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Perfusion and electromagnetic stimulation bioreactor for bone tissue engineering: optimization, characterization and validation tests

Beatrice Masante

Perfusion and electromagnetic stimulation bioreactor for bone tissue engineering: optimization, characterization and validation tests.

Rel. Diana Nada Caterina Massai, Cristina Bignardi, Stefano Gabetti. Politecnico di Torino, Corso di laurea magistrale in Ingegneria Biomedica, 2021


Non-invasive pulsed electromagnetic field (PEMF) stimulation is largely adopted in clinical practice for boosting the endogenous repair of bone fractures. However PEMF is applied empirically in clinics, without standardized guidelines, since many biological mechanisms and signalling pathways modulated by PEMF stimulation remain unknown. With a view to develop a reliable in vitro investigation methodology for studying the precise combination of physical parameters needed to reach a desired biological effect, the aim of this thesis was the optimization and validation of a versatile bioreactor that combines tunable direct perfusion and PEMF stimulation for in vitro investigating three-dimensional (3D) bone-like tissues under defined physical stimuli. The bioreactor, to be incubated, is composed of: 1) culture chamber; 2) perfusion unit; 3) PEMF stimulation unit. The 3D-printed culture chamber (priming volume=2.5ml) consists of two screwable cylindrical parts and allows housing scaffolds of different sizes press-fit in tailored silicone holders, for guaranteeing direct perfusion. The culture chamber is part of a closed-loop perfusion circuit, based on a peristaltic pump and controlled by a customized Arduino-equipped control box, which enables uni- or bi-directional flow (0.006-24ml/min) for scaffold cell seeding or perfusion culture. The PEMF stimulation is provided by a commercial device (1.5mT, 75Hz). To support the design optimization phase, multiphysics simulations (COMSOL Multiphysics) were performed, enabling characterizing the flow and magnetic fields developing within the culture chamber. For the 2D stationary computational fluid dynamic (CFD) analysis, the construct was modelled as a porous medium, and flow rates of 0.1 and 0.3 ml/min were imposed. For the 3D stationary and 2D time-dependent magnetic field modeling, 6 domains with different electrical properties were set. As exploratory biological tests, human mesenchymal stem cells were seeded into commercial porous scaffolds and, after 48 h of static culture, the constructs were transferred into the culture chamber and exposed to unidirectional flow (0.3ml/min, 2h/day w/o PEMF stimulation) for additional 15 days. Control constructs were statically cultured for 15 days. During the culture, cell viability and release of the early osteogenic marker alkaline phosphatase (ALP) were assessed. Adopting an iterative process of design, modeling, prototyping, and testing, the bioreactor was optimized and validated. Within the culture chamber, the culture medium flows through the construct following an S-shaped path that prevents air bubbles and stagnation/recirculation regions, as confirmed by the CFD results. Magnetic field modeling showed that the construct is exposed to a homogeneous magnetic field of 1.5mT. The cell viability analysis showed no significant differences between perfused and control constructs, while direct perfusion significantly increased the ALP release compared to control, promoting the formation of a bone-like construct. Systematic tests on bone-like constructs cultured under individual/combined biophysical stimuli are ongoing. The proposed bioreactor provides, for the first time, a controlled 3D dynamic culture environment in which direct perfusion and PEMF stimulation can be combined, allowing an in-depth investigation of the biological effects exerted by PEMF exposure on bone tissue, with the final goal of providing guidelines for improved therapeutic treatments.

Relators: Diana Nada Caterina Massai, Cristina Bignardi, Stefano Gabetti
Academic year: 2020/21
Publication type: Electronic
Number of Pages: 81
Additional Information: Tesi secretata. Fulltext non presente
Corso di laurea: Corso di laurea magistrale in Ingegneria Biomedica
Classe di laurea: New organization > Master science > LM-21 - BIOMEDICAL ENGINEERING
Aziende collaboratrici: Politecnico di Torino
URI: http://webthesis.biblio.polito.it/id/eprint/19651
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