Design, Fabrication and Characterization of a Multimodal Cell Stimulation Bioreactor for Bone Tissue Engineering

Bone tissue disorders arising from trauma or pathological issues pose a significant clinical burden worldwide, due to the growth and aging of the global population and the cost of associated medical interventions. The gold standard of surgical procedures for bone disease are bone grafts (autografts, allografts or xenografts) that consist in replacing the damaged tissue with a healthy one, explanted from the patient or from a donor. However, these treatments are restricted by the limited availability of bone and by infections that follow the harvesting of the tissue, as well as the risk of the graft rejection. In this context the Bone Tissue Engineering represent a possible alternative strategy, heading to the in vitro fabrication of viable engineered tissue substitutes, characterized by controlled mechanical, topographical, chemical and electrical properties. It mainly involves 1) a cell osteo-competent source; 2) a compliant biomaterial as a scaffold for cell adhesion and proliferation; 3) a proper biochemical and biophysical in vitro stimulation of the artificial biological construct to allow for cell differentiation into osteoblast before the implantation in vivo. This engineered approach leads to the development of a highly controlled and automated system- the Bioreactor- that provides proper cell/scaffold culture conditions and integrates biomimetic stimulation systems to induce the desired differentiation bioprocess. Regarding the bone tissue development, the main biophysical stimulations that should be integrated into the Bioreactor, are 1) the hydrodynamic shear-stress of the interstitial fluid flow, 2) the mechanical bending and compression exerted during physical activity, 3) the electric impulse arising from the nervous system and from the bone matrix due to the piezoelectricity of its components. The complexity of integrating many different stimulating systems into a single multimodal device, has led to the development of several unimodal Bioreactors, that can exert one or at least two of the previously described stimuli, therefore missing the biophysical mimicry of the culture environment. To overcome this issue, in this thesis work I develop a multimodal stimulation bioreactor system, inspired to a two-dimensional parallel plate system, for bone-derived cells proliferation and differentiation, projected ad hoc for hosting a piezoelectric membrane as cell culture substrate. The membrane is excited both with a fluidic system and with an integrated immersion ultrasound probe, that generates a pressure field able to vibrate the membrane and thus activating its piezoelectricity. This study aims first at pointing out the effects of these stimuli individually, thus enabling to switch on one stimulation at a time, and then, to combine them to induce an optimal osteogenic answer. To achieve a high control over the exerted stimulus, I take advantages from a FEM software-COMSOL Multiphysics- and I simulate the physical environments according to the initial conditions and the boundary conditions that I set up for the experimental testing. I conclude with suggestion for further experimental testing that more strongly validate the potentiality of the Bioreactor, and for further developments of the invention to make it suitable for other applications.