Com o aumento da expectativa de vida e o envelhecimento da população, a medicina regenerativa vem ocupando um importante espaço visando manter a qualidade de vida da população. A engenharia de tecidos, apoiada nos avanços da biotecnologia e da nanotecnologia, vem se configurando como alternativa mais versátil e menos custosa ao reparo e transplante de tecidos e órgãos. Os arcabouços para engenharia tecidual constituídos de nanofibras têm o potencial para mimetizar a arquitetura nanométrica dos tecidos humanos, especialmente devido à grande área superficial e elevada porosidade. Para a fabricação de arcabouços de nanofibras, a técnica mais utilizada é a de eletrofiação, devido à sua alta versatilidade, e os materiais mais estudados são os polímeros biodegradáveis e biocompatíveis, que são os mais desejados para fins biomédicos. A esterilização é uma etapa crítica no processo de fabricação de produto médico implantável e pode ter impacto no desempenho dos arcabouços poliméricos. Desta forma, o objetivo deste estudo foi avaliar o impacto da esterilização por gás ozônio em arcabouços de nanofibras poliméricas eletrofiadas para engenharia de tecidos. A esterilização por ozônio foi eficaz na inativação do indicador biológico G. stearothermophilus, caracterizando eficácia na letalidade microbiana; igualmente, não se detectou crescimento microbiano no teste de esterilidade. Os arcabouços de nanofibras de poli(ácido láctico-co-ácido glicólico) tiveram suas propriedades físico-químicas, mecânicas e biológicas preservadas, mantendo o mesmo desempenho como suporte para o crescimento de fibroblastos NIH3T3 após a esterilização. Já os arcabouços de poli-caprolactona, tiveram suas propriedades alteradas e apresentaram um melhor desempenho na proliferação celular de fibroblastos L929 após a esterilização. Assim, o gás ozônio se mostrou como um método alternativo para a esterilização de nanofibras poliméricas para engenharia tecidual. / Since world population is ageing, regenerative medicine has become a growing area in the medical field in order to maintain the life quality of population. With the advance of biotechnology and nanotechnology, tissue engineering has emerged as a more versatile and less costly alternative to tissue repair and transplantation. Nanofibers have the potential to mimic the human tissue architecture at the nanometer scale, especially due to their large surface area and high porosity. Electrospinning is the most applied technique to fabricate nanofibers scaffolds mainly because of its powerful and high versatility. Many polymers can be used on the fabrication of nanofibers scaffolds; however, the biodegradable and biocompatible polymers are the most desired ones for biomedical purposes. Sterilization is a critical step in the fabrication process and might impact the performance of polymeric scaffolds. Therefore, the aim of this study was to evaluate the impact of sterilization by ozone gas on polymeric electrospun nanofibers scaffolds for tissue engineering. Ozone gas sterilization was efficient in killing the G. stearothermophilus spores, a common biological indicator used for validation of sterilization processes. The sterilization method preserved the physico-chemical, mechanical, and biological properties of poly(lactic-co-glycolic) acid nanofibers, keeping the performance of NIH3T3 proliferation on the scaffolds. On the other hand, the same sterilization method altered some properties of poly-caprolactone electrospun scaffolds, what improved L929 fibroblasts proliferation on the scaffolds after sterilization. Therefore, ozone gas was found to be a benign sterilization method for polymeric electrospun scaffolds for tissue engineering.

Para se avaliar a citoxicidade de materiais endodônticos, uma das metodologias utilizadas é a implantação de amostras dos materiais no tecido conjuntivo de pequenos animais, e para essa implantação são utilizados acondicionadores. O propósito deste estudo foi comparar as reações teciduais de alguns acondicionadores utilizados neste tipo de pesquisa, propondo desta maneira uma melhor metodologia para ser utilizada em trabalhos futuros. Foram implantados no tecido subcutâneo de 54 ratos, tubos de polietileno, bastões de dentina e cápsulas de colágeno, formando 3 grupos que permaneceram com os implantes pelos períodos de 15, 30 e 60 dias. Os espécimes dos grupos I e II foram analisados pela microscopia óptica de modo descritivo e morfométrico, considerando 5 critérios morfológicos: fibras colágenas, fibroblastos, vasos sanguíneos, células inflamatórias e outros componentes. O grupo III foi submetido, apenas, à análise descritiva. Os valores médios encontrados nos grupos I e II foram submetidos ao Teste t de Student para comparação entre os grupos nos períodos experimentais. O Teste ANOVA foi aplicado para comparar os períodos, nos grupos experimentais, e os valores significantes foram submetidos ao teste de Tukey. Os resultados obtidos revelaram que os tubos de polietileno e bastões de dentina induziram reações teciduais semelhantes, demonstrando, aos 60 dias, um comportamento de reparo. As cápsulas de colágeno foram reabsorvidas e houve recomposição morfológica do tecido subcutâneo. / With the purpose of evaluating the citotoxicity of some endodontic materials, their implantation in subcutaneous tissue of small animals is performed, with the aid of containers, which are used to carry the testing materials. The aim of this study was to compare the subcutaneous tissue response related to the implantation of polyethylene tubes, dentin tubes and collagen capsules usually used as carrier materials in this kind of methodology. 54 rats were divided in 3 groups: polyethylene tubes (Group I), dentin tubes (Group II) and collagen capsules (Group III). After 15, 30 and 60 days of implantation, the animals were killed and the specimens of group I and II were prepared for descriptive and morphometric analysis considering: collagen fibers, fibroblasts, vessels, inflammatory cells and other components. The group III (collagen capsules) was only evaluated through descriptive analyses. The average values of group I and II were submitted to T student test for comparison between the groups in the experimental periods. ANOVA-Tukey test was used to show a difference in the tissue response in the different periods to each material. The results showed similar tissue reaction between polyethylene and dentin tubes, showing organization of the tissue at 60 days. The collagen capsules were resorbed showing morphological recomposition of the subcutaneous tissue.

Dans le but de réduire la morbidité et la durée d’hospitalisation, la médecine régénérative progresse de nos jours vers le développement de techniques chirurgicales moins invasives. Cette recherche en chirurgie mini-invasive a motivé le développement de matrices injectables pour l’ingénierie tissulaire osseuse. Ces matrices doivent aussi être capables de durcir une fois injectées in situ, acquérir la forme souhaitée ainsi que des propriétés mécaniques compatibles avec le tissu hôte qu’elles doivent réparer. De nombreux hydrogels sont déjà employés pour cette application mais aucun ne remplit complètement les propriétés requises. L’objectif de cette thèse est de développer de nouveaux substituts de greffe osseuse : des hydrogels à base de biopolymères associés à des cellules osseuses pour obtenir des greffons mi-synthétiques, mi-biologiques. Des nanoparticules de phosphates de calcium sont ajoutées pour améliorer les propriétés biologiques et mécaniques des hydrogels. L’hydroxyapatite, le phosphate de calcium choisi, est attrayante à cause de ses similitudes chimiques et structurales au constituent minéral de l’os humain. Le but de ce travail est de synthétiser des nanofils d’hydroxyapatite par la méthode template et des nanopoudres d’hydroxyapatite de taille contrôlée par sonoélectrochimie pulsée déphasée. Ensuite pour améliorer les propriétés intrinsèques des structures 3D, ces nanoparticules de phosphates de calcium seront insérées dans des matrices d’hydrogel synthétisées par le laboratoire d’ingénierie ostéo-articulaire et dentaire (LIOAD) de Nantes. Des mesures de coefficient de diffusion seront suivies par des tests de cytotoxicité et de biocompatibilité de ces matériaux. Des études en sous-cutané et après implantation en milieu osseux suivront. / In order to reduce morbidity and hospital stay, regenerative medicine is nowadays moving towards the development of less invasive surgical techniques. This search for a minimally invasive surgery has motivated the development of injectable matrices for bone tissue engineering. These matrices must also be able to harden in situ once injected, acquire the desired shape and mechanical properties compatible with the host tissue it intends to repair. Many hydrogels are already used for this application but none fully meets the required properties. The objective of this thesis is to develop new bone graft substitutes: hydrogels based on biopolymers associated with bone cells to achieve half synthetic and half biological grafts. Nanoparticles of calcium phosphates are added to improve the biological and mechanical properties of hydrogels. Hydroxyapatite, calcium phosphate chosen, has attracted much attention because of its chemical and structural similarity to the mineral constituent of human bone. The aim of this work is to synthesize firstly hydroxyapatite nanowires by the template method and secondly size controlled hydroxyapatite nanopowders by out-of-phase pulsed sonoelectrochemistry. Thirdly to improve the intrinsic properties of these three-dimensional structures, those nanoparticles of calcium phosphates will be added in the matrices of hydrogel synthesized by the LIOAD. Measurements of diffusion coefficient will be followed by testing cytotoxicity and biocompatibility of those materials. A subcutaneous study and bone model study will follow.

<p>Transplantation of islets of Langerhans is a potential treatment of type 1 diabetes that aims to restore normal glucose homeostasis. Microencapsulation of islets could enable transplantation in the absence of immunosuppression, which would be beneficial as the side effects associated with immunosuppression outweigh the potential benefits of islet transplantation. Alginate is a polysaccharide that can be harvested from brown algae and is often used for microencapsulation of cells.</p><p>The aim of this study was to evaluate alginate/poly-L-lysine/alginate capsules with regard to their biocompatibility and permeability to cytokines. Moreover, the function of microencapsulated islets was studied <i>in vitro</i> as well as their ability to reverse hyperglycaemia in diabetic mice.</p><p>Microencapsulated rodent islets functioned well <i>in vitro</i>, with similar insulin release rates and glucose oxidation rates as naked islets. However, when cultured with interleukin-1β and tumour necrosis factor-α, microencapsulated islets were functionally suppressed, showing that the capsules are permeable to these cytokines. The biocompatibility of capsules varied depending on their composition. The presence of poly-L-lysine in the capsule decreased the biocompatibility. However, the biocompatibility of the capsules was improved when the coating alginate had been epimerised, i.e. enyzmatically tailored. Transplantation of microencapsulated allogeneic islets to immune competent mice lowered blood glucose concentrations up to 1 month after implantation. The success of the microencapsulated islet graft depended on the composition of the alginate/poly-L-lysine/alginate capsule used, as capsules that had poor biocompatibility failed to reverse hyperglycaemia more than transiently in athymic nude mice.</p><p>In conclusion, alginate/poly-L-lysine/alginate capsules can protect islets of Langerhans from allogeneic rejection in mice. However, the composition of the capsule is of critical importance in the success of transplantation. Epimerised alginates may provide a novel capsule with ideal properties for microencapsulation of islets of Langerhans.</p>

For decades, Metallic Glass, with its isotropic featureless structure while exhibiting outstanding mechanical properties was possible only at a high rate of quenching and with at least one dimension in the submicron regime.  This limitation was overcome with the discovery of Bulk Metallic glasses, BMGs, containing three or more elements following the additional two empirical rules of optimum geometric size differences and negative energy of mixing among the constituent elements. Since then thousands of Fe-, Ni-, Al-, Mg-, Ti- based BMGs have been discovered and comprehensively investigated mainly by groups in Japan and USA. Yet the discovery of new combinations of elements for BMGs is alchemy. We do not know with certainty which element when added will make possible a transition from being a ribbon to a bulk rod.    In this thesis we report a discovery of castable BMGs rods on substitution of Fe by nickel in an alloy of FeBNb which could otherwise have been only melt-spun into ribbons.  For example, we find that substitution of just 6 at.% of Fe raises the glass forming range, GFA, to as much as ∆Tx =40K while the other parameters for GFA like Trg, γ, and δ reach enhanced values 0.57, 0.38, and 1.40 respectively.  Furthermore, the electrical conductivity is found to increase by almost a factor of two.  Magnetically it becomes softer with coercivity 260mOe which further reduces to much lower values on stress relaxation.  Ni does not seem to carry a magnetic moment while it enhances the magnetic transition temperature linearly with Ni concentration. We have investigated the role of Ni in another more stable BMGs based FeBNbY system in which case ∆Tx becomes as large as 94K with comparable enhancement in the other GFA parameters. Due to the exceptional soft magnetic properties, Fe-based bulk metallic glasses are considered potential candidate for their use in energy transferring devices. Thus the effect of Ni substitution on bulk forming ability, magnetic and electrical transport properties have been studied for FeBNb and FeBNbY alloy systems. The role of Ni in these systems is densification of the atomic structure and its consequence. We have exploited the superior mechanical properties of BMGs by fabricating structures that are thin and sustainable.  We have therefore investigated studies on the thin films of these materials retaining their excellent mechanical properties. Magnetic properties of FeBNb alloy were investigated in thin films form (~200-400nm) in the temperature range of 5-300K. These Pulsed Laser deposited amorphous films exhibit soft magnetism at room temperature, a characteristic of amorphous metals, while they reveal a shift in hysteresis loop (exchange anisotropy, HEB=18-25Oe), at liquid helium temperature. When thickness of films is reduced to few nanometers (~8-11nm), they exhibit high transparency (&gt;60%) in optical spectrum and show appreciably high saturation Faraday rotation (12o/μm, λ= 611nm). Thin films (~200-400nm) of Ni substituted alloy (FeNiBNb) reveal spontaneous perpendicular magnetization at room temperature. Spin-reorientation transition was observed as a function of film thickness (25-400nm) and temperature (200-300K), and correlated to the order/disorder of ferromagnetic amorphous matrix as a function of temperature. These two phase films exhibits increased value of coercivity, magnetic hardening, below 25K and attributed to the spin glass state of the system.    Using the bulk and thin films we have developed prototypes of sensors, current meters and such simple devices although not discussed in this Thesis.                                         Ti-based bulk metallic glasses have been attracting significant attention due to their lower density and high specific strength from structural application point of view. High mechanical strength, lower values of young’s modulus, high yield strength along with excellent chemical behaviors of toxic free (Ni, Al, Be) Ti-based glassy metals make them attractive for biomedical applications. In the present work, toxic free Ti-Zr-Cu-Pd-Sn alloys were studied to optimize their bulk forming ability and we successfully developed glassy rods of at least 14mm diameter by Cu-mold casting. Along with high glass forming ability, as-casted BMGs exhibit excellent plasticity. One of the studied alloy (Ti41.5Zr10Cu35Pd11Sn2.5) exhibits distinct plasticity under uniaxial compression tests (12.63%) with strain hardening before failure which is not commonly seen in monolithic bulk metallic glasses. / <p>QC 20120906</p> / Hero-m

Transplantation of islets of Langerhans is a potential treatment of type 1 diabetes that aims to restore normal glucose homeostasis. Microencapsulation of islets could enable transplantation in the absence of immunosuppression, which would be beneficial as the side effects associated with immunosuppression outweigh the potential benefits of islet transplantation. Alginate is a polysaccharide that can be harvested from brown algae and is often used for microencapsulation of cells. The aim of this study was to evaluate alginate/poly-L-lysine/alginate capsules with regard to their biocompatibility and permeability to cytokines. Moreover, the function of microencapsulated islets was studied in vitro as well as their ability to reverse hyperglycaemia in diabetic mice. Microencapsulated rodent islets functioned well in vitro, with similar insulin release rates and glucose oxidation rates as naked islets. However, when cultured with interleukin-1β and tumour necrosis factor-α, microencapsulated islets were functionally suppressed, showing that the capsules are permeable to these cytokines. The biocompatibility of capsules varied depending on their composition. The presence of poly-L-lysine in the capsule decreased the biocompatibility. However, the biocompatibility of the capsules was improved when the coating alginate had been epimerised, i.e. enyzmatically tailored. Transplantation of microencapsulated allogeneic islets to immune competent mice lowered blood glucose concentrations up to 1 month after implantation. The success of the microencapsulated islet graft depended on the composition of the alginate/poly-L-lysine/alginate capsule used, as capsules that had poor biocompatibility failed to reverse hyperglycaemia more than transiently in athymic nude mice. In conclusion, alginate/poly-L-lysine/alginate capsules can protect islets of Langerhans from allogeneic rejection in mice. However, the composition of the capsule is of critical importance in the success of transplantation. Epimerised alginates may provide a novel capsule with ideal properties for microencapsulation of islets of Langerhans.

This study was aimed to synthesize low molecular weight hydroxyl terminated polyurethane acrylate polymers that can be used in biomedical applications. Acrylate end capping via inter-esterification reaction was successfully achieved with the methacryloyl chloride addition to the hydroxyl ends of the polyurethane at low temperatures. Isocyanate terminated polyurethane acrylates were also synthesized for the sake of comparison. TDI, HDI and MDI were used as diisocyanates for urethane synthesis and they were end capped with MMA and HEMA. Nature of the monomers used had an effect on thermal, morphological, and rheological properties that were interpreted in terms of the level of hydrogen bonding and degree of phase separation. Synthesized polymers were characterized by NMR, FTIR-ATR, DSC, TGA, GPC, Mass Spectroscopy, SEM and rheometry. In the literature, polyurethane acrylate polymers have been synthesized from the isocyanate terminated polyurethanes in which the urethane chains were ended with isocyanate groups. However, the toxicity of the isocyanate groups limited their biomedical applications especially in prosthetic dentistry as a soft lining material. Therefore, it is inevitable to explore the cytotoxicity of polyurethane acrylate polymers. For this purpose, silver nanoparticles that have an average particle size of 40 nm, were incorporated to the synthesized polymers. This addition, which intends to improve the degree of cytotoxicity, was successful to a certain extent.

This study aimed synthesizing hydroxylapatite (HA) and Zn2+, F- and Cl- doped HA by solution precipitation method. The synthesized compounds were sintered at 1100&deg / C for 1h. For structural characterization, density of the samples were measured by Archimedes&rsquo / method. It was observed that Zn2+ addition increased the density significantly whereas F- caused a decrease and Cl- increased the density with a little amount. XRD was applied to the samples and it was found that co-doping of Zn2+ and F- ions decreased the unit cell volume of HA with F- addition. Other compositions gave fluctuated results in terms of unit cell volumes. HA phase and a little amount of CaO phase were detected in some samples. FTIR spectroscopy was used to detect whether Zn2+, F- and Cl- ions were incorporated to the HA structure or not by observing the bands corresponding to the bonds in the molecules. The amount of addition was also detected by FTIR. Results showed that ion incorporation to the HA structure was done successfully. SEM images were v analyzed and grain sizes of samples were calculated by Rietvelt analysis. Grain sizes of the samples increased by Cl- addition and decreased by Zn2+ and/or F- addition. For mechanical characterization, Vickers microhardness test was applied. Fracture toughness was calculated from Vickers microhardness results. According to the results, the highest microhardness values were found for F- and Zn2+ co-doped samples. It was also shown that fracture toughness decreased by Zn2+ addition. However, Zn2+ and F- co-doped samples gave higher fracture toughness results when compared with pure HA. Cl- addition also decreased the fracture toughness. The best compositions in terms of structural and mechanical properties was chosen as Zn2+ and F- co-doped samples and biological characterization was applied to these samples. Saos-2 cell line was used in biological examinations. For biological characterizations, Alamar Blue&trade / assay to detect viability and alkaline phosphatase activity (ALP) assay to detect differentiation were done. It was observed that 2 mol.% Zn2+ addition increased the cell viability and alkaline phosphatase activity. 1 mol.% F- addition also improved cell viability and alkaline phosphatase activiy. SEM images were analyzed to observe the morphology of the cells on HA and selected doped HA discs. In accordance with Alamar Blue&trade / assay and alkaline phosphatase activity assay, cells showed dendritic shapes on 2 Zn and 2 Zn 1 F sample which was the indicator of good material-cell interaction. Dissolution test was also applied by immersing the samples in simulated body fluid (SBF). pH change and SEM images for Ca2+ deposition were investigated. Increase in pH change with time was observed. F- included samples gave the lowest pH change results, especially 2 Zn 1 F. Dissolution pits and some apatitic formations were observed in SEM images.

Silicon carbide (SiC) has been around for more than 100 years as an industrial material and has found wide and varied applications because of its unique electrical and thermal properties. In recent years there has been increased attention on SiC as a viable material for biomedical applications. Among these applications are those where SiC is used as a substrate material for biosensors and biotransducers, taking advantage of its surface chemical, tribological and electrical properties. In this work we have used the proven bio- and hema-compatibility of SiC to develop a viable biorecognition interface using SiC as the substrate material for myocardial infarction detection. The approach followed included the development of an electrochemical-based sensor in which 3C-SiC is used as the active electrode and where flat band potential energy changes are monitored after successive modification of the SiC with aminopropyltriethoxysilane, anti-myoglobin and myoglobin incubation. We have studied the quality of self assembled monolayers obtained by surface modification of SiC using organosilanes such as aminopropyltriethoxysilane and octadecene, which is the starting point for the immobilization of cells or proteins on a substrate. We employed this technique on 6H-SiC where we were able to control the proliferation of H4 human neuroglioma and PC12 rat pheochromocytoma cells in vitro. Finally, aminopropyltriethoxysilane (APTES) was successfully used to immobilize anti-myoglobin on the 3C-SiC electrodes as demonstrated by fluorescence microscopy results. The electrical characterization of the surfaces was performed via impedance spectroscopy and by measuring changes in flat band potential using the Mott-Schottky plot technique. Changes in flat band and impedance of the SiC/antibody/protein interface would allow us to detect changes in the space charge region of the semiconductor. However, we believe that because of the presence of surface states and different crystal defects on the 3C-SiC we did not observed repeatable results that allowed us to identify the presence of myoglobin in solution. In addition, certain modifications need to be performed to the electrochemical cell in order to confirm the presence of the myoglobin immobilized on the functionalized SiC surfaces.

published_or_final_version / Anatomy / Master / Master of Philosophy