Interface integration of layered collagen scaffolds with defined matrix stiffness: implications for sheet-based tissue engineering.

Successful application of sheet-based engineering for complex tissue reconstruction requires optimal integration of construct components. An important regulator of cellular responses (such as migration and collagen deposition) mediating interface integration is matrix stiffness. In this study we developed a sheet-based 3D model of interface integration that allows control of interface matrix stiffness. Fluid was removed from acellular or fibroblast-seeded bilayer collagen hydrogel constructs, using plastic compression to increase collagen density and matrix stiffness. Cell-seeded constructs were either compressed at day 0 and cultured for 7 days (compressed culture, high stiffness) or left uncompressed during culture and compressed on day 7 (compliant-compressed culture, low stiffness). Constructs were fitted onto a mechanical testing system to measure interface adhesive strength. Analysis of stresses by finite element modelling predicted a sharp rise of stress and rapid failure at the interface. While cell-seeded constructs showed a six-fold increase in interface adhesive strength compared to acellular control constructs (p < 0.05), there was no significant difference between low- and high-stiffness cultures after 1 week. Cell migration across the interface was greater in low- compared to high-stiffness constructs at 24 h (p < 0.05); however, no significant difference was observed after 1 week. Visualization of interfaces showed fusion of the two layers in low- but not in high-stiffness constructs after 1 week of culture. The ability to regulate cellular behaviour at an interface by controlling matrix stiffness could provide an important tool for modelling the integration of sheet-based bioengineered tissues in bioreactor culture or post-implantation.