Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Materials based on polyhydroxybutyrate (PHB), chitosan (CHI) and fibroin (SF) are biocompatible and attractive for applications in bone tissue engineering. In this work, 3D scaffolds of PHB/CHI and PHB/CHI/SF in different proportions was prepared, characterized and evaluated the in vitro behavior: group I: PHB/CHI (50:50 wt.%), group II: PHB/CHI/SF (50:45:5 wt.%) and group III: PHB/CHI/SF (50:35:15 wt.%). The scaffolds were produced by the lyophilization method of the components mixtures. The physical-chemical characterization of the scaffolds was performed by X-ray diffraction, infrared spectroscopy, scanning electron microscopy and thermogravimetric analysis. A highly porous nature was revealed by SEM analysis of the scaffolds. The FTIR analysis revealed that the constituents used in the preparation of the scaffolds interacted chemically with each other. Thermal analysis showed that fibroin increases the thermal stability of the scaffolds. The viability and cell proliferation were assessed by the MTT method and the cytotoxicity results showed that all scaffolds are non- cytotoxic. In addition, the scaffolds stimulated cell proliferation and are promising for tissue engineering applications. / Materiais à base de polihidroxibutirato (PHB), quitosana (QUI) e fibroína (SF) são biocompatíveis e atrativos para aplicações na engenharia tecidual óssea. Neste trabalho foi preparado, caracterizado e avaliado o comportamento in vitro de arcabouços 3D de PHB/QUI/SF em diferentes proporções: grupo I: PHB/QUI (50:50 % em massa), grupo II-PHB/QUI/SF (50:45:5 % em massa) e grupo III- PHB/QUI/SF (50: 35:15 % em massa). Os arcabouços foram produzidos pelo método da liofilização das misturas dos componentes dos três grupos. A caracterização físico-química dos arcabouços foi realizada por difração de raios X, espectroscopia no infravermelho, microscopia eletrônica de varredura e análise termogravimétrica. A análise de MEV mostrou que os arcabouços 3D apresentam uma boa porosidade. Por FTIR observou- se que os componentes utilizados na preparação dos arcabouços interagiram quimicamente entre si. As análises térmicas indicam que a fibroína aumenta a estabilidade térmica dos arcabouços. A viabilidade e a proliferação celular foram avaliadas pelo método do MTT e os resultados de citotoxicidade mostraram que ambos os arcabouços não são citotóxicos. Além disso, os arcabouços estimularam a proliferação celular, sendo promissores para aplicações em engenharia tecidual.

Made available in DSpace on 2017-07-10T18:08:02Z (GMT). No. of bitstreams: 1 Vinicius Muller.pdf: 5054743 bytes, checksum: 4684dd9325f819006bfd84424d666674 (MD5) Previous issue date: 2014-07-04 / Electrospinning technique was applied in the covering of fused silica fibers by regenerated silk fibroin nanofibers (RSF). The parameters of electrospinning process were evaluated through factorial experimental design 22. This study showed the variables flow of solution and capillary-collector distance were statistically significant in the medium diameter response. The model obtained was validated through variance analysis (ANOVA) and response surface methodology. The material was used at solid phase micro extraction (SPME) in the extraction of samples containing a small chain alcohol, applied at gas chromatography (GC). The recovered device was characterized through Fourier transform infrared spectroscopy with attenuated total reflectance (FTIR-ATR) and thermal analysis (DSC and TGA). The RSF nanofibers covered fibers were morphologic evaluated through scanning electronic spectroscopy (MEV). The results of MEV showed that was possible to obtain fibers in nanoscale, medium diameter around 304 ± 46 nm. The fibers were also submitted to simulated thermal treatment (100°C to 250°C). There was no fusion between the fibers until 250°C (4 hours of treatment), and a little structural damage. The thermal analysis of FRS and covered FRS nanofibers device showed that the electrospun material maintained the thermal stability, with loss due thermal degradation from 250°C. The GC assay demonstrated the standard SPME covered device didn´t interfere on the suggested method to the alcohol isopropyl analysis (AIS). The times of extraction and desorption were optimized in 20 minutes and 10 minutes, respectively. The methodology was evaluated about linearity and specificity. In the concentration between 10 to 500 ppm of AIS, the method was linear, with R2= 0,9927 and selective, with high resolution between peaks of methanol, AIS and ethyl acetate. Electrospinning process of RSF as covering device showed great potential to use in SPME due its high surface area, thermal stability and easiness process, as well as potential to application in extraction of alcohols and GC analysis. / A técnica de eletrofiação foi aplicada no revestimento de fibras de sílica fundida por nanofibras de fibroína de seda regenerada (FSR). Os parâmetros do processo de eletrofiação foram avaliados por meio de delineamento experimental fatorial 2². Este estudo mostrou que as variáveis vazão da solução e distância entre capilar-coletor foram estatisticamente significativas na resposta diâmetro médio. O modelo obtido foi validado através da análise de variância (ANOVA) e metodologia de superfície de resposta. O material foi empregado em Microextração em Fase Sólida (Solid Phase Microextraction SPME ) na extração de amostras contendo um álcool de cadeia pequena, aplicado em cromatografia gasosa (CG). O dispositivo recoberto foi caracterizado por meio de espectroscopia na região do infravermelho (FTIR-ATR) e análises térmicas (DSC e TGA). As fibras recobertas com nanofibras de FSR foram avaliadas morfologicamente através de microscopia eletrônica de varredura (MEV). Por meio dos resultados das análises de MEV, observou-se que foi possível obter fibras em escala nanométrica, com diâmetro médio em torno de 304 ± 46 nm. As fibras também foram submetidas a tratamento térmico em forno (100°C a 250°C). Não houve fusão das fibras até 250°C (até 4 h de tratamento), e poucos danos estruturais. As análises térmicas da FSR e do dispositivo recoberto com as nanofibras de FSR mostraram que o material eletrofiado manteve a estabilidade térmica, com perda por degradação térmica a partir de 250 °C. Os ensaios em CG demonstraram que o branco com o dispositivo de SPME recoberto não interferiu no método proposto para a análise de álcool isopropílico (AIS). Os tempos de extração e dessorção do analito foram otimizados em 20 min e 10 min, respectivamente. A metodologia foi avaliada quanto à linearidade e especificidade. Na faixa de concentração de 10 a 500 ppm de AIS, o método foi linear, com R=0,9927 e seletivo, apresentando alta resolução entre os picos de metanol, AIS e acetato de etila. O processo de eletrofiação da FSR como recobrimento do dispositivo mostrou grande potencial para emprego em SPME devido a sua grande área superficial, estabilidade térmica e facilidade no processo, bem como potencial para aplicação em extração de alcoóis e análise em CG.

Une lésion au niveau d’un nerf périphérique peut provoquer la perte de fonction sensorielle et motrice, et dans le cas de neurotmésis, la régénération spontanée ne se produira pas. De plus, si l’espace entre les deux segments de nerf est trop important, une suture directe n’est pas possible et l’implantation d’une greffe est nécessaire afin de créer une liaison entre les deux segments de nerf. L’autogreffe de nerf est le « gold standard » pour des procédés de réparation nerveuse : une portion d’un nerf sein (qui est considéré comme un nerf moins important) est prise du même patient et implantée au site de la lésion. Cependant, il existe plusieurs désavantages avec ce procédé comme une deuxième chirurgie, la perte de fonction au site du don, la possibilité de développer un neurome sur ce même site, ainsi qu’un taux de réussite de 50% dans les cas où l’espace entre les deux segments de nerf est très important. Il reste donc, un besoin de trouver un procédé alternatif afin d’augmenter le taux de réussite et d’éliminer les désavantages de l’autogreffe. L’objectif de cette étude est d’avancer vers une solution alternative de l’autogreffon en utilisant des biomatériaux. Cette thèse se divise en trois parties. La première se focalise sur le développement d’un modèle de guide nerveux basé sur des nanofibres de fibroïne de soie. Ce matériau est composé d’une organisation complexe qui inclut deux surfaces de nanofibres alignées avec une couche de nanofibres aléatoires à l’intérieur afin d’améliorer des propriétés mécaniques du matériau sans la perte d’orientation des fibres pour la régénération nerveuse. Le matériau est ensuite manipulé pour fabriquer un tube, multi-canaux avec une « enveloppe » supplémentaire afin de faciliter le procédé d’implantation chirurgicale. Ce guide nerveux a été soumis pour l’obtention d’un brevet européen le 12 juillet 2017 et cela est le sujet d’un deuxième article qui a été soumis pour publication. La deuxième partie de cette étude explore des possibilités d’une fonctionnalisation du matériau afin d’améliorer son efficacité pour la régénération nerveuse. Cette étude explore la fonctionnalisation de la fibroïne de soie avec une deuxième protéine, plusieurs facteurs de croissance, et des nanoparticules. Chacune de ces fonctionnalisations donne une possibilité d’ajouter des propriétés favorables à la fibroïne de soie, un matériau naturel et biocompatible. La troisième partie de cette étude examine l’efficacité d’un guide nerveux composé de la fibroïne de soie fonctionnalisée avec des facteurs de croissance pour la régénération nerveuse périphérique en comparaison avec un guide nerveux composé de la fibroïne de soie sans aucune fonctionnalisation et une suture direct (qui simule une autogreffe). Trois techniques d’évaluation différentes de la régénération nerveuse ont été réalisées afin d’obtenir une analyse plus complète. Il y a de nombreux mécanismes impliqués dans la régénération nerveuse, il est donc nécessaire d’étudier différents paramètres pour analyser l’efficacité de régénération. Les résultats d’analyses histologiques, d’électromyographie, et de capture de mouvement, ont été considérées ensemble afin d’arriver à une conclusion sur la réussite d’une régénération nerveuse pendant cette étude. Pour conclure cette étude, les guides nerveux fonctionnalisés avec une combinaison de facteurs de croissance démontrent une meilleure régénération nerveuse et une récupération de fonction supérieure. / Injury to a peripheral nerve can cause loss of sensory and motor function, and if the injury is very severe where the nerve undergoes neurotmesis, unassisted nerve regeneration may not occur. In this case, where the gap between nerve segments is too large to carry out a direct end to end suture, a graft is sutured to bridge the gap between sectioned nerve segments. The autologous nerve graft, where a portion of a less important nerve from the same patient is removed and grafted between nerve segments, continues to be the gold standard procedure for nerve repair. However, there are several drawbacks of this technique including a second surgical procedure, loss of function at the donor site, possibility of developing a painful neuroma at the donor site, and the 50% success rate of autografts used in large gaps. There is therefore a need for a tissue engineered nerve graft that can replace the autograft, and this study aims to advance toward an effective autograft alternative. This PhD is presented as a three part study consisting first of the development of a novel nerve guidance conduit based on a tri-layered silk fibroin nanofiber material comprised of a complex organization including two aligned fiber surfaces and a randomly deposited fiber interior to improve the mechanical properties of the material while not compromising the guidance capabilities of aligned nanofibers for nerve regeneration. The material is then used to fabricate a multi-channeled tube with an additional “jacket layer” in order to facilitate surgical implantation. This NGC has been submitted to be patented on July 12, 2017 and is the subject of the second article submitted for review for publication. The second part of this study explores the different possibilities of the functionalization of the material in order to improve the effectiveness for nerve regeneration. This study explores functionalizing the silk fibroin with a second protein, several growth factors, and nanoparticles that all have potential to add favorable properties to the natural biocompatible silk fibroin material. The final part of this study tests the effectiveness of growth factor-embedded silk fibroin NGCs for peripheral nerve regeneration in comparison with non-functionalized silk fibroin devices and a direct suture to simulate results obtained with an autograft. Three different techniques for the evaluation of nerve regeneration were used in order to produce a more comprehensive analysis. As there are many mechanisms involved in nerve regeneration, only one or two analysis techniques cannot paint a complete picture of the success of nerve regeneration. Therefore, histological analyses, electromyography analyses, and motion capture analyses were carried out and considered together in order to make a conclusion on the level of nerve regeneration success during this study. The conclusions from this study were that a NGC functionalized with a combination of growth factors appeared to exhibit the most successful nerve regeneration and functional recovery.

Cellulose is the most abundant biopolymer on Earth as it is found in every plant cell wall; therefore, it represents one of the most promising natural resources for the fabrication of sustainable materials. In plants, cellulose is mainly used for structural integrity, however, some species organise cellulose in helicoidal nano-architectures generating strong iridescent colours. Recent research has shown that cellulose nanocrystals, CNCs, isolated from natural fibres, can spontaneously self-assemble into architectures that resemble the one producing colouration in plants. Therefore, CNCs are an ideal candidate for the development of new photonic materials that can find use to substitute conventional pigments, which are often harmful to humans and to the environment. However, various obstacles still prevent a widespread use of cellulose-based photonic structures. For instance, while the CNC films can display a wide range of colours, a precise control of the optical appearance is still difficult to achieve. The intrinsic low thermal stability and brittleness of cellulose-based films strongly limit their use as photonic pigments at the industrial scale. Moreover, it is challenging to integrate them into composites to obtain further functionality while preserving their optical response. In this thesis, I present a series of research contributions that make progress towards addressing these challenges. First, I use an external magnetic field to tune the CNC films scattering response. Then, I demonstrate how it is possible to tailor the optical appearance and the mechanical properties of the films as well as to enhance their functionality, by combining CNCs with other polymers. Finally, I study the thermal properties of CNC films to improve the retention of the helicoidal arrangement at high temperatures and to explore the potential use of this material in industrial fabrication processes, such as hot-melt extrusion.