Minimal contraction for tissue-engineered skin substitutes when matured at the air–liquid interface

The structural stability of skin substitutes is critical to avoid aesthetic and functional problems after grafting, such as contractures and hypertrophic scars. The present study was designed to assess the production steps having an influence on the contractile behaviour of the tissue-engineered skin made by the self-assembly approach, where keratinocytes are cultured on tissue-engineered dermis comprised of fibroblasts and the endogenous extracellular matrix they organized. Thus, different aspects were investigated, such as the assembly method of the engineered dermis (various sizes and anchoring designs) and the impact of epithelial cell differentiation (culture submerged in the medium or at the air–liquid interface). To evaluate the structural stability at the end of the production, the substitutes were detached from their anchorages and deposited on a soft substrate, and contraction was monitored over 1 week. Collected data were analysed using a mathematical model to characterize contraction. We observed that the presence of a differentiated epidermis significantly reduced the amount of contraction experienced by the engineered tissues, independently of the assembly method used for their production. When the epidermis was terminally differentiated, the average contraction was only 24 4% and most of the contraction occurred within the first 12 h following deposition on the substrate. This is 2.2-fold less compared to when the epidermis was cultured under the submerged condition, or when tissue-engineered dermis was not overlaid with epithelial cells. This study highlights that the maturation at the air–liquid interface is a critical step in the reconstruction of a tissueengineered skin that possesses high structural stabilit