In vitro reconstruction of an endothelialized skin substitute provided with a microcapillary network using biopolymer scaffolds

Successful in vitro reconstruction of skin requires the inclusion of several cell types that give rise in coculture to the specific elements present in native skin, and the appropriate scaffolding structure to house and support these cells. In addition to the two main structural components, epidermis and dermis, one critical apparatus of the skin is a capillary network that guarantees adequate perfusion of nutrients and oxygen. The aim of the present study was to develop an in vitro coculture system that assumed the human dermal-epidermal architecture and included a microcapillary network in a three-dimensional biomaterial that guaranteed ease of handling in a clinical setting. Endothelialized skin (ES) was prepared by coculturing three human cell types: keratinocytes, fibroblasts, and endothelial cells, obtained from human full-thickness skin samples, in scaffolds produced from modified hyaluronic acid. Results were evaluated by histological and immunohistochemical analyses at different time points. In vitro, engineered skin obtained with this composite culture developed into a well-differentiated upper layer of stratified keratinocytes lining a dermal-like structure, in which fibroblasts, extracellular matrix and a microvascular network were present. Furthermore, the biodegradable fabric produced from hyaluronic acid and used as the scaffolding support for this in vitro constructed skin graft greatly facilitated handling in the perioperative period.