Le Poly Vinyl Alcohol (PVA) a été associé aux verres élaborés dans un système quaternaire (BG) 46S6 par les procédés cités (fusion, sol-gel et sacffolds). Différents paramètres intervenant dans les synthèses des verres bioactifs ont été étudiés, nous citons à titre d’exemple : la température, le pH, la taille des particules, le rapport Polymère / verres, la microstructure, la porosité et la biodégradation. Les caractéristiques thermiques des verres élaborés ont été également déterminées après chaque synthèse par analyse thermique différentielle (DSC/TG, DTA/TG). Ainsi, la température de fusion, la température de transition vitreuse et la température de cristallisation ont été élucidées. Ces caractéristiques thermiques changent lorsque la composition chimique du verre est modifiée. A ce titre, les compositions chimiques ont été étudiées par Fluorescence (XRF) et Inductively Coupled Plasma-Opticale Emission Spectroscopy (ICP-OES) après chaque synthèse pour s’assurer de la pureté des verres bioactifs élaborés et destinés à des applications médicales. Plusieurs techniques physico chimiques d’analyses (DRX, MEB, MET, FT-IR, XRF, ICPOES) ont été mises en oeuvre pour déterminer les propriétés physico chimiques de nos verres bioactifs avant et après expérimentations « in vitro ». Le nano composite Polymère-Verres scaffolds que nous avons obtenu présente des particules de tailles comprises entre 40 et 61 nm et une porosité d’environ 85%. La biodégradation des verres scaffolds décroît lorsque la teneur en verre scaffolds dans le nano composite croît. Les expérimentations « in vitro » montrent qu’après immersion de ces nano composites dans un liquide physiologique synthétique (SBF), une couche d’apatite (phosphate de calcium) se forme à leur surface. L’épaisseur de la couche formée dépend clairement de la taille des particules et du rapport polymère / verre scaffolds. / The aim of the present work is the preparation of Bioactive Glass (BG) 46S6 by different techniques. Fabrication of composite scaffolds by using of Poly Vinyl Alcohol (PVA) and quaternary BG (two methods melting and sol-gel) with different ratios to the prepared scaffolds was carried out. Different factor affecting the final properties of the prepared composite scaffolds were investigated in this study, such as; temperature of treatment, BG particle size, polymer/glass ratio, microstructure, porosity, biodegradation, bioactivity, and drug release. The thermal behavior of the prepared bioactive glass by sol-gel and melting techniques were identified using Differential Scanning Calorimetric/Thermo Gravimetric (DSC/TG) or Differential Thermal Analysis/Thermo Gravimetric (DTA /TG). The elemental composition of the prepared bioactive glasses was determined by X-rays Fluorescence (XRF) to confirm that the prepared bioactive glasses have the same elemental compositions and high purity for biomedical applications. The particle size of the prepared bioactive glass was determined by Transmission Electron Microscopic (TEM). Nano-bioactive glass could be obtained by modified sol-gel and the obtained particle size ranged between 40 to 61 nm. The prepared bioactive glass by both applied methods has the same amorphous phase and all identified groups as well as. The porous scaffold has 85% porosity with a slight decrease by increasing the glass contents. The degradation rate decreased by increasing of glass content in the prepared scaffolds. The bioactivity of the prepared composite scaffolds was evaluated by XRD, FTIR, SEM coupled with EDX and Inductively Coupled Plasma-Optical Emission Spectroscopic (ICP-OES). It has been observed that after soaking in Simulated Body Fluid (SBF), there was an apatite layer formed on the surface of the prepared samples with different thickness depending on the glass particle size and polymer/glass ratio.

No description available.

Hydrogels are popular materials for biological applications since they exhibit properties like that of natural soft tissue and have tunable properties. Biodegradable hydrogels provide an added advantage in that they degrade in an aqueous environment thereby avoiding the need for removal after the useful lifetime. In this work, we investigated poly(β-amino ester) (PBAE) biodegradable hydrogel systems. To begin, the factors affecting the macromer synthesis procedure were studied to optimize the reproducibility of the resulting hydrogels made and create new methods of tuning the properties. Hydrogel behavior was then tuned by altering the hydrophilic/hydrophobic balance of the chemicals used in the synthesis to develop systems with linear and two-phase degradation profiles. The goal of the research was to better understand methods of controlling hydrogel properties to develop systems for several biomedical applications. Several systems with a range of properties were synthesized, and their in vitro behavior was characterized (degradation, mechanical properties, cellular response, etc.). From these studies, materials were chosen to serve as porogen materials and an outer matrix material to create a composite scaffold for tissue engineering. In most cases, a porous three dimensional scaffold is ideal for cellular growth and infiltration. In this work, a composite with a slow degrading outer matrix PBAE with fast degrading PBAE microparticles was created. First, a procedure for developing porogen particles of controlled size from a fast-degrading hydrogel material was developed. Porogen particles were then entrapped in the outer hydrogel matrix during polymerization. The resulting composite systems were degraded and the viability of these systems as tissue engineering scaffolds was studied. In a second area of work, two polymer systems, one PBAE hydrogel and one sol-gel material were altered through the addition of iron oxide nanoparticles to create materials with remote controlled properties. Iron oxide nanoparticles have the ability to heat in an alternating magnetic field due to the relaxation processes. The incorporation of these nanoscale heating sources into thermosensitive polymer systems allowed remote actuation of the physical properties. These materials would be ideal for use in applications where the system can be changed externally such as in remote controlled drug delivery.

Sterilization of tissue engineered scaffolds is an important regulatory issue and is at the heart of patient safety. With the introduction of new biomaterials and micro/nano structured scaffolds, it is critical that the mode of sterilization preserve these built-in features. Conventional sterilization methods are not optimal for engineered polymeric systems and hence alternate systems need to be identified and validated. PCL is polyester with a low melting point (heat labile), susceptible to hydrolysis and is popular in tissue engineering. Electrospinning generates some nanoscale features within the scaffold, the integrity of which can be affected by sterilization method. Chapter 1 explores the possibility of using Peracetic acid (PAA) to sterilize polymeric scaffolds that are sensitive to heat or moisture. PAA is a strong oxidizing agent that has been approved for sterilizing catheters and endoscopes. The ability of PAA to sterilize at room temperature, its breakdown into non-toxic end products and effectiveness at low concentrations are major advantages. Chapter 2 evaluates the ability of PAA-sterilized PCL scaffolds (PAA-PCL) to support survival and proliferation of mouse calvarial osteoblast cell line, MC3T3. While Ctrl-PCL scaffolds (ethanol-disinfected) showed robust cell survival, PAA-PCL scaffolds induced massive cell death. Following interrelated hypotheses are tested: the observed cytotoxicity was due adsorption of PAA and/or hydrogen peroxide onto PCL fibers during sterilization; and elimination of adsorbed residues will restore scaffold cytocompatibility. Neither extensive aeration nor chemical neutralization with sodium thiosulfate and catalase were effective in improving cell survival. However, quenching PAA-PCL scaffolds in 70% ethanol for 30 minutes effectively removed adsorbed PAA residues and completely restored cell viability and proliferation over a 7 day period. In order to test if PAA-induced toxicity was limited to electrospun PCL scaffolds, commercially available, porous polystyrene scaffolds (Alvetex®) was treated with PAA. Interestingly, a statistically significant increase in cell survival and proliferation resulted with PAA treatment and this was abolished by ethanol quenching. Combined, these results illustrate that PAA treatment can produce diametrically opposite effects on cell survival depending on substrate chemistry and that PAA can be used to effectively sterilize tissue engineering scaffolds without compromising cell viability.

Advanced biomaterials and sophisticated processing technologies aim to fabricate tissue-engineering scaffolds that can predictably interact within a biological environment at a cellular level. Sterilization of such scaffolds is at the core of patient safety and is an important regulatory issue that needs to be addressed prior to clinical translation. In addition, it is crucial that meticulously engineered micro- and nano- structures are preserved after sterilization. Conventional sterilization methods involving heat, steam and radiation are not compatible with engineered polymeric systems because of scaffold degradation and loss of architecture. Using electrospun scaffolds made from polycaprolactone (PCL), a low melting polymer, and employing spores of Bacillus atrophaeus as biological indicators, we compared ethylene oxide, autoclaving and 80% ethanol to a known chemical sterilant, peracetic acid (PAA), for their ability to sterilize as well as their effects on scaffold properties. PAA diluted in 20% ethanol to 1000 ppm or above, sterilized electrospun scaffolds in 15 min at room temperature while maintaining nano-architecture and mechanical properties. Scaffolds treated with PAA at 5000 ppm were rendered hydrophilic, with contact angles reduced to zero degrees. Therefore, PAA can provide economical, rapid and effective sterilization of heat-sensitive polymeric electrospun scaffolds used in tissue-engineering.

A engenharia de tecidos e a medicina regenerativa possuem como objetivo principal o restabelecimento morfológico/funcional de tecidos e órgãos lesionados com a utilização de células, matrizes celulares e células tronco, controlando as respostas imunológicas/bioquímicas promovidas pelo organismo. Adicionalmente, a ciência dos materiais busca desenvolver biomateriais biocompatíveis que não promovam reações imunológicas indesejadas e proporcionem o reestabelecimento das funções do tecido/órgão. Polímeros de origem natural destacam-se como biomateriais por assemelharem-se a macromoléculas biológicas, similaridade com a matriz extracelular, menor possibilidade de estimulação de inflamação crônica e baixa ou ausência de toxicidade. O presente trabalho teve como objetivo desenvolver matrizes macromoleculares originadas do óleo de soja epoxidado (OSE), analisando a relação estrutura química/atividade biológica das matrizes macromoleculares para uso como biomaterial na engenharia de tecidos. A síntese do OSE foi efetuada pela rota oleoquímica, cuja eficiência foi determinada por espectroscopia de infravermelho e o rendimento da reação de 85% determinado por ressonância magnética nuclear de prótons. A partir da análise por calorimetria exploratória diferencial, detectou-se uma diminuição da temperatura de transição vítrea do polímero do óleo de soja (POSE) em relação ao OSE, sugerindo aumento do crescimento das cadeias poliméricas do POSE. Através da análise termogravimétrica, foi possível definir o perfil de degradação do OSE, com degradação em duas etapas, e do POSE, que degrada em apenas uma etapa e demonstra maior estabilidade térmica do POSE pelo aumento das interações moleculares. A reticulação e a hidrofilicidade do POSE foram promovidas com a adição de metacrilato de 2-hidroxietila (HEMA) à formulação por enxertia do monômero pela irradiação gama. Os resultados obtidos identificaram aumento da estabilidade mecânica, da gelificação e da absorção de água com o aumento do conteúdo de HEMA. Por fim, o grau de cristalinidade estimado para esses polímeros enxertados com HEMA de 27,5% foi definido através da difratometria de raios-X. A segunda etapa caracterizou-se pelo (i) desenvolvimento de POSEs com a enxertia de HEMA nas proporções OSE/HEMA 90:10 e 65:35 com irradiação por raios gama nas doses de 50 e 100kGy, (ii) caracterização físico-química dos POSE-HEMA e (iii) análise biológica desses materiais. Através da espectroscopia de infravermelho, pode-se detectar as regiões epoxidadas do POSE, assim como o sucesso da enxertia do monômero HEMA em todas as concentrações e doses de radiação utilizadas. Através da calorimetria exploratória diferencial, calculou-se a energia de ativação (Ea) dos polímeros. A cristalinidade dos materiais foi definida por difratometria de raios-X, mostrando caráter amorfo do material, bem como um pequeno incremento na porcentagem da cristalinidade com o aumento da intensidade das doses de radiação durante a síntese e um decréscimo dessa cristalinidade com o aumento na concentração de HEMA. A análise da citotoxicidade das amostras mostrou a ausência de toxicidade dos POSE-HEMA, confirmando a eficiência das lavagens dos polímeros para retirada de resíduos do processamento. A análise da hemocompatibilidade mostrou ausência de adesão de plaquetas e os testes de crescimento celular nas matrizes foram positivos. Através dos resultados obtidos nesta pesquisa, pôde-se concluir pelo potencial de utilização dos POSE-HEMA na engenharia de tecidos. / Tissue engineering and regenerative medicine have as main objective the morphologic/functional reestablishment of injured tissues and organs using cells, scaffolds, stem cells and control of immunological/biochemical responses promoted by the body. In addition, materials science seeks to develop biocompatible biomaterials that do not promote unwanted immune responses and provide the re-establishment of the functions of the tissue/organ. Polymers of natural origin stand out as biomaterials to resemble biological macromolecules, similarity to the extracellular matrix, reduced chance of inflammation and chronic pacing low or no toxicity. This study aimed the development of macromolecular arrays originated from epoxidized soybean oil (OSE), analyzing the relationship between the chemical structure/biological activity of the macromolecular arrays for use as biomaterials in tissue engineering. The synthesis of OSE was performed through the oleochemical route, whose efficiency was determined by infrared spectroscopy and the reaction yield of 85%, determined by nuclear magnetic resonance spectroscopy. From the analysis by differential scanning calorimetry, it was detected a decrease of the glass transition temperature of the epoxidized soybean oil polymer (POSE) compared with OSE, suggesting an increase of the growth of polymer chains of POSE. Thermogravimetric analysis was performed to define the OSE degradation profile, which degrades in two steps. The POSE degrades in just one step and shows higher thermal stability by the increased molecular interactions. The hydrophilicity and crosslinking of POSE was promoted by the addition of 2-hydroxyethyl methacrylate (HEMA) with the monomer grafting by gamma irradiation. The results showed an increased mechanical stability, gelation and water absorption with the HEMA content increasing. Finally, the degree of crystallinity for such polymers grafted with HEMA was 27.5%, estimated by X-ray diffractometry. The second stage was characterized by (i) developing POSEs with the grafting of HEMA in the proportions OSE / HEMA 90:10 and 65:35 irradiated by gamma rays at doses of 50 and 100kGy, (ii) physico-chemical characterization of POSE-HEMA and (iii) analysis of biological materials. By infrared spectroscopy, it was detect the epoxidized regions of POSE, as well as the successful grafting of the monomer HEMA concentrations with all radiation doses. By differential scanning calorimetry, the activation energy was calculated (Ea) of the polymers. The crystallinity of the material was defined by X-ray diffraction, showing tendency of amorphous material as well as a small percentage of the increase in crystallinity with increasing intensity of radiation doses during this synthesis and a decrease in crystallinity with the increasing concentration of HEMA. The analysis of the samples did not show cytotoxicity on POSE-HEMA and confirmed the efficiency of polymer washings to remove the processing waste. The analysis of hemocompatibility showed any platelet adhesion and the cell growth on the scaffolds was positive. From the results obtained in this research, we concluded by the potential use of POSE-HEMA in tissue engineering.

Um novo método de modificação da superfície de nanopartículas de hidroxiapatita (HAP) por reação com cloreto de lauroíla foi desenvolvido, gerando a nanopartícula funcionalizada por laurato (HAP-CL). A superfície modificada da HAP foi confirmada por infravermelho, termogravimetria, ressonância magnética nuclear e análise elementar. Provamos por testes mecânicos a capacidade de criar compósitos com alto teor de HAP-CL em matrizes poliméricas de poli(L-acido láctico) (PLLA) e poli(succinato de isosorbídeo-b-L-lactídeo) (PLLA-co-PIS) sem perda significativa de propriedades mecânicas. Diferentes quantidades de HAP-CL, HAP enxertado com PLLA (PLLA-g-HAP) e HAP puro foram dispersadas em soluções de PLLA para formar fibras eletrofiadas. Para comparar a dispersão destas nanopartículas nas fibras e a sua morfologia, análise de microscopia eletrônica de varredura e de transmissão foram empregadas. A HAP-CL exibiu melhor dispersão na matriz polimérica do que o PLLA-g-HAP e HAP, e permitiu a produção de fibras com grande quantidade de HAP-CL (até 30% massa da fase mineral em relação à massa do polímero(mm/mp)), tanto para o PLLA como para o PLLA-co-PIS. Células tronco mesenquimais derivadas de polpa de dente foram cultivadas em fibras de PLLA com alto teor de HAP-CL, resultando em um aumento significativo da atividade de fosfatase alcalina (ALP), nos dias 14 e 21 (p <0,001) quando comparados com conteúdos mais baixos de HAP-CL, assim como um melhor processo de mineralização apresentado pelos teste de vermelho de alizarina depois de 21 dias (p <0,001). As células cultivadas nas fibras de PLLA-co-PIS contendo 30% de HAP-CL (mm/mp) apresentaram maior atividade de ALP após 21 dias (p <0,05), e melhor processo de mineralização, depois de 14 e 21 dias (p <0,05) do que as fibras de PLLA com 30% de HAP-CL (mm/mp). PLLA e PLLA-co-PIS, ambos contendo 30% de HAP-CL (mm/mp) induziram uma maior expressão de osteocalcina e osteopontina, dois marcadores de diferenciação osteoblástica, quando comparados ao PLLA e PLLA-co-PIS (controle). Finalmente, em experimentos in vivo, as fibras de PLLA-co-PIS contendo 30% de HAP-CL (mm/mp) apresentaram um desempenho superior no processo de neoformação óssea do que as fibras de PLLA com o mesmo conteúdo de nanopartículas. Em conclusão, os nossos resultados in vitro demonstraram que os suportes construídos de compósitos de PLLA-co-PIS contendo 30% de HAP-CL (mm/mp) se mostraram superiores tanto na adesão e proliferação quanto na diferenciação de células mesenquimais de polpa de dente em osteoblastos. Além disso, experimentos in vivo confirmaram estes resultados, demonstrando que estes nanocompósitos são excelentes modelos para implantes destinados a regeneração óssea. / A new method of surface modification of hydroxyapatite nanoparticles (HAP) by reaction with lauroyl chloride was developed, producing the laurate functionalized nanoparticle (HAP-CL). The surface modified HAP was confirmed by infrared, thermogravimetric analysis, nuclear magnetic resonance and elemental analysis. We proved by mechanical tests the ability to create composites with high HAP-CL content in poly(L-lactic acid) (PLLA) and poly(isosorbide succinate-b-L-lactide) (PLLA-co-PIS) polymeric matrixes without significant loss of mechanical properties. Different amounts of HAP-CL, HAP grafted with PLLA (PLLA-g-HAP) and HAP were dispersed in pure PLLA solutions to form nanofibers. To compare the dispersion of these nanoparticles in the fibers and their morphology, scanning and transmission electron microscopies were employed. HAP-CL showed better dispersion in the polymer matrix than the PLLA-g-HAP and HAP, and allowed fiber production with large amounts of HAP-CL (up to 30% mineral to polymer weight (wm/wp)) for both PLLA and PLLA-co-PIS. Mesenchymal stem cells derived from dental pulp were cultured in PLLA fibers with high levels of HAP-CL, resulting in a significant increase in alkaline phosphatase activity (ALP), on days 14 and 21 (p <0.001) as compared to those with lower content HAP-CL, as well as a better mineralization process shown by alizarin red test after 21 days (p <0.001). Cells grown in PLLA-co-PIS fibers containing 30% of HAP-CL showed higher ALP activity after 21 days (p <0.05) and a better mineralization process, after 14 and 21 days (p <0.05 ) than fibers of PLLA with 30% HAP-CL. PLLA and PLLA-co-PIS, both containing 30% of HAP-CL (wm/wp) induced a higher expression of osteocalcin and osteopontin, two osteoblast differentiation markers when compared with PLLA and PLLA-co-PIS (control). Finally, in the in vivo experiments, the PLLA-co-PIS fibers containing 30% HAP-CL (wm/wp) outperformed the process of bone formation than PLLA fibers with the same content of nanoparticles. In conclusion, our in vitro results demonstrated that scaffolds from composites of PLLA-co-PIS containing 30% HAP-CL (wm/wp) were superior both in adhesion and in the differentiation and proliferation of dental pulp stem cells in osteoblasts. Furthermore, in vivo experiments confirmed these results, demonstrating that these nanocomposites are excellent models for implants for bone regeneration

Um novo método de modificação da superfície de nanopartículas de hidroxiapatita (HAP) por reação com cloreto de lauroíla foi desenvolvido, gerando a nanopartícula funcionalizada por laurato (HAP-CL). A superfície modificada da HAP foi confirmada por infravermelho, termogravimetria, ressonância magnética nuclear e análise elementar. Provamos por testes mecânicos a capacidade de criar compósitos com alto teor de HAP-CL em matrizes poliméricas de poli(L-acido láctico) (PLLA) e poli(succinato de isosorbídeo-b-L-lactídeo) (PLLA-co-PIS) sem perda significativa de propriedades mecânicas. Diferentes quantidades de HAP-CL, HAP enxertado com PLLA (PLLA-g-HAP) e HAP puro foram dispersadas em soluções de PLLA para formar fibras eletrofiadas. Para comparar a dispersão destas nanopartículas nas fibras e a sua morfologia, análise de microscopia eletrônica de varredura e de transmissão foram empregadas. A HAP-CL exibiu melhor dispersão na matriz polimérica do que o PLLA-g-HAP e HAP, e permitiu a produção de fibras com grande quantidade de HAP-CL (até 30% massa da fase mineral em relação à massa do polímero(mm/mp)), tanto para o PLLA como para o PLLA-co-PIS. Células tronco mesenquimais derivadas de polpa de dente foram cultivadas em fibras de PLLA com alto teor de HAP-CL, resultando em um aumento significativo da atividade de fosfatase alcalina (ALP), nos dias 14 e 21 (p <0,001) quando comparados com conteúdos mais baixos de HAP-CL, assim como um melhor processo de mineralização apresentado pelos teste de vermelho de alizarina depois de 21 dias (p <0,001). As células cultivadas nas fibras de PLLA-co-PIS contendo 30% de HAP-CL (mm/mp) apresentaram maior atividade de ALP após 21 dias (p <0,05), e melhor processo de mineralização, depois de 14 e 21 dias (p <0,05) do que as fibras de PLLA com 30% de HAP-CL (mm/mp). PLLA e PLLA-co-PIS, ambos contendo 30% de HAP-CL (mm/mp) induziram uma maior expressão de osteocalcina e osteopontina, dois marcadores de diferenciação osteoblástica, quando comparados ao PLLA e PLLA-co-PIS (controle). Finalmente, em experimentos in vivo, as fibras de PLLA-co-PIS contendo 30% de HAP-CL (mm/mp) apresentaram um desempenho superior no processo de neoformação óssea do que as fibras de PLLA com o mesmo conteúdo de nanopartículas. Em conclusão, os nossos resultados in vitro demonstraram que os suportes construídos de compósitos de PLLA-co-PIS contendo 30% de HAP-CL (mm/mp) se mostraram superiores tanto na adesão e proliferação quanto na diferenciação de células mesenquimais de polpa de dente em osteoblastos. Além disso, experimentos in vivo confirmaram estes resultados, demonstrando que estes nanocompósitos são excelentes modelos para implantes destinados a regeneração óssea. / A new method of surface modification of hydroxyapatite nanoparticles (HAP) by reaction with lauroyl chloride was developed, producing the laurate functionalized nanoparticle (HAP-CL). The surface modified HAP was confirmed by infrared, thermogravimetric analysis, nuclear magnetic resonance and elemental analysis. We proved by mechanical tests the ability to create composites with high HAP-CL content in poly(L-lactic acid) (PLLA) and poly(isosorbide succinate-b-L-lactide) (PLLA-co-PIS) polymeric matrixes without significant loss of mechanical properties. Different amounts of HAP-CL, HAP grafted with PLLA (PLLA-g-HAP) and HAP were dispersed in pure PLLA solutions to form nanofibers. To compare the dispersion of these nanoparticles in the fibers and their morphology, scanning and transmission electron microscopies were employed. HAP-CL showed better dispersion in the polymer matrix than the PLLA-g-HAP and HAP, and allowed fiber production with large amounts of HAP-CL (up to 30% mineral to polymer weight (wm/wp)) for both PLLA and PLLA-co-PIS. Mesenchymal stem cells derived from dental pulp were cultured in PLLA fibers with high levels of HAP-CL, resulting in a significant increase in alkaline phosphatase activity (ALP), on days 14 and 21 (p <0.001) as compared to those with lower content HAP-CL, as well as a better mineralization process shown by alizarin red test after 21 days (p <0.001). Cells grown in PLLA-co-PIS fibers containing 30% of HAP-CL showed higher ALP activity after 21 days (p <0.05) and a better mineralization process, after 14 and 21 days (p <0.05 ) than fibers of PLLA with 30% HAP-CL. PLLA and PLLA-co-PIS, both containing 30% of HAP-CL (wm/wp) induced a higher expression of osteocalcin and osteopontin, two osteoblast differentiation markers when compared with PLLA and PLLA-co-PIS (control). Finally, in the in vivo experiments, the PLLA-co-PIS fibers containing 30% HAP-CL (wm/wp) outperformed the process of bone formation than PLLA fibers with the same content of nanoparticles. In conclusion, our in vitro results demonstrated that scaffolds from composites of PLLA-co-PIS containing 30% HAP-CL (wm/wp) were superior both in adhesion and in the differentiation and proliferation of dental pulp stem cells in osteoblasts. Furthermore, in vivo experiments confirmed these results, demonstrating that these nanocomposites are excellent models for implants for bone regeneration

A eletrofiação é uma celebrada técnica de processamento de polímeros, capaz de produzir fibras de diâmetro nanométrico. A montagem comum do sistema de eletrofiação permite a captação de fibras aleatórias sob a forma de um não-tecido. Diversas modificações nessa montagem permitem a obtenção de diferentes morfologias de fibras. Tais modificações são revisadas e discutidas neste trabalho. Na produção de suportes de crescimento de células neurais, é interessante que seja incorporada alguma anisotropia no meio. Assim, um aparato de eletrofiação, capaz de produzir fibras alinhadas, foi construído e a variação dos parâmetros de seu processamento permitiu a obtenção de diferentes qualidades de alinhamento das fibras para dois polímeros biodegradáveis. Diversos parâmetros influenciaram a qualidade desse alinhamento, porém a velocidade de captação das fibras mostrou ser o mais impactante, em acordo com dados reportados na literatura. A morfologia das fibras foi avaliada quanto ao seu diâmetro, com o auxílio de micrografias de MEV e do software de edição de imagens ImageJ. Adicionalmente buscou-se avaliar a qualidade do alinhamento de tais fibras. Para tanto, foi desenvolvida uma metodologia de quantificação de qualidade de alinhamento de fibras, baseado nas micrografias e na ferramenta de FFT do ImageJ. A metodologia proposta foi capaz de ordenar de maneira objetiva e consistente a qualidade do alinhamento das fibras obtidas, mesmo quando a análise visual (usada como referência) se provava ineficiente. A metodologia proposta foi incorporada num plugin para ImageJ, via algoritmo computacional escrito em Java. Com o uso do plugin, foi possível processar diversas micrografias, obtidas em diferentes pontos das malhas eletrofiadas e com variadas magnificações, a fim de se criar uma estatística dos resultados obtidos para qualidade de alinhamento das fibras, algo inédito na literatura. Malhas eletrofiadas com diferentes qualidades de alinhamento de suas fibras foram utilizadas como substrato na cultura de células precursoras neurais, provenientes de neuroesferas. Foi feita a cultura de células progenitoras neurais, provenientes de neuroesferas, tendo como substrato malhas eletrofiadas com diferentes qualidades de alinhamento, a fim de se avaliar o impacto dos contatos físicos das fibras sobre a migração e diferenciação de tais células. / Electrospinning is a celebrated technique of polymer processing, able to produce fibers with nanometric diameter. Common assembly of electrospinning apparatus allows collection of random fibers in a non-woven matt. Several modifications on this assembly enable different fiber morphologies to be obtained. Such modifications are revised and discussed in this work. In the production of cell growth scaffolds, its interesting that some anisotropy is incorporated in the medium. Therefore, an electrospinning apparatus capable of producing aligned fibers was constructed. Variation of processing parameters of said apparatus enabled different alignment qualities of fibers to be attained for two biodegradable polymers. Many parameters influenced on the quality of said alignment; fiber collection speed, however, proved more impacting, in accordance with literature data. Fiber morphology was assessed in regard to its diameter with the aid of MEV micrographs and ImageJ software. Furthermore, assessment of fiber alignment quality was sought. For this matter, it has been developed a quantification methodology for fiber alignment quality, based on micrographs and ImageJ\'s FFT tool. The proposed methodology was able to objectively and consistently rank fiber alignment quality, even when visual analysis (used as reference) failed to do so. This methodology was incorporated in a plugin for ImageJ, via Java script algorithm. With the aid of this plugin it was feasible to process several micrographs, taken from electrospun mats at different spots and magnifications. This helped create statistics about obtained results of fiber alignment quality, on an unprecedented approach in written literature. Electrospun mats with varying quality in fiber alignment were used as substrate in the culture of neural precursor cells from neurospheres to assess the influence of contact guidance on migration and differentiation of such cells

Collagen scaffolds are porous structures which are used in bioreactors and in a wide range of tissue engineering applications. In these contexts, the scaffolds may be subjected to conditions in which fluid is forced through the structure and the scaffold is simultaneously compressed. It is clear that fluid transport within collagen scaffolds, and the inter-relationships between permeability, scaffold structure, fluid pressure and scaffold deformation are of key importance. However, these relationships remain poorly understood. In this thesis, a series of isotropic collagen structures were produced using a freeze-drying technique from aqueous slurry concentrations 0.5, 0.75 and 1 wt%, and fully characterised using X-ray micro-tomography and compression testing. It was found that collagen wt% influenced structural parameters such as pore size, porosity, relative density and mechanical properties. Percolation theory was used to investigate the pore interconnectivity of each scaffold. Structures with lower collagen fraction resulted in larger percolation diameters, but lower mechanical stiffness. Aligned collagen scaffolds were also produced by altering the freeze-drying protocol and using different types of mould materials and designs. It was found that a polycarbonate mould with stainless base resulted in vertically aligned structures with low angular variation. When compared with isotropic scaffolds from slurry of the same concentration, aligned scaffolds had a larger percolation diameter. Tortuosity was used as a mathematical tool to characterise the interconnected pathways within each porous structure. The effect of the size of the region of interest (ROI) chosen and the size of the virtual probe particle used in the analysis on the values of tortuosity calculated were determined and an optimised calculation methodology developed. Increasing the collagen fraction within isotropic scaffolds increased the tortuosity, and aligned structures had smaller tortuosity values than their isotropic counterparts. Permeability studies were conducted using two complementary experimental rigs designed to cover a range of pressure regimes and the results were compared with predictions from mathematical models and computational simulations. At low pressures, it was found that the lower collagen fraction structures, which had more open morphologies, had higher permeabilities. Alignment of the structure also enhanced permeability. The scaffolds all experienced deformation at high pressures resulting in a restriction of fluid flow. The lower collagen fraction scaffolds experienced a sharper decrease in permeability with increased pressure and aligned structures were more responsive to deformation than their isotropic counterparts. The inter-relationships between permeability, scaffold structure, fluid pressure and deformation of collagen scaffolds were explored. For isotropic samples, permeability followed a broad $(1- \epsilon)^2$ behaviour with strain as predicted by a tetrakaidecahedral structural model, with the constant of proportionality changing with collagen fraction. In contrast, the aligned structures did not follow this behaviour with the permeability dropping much more sharply in the early stages of compression. Open-cell polyurethane (PU) foams, sometimes used as dressings in wound healing applications, are often compared with collagen scaffolds in permeability models and were used in this thesis as a comparison structure. The foam had a higher permeability than the scaffolds due to its larger pore sizes and higher interconnectivity. In the light of the effects of compression on permeability, the changes in porous structure with compression were explored in isotropic and aligned 0.75 wt% scaffolds. Unlike the fluid flow experiments, these experiments were carried out in the dry state. Deformation in simple linear compression and in step-wise compression was studied, and the stress relaxation behaviour of the scaffolds characterised. A methodology was developed to characterise the structural changes accompanying compression using X-ray micro-tomography with an in situ compression stage. The methodology accounted for the need for samples to remain unchanged during the scan collection period for stable image reconstruction. The scaffolds were studied in uniaxial compression and biaxial compression and it was found that pore size and percolation diameter decreased with increasing compressive strain, while the tortuosity increased. The aligned structure was less affected than the isotropic at low compressions, in contrast to the results from the permeability study in which the aligned structure was more responsive to strain. This suggests that the degree of hydration may affect the structural changes observed. The insights gained in this study of the inter-relationships between microstructure, fluid dynamics and deformation in collagen scaffolds are of relevance to the informed design of porous structures for medical applications.