Development of biomimetic environments for physiologically relevant in-vitro models

The aim of this thesis was to design and develop different systems and methods for in-vitro research, in order to create different dynamic environments for simulating different organs. This work is organized in two parts: the first part describes the optimization of a control unit able to monitor and actively control all the main parameters for performing cell cultures (i.e. temperature, pH and pressure) and its use for creating physio-pathological models. The second part concerns the development of a stand-alone and sensorized bioreactor for mechanical stimulation of engineered constructs. This system was then used to develop an in-vitro model of cardiac tissue. Firstly, an existing control unite (SUITE: Supervising Unit for In-vitro TEsting) was improved and optimized: real-time monitoring of the oxygen concentration was introduced, and the control algorithm modified in order to assure the generation of different hydrostatic pressures for long time. Moreover, additional tools were developed for allowing the connection of the control system to different bioreactors with fluid flow (i.e. single flow, double-flow with membrane interface, applying mechanical stimuli). Then, the system was used to simulate two different physio-pathological conditions: a pathological liver model for portal hypertension, testing the ability of the control system to create a controlled hydrostatic pressure in a single flow bioreactor, and a physiological model of the intestine, taking advantage of the pH control and regulation applied by SUITE, and using a double flow bioreactor with membrane. In the latter model, the physical environment inside the bioreactor chamber will also be carefully characterized in term of fluid-induced forces on the intestinal epithelium, in order to evaluate the effect of such stimulus on the cell barrier. In the second part, a Sensorized Squeeze PRessure bioreactor (S2PR) was designed, tested and finally validated on cardiac cells. The system is able to create a cyclic hydrodynamic pressure on the cell-seeded construct by the controlled movement of a piston inside a fluid-filled chamber. In order to assure high usability of the system, the bioreactor was developed to be stand-alone, automatically finding the starting position of the piston to apply the desired stimulus according to a user-defined cycle. The flexibility of the S2PR was evaluated using cardiac cells seeded on different constructs, as dynamic cardiac model with fluid-induced forces and pressures. Moreover, the device is connectable with the previous optimized control unit, to create complex patterns of physical stimuli with combination of hydrostatic and hydrodynamic stimuli, pH control and oxygen monitoring. The results of this study can be applied to several fields of biological sciences, like drug discovery and testing, toxicology, tissue engineering and regeneration, or development of disease models or personalized therapy. These areas critically need more relevant in-vitro models to better predict human response to external stimuli such as chemical substances or physical environmental changes, in order to improve the results of in-vitro studies and reduce the animal use in research