| The continuous assessment of developing tissue substitutes is crucial to understand their evolution over time. However, this represents quite a challenge when thick samples must be evaluated with standard microscopy techniques. Common characterization methods are time consuming and usually result in the destruction of the culture. Real-time, in situ, non-invasive and non-destructives methods are needed to monitor the growth of large non-transparent constructs in tissue engineering. Medical imaging modalities, which can provide information on the structure and function of internal organs and tissues in living organisms, have the potential of allowing repetitive monitoring of these 3D cultures in vitro. The working hypothesis of this thesis was to establish standard noninvasive and nondestructive real-time bioreactor imaging protocols for in situ monitoring of the viability and metabolism of endothelial cells when grown in perfused 3D fibrin gel scaffolds. To achieve this goal, a culture chamber with hollow fibers was designed and a pulsatile perfusion bioreactor system, able to promote cell survival and proliferation, was constructed and validated. Standard imaging protocols in Positron Emission Tomography (PET) are not adapted to image bioreactor systems. A suitable method had to be devised using the well-known radiotracer 18F-fluorodeoxyglucose ( 18FDG), a marker of glucose metabolism. Optimal uptake conditions were determined using cell monolayers and the best parameters were then applied on perfused 3D cultures to evaluate perfusion, cell viability and emerging cell structures. After only 12 hours of culture, the cell density could be estimated and cell structures were localized within the fibrin gels after 1-2 weeks of culture. PET is a promising tool for tissue engineering with many specific tracers available that might eventually be able to reveal new information on tissue development.;Key words: Endothelial cells, Perfusion bioreactor, Positron Emission Tomography (PET), 18F-fluorodeoxyglucose ( 18FDG), Tissue Engineering, 3D cultures, Fibrin. |
