Pour le comblement d'une perte de tissu, la greffe reste une méthode usitée en chirurgie osseuse mais une lacune importante conduit à l'emploi de matériaux artificiels. Alternative aux implants métalliques, les substituts osseux biocéramiques ont une efficacité insuffisante dans de nombreuses indications : problème d'intégration d'un volume important, résistance inadéquate à long terme, faible bioréactivité. Doper ces biocéramiques est une voie intéressante mais nombre de dopants ont abouti à des effets cytotoxiques voire à la dégradation des propriétés mécaniques. Le magnésium a été étudié comme dopant de l'hydroxyapatite pour des faibles teneurs, apportant des améliorations sans affecter la biocompatibilité. Cependant, les fortes teneurs et l'optimisation de la synthèse d'une biocéramique dopée ont été peu étudiées. Nous avons étudié et optimisé la synthèse d’hydroxyapatite par la méthode de précipitation aqueuse pour un dopage au magnésium de 1, 2, 5 et 10 % en masse. Caractérisations physicochimiques (DRX, MEB, densité), biologiques (cytotoxicité) et mécaniques (microdureté, élasticité) ont été menées (poudres ou pastilles frittées ou non). Nous avons montré l'apparition de TCP à partir de 2 % Mg, la densité des échantillons diminuant quand la teneur croît. Le dopage accroît la microdureté et le module d’Young. Aucune cytotoxicité n'a été révélée mais une importante baisse d’activité cellulaire a été remarquée pour 10 % de magnésium. Une légère augmentation a été observée pour 1 %. L'aptitude à la mise en forme a été appréciée via le coulage d'une réplique d'un implant intervertébral. Le dopage d'HA à 1 % Mg s'avère être à tout point de vue le compromis optimal. / For the filling of a loss of tissue, the graft is a current process in bone surgery, but a significant deficiency leads to the use of artificial materials. Alternative to metallic implants, bone substitutes bioceramics have insufficient efficacy in many indications: problem of integrating a large volume, inadequate long-term resistance, low bioreactivity. Doping these bioceramics is an interesting way but many of dopants have conducted to cytotoxic effects or to the degradation of mechanical properties. Magnesium has been studied as a dopant of hydroxyapatite for low contents, improving thebioceramic without affecting its biocompatibility. However, high contents and the optimization of the synthesis of such a doped bioceramic have been little studied. We investigated and optimized the synthesis of hydroxyapatite by the aqueousprecipitation process for magnesium doping of 1, 2, 5 and 10 wt%. Physico-chemical (XRD, SEM, density), biological (cytotoxicity) and mechanical (microhardness, elasticity) characterizations were conducted (on powders or pellets sintered or not). We have shown the occurrence of TCP from 2% Mg, the density of samples decreasing when the content grows. The doping increases the microhardness and the Young’s modulus. No cytotoxicity was revealed but a significant decrease in cellular activity was noted for 10% magnesium. A slight increase was observed for 1%. The ability to form an implant was assessed via the slip casting of a replica of an intervertebral implant. Doping HA to 1% Mg was found to be at any point of view the optimum composition.

Em procedimentos de regeneração óssea guiada (ROG) as membranas de colágeno são os materiais mais utilizados como barreira; porém, sua tendência ao colapso faz indispensável a utilização de materiais de suporte. Para este propósito, os xenoenxertos são substitutos ósseos considerados padrão-ouro, embora apresentem um período longo de reabsorção que impede grande formação do osso. Em contrapartida o ?-fosfato tricálcico (?-TCP) permite boa formação do osso, mas é reabsorvido rapidamente e fracassa quando precisa dar suporte à membrana. O alendronato, um bisfosfonato nitrogenado, é uma droga antirreabsortiva para o tratamento da osteoporose e outras doenças ósseas porque inibe a função dos osteoclastos. O plasma rico em fibrina (PRF) é um concentrado de fibrina sem adição de químicos que consegue estimular processos de cicatrização pelos fatores que fazem parte de sua composição. Neste estudo qualitativo de ROG em defeitos de 5 mm no osso parietal de ratos foi avaliado: 1) o efeito na formação óssea da administração local de 1g/ml de alendronato sódico adicionado a xenoenxerto porcino e a ?-TCP; 2) a adição local de alendronato e PRF a ?-TCP na possibilidade de diminuir a rápida reabsorção do material e impedir o colapso da membrana. Foram usados 100 ratos adultos Wistar distribuídos em 5 grupos (n=20): Xenoenxerto controle (XE-C); xenoenxerto adicionado com alendronato (XE-AL); ?-TCP controle (TCP-C); ?-TCP adicionado com alendronato (TCP-AL); e, ?-TCP adicionado com PRF (TCP-F). Em todos os grupos o enxerto foi coberto com membrana. Dois tempos de estudo de quatro e oito semanas foram considerados para cada grupo (n=10). Ao final de cada tempo, os animais foram sacrificados e as amostras foram fixadas, descalcificadas e processadas para seu estudo em microscopia de luz por meio de análise histológica, histoquímica TRAP e imuno-histoquímica para osteopontina (OPN). Os resultados mostraram maior formação do osso tanto para xenoenxerto como para ?-TCP quando foram adicionados com alendronato local, em ambos tempos de estudo. Nos grupos do ?-TCP a adição de alendronato local permitiu diminuir a reabsorção dos grânulos, melhorando o suporte à membrana ao final dos tempos de estudo; no entanto, no grupo do PRF a reabsorção foi maior e teve pouca formação de osso, provocando colapso da membrana. Adicionalmente, regiões de osso primário subjacentes à membrana de colágeno foram observadas em todos os grupos. / Collagen membranes are the most used materials as a barrier in guided bone regeneration (GBR) procedures; however, its tendency to collapse makes indispensable the use of support materials. For this purpose, xenografts, which are bone substitutes, although they have a long period of resorption that prevents large bone formation, are still considered the gold standard support material. In contrast, tricalcium ?-phosphate (?-TCP) allows good bone formation, but is rapidly reabsorbed and fails when it needs to support the membrane. Alendronate, a nitrogenated bisphosphonate, is an anti-resorptive drug for treatment of osteoporosis and other bone diseases because it inhibits the function of osteoclasts. Fibrin-rich plasma (FRP) is a fibrin concentrate with no added chemicals that can stimulate healing processes by the factors that are part of its composition. In this qualitative study of ROG in 5 mm defects in the rat parietal bone, was evaluated: 1) the effect on bone formation of local administration of 1g / ml sodium alendronate added to porcine xenograft and ?-TCP; 2) the local addition of alendronate and PRF to ?-TCP in the possibility of diminishing the rapid reabsorption of the material and preventing the collapse of the membrane. A 100 adult Wistar rats distributed in 5 groups was used (n = 20): Xenograft control (XE-C); xenograft added with alendronate (XE-AL); ?-TCP control (TCP-C); ?-TCP added with alendronate (TCP-AL); and, ?-TCP added with PRF (TCP-F). In all groups the graft was covered with membrane. Two study times of four and eight weeks were considered for each group (n = 10). At the end of each time, the animals were sacrificed and the samples were fixed, decalcified and processed for light microscopy by histological analysis, TRAP histochemistry and immunohistochemistry for osteopontin (OPN). Results showed higher bone formation for both xenograft and ?-TCP when added with local alendronate at both study times. In the ?-TCP groups the addition of local alendronate allowed to decrease grain resorption, improving membrane support at the end of the study times; however, in the PRF group the resorption was greater and had little bone formation, causing membrane collapse. In addition, primary bone formed in the underlying collagen membrane were observed in all groups.

Avec de nombreuses qualités exceptionnelles (biocompatible et ostéogénique), la nacre représente un biomatériau naturel comme substitut osseux. Mais les composés ostéogéniques dans la nacre ne sont pas encore connus. Nos travaux visent à l’identification des composés ostéogéniques de la nacre. L’ESM (éthanol soluble matrix) est un extrait de la nacre qui est démontré ostéogénique. A partir d’ESM, nous avons essayé plusieurs approches pour cibler et identifier ces composés. Grâce au couplage des cellules MC3T3-E1 et d’ostéoblastes humains arthrosiques, nous avons démontré que la partie cationique d’ESM est ostéogénique, sans interaction avec la partie anionique. Le calcium joue un rôle dans l’activité ostéogénique d’ESM. Ensuite, nous avons créé une lignée cellulaire exprimant de manière stable un plasmide contenant un gène rapporteur ostéogénique (ATDC5 pMetLuc2 ColX promoteur). Grâce à cette lignée, nous avons découvert que les lipides et les sucres présents dans l’ESM ont un effet ostéogénique. Les peptides précipités par TCA sont aussi démontrés ostéogéniques, et ont conduit à leur identification partielle par LC-MS. Ces résultats nous permettent d’avancer plus loin et plus rapidement vers l’identification des composés ostéogéniques de la nacre et vers les applications de la nacre en orthopédie clinique / With many exceptional qualities (biocompatible and osteogenic), nacre represents a natural biomaterial as a bone substitute. However, the osteogenic compounds in nacre are not yet known. Our work aims at the identification of the osteogenic compounds in nacre. The ESM (soluble ethanol matrix) is an extract of nacre that is shown to be osteogenic. From the ESM, we have tried several approaches to target and identify these compounds. Thanks to the coupling of MC3T3-E1 cells and the human osteoarthritis osteoblasts, we demonstrated that the cationic part of the ESM is osteogenic, without interaction with the anionic part. Calcium plays a role in the osteogenic activity of the ESM. Then, we created a cell line stably expressing a plasmid containing an osteogenic reporter gene (ATDC5 pMetLuc2 ColX promoter). Thanks to this cell line, we found out that the lipids and sugars in the ESM have an osteogenic effect. The peptides precipitated by TCA are also demonstrated to be osteogenic, which have led to their partial identification by LC-MS. These results allow us to move farther and faster towards the identification of osteogenic compounds in nacre and the applications of nacre in clinical orthopaedics

A cicatrização de um alvéolo após a extração dental é uma resposta reparadora. Após a exodontia, a cicatrização espontânea irá levar à perda do volume e do formato do rebordo original. A preservação alveolar envolve qualquer procedimento que limite os efeitos da reabsorção pós-extração. O objetivo deste estudo foi avaliar o uso de um substituto ósseo xenogênico na preservação alveolar pós-extração dental comparado com a cicatrização espontânea em dentes anteriores da maxila com defeitos ósseos da parede vestibular maiores do que 50%. Foram tratados 13 pacientes com necessidade de exodontia de dente anterior da maxila com perda >50% da parede vestibular. Todas as exodontias foram realizadas sem retalho. Os pacientes foram alocados aleatoriamente em grupo teste: preservação alveolar com substituto ósseo (7 participantes) e grupo controle: cicatrização espontânea (6 participantes). O desfecho primário foi a alteração dimensional do rebordo alveolar calculada entre as imagens tomográficas de início (pós-exodontia) e depois de 4 meses da intervenção, na espessura 1mm abaixo da porção coronal da crista (HW-1). Os desfechos secundários foram as medidas vestibular e palatina, e os segmentos transversais HW-3 e HW-5. A medida HW-1 reduziu 51,21 % no grupo teste e 82,80% no grupo controle, com uma diferença intergrupos estatisticamente significante de 31,59%. Para as medidas transversais, houve redução estatisticamente significante tanto para HW-3 quanto para HW-5 entre os dois grupos. Em alvéolos com perda da parede vestibular maior que 50%, a utilização do substituto ósseo promoveu uma menor perda horizontal na ordem de 30%. / The healing process of post-extraction sockets is bone resorption. After tooth extraction, spontaneous healing will lead to loss of volume and shape of the original ridge. Alveolar ridge preservation involves any procedure that limits the effects of post-extraction resorption. The objective of this study was to evaluate the use of the xenogenic bone substitute in the post-extraction alveolar preservation compared to spontaneous healing in anterior maxilla teeth with vestibular wall defects greater than 50%. Thirteen patients with anterior maxillary tooth extraction with a > 50% buccal wall loss were treated. All the surgeries were made flapless. Patients were randomly assigned to a test group: alveolar preservation with bone substitute (7 participants); and control group: spontaneous healing (6 participants). The primary outcome was the dimensional alteration of the alveolar ridge calculated between the tomographic images at the beginning (post-extraction) and after 4 months of the intervention, at the thickness 1mm below the coronal portion of the crest (HW-1). Secondary outcomes were measures buccal and palatine, and HW-3 and HW-5 cross-sections. The HW-1 measure reduced 51.21% in the test group and 82.80% in the control group, with a statistically significant intergroup difference of 31.59%. For the other horizontal measurements, there was a statistically significant reduction for both HW-3 and HW-5 between the two groups. In sockets with buccal wall loss greater than 50%, the use of the bone substitute provided a 30% lower horizontal loss.

A cicatrização de um alvéolo após a extração dental é uma resposta reparadora. Após a exodontia, a cicatrização espontânea irá levar à perda do volume e do formato do rebordo original. A preservação alveolar envolve qualquer procedimento que limite os efeitos da reabsorção pós-extração. O objetivo deste estudo foi avaliar o uso de um substituto ósseo xenogênico na preservação alveolar pós-extração dental comparado com a cicatrização espontânea em dentes anteriores da maxila com defeitos ósseos da parede vestibular maiores do que 50%. Foram tratados 13 pacientes com necessidade de exodontia de dente anterior da maxila com perda >50% da parede vestibular. Todas as exodontias foram realizadas sem retalho. Os pacientes foram alocados aleatoriamente em grupo teste: preservação alveolar com substituto ósseo (7 participantes) e grupo controle: cicatrização espontânea (6 participantes). O desfecho primário foi a alteração dimensional do rebordo alveolar calculada entre as imagens tomográficas de início (pós-exodontia) e depois de 4 meses da intervenção, na espessura 1mm abaixo da porção coronal da crista (HW-1). Os desfechos secundários foram as medidas vestibular e palatina, e os segmentos transversais HW-3 e HW-5. A medida HW-1 reduziu 51,21 % no grupo teste e 82,80% no grupo controle, com uma diferença intergrupos estatisticamente significante de 31,59%. Para as medidas transversais, houve redução estatisticamente significante tanto para HW-3 quanto para HW-5 entre os dois grupos. Em alvéolos com perda da parede vestibular maior que 50%, a utilização do substituto ósseo promoveu uma menor perda horizontal na ordem de 30%. / The healing process of post-extraction sockets is bone resorption. After tooth extraction, spontaneous healing will lead to loss of volume and shape of the original ridge. Alveolar ridge preservation involves any procedure that limits the effects of post-extraction resorption. The objective of this study was to evaluate the use of the xenogenic bone substitute in the post-extraction alveolar preservation compared to spontaneous healing in anterior maxilla teeth with vestibular wall defects greater than 50%. Thirteen patients with anterior maxillary tooth extraction with a > 50% buccal wall loss were treated. All the surgeries were made flapless. Patients were randomly assigned to a test group: alveolar preservation with bone substitute (7 participants); and control group: spontaneous healing (6 participants). The primary outcome was the dimensional alteration of the alveolar ridge calculated between the tomographic images at the beginning (post-extraction) and after 4 months of the intervention, at the thickness 1mm below the coronal portion of the crest (HW-1). Secondary outcomes were measures buccal and palatine, and HW-3 and HW-5 cross-sections. The HW-1 measure reduced 51.21% in the test group and 82.80% in the control group, with a statistically significant intergroup difference of 31.59%. For the other horizontal measurements, there was a statistically significant reduction for both HW-3 and HW-5 between the two groups. In sockets with buccal wall loss greater than 50%, the use of the bone substitute provided a 30% lower horizontal loss.

Ce travail de thèse a eu comme objectif la mise en place et la validation d’une méthodologie expérimentale pour le suivi de l’évolution in-vitro de substituts osseux à base de phosphates de calcium. Du phosphate dicalcique dihydraté (DCPD, CaHPO4·2H2O) et du phosphate tricalcique bêta (β-TCP, β-Ca3(PO4)2) ont été choisis comme matériaux modèles de deux grandes classes de substituts osseux: les “biosolubles” (sujets à dissolution après implantation) et les “biorésorbables” (sujets à résorption cellulaire après implantation). Pour l’étude des phénomènes de dissolution et de reprécipitation observés lorsque les phosphates de calcium sont plongés en solution, ces matériaux ont été produits sous forme d’échantillons microporeux (60% de porosité pour le DCPD, 75% pour le β-TCP) et soumis à des tests de dissolution in-vitro en conditions statiques ou dynamiques (sans ou avec renouvellement du liquide) dans différentes solutions tamponnées à pH physiologique (TRIS et PBS) et pour des durées s’étalant entre 30 minutes et 2 mois. L’analyse des propriétés physico-chimiques, microstructurales et mécaniques des échantillons avant et après immersion a permis d’évaluer l’influence du type de milieu et des conditions de test choisies sur l’évolution des échantillons. Une attention particulière a été prêtée à la caractérisation mécanique: la technique de micro-indentation instrumentée sphérique a été préférée à autres essais plus conventionnels. Cette technique permet d’évaluer plusieurs paramètres tels que la dureté et le module de Young de façon quasi non-destructive et à une échelle locale. En conséquence, l’utilisation de la micro-indentation s’est avérée d’une grande aide pour le suivi des caractéristiques d’échantillons dégradés qui présentaient un gradient de propriétés entre la surface (où le processus de dégradation commence) et le cœur (sujet à des changements sur plus long terme). La dernière partie de cette étude a été dédiée à l’étude du deuxième phénomène qui entraine la résorption de substituts osseux in-vivo, à savoir la résorption cellulaire. Pour cela des essais cellulaires avec des cellules précurseurs d’ostéoclastes ont été réalisés sur des échantillons denses de β-TCP pur ou dopé avec 5% molaire de magnésium. L’addition de cet élément est censée modifier les propriétés du matériau (notamment sa solubilité) et, par conséquence, modifier le comportement cellulaire. Les résultats des tests ont confirmé la cytocompatibilité des deux types de β-TCP, mais ont également mis en avant une difficulté d’activation des ostéoclastes. Deux des causes possibles seraient liées à la topographie de surface des échantillons et au relargage des ions calcium suite à la dissolution du matériau. / The present Ph.D. thesis work was aimed to establish and assess an experimental methodology to monitor the in-vitro evolution of calcium phosphate (CaP) bone substitutes. Dicalcium phosphate dihydrate and beta-tricalcium phosphate were chosen as model of two main classes of bone substitutes: “biosoluble” ones (which undergo dissolution after implantation) and “bioresorbable” ones (which undergo cellular resorption after implantation). In order to study the dissolution and precipitation phenomena, which take place once CaPs are immersed in solution, these materials were produced in the form of micro-porous samples (60% of porosity for DCPD, 75% for β-TCP) and used for dissolution tests in-vitro in static and dynamic conditions (without or with liquid renewal) in different buffered solutions at physiologic pH (TRIS and PBS) and for periods of time ranging between 30 minutes and 2 months. The analysis of the physico-chemical, microstructural and mechanical properties of the samples before and after immersion allowed to evaluate the influence of the chosen medium and immersion conditions on the evolution of the specimens. Particular attention was paid to the mechanical characterisation: instrumented spherical micro-indentation was preferred to other more conventional tests. This technique enables the evaluation of several parameters such as the hardness and the Young’s modulus in a quasi-non-destructive way and on a local scale. As a consequence, the use of micro-indentation proved to be of great help for monitoring the characteristics of the degraded specimens, which presented a gradient of properties between the surface (where the degradation process starts) and the core (subject to changes on a longer period). The last part of this work was focused on the study of the second main phenomenon, which takes part in the in-vivo resorption of bone substitutes, that is to say the cellular resorption. For this purpose, cellular tests with osteoclast-precursor cells were carried out on dense samples made of pure and magnesium-doped β-TCP (5 mol.% of Mg). The addition of magnesium was aimed to modify the properties of the material and, as a consequence, the cellular behavior. The results confirmed the cytocompatibility of both types of β-TCP, but they also showed a difficult activation of osteoclasts. Two of the possible causes would be linked to the topography of the surface of the specimens and to the release of calcium ions due to the dissolution of the material.

Les substituts osseux synthétiques servent au remplacement temporaire des tissus osseux, favorisent la formation, la croissance et la survie de l’os et sont biorésorbables. Aucun matériau monophasé ne remplissant complètement ces exigences, un matériau composite bioinspiré est une alternative possible. L’objectif de cette thèse était par conséquent d’étudier la synthèse et les propriétés de deux composites biocéramiques/biopolymères : des hydrogels physiques de chitosane minéralisés avec de l’apatite, et une hardystonite architecturée imprégnée par des hydrogels physiques de chitosane. Afin d’obtenir le premier matériau, deux approches ont été développées. La première a consisté à fabriquer des hydrogels physiques de chitosane puis à les minéraliser avec de l’apatite ; la formation de microcapillaires se produit avec des conditions de synthèse spécifiques, et les précipités d’apatite ont été trouvés uniquement à la surface des hydrogels. La seconde approche consiste à convertir des suspensions homogènes contenant le phosphate de calcium et le chitosane en hydrogels de chitosane minéralisés par l’apatite. Les suspensions ont été préparées soit avec un mélange simultané, soit avec des mélanges successifs de suspensions phosphates de calcium avec les solutions de chitosane. Des agrégats minéraux plus petits avec une distribution plus uniforme ont été formés avec la méthode des mélanges successifs. Cela est attribué à une meilleure homogénéité, une viscosité plus faible et l’absence de chitosane. De manière générale, trois paramètres influencent les propriétés mécaniques d’hydrogels de chitosane minéralisés : la base utilisée pour la gélification (déterminant la vitesse de gélification : une grande vitesse conserve l’enchevêtrement des chaînes, résultant en une meilleure élasticité) ; la densité de la réticulation physique (cela induit un module de conservation plus important) et la force ionique (qui mène au désenchevêtrement des chaînes de chitosane, donc, à un faible module de conservation). Cette compréhension a permis l’utilisation de ces suspensions de phosphate de calcium-chitosane en tant qu’encre pour l’impression 3D. Les hydrogels de chitosane et les hydrogels minéralisés ne sont pas cytotoxiques. Pour fabriquer le second matériau, une encre pré-céramique a été imprimée en 3D puis frittée pour former une céramique d’hardystonite cristalline. Les scaffolds d’hardystonite ont été imprégnés par la solution de chitosane, converties ensuite en hydrogels physiques de chitosane. A plus forte concentration de chitosane, la viscosité de la solution était plus grande et l’imprégnation de la matrice plus lente. Avec une vitesse de gélification plus importante, qui dépend de la base utilisée pour la gélification, la perte de poids est plus faible pendant la gélification. L’hydrogel de chitosane a partiellement rempli les pores participant au support de charges externes et à la dissipation d’énergie par rupture. / Bone substitutes, an approach to attend social demand for bone healing and reparation, are temporary replacements of bone tissue, promote bone formation and growth and finally are bioresorbed. No single material meets these requirements; an alternative is a bioinspired composite material. The objective of this thesis was thus to study the synthesis and properties of two bioceramics/biopolymer composites: chitosan physical hydrogels mineralized with apatite and hardystonite scaffolds impregnated with chitosan physical hydrogels. To obtain the first material, two strategies were developed. The first one consisted in the fabrication of chitosan physical hydrogels and its subsequent mineralization with apatite; the formation of micro-capillaries occurred under particular synthesis conditions, and apatite precipitates were found only on the surface of hydrogels. The second strategy consisted in a simultaneous conversion of chitosan-calcium phosphate suspensions into chitosan-apatite hydrogels. The suspensions were prepared by sequential or simultaneous mixing of calcium and phosphate suspensions with chitosan solutions. Smaller and more uniformly distributed mineral aggregates were formed following sequential mixing, attributed to higher homogeneity, lower viscosity and no-presence of chitosan. This enabled the use of these chitosan-calcium phosphate suspensions as inks for 3-D printing. In general, three factors impacted the mechanical properties of mineralized chitosan hydrogels: the base used for gelation (determining the gelation rate: a higher rate preserved chain entanglement, resulting in higher elasticity); the density of physical crosslinks (hence a higher storage modulus) and the ionic strength (that led to chitosan chain disentanglements, thus, low storage modulus). Chitosan hydrogels and mineralized hydrogels were not cytotoxic, having no deleterious effects on osteoblasts proliferation. To fabricate the second material, pre-ceramic ink was 3-D printed and then sintered to form crystalline hardystonite ceramic. Hardystonite scaffolds were impregnated with chitosan solution that was, next, converted to chitosan physical hydrogel. At higher chitosan concentration, viscosity of solution was higher and scaffold impregnation was lower. At higher gelation rate, which depend on base used for gelation, lower weight loss during gelation. Chitosan hydrogel partially filled the pores contributing to bearing of external loads and to energy dissipated by fracture.

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Agradecemos a compreensão. on 2016-09-05T20:04:15Z (GMT) / Submitted by RODRIGO DOS SANTOS PEREIRA null (rd2502@yahoo.com.br) on 2016-09-13T00:17:32Z No. of bitstreams: 1 Tese Rodrigo.pdf: 108582918 bytes, checksum: 2c7b303850ee3479eadff15a0275764a (MD5) / Approved for entry into archive by Juliano Benedito Ferreira (julianoferreira@reitoria.unesp.br) on 2016-09-13T14:30:15Z (GMT) No. of bitstreams: 1 pereira_rs_dr_arafo.pdf: 108582918 bytes, checksum: 2c7b303850ee3479eadff15a0275764a (MD5) / Made available in DSpace on 2016-09-13T14:30:15Z (GMT). No. of bitstreams: 1 pereira_rs_dr_arafo.pdf: 108582918 bytes, checksum: 2c7b303850ee3479eadff15a0275764a (MD5) Previous issue date: 2016-07-19 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / A deficiência óssea vertical do seio maxilar impossibilita a instalação de implantes dentais necessários para a reabilitação protética contudo, técnicas cirúrgicas para a elevação da membrana sinusal e o uso dos biomateriais para enxertia óssea permitiram alterar esta condição. O ósseo autógeno é considerado o mais previsível e o padrão ouro para tal finalidade porém, biomateriais como o vidro bioativo, amplamente utilizado na odontologia, permitem substituí-lo. O objetivo deste estudo foi avaliar formação óssea, o comportamento celular e a taxa de reabsorção do vidro bioativo em 2 proporções: puro (grupo 1); adicionado ao osso autógeno 1:1 (grupo 2), comparando com o osso autógeno (grupo 3) em seios maxilares de humanos. Com 15 dias de operados os pacientes realizaram um tomografia computadorizada (TC) cone beam para determinar o volume inicial do enxerto (T1). Após 6 meses uma nova TC foi realizada determinando o volume ósseo final (T2) e também, a coleta das biópsias com a instalação simultânea de implantes dentais. As amostras foram divididas em 3 áreas: leito, intermediária e apical onde foram avaliadas a histomorfometria e imunomarcações para Runx2, VEGF, osteocalcina e TRAP. No grupo 1 os resultados foram de 42.6%, 44.5% e 48% de formação óssea no leito, intermediária e apical respectivamente. No grupo 2 as taxas encontradas foram de 36.6% no leito, 33.2% na intermediária e 45.8% na apical. No grupo 3 foi de 34.4% para o leito, 35.0 na intermediária e 42.0% na apical. (p>0.05) Os 3 grupos apresentaram-se com comportamento semelhante nas imunomarcações realizadas mostrando estar maturado e calcificado o suficiente para receber implantes dentais. As taxas de reabsorção também mostraram-se semelhantes com 44.2% para o grupo 1, 37.9% para o grupo 2 e 45.7% no grupo 3. (p>0.05) Assim, podemos concluir que o vidro bioativo possui resultados equiparados ao osso autógeno. / Vertical bone deficit due maxillary sinus pneumatization forbid the dental implants placement required for prosthetic rehabilitation however, surgical techniques to elevate maxillary sinus membrane and studies about biomaterials, allows repair these situation. Autogenous bone graft is considered the most predictable and the gold standard for this nevertheless, biomaterials as bioactive glass, has been widely used in dental surgery and allows substitute it. The aim of this study was evaluate the bone formation, the cellular behavior and the resorption rates of bioactive glass in 2 proportions: pure (group 1); add to autogenous bone graft (group 3), comparing then with autogenous bone graft in human maxillary sinus. Post-operative cone beam computed tomography (CT) was used to measure the initial graft volume after 15 days (T1). Six months later, another CT was performed to evaluate the final graft volume (T2) and determine the graft resorption rate and the harvest of biopsies with dental implants placement simultaneously. The samples were divided in 3 areas: bed, intermediately and apical which were evaluated to histomorphometric and immunostaining to Runx2, VEGF, osteocalcin and TRAP. In group 1 the bone formation were 42.6%, 44.5 and 48.0% for bed, intermediately and apical respectively. In group 2 the rates were 36.6% in bed, 33.2% in intermediately and 45.8% in apical. In group 3 were 34.4% in bed, 35.0% in intermediately and 42.0% in apical. (p>0.05) the 3 groups had the same behavior for the 4 proteins showing be matured and calcified to receive dental implants. The resorption rates were similar for the 3 groups with 44.2% % for group 1, 37.9% for group 2 and 45.7% for group 3. (p>0.05) In conclusion, the bioactive glass is a good substitute to autogenous bone graft in both proportions evaluated.

No description available.

One of the greatest obstacles to clinical translation of bone tissue engineering is the inability to effectively and efficiently vascularise scaffolds. This limits the size of defects that can be repaired, as blood perfusion is necessary to provide nutrient and waste exchange to tissue at the core of scaffolds. The goal of this work was to systematically explore whether architecture, at a scale of hundreds of microns, can be used to direct the growth of microvessels into the core of scaffolds. A pipeline was developed for the production of hydroxyapatite surfaces with controlled architecture. Three batches of hydroxyapatite were used with two different particle morphologies and size distributions. On sintering, one batch remained phase pure and the other two batches were biphasic mixtures of α-tricalcium phosphate (α-TCP) and hydroxyapatite. Sample production methods based on slip casting of a hydroxyapatite-gelatin slurry were explored. The most successful of these involved the use of curable silicone to produce moulds of high-resolution, three dimensional (3D) printed parts with the desired design. Parts were dried and sintered to produce patterned surfaces with higher resolution than obtainable through conventional 3D printing techniques. Given the difficulties associated with the structural reproducibility of concave pores architectures in 3D reported in the literature, in this work, a 2.5D model has been developed that varies architectural parameters in a controlled manner. Six contrasting architectures consisting of semi-circular ridges and grooves were produced. Grooves and ridges were designed to have widths of 330 μm and 660 μm, with periodicities, respectively, of 1240 μm and 630 μm. Groove depth was varied between 150 μm and 585 μm. Co-cultures of endothelial cells and osteoblasts were optimised and used to grow microcapillary-like structures (referred to as "microvessels") on substrates. Literature shows that these precursors to microcapillaries contain lumina and can produce functional vasculature, demonstrating their clinical promise. The effects of the composition and surface texture of grooved samples on microvessel formation were studied. It was found that surface microtopography and phase purity (α-TCP content) did not affect microvessel formation. However, hydroxyapatite architecture was found to significantly affect microvessel location and orientation. Microvessels were found to form predominantly in grooves or between convexities. Two metrics - the degree of alignment (DOA) and the degree of containment (DOC) - were developed to measure the alignment of endothelial cell structures and their localisation in grooves. For all patterned samples, the CD31 (an endothelial cell marker) signal was at least 2.5 times higher along grooves versus perpendicular to grooves. In addition, the average signal was at least two times higher within grooves than outside grooves for all samples. Small deep grooves had the highest DOA and DOC (6.13 and 4.05 respectively), and individual, highly aligned microvessels were formed. An image analysis method that compares sample X-ray microtomography sections to original designs to quantify architectural distortion was developed. This method will serve as a useful tool for improvements to architectural control for future studies. This body of work shows the crucial influence of architecture on microvessel self-assembly at the hundreds of micron scale. It also highlights that microvessel formation has a relatively low sensitivity to phase composition and microtopography. These findings have important implications for the design of porous scaffolds and the refinement of fabrication technologies. While important results were shown for six preliminary architectures, this work represents a toolkit that can be applied to screen any 2.5D architecture for its angiogenic potential. This work has laid the foundations that will allow elucidating the precise correspondence between architecture and microvessel organisation, ultimately enabling the "engineering" of microvasculature by tuning local scaffold design to achieve desirable microvessel properties.