Dynamic mechanical stimulations induce anisotropy and improve the tensile properties of engineered tissues produced without exogenous scaffolding.

Mechanical strength and the production of extracellular matrix (ECM) are essential characteristics for engineered tissues designed to repair and replace connective tissues that are subject to stress and strain. In this study, dynamic mechanical stimulation (DMS) was investigated as a method to improve the mechanical properties of engineered tissues produced without the use of an exogenous scaffold, referred to as the self-assembly approach. This method, based exclusively on the use of human cells without any exogenous scaffolding, allows for the production of a tissue sheet comprised of cells and ECM components synthesized by dermal fibroblasts in vitro. A bioreactor chamber was designed to apply cyclic strain to engineered tissues in order to determine if dynamic culture had an impact on their mechanical properties and ECM organization. Fibroblasts were cultured in the presence of ascorbic acid for 35 days to promote ECM production and allow the formation of a tissue sheet. This sheet was grown on a custom-built anchoring system allowing for easy manipulation and fixation of the tissue in the bioreactor. Following the 35 day period, tissues were maintained for 3 days in static culture (SC), or subjected either to a static mechanical stimulation of 10% strain, or a dynamic DMS with a duty cycle of 10% uniaxial cyclic strain at 1Hz. ECM was characterized by histology, immunofluorescence labeling and Western blotting. Both static and dynamic mechanical stimulation induced the alignment of assessed cytoskeletal proteins and ECM components parallel to the axis of applied strain and increased the ECM content of the tissues compared to SC. Measurement of the tensile mechanical properties revealed that mechanical stimulation significantly increases both the ultimate tensile strength and tensile modulus of the engineered tissues when compared to the non-stimulated control. Moreover, we demonstrated that cyclic strain significantly increases these parameters when compared to a static-loading stimulation and that mechanical stimulation contributes to the establishment of anisotropy in the structural and mechanical properties of self-assembled tissue sheets.