Tissue engineering is a multidisciplinary field that is rapidly emerging as a promising approach for tissue repair and regeneration. In this approach, scaffolds which allow cells to invade the construct and guide the cells grow into specific tissue play a pivotal role. Electrospinning has gained popularity recently as a simple and versatile method to produce fibrous structures with nano- to microscale dimensions. These electrospun fibers have been extensively applied to create nanofiber scaffolds for tissue engineering applications. Specifically for bone and cartilage tissue engineering, polymeric materials have some attractive properties such as the biodegradability. Ceramic scaffolds and implant coatings, such as hydroxyapatite and silica-based bioglass have also been considered as bone graft substitutes for bone repair because of their bioactivity and, in some cases, tunable resorbability. Besides tissue engineering scaffolds, for clinical application, especially for load-bearing artificial implants, metallic materials such as titanium are the most commonly used material. Osseointegration between bone and implants is very essential for implant success. To achieve better osseointegration between bone and the implant surface, three dimensional porous structures can provide enhanced fixation with bone by allowing tissue to grow into the pores. In this study, pre-3D electrospun polymer and ceramic scaffolds with peptide conjugation and 3D titanium scaffolds with different surface morphology were fabricated to testify the osteoblast and mensechymal stem cell attachment and differentiation. The overall goal of this thesis is to determine if the peptide functionalization of polymeric scaffolds and physical parameters of ceramic and metallic scaffold can promote osteoblast maturation and mesenchymal stem cell differentiation in vitro to achieve an optimal scaffold design for greater osseointegration. The results of the studies showed with functionalization of MSC- specific peptide, polymer scaffolds behaved with higher biocompatibility and MSC affinity. For the ceramic and metallic scaffolds, microstructures and nanostructures can synergistically promote osteoblast maturation and 3D micro-environment with micro-roughness is a promising design for osteoblast maturation and MSC differentiation in vitro compared to 2D surfaces.

In the first part of these studies, we have performed standard microsurgical procedures provide a solution for long standing bone and soft tissue defects, even in cases of longstanding osteomyelitis of long bones. When long bony segments are missing, the microvascular bone transfer provides a reliable method. In smaller soft tissue and bone defects, the application of a descending genicular osteomyocutaneous flap provides an option with low donor site morbidity. In the second part, we have focussed on reducing the donor site morbidity and expanded on the application of tissue engineering methods. MSCs derived from bone marrow can be injected percutaneous or be combined with an autologous bony scaffold for treatment of delayed union and nonunion. The outcome of our studies, however, limited in number of patients, clearly showed the possibilities and advantages of this new approach. A multimodality approach is essential, but it can provide promising solutions. Well-established microvascular and modern biotechnology methods will improve patient satisfaction and functional recovery in severe limb trauma, often the result of high-energy motorcycle accidents. / Doctorat en Sciences médicales (Médecine) / info:eu-repo/semantics/nonPublished

Les traitements actuels des lésions osseuses maxillo-faciales ont été éprouvés. La greffe autogène présente les propriétés idéales, mais montre des inconvénients : douleurs chroniques, infection… Certains comblements proposés ne permettent pas une néoformation vasculaire, garante de la viabilité des tissus régénérés pour des lésions importantes. Il faut alors développer des implants avec les caractéristiques recherchées et trouver les moyens de lutter contre le risque infectieux. Ce travail présente les résultats de recherches menées sur la fabrication d’implants nanofibreux mimant la MEC du tissu osseux, dotés d’une porosité favorable à une formation vasculaire et pouvant être fonctionnalisés par des facteurs de croissance / des cellules. Une réflexion éthique est menée sur le développement de ces avancées et sur leurs applications afin de garantir qu’elles constituent un réel progrès pour les patients. Il est aussi montré que l’on peut améliorer la sécurité des soins dans le traitement des lésions osseuses maxillo-faciales en développant des équipements dans le champ de l’hygiène et par la mise en place de procédures visant à évaluer leur efficacité. / Current treatments of maxillofacial bone defects have now been proven. Only the autogenous graft presents the ideal properties but shows complications: chronic pain, infection... Some bone filling techniques that are currently available do not allow the formation of blood vessels, guaranteeing the sustainability of the regenerated tissue for large lesions. It is then necessary to develop implants in that way, and to find ways to fight effectively the risk of infection. This work presents the results of research conducted on the fabrication of nanofibrous implants mimicking the ECM of bone tissue, with a porosity that is favorable to a vascular formation. These implants can be functionalized with growth factors / cells. Ethical considerations are provided on the development of these advances, but also on their applications to ensure that these developments constitute a real progress in the interest of patients. Moreover, this work shows that it is possible to improve the safety of care in the treatment of maxillofacial bone defects, with the development of equipment in the field of hygiene and the establishment of procedures to assess their effectiveness.

L'objectif de ce travail de thèse est de dégager des méthodes et des outils permettant de mieux comprendre le rôle joué par les phénomènes de transport (cellulaire, hydraulique et chimique) dans la mise au point de stratégies thérapeutiques de réparation osseuse. Pour cela, nous avons choisi d'associer deux approches : la réalisation d'études expérimentales et la mise au point de modèles numériques. Nous avons ainsi pu, lors d'une première étude présentée dans le chapitre 2 de ce document, relier la perméabilité intrinsèque d'un milieu poreux, paramètre déterminant dans l'étude du transport de fluide en son sein, à la structure géométrique de ses pores. Nous avons également mis en évidence l'importance des interactions électrochimiques lors de la progression d'une solution ionique (telle que les fluides physiologiques) à travers le tissu osseux, en raison de la structure poreuse et de la composition chimique (présence de fibres de collagènes chargées par exemple) de ce dernier. Ces outils ont ensuite permis d'analyser, en première approche, les résultats expérimentaux obtenus lors de la réalisation de tests de perméabilité sur des échantillons de périoste fémoral ovin, dans le but d'identifier les phénomènes physico-chimiques à l'origine du comportement particulier de cette membrane (chapitre 5). Nous nous sommes par ailleurs intéressés au développement d'implants osseux associant un substrat minéral biocompatible et des cellules souches mésenchymateuses, afin de favoriser une reconstruction tissulaire en volume des lésions de grande taille. Nous avons ainsi pu mettre en place, dans le chapitre 3, un dispositif expérimental permettant de réaliser de manière reproductible un test d'ensemencement cellulaire et d'évaluer le nombre, la répartition et le taux de viabilité des cellules greffées sur le biomatériau utilisé. A partir des résultats expérimentaux issus des tests d'ensemencement cellulaire, nous avons ensuite développé un modèle numérique dans le chapitre 4, pour dégager un ensemble de critères à respecter dans l'élaboration d'un substitut osseux qui favoriserait un développement tissulaire homogène contrôlé lors des premières étapes de la culture in vitro de ce type d'implants. Ce modèle constitue une première étape dans la détermination d'un cahier des charges géométrique de tels substrats / This study aims to set up methods and tools to improve our understanding of the role played by transport phenomena (transport of cells, fluid and chemical species) in the development of new therapeutic protocols for bone reconstruction, using a double approach: experimental studies and numerical simulations. Hence, in the second chapter of this document, we have been able to link the intrinsic permeability of a porous medium – a key parameter regarding fluid transport through porous media – to the geometric structure of its pores. We have also highlighted the influence of electrochemical interactions on the flow of an ionic solution (such as physiologic fluids) through cortical bone, due to its porous structure and its chemical composition (presence of electrically charged fibers). These tools have then enabled us to analyze, at first glance, the experimental results of permeability tests conducted on ovin femoral periosteum, to identify the chemical-physical phenomena responsible for the specific behavior of this membrane (chapter 5). We also focused on the development of large bone implants coupling a mineral substitute and mesenchymal stem cells to enhance a volumic reconstruction of critical-sized bone defects. We have therefore designed, in chapter 3, a custom experimental set up that allows one to perform a reproducible cell seeding test on a porous scaffold and quantify the number of seeded cells as well as their viability rate. The experimental results provided by these tests have then initiated the numerical model exposed in chapter 4, that aims to highlight criteria to meet regarding the design of new bone substitutes that would enhance a homogeneous volumic tissue growth during the first stages of the extit [in vitro} development of coupled implants

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Magnetic resonance imaging is a non-invasive imaging modality that is used to produce detailed images of soft tissues within the human body. Typically, MRI scanners used in the clinical setting are high field systems because they have a magnetic field strength greater than 1.5 Tesla. The high magnetic field offers the benefit of high spatial resolution and high SNR. However, low filed systems can also produce high resolution MR images with the added benefit of imaging stiffer samples. In this study, a low field 0.5 T MR system was used to image various samples to demonstrate the capability of the low field system in acquiring MR images with resolution comparable to high field systems. Furthermore, the MR system was modified to one capable of performing low field MR Elastography (MRE), a technique that can non-destructively measure the mechanical properties of soft samples. Agarose gel phantom of 0.5% wt. and 1.0% wt. were used to validate the MRE system. Additionally, a rat brain was used to assess the sensitivity of the MRE system in measuring the mechanical properties of small tissues. The results illustrated that the low field MR system can acquire high resolution images and provide sufficient tissue contrast (e.g through long TE times (80 ms), which is not possible with high field systems). MRE results on gel phantoms illustrated the capability of the low field system to accurately measure the mechanical properties and the MRE testing of rat brain demonstrated the potential of the system to study biological tissues. Finally, the capability of low field MRI and MRE to assess the growth of tissue engineered bone has the potential to transform the field of tissue engineering.

The ever increasing need for technology development requires the integration of inexpensive, light weight and high strength materials which are able to meet the high standards and specifications for various engineering applications. The intention of this work is to show that the suitable material selection and the utilization of plasma spray processing can be of potential interest to a large number of industrial, biomedical and everyday life applications. This research demonstrates also that plasma processing is a promising engineering tool for multifunctional coatings and near-net-shape manufacturing. Further, the theoretical and experimental results are combined in order to explain the mechanisms behind nanostructure retention and enhanced properties. Proper design of experiments, an appropriate material selection and experimental methodology are discussed herein. The experimental conditions were optimized in order to achieve the best materials properties according to their explicit properties and functions. Specific materials were consolidated according to their prospective performance and applications: 1) Plasma spraying of nano-Ceria-stabilized Zirconia free form part for stem cells scaffolds, 2) Plasma spraying of FeCrAlY on Ti-alloy plate, additionally coated with nano-size Hydroxyapatite for bone tissue engineering, 3) Wire-arc spraying of nano-based steel wires for aerospace and automotive applications. The performance and characteristics of all of the developed coatings and free-form-parts are evaluated using state-of-the art characterization techniques.

Biological hydrogels are highly promising materials for bone tissue engineering (BTE) due to their high biocompatibility and biomimetic characteristics. However, for advanced and customized BTE, precise tools for material stabilization and tuning material properties are desired while optimal mineralisation must be ensured. Therefore, reagent-free crosslinking techniques such as high energy electron beam treatment promise effective material modifications without formation of cytotoxic by-products. In the case of the hydrogel gelatin, electron beam crosslinking further induces thermal stability enabling biomedical application at physiological temperatures. In the case of enzymatic mineralisation, induced by Alkaline Phosphatase (ALP) and mediated by Calcium Glycerophosphate (CaGP), it is necessary to investigate if electron beam treatment before mineralisation has an influence on the enzymatic activity and thus affects the mineralisation process. The presented study investigates electron beam-treated gelatin hydrogels with previously incorporated ALP and successive mineralisation via incubation in a medium containing CaGP. It could be shown that electron beam treatment optimally maintains enzymatic activity of ALP which allows mineralisation. Furthermore, the precise tuning of material properties such as increasing compressive modulus is possible. This study characterizes the mineralised hydrogels in terms of mineral formation and demonstrates the formation of CaP in dependence of ALP concentration and electron dose. Furthermore, investigations of uniaxial compression stability indicate increased compression moduli for mineralised electron beam-treated gelatin hydrogels. In summary, electron beam-treated mineralized gelatin hydrogels reveal good cytocompatibility for MG-63 osteoblast like cells indicating a high potential for BTE applications.

In this study, we integrated different calcium sources into sol-gel hybrid glass scaffolds with the aim of producing implants with long-lasting calcium release while maintaining mechanical strength of the implant. Calcium(II)-release was used to introduce bioactivity to the material and eventually support implant integration into a bone tissue defect. Tetraethyl orthosilicate (TEOS) derived silica sols were cross-linked with an ethoxysilylated 4-armed macromer, pentaerythritol ethoxylate and processed into macroporous scaffolds with defined pore structure by indirect rapid prototyping. Triethyl phosphate (TEP) was shown to function as silica sol solvent. In a first approach, we investigated the integration of 1 to 10% CaCl2 in order to test the hypothesis that small CaCl2 amounts can be physically entrapped and slowly released from hybrid glass scaffolds. With 5 and 10% CaCl2 we observed an extensive burst release, whereas slightly improved release profiles were found for lower Calcium(II) contents. In contrast, introduction of melt-derived bioactive 45S5 glass microparticles (BG-MP) into the hybrid glass scaffolds as another Calcium(II) source led to an approximately linear release of Calcium(II) in Tris(hydroxymethyl)aminomethane (TRIS) buffer over 12 weeks. pH increase caused by BG-MP could be controlled by their amount integrated into the scaffolds. Compression strength remained unchanged compared to scaffolds without BG-MP. In cell culture medium as well as in simulated body fluid, we observed a rapid formation of a carbonated hydroxyapatite layer on BG-MP containing scaffolds. However, this mineral layer consumed the released Calcium(II) ions and prevented an additional increase in Calcium(II) concentration in the cell culture medium. Cell culture studies on the different scaffolds with osteoblast-like SaOS-2 cells as well as bone marrow derived mesenchymal stem cells (hMSC) did not show any advantages concerning osteogenic differentiation due to the integration of BG-MP into the scaffolds. Nonetheless, via the formation of a hydroxyapatite layer and the ability to control the pH increase, we speculate that implant integration in vivo and bone regeneration may benefit from this concept.

Characterization of 3D printed polyester scaffolds modified by nano-hydroxyapatite for bone tissue engineering