Replicating the 3D cardiomyocyte environment in the squeeze pressure bioreactor

Cardiac cells are normally subject to mechanical stresses during hemodynamic loading/unloading in the heart, modulating their physiological development and functions. Furthermore, a 3D structure can increase differentiation of cardiac cells. Here, the SQPR (SQueeze PRes- sure) bioreactor is used to apply a cyclic hydrodynamic non-contact pressure on a H9c2 murine cardiac cell line in combination with scaffolds, to recreate a functional cardiac environment. The system is composed of a control unit and a chamber with a piston which moves up and down, creating a local overpressure on the base of bioreactor, where the cell construct lies. A CFD model identified pressure peak values around 1 kPa. The system was set to a 1 Hz cyclic loading frequency for 24 h. 2D (i.e. collagen sandwich) and 3D hydrogel (i.e. gelatin porous) scaffolds were fabricated and compressive mechanical tests were performed to evaluate the mechanical properties of both constructs. Typical values range within 1 to 10 kPa. Cellular experiments were performed seeding the 2D scaffolds with 6 · 104 cells/cm2 and the 3D constructs with 1 · 106 cells/cm3. The results showed that stimulated cells have good viability compared to controls, indicating that the SQPR does not damage the cell culture. Moreover, morphological and histological analyses show that after stimulation cells adopt a more cardiotypic phenotype with an organized cytoskeletal structure and elongated F-actin fibers.