In this thesis, several types of scaffolds composed of different materials and designed structures and functions were fabricated by 3D plotting under mild conditions (room temperature and without using any organic solvent). Broad biomaterials including inorganic (such as calcium phosphate cement and mesoporous bioglass), organic (such as alginate and gelatin) and composite materials were prepared into printable pastes to plot as 3D scaffolds for bone tissue engineering. Organic/inorganic biphasic and bipartite structure, core/shell alginate/nano-hydroxyapatite and hollow fiber structure were designed and realized. Scaffolds with multi functions including suitable mechanical properties, sustained drug/protein delivery and in vitro vascularization were achievable. 3D plotting provided great achievements in the field of tissue engineering by preparing advanced scaffolds, as well as by plotting cell/matrix constructs, and even complex tissues and organs.

Le but de notre étude visait à déterminer si les matrices ECM (extracellular matrix) préparés à partir d'un modèle vertébré (Axolotl) capables de régénérer ses tissus suite à une blessure sont plus efficaces pour stimuler les réponses régénératives chez les animaux non régénérant (par exemple les mammifères). Nous avons testé la capacité de matrice ECM axolotl à améliorer la guérison des plaies cutanées dans des souris et nous les avons comparés à une matrice disponible commercialement (échafaudage Symbios PerioDerm) pour leur efficacité à favoriser la guérison des plaies. Des lésions d'excision ont été créées sur le dos de souris et les animaux ont été regroupés dans différents groupes; a-) ECM de peau axolotl décellularisée (groupe Axolotl), b-) matrice de derme acellulaire Symbios Perioderm (groupe PerioDerm), c-) grillage en titane (groupe témoin); respectivement. Les tissus des plaies ont été récoltés à des moments précis : 7 jours et 30 jours après la blessure pour évaluer la guérison des plaies. La guérison des blessures ayant reçu les différentes matrices a été comparées entre elles en utilisant le test de transillumination et des analyses histologiques. Les résultats indiquent que la ECM de peau d’axolotl décellularisée est bien tolérée par les souris, car aucun rejet n'a été observé. Le groupe qui a reçu l'ECM de la peau axolotl décellularisé a démontré une réépithélialisation, une densité cellulaire, une teneur en collagène (avec une organisation similaire à un tissu intact) et une vascularisation (angiogenèse) élevées par rapport aux groupes PerioDerm et témoins. La présence de follicules pileux était également observé dans le groupe axolotl (qui n'est pas présent dans PerioDerm et groupes de contrôle). Sur la base de nos résultats, l'hypothèse de base semble être correcte en ce qu'une matrice ECM provenant d'un régénérateur puissant semble favoriser la guérison plus efficacement chez les animaux normalement non régénérants. Cependant, des recherches supplémentaires devront être menées pour confirmer ces résultats. / The aim of our study sought to determine whether ECM scaffolds prepared from a vertebrate model (Axolotl) capable of regenerating tissues following injury are more effective at stimulating regenerative responses in non-regenerating animals (e.g., mammals). We tested the ability of axolotl decellularized ECM scaffolds to improve skin wound healing in mammalian models and compare the axolotl skin ECM scaffold to a commercially available one (Symbios PerioDerm scaffold) for efficiency in promoting wound healing. Excisional lesions were created on the back of mice, and animals in different groups were treated by; a-) decellularized axolotl skin ECM (Axolotl group), b-) Symbios Perioderm acellular dermis scaffold (PerioDerm group), d-) Titanized mesh only (Control group); respectively. Wound tissues were harvested at time points: 7- and 30-days post-wounding to assess the scaffolds impact on wound healing. Wound healing was compared between the Axolotl, PerioDerm and Control groups using transillumination test and histological analyses, Results indicate that the decellularized axolotl skin ECM is well tolerated by mammalian models, as no immune rejection was observed. The axolotl group that received the decellularized Axolotl Skin ECM demonstrated high reepithelialization, cellular density, collagen content (in a porous pattern similar to intact skin), vascularization (angiogenesis) compared to PerioDerm and control groups. The presence of hair follicles was also observed in the axolotl group (which is not present in PerioDerm and control groups). Based on our results, the basic hypothesis appears to be correct in that an ECM scaffold from a strong regenerator seems to promote healing more efficiently in non-regenerating animals. However, further research should be conducted to confirm these findings.

Annually, about 80,000 people die in the United Kingdom due to myocardial infarction, congestive heart failure, stroke, or from other diseases related to blood vessels. The current gold standard treatment for replacing the damaged blood vessel is by autograft procedure, during which the internal mammary artery (IMA) graft or saphenous vein graft (SVG) are usually employed. However, some limitations are associated with this type of treatment, such as lack of donor site and post-surgery problems that could negatively affect the patient’s health. Therefore, this present work aims to fabricate a synthetic blood vessel that mimics the natural arteries and to be used as an alternative method for blood vessel replacement. Polymeric materials intended to be used for this purpose must possess several characteristics including: (1) Polymers must be biocompatible; (2) Biodegradable with adequate degradation rate; (3) Must maintain its structural integrity throughout intended use; (4) Must have ideal mechanical properties; and (5) Must encourage and enhance the proliferation of the cells. The feasibility of using synthetic biodegradable polymers such as poly (ε- caprolactone) (PCL) and poly (lactide-co-glycolic acid) (PLGA) for fabricating tubular vascular grafts was extensively investigated in this work. Many fundamental experiments were performed to develop porous tissue- engineered polymeric membranes for vascular graft purposes through electrospinning technique to achieve the main aim. Electrospinning was selected since the scaffolds produced by this method usually resemble structural morphology similar to the extracellular matrix (ECM). Hence, four 6mm in diameter tubular shape vascular grafts PCL only, PLGA only, coaxial (core-PCL and shell-PLGA), and bilayer (inner layer-PCL and outer layer-PLGA) was designed and fabricated successfully. The structure and properties of each scaffold membrane were observed by scanning electron microscopy (SEM), and these scaffolds were fully characterized by Fourier-transform infrared spectroscopy (FT-IR), X-ray diffraction (XRD), thermogravimetric analysis (TGA), water contact angle measurements, mechanical tensile test, as well as cell culture studies were carried out by seeding human umbilical vein cells (HUVEC) and human vascular Fibroblast cells (HVF). Moreover, all polymeric grafts underwent degradation process, and the change in their morphological structure properties was studied over 12 weeks at room temperature. All scaffolds were also exposed to a controlled temperature of 37°C for four weeks, in phosphate-buffered saline solution (pH, 7.3). It was found that all scaffolds displayed exceptional fibre structure and excellent degradability with adequate steady weight-loss confirming the suitability of the fabricated scaffolds for tissue engineering applications. The coaxial and bilayer scaffolds degraded at a much slower (and steadier) rate than the singular PCL and PLGA tubular scaffolds. Coaxial grafts fabricated via coaxial needle showed an increase in their fibre diameter and pore size volume than other membranes, but also showed to have significant tensile strength, elongation at fracture, and Young’s modulus. To conclude, all scaffolds have demonstrated to be reliable to adhere and proliferate HUVEC, and HVF cells, but these cells were attracted to the PLGA membrane more than other fabricated membranes.

Indiana University-Purdue University Indianapolis (IUPUI) / Dicalcium phosphate dihydrate (DCPD) cements have been used for bone repair due to its excellent biocompatibility and resorbability. However, DCPD cements are typically weak and brittle. To overcome these limitations, the sodium citrate used as a setting regulator and the coating of poly-L-lactide acid (PLLA) technique have been proposed in this study. The first purpose of this thesis is to develop composite PLLA/DCPD scaffolds with enhanced toughness by PLLA coating. The second purpose is to examine the biocompatibility of the scaffolds. The final purpose is to investigate the degradation behaviors of DCPD and PLLA/DCPD scaffolds. In this experiment, DCPD cements were synthesized from monocalcium phosphate monohydrate (MCPM) and 𝛽-tricalcium phosphate (𝛽 –TCP) by using deionized water and sodium citrate as liquid components. The samples were prepared with powder to liquid ratio (P/L) at 1.00, 1.25 and 1.50. To fabricate the PLLA/DCPD composite samples, DCPD samples were coated with 5 % PLLA. The samples were characterized mechanical properties, such as porosity, diametral tensile strength, and fracture energy. The mechanical properties of DCPD scaffolds with and without PLLA coating after the in vitro static degradation (day 1, week1, 4, and 6) and in vitro dynamic degradation (day 1, week 1, 2, 4, 6, and 8) were investigated by measuring their weight loss, fracture energy, and pH of phosphate buffer solution. In addition, the dog bone marrow stromal stem cells (dBMSCs) adhesion on DCPD and PLLA/DCPD composite samples were examined by scanning electron microscopy. The cell proliferation and differentiation in the medium conditioned with DCPD and PLLA/DCPD composite samples were studied by XTT (2,3-Bis(2-methoxy-4- nitro-5-sulfophenyl)-2H-tetrazolium-5-carboxanilide inner salt), and alkaline phosphatase (ALP) assay, respectively. The addition of sodium citrate and PLLA coating played a crucial role in improving the mechanical properties of the samples by increasing the diametral tensile strength from 0.50 ± 0.15 MPa to 2.70 ± 0.54 MPa and increasing the fracture energy from 0.76 ± 0.18 N-mm to 12.67 ± 4.97 N-mm. The DCPD and PLLA/DCPD composite samples were compatible with dBMSCs and the cells were able to proliferate and differentiate in the conditioned medium. The degradation rate of DCPD and PLLA/DCPD samples were not significant different (p > 0.05). However, the DCPD and PLLA/DCPD composite samples those used sodium citrate as a liquid component was found to degrade faster than the groups that use deionized water as liquid component

Le travail présenté dans ce manuscrit concerne la fabrication de biomatériaux poreux à base d’acide polylactique pour les tissus conjonctifs et calcifiés en utilisant des procédés de chimie verte. Le but de cette thèse est de corréler l’influence de certains paramètres de procédés à la structure morphologique et les propriétés des mousses générées. Nous avons étudié, d’un côté, les effets de mélange d’acide hyaluronique et d’acides polylactiques afin d’améliorer les propriétés d’adhésion de ces biomatériaux. Nos résultats montrent bien une augmentation de l’énergie d’adhésion mais aussi une diminution de la taille équivalente des pores et de la porosité des biomatériaux poreux après moussage par les fluides supercritiques. D’un autre côté, nous avons étudié les effets de mélanges des triphosphates de calcium et d’acides polylactiques en tant que substitut osseux. L’influence d’un ajout de cires en tant qu’agent porogène a été discutée et les méthodes de préparation des pastilles (voie sèche ou humide) ont été analysées. Dans cette optique la fabrication semi-industrielle de biomatériaux poreux a été testée en fixant les paramètres du procédé de moussage par le CO2 supercritique (pression, température et temps de saturation, vitesse de dépressurisation) et nous avons contrôlé les mousses de formulations optimisées en termes de porosité et de distribution des pores. En conclusion, ce travail rend possible d’adapter les paramètres des procédés de CO2 supercritique et de co-broyage aux propriétés des biomatériaux poreux. En perspective, cette ouvre la voie à de nouvelles recherches à la fois dans les domaines des modèles 3D tumoraux et d’ingénierie tissulaire. / The work presented in this manuscript concerns the production of scaffolds based polylactides for connective tissues and bone regeneration by adapting green technology. The aim of this thesis was to correlate the influence of different process parameters on the morphological structures and properties of the scaffold generated. On one hand, we studied effect of the blending of hyaluronic acid and polylactides to enhance the surface adhesion properties of scaffolds. Our results relate to an increase in surface properties but a decrease of equivalent pore size and porosity after foaming scaffolds by supercritical process. Calcium Tri-Phosphate On other hand, we studied the effect of the blending of calcium tri-phosphates and polylactides as bone substitute. Influence of adding wax as porogen agent has been discussed and a comparison between wet and dry methods to generate scaffolds has been analyzed. For this purpose, semi-industrial fabrication of porous biomaterials has been tested by blocking supercritical CO2 parameters (saturation pressure, temperature and time, depressurization rate) and you have control the optimized formulation composite scaffold, in term of porosity and distribution of pores. In conclusion, this work made it possible to adapt the process parameters of supercritical CO2 and co-grinding at the properties of scaffolds. In perspective, this research opens new development ways in scaffolds, in both domains of 3D tumoral model and tissue engineering.

Thesis (Ph. D.)--University of South Florida, 2008. / Title from PDF of title page. Document formatted into pages; contains 133 pages. Includes vita. Includes bibliographical references.

La protéine kinase DYRK1A fait partie du groupe des CMGC kinases et est impliquée dans divers processus neurodégénératifs tels que la maladie d’Alzheimer.Dans ce cadre, une étude topologique a été menée autour d’un hit imidazo[1,2-B]pyridazine identifié par un criblage biologique. Ce composé a servi à concevoir des inhibiteurs ATP-Compétitifs de DYRK1A par l’utilisation de méthodes métallo-Catalysées (Pd, Cu) pour introduire divers fragments fonctionnalisés.Sur les soixante dérivés imidazo[1,2-X]azine synthétisés, sept composés ont montré une affinité nanomolaire pour DYRK1A (IC50 = 41-130 nM). En parallèle de ce travail de pharmacochimie, le développement de nouvelles méthodologies de synthèse a visé à la polysubstitution régiosélective du cycle pyridazine.En dernier lieu, nous avons donné les éléments et concepts permettant la construction de chimiothèques virtuelles dérivées de pyridazines et destinées à être criblées in silico. / DYRK1A protein kinase belongs to the CMGC group and is involved in neurodegenerative disorders such as Alzheimer’s disease.In this context we examined an imidazo[1,2-B]pyridazine hit identified by biological screening, through detailed structure-Activity relationship studies. This compound was used to synthesize DYRK1A ATP-Competitive inhibitors by using metallo-Catalyzed methodologies (Pd, Cu) in order to introduce various functionalized moieties.Out of the 60 derivatives synthesized, 7 compounds showed nanomolar activities (IC50 = 41-130 nM).Beside this work of medicinal chemistry, new synthetic methodologies has been developed to regioselectively access polysubstituted pyridazine derivatives. Finally, we developed data and concepts to establish virtual pyridazine libraries for in silico screening.

This mixed methods study compared distance and blended undergraduate environmental students at Royal Roads University (RRU) as they participated in online asynchronous PBL case discussion forums as part of an Ecotoxicology course. This study examined the differences between distance and blended teams in their activity, approaches, and levels of critical thinking in an online PBL activity. Critical thinking was evaluated using the cognitive presence indicators of the community of inquiry framework developed by Garrison, Anderson and Archer (2001). An organization indicator was added to the framework to capture posts that organized the discussion forum layout or the team and the distribution of work. The use of the organization indicator in the thread map analysis revealed that teams adopted one of two approaches to the online PBL activity, either an organic approach or an organizational scaffold approach. An open coding approach to content analysis of the posts was used to develop two coding schemes to capture the use of learning scaffolds and degree of online collaboration respectively. These coding schemes were used to compare scaffolding and collaboration behaviours of distance and blended students during the online PBL activity. The study found that whether teams used the online discussion forums or face-to-face discussion as their primary communication method influenced both the timing and the critical thinking content of the online discussion forums. Student moderators’ choices influenced the structure and approach to the PBL activity, as well as the form of document assembly that was observed in the online discussion forums. The learning scaffolds coding scheme demonstrated that both distance and blended students were reading beyond the assigned reading list. Both distance and blended students appeared to develop skills in identifying information gaps over the progression of the PBL case problems as their observable level of critical thinking remained consistent as the problem scaffolding was faded. Although both environmental and non-environmental work experience may be used to scaffold team learning, they are used differently. Online PBL is a good fit for the Royal Roads University Learning and Teaching Model and may be used to provide some consistency across blended and online course content. / Graduate

D. Tech. (Department of Biotechnology and Chemistry, Faculty of Applied and Computer Sciences), Vaal University of Technology. / The synthesis of Ag and Cu nanoparticles was carried out using the thermal decomposition method in the presence of oleylamine as a capping agent. This method was used because it can produce uniform and monodisperse nanoparticles with controlled size distribution. The nanoparticles synthesized under various conditions were characterized by transmission electron microscopy (TEM), UV/Vis spectroscopy, photoluminescence spectroscopy (PL), and X-ray diffraction (XRD). The effect of precursor concentration on the morphology and size of the nanoparticles was investigated. It was observed that an increase in the precursor concentration resulted in an increase in particle sizes with different morphologies for both Ag and Cu nanoparticles. The increase in particle sizes for Ag nanoparticles was due to Ostwald ripening, while for Cu nanoparticles it was due to agglomeration, as Cu tends to oxidize in the atmosphere, leading to a change in particle size and shape. However, the ability to control and manipulate their physical and chemical properties depends on tuning their size and shape. Therefore, varying the precursor concentration helped in selecting the optimal concentration for this study. The nanoparticles produced were used in another study as fillers or additives for the production of nanofiber composites. The development of nanofibers by electrospinning process has led to potential applications in filtration, tissue engineering scaffolds, drug delivery, wound dressing and etc. The current study is an attempt to fabricate composite nanofibers that can be used as wound dressing material for effective wound healing. The approach involves the blending of two different polymers both being biocompatible and biodegradable were one is a natural polymer and the other is a synthetic polymer. In this study, different weight ratios of CS/PVA blends, Ag/Cu/CS/PVA, Ag/CS/PVA and Cu/CS/PVA composite fibers have been successfully prepared by the electrospinning process. The tip-to-collector distance was kept at 15 cm and the applied voltage was varied from 15 to 25 kV. The effects of the weight ratios applied voltage and the nanoparticles loading on the morphology and diameter of the fibers were investigated. The resultant fibers were characterized using scanning electron microscopy (SEM), XRD, Fourier Transform Infrared Spectroscopy (FTIR), Thermogravimetric analysis (TGA) and UV-Vis spectroscopy. The SEM results showed that an increase the amount of chitosan in the CS/PVA blend resulted in a decrease in the fiber diameters while an increase in the voltage from 15 to 25 kV led to a decrease in the fiber diameters. Furthermore, an increase in fiber diameters was observed with irregular morphologies upon addition of Ag/Cu nanoparticles into the blend. The latter changes are perceived to be as a result of an increased conductivity and a higher charge density. The XRD results showed peaks which correspond to Ag in the face centred cubic. Ag peaks are more dominant than Cu peaks in the XRD of the mixed nanoparticles. The FTIR spectra of the Ag/Cu/CS/PVA composite fibers gave almost identical features as the blend. This proves that there was an interaction between CS and PVA polymer due to intermolecular hydrogen bonding. The TGA curves showed no significant effect on the thermal stability of the composite fibers upon addition of different nanoparticles loadings. The absorption spectra of the composite fibers showed an improved optical properties compared to the blend. For Ag and Cu nanoparticles composite fibers it was observed that addition of Ag nanoparticles in the blend resulted in an increase in fiber diameters with uniform morphology whereas for Cu resulted in a decrease in fiber diameters. Both Ag and Cu composite fibers showed an improved optical properties. The effect of CS/PVA, Ag/Cu, Ag, and Cu nanofibers on the selected microorganism (K.pneumoniae, S. aureus, P. aeruginosa, and E.coli) was evaluated using the disk diffusion method. It was observed that Ag/Cu/CS/PVA composite fibers showed greater activity against all microorganisms compared to Ag and Cu composite fibers. The alamar blue and Pierce Lactase dehydrogenase (LDH) assay were used to assess the effect of the blend and the composite fibers on cell viability and cytotoxicity, respectively. The results show that the prepared blend and the composite fibers did not have any toxic effect on human adipose derived stem cells (hADSC). The results also showed that as the concentration of Ag/Cu nanoparticles was increased the viability of the cells also increased after 24 hour incubation. More proliferation was observed in day 1 compared to day 3. The 30/70 blend showed more viable cell compared to the negative control. For Ag and Cu composite fibers the 30/70 CS/PVA blend increased cell proliferation after 3 days with 17% more viable cells compared to the negative control. These results show that the prepared blend with its composite fibers are biocompatible with human (ADSC) and may be suitable for use in biomedical application such as wound dressing.

Stem cell bioengineering and therapy require different model systems and materials in different stages of development. If a chemically defined biomatrix system can fulfill most tasks, it can minimize the discrepancy among various setups. By screening biomaterials synthesized through a coacervation-mediated self-assembling mechanism, a biomatrix system optimal for 2D human mesenchymal stromal cell (hMSC) culture and osteogenesis is identified. Its utility for hMSC bioengineering is further demonstrated in coating porous bioactive glass scaffolds and nanoparticle synthesis for esiRNA delivery to knock down the SOX-9 gene with high delivery efficiency. The self-assembled injectable system is further utilized for 3D cell culture, segregated co-culture of hMSC with human umbilical vein endothelial cells (HUVEC) as an angiogenesis model, and 3D bioprinting. Most interestingly, the coating of bioactive glass with the self-assembled biomatrix not only supports the proliferation and osteogenesis of hMSC in the 3D scaffold but also induces the amorphous bioactive glass (BG) scaffold surface to form new apatite crystals resembling bone-shaped plate structures. Thus, the self-assembled biomatrix system can be utilized in various dimensions, scales, and geometries for many different bioengineering applications.