmune skin disease pemphigus vulgaris (PV), which targets the cell-cell junctions at the interface of the basal and suprabasal layers. Inspired by epithelial regeneration in wound healing, we develop a method that combines 3D bioprinting and spatially guided self-reorganization of keratinocytes to recapture the fine structural hierarchy that lies in the deep layers of the epidermis. Here, keratinocyte-laden fibrin hydrogels are bioprinted to create geographical cues, guiding dynamic self-reorganization of cells through collective migration, keratinocyte differentiation and vertical expansion. This process results in a region of self-organized multilayers (SOMs) that contain the basal to suprabasal transition, marked by the expressed levels of different types of keratins that indicate differentiation. Finally, we demonstrate the reconstructed skin tissue as an in vitro platform to study the pathogenic effects of PV and observe a significant difference in cell-cell junction dissociation from PV antibodies in different epidermis layers, indicating their applications in the preclinical test of possible therapies.