In open angle glaucoma, dysfunction of the trabecular meshwork (TM) results in impaired aqueous humour outflow leading to an elevated intraocular pressure (IOP) that underlies optic nerve damage and irreversible blindness. Currently, no curative treatment is available for the disease. Indeed, most pharmacological and surgical interventions usually provide only temporary relief from elevated IOP while little progress has been made in targeting the root cause of this disease: correcting the dysfunctional TM. In this context, we hypothesized that regeneration/refunctionalization of the TM may represent an effective therapeutic option to halt disease progression or even reverse the pathologic process. We previously demonstrated in a rat model of glaucoma that the injection of mesenchymal stromal cells (MSCs) cultured under hypoxic conditions or their conditioned media (MSC-CM) into laser-damaged TM area results in tissue regeneration. Injection of MSC or conditioned media in our glaucoma model led to activation and proliferation of ocular progenitor cells culminating in TM regeneration and a decrease in IOP. However, the mechanistic basis for this regenerative process remained elusive. Thus, the aim of this thesis is to elucidate the mechanistic basis of MSC secretome-mediated TM regeneration and the subsequent decrease in IOP. We now demonstrate that injection of hypoxic MSC-CM into laser-induced glaucomatous eyes resulted in massive immune cell recruitment. We also demonstrate that these hypoxic MSC-CM conditioned cells produced pro-regenerative factors in vitro and in vivo. Next, employing a proteomic approach, we identified and verified the pro-regenerative effect of several factors secreted by hypoxic MSC-CM recruited cells, which in turn induced the activation/proliferation of ocular progenitor cells leading to TM regeneration and decreased IOP. Upon individual injection of the purified factors into glaucomatous rat eyes, we observed a partial and delayed but significant decrease in IOP that correlated with an increase in the activation and proliferation of neuronal progenitor cells in the TM area. The co-injection of these factors resulted in a significant decrease in IOP compared with individual factor injection. Importantly, this drop in IOP was associated with restoration of retinal functionality, thus demonstrating the importance of these factors in the TM regeneration process and disease control. The findings presented in this thesis provide a novel acellular therapeutic approach for glaucoma treatment via in situ TM regeneration. Moreover, the knowledge gained here could have a lasting impact on how we induce tissue regeneration in other degenerative diseases and lead to novel therapeutic advances in regenerative medicine. / Dans le glaucome à angle ouvert, le dysfonctionnement du trabéculum (TM), un tissu nécessaire à la filtration de l'humeur aqueuse, entraîne une élévation de la pression intraoculaire (PIO). Ceci cause des lésions au niveau du nerf optique et une cécité irréversible. Présentement, aucun traitement curatif n'a été développé pour cette maladie. Nous émettons l'hypothèse que la régénération et re-fonctionnalisation du trabéculum peut représenter une option thérapeutique efficace pour arrêter ou inverser la progression de la maladie dans de nombreux cas de glaucome. Nous avons précédemment démontré les effets régénérateurs des cellules mésenchymateuses (MSCs) et de leurs milieux conditionnés par l'hypoxie (MSC-CM) dans la régénération du TM suite à un dommage par laser. Ce processus a conduit à l'activation et à la prolifération des cellules progénitrices oculaires résultant en une diminution de la PIO dans un modèle de glaucome induit par laser chez le rat. Cependant, la base mécanistique de ce processus de régénération reste encore inconnue. Ainsi, le but de cette thèse de recherche est d'élucider cette base mécanistique de la régénération du TM médiée par le sécrétome des MSC et la diminution subséquente de la PIO. À cette fin, l'injection de MSC-CM hypoxique dans les yeux glaucomateux induits par laser a entraîné un important recrutement de cellules immunitaires. Sous l’action du MSC-CM, ces cellules produisent des facteurs pro-régénératifs in vitro et in vivo. Ensuite, nous avons utilisé une approche protéomique et vérifié l'effet pro-régénératif des facteurs sécrétés par ces cellules exposées au MSC-CM hypoxique, sur l'activation et la prolifération des cellules progénitrices oculaires et la PIO. Lors de l'injection de ces facteurs chez le rat glaucomateux, nous avons observé une augmentation significative de l'activation et de la prolifération des cellules progénitrices neuronales présentes dans la zone du TM, résultant en une diminution de la PIO. De plus, l’injection combinée de ces facteurs résulte en une diminution synergique importante de la PIO. Cette baisse de la PIO était associée à une restauration de la fonction rétinienne, démontrant ainsi l'importance de ces facteurs dans le processus de régénération du TM et de contrôle de la maladie. Les résultats présentés dans cette thèse pourraient amener à une nouvelle approche thérapeutique acellulaire pour le traitement du glaucome via la régénération du TM. De plus, les connaissances acquises au cours de cette thèse pourraient avoir un impact durable sur la manière d’aborder la régénération tissulaire dans d'autres maladies dégénératives et amener des avancées thérapeutiques nouvelles en médecine régénératrice

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[ES] En esta tesis, el principal objetivo es analizar el efecto de la osteoporosis en el comportamiento mecánico del hueso trabecular. El comportamiento del hueso trabecular se ha abordado desde diferentes enfoques: ensayos experimentales, modelos de elementos finitos (EF) e imagen médica obtenida por micro-CT. El impacto de la osteoporosis se ha estudiado en distintas escalas. Se ha abordado un estudio a nivel de tejido lamelar del hueso trabecular para estimar las ecuaciones que nos permitan definir las propiedades elásticas y resistentes en función de la densidad mineral ósea y la porosidad del tejido. Estas ecuaciones inferidas en los modelos de EF permitirán estudiar el comportamiento del tejido óseo lamelar abarcando un amplio rango de porosidades y DMOs. Se ha realizado un estudio de la estructura trabecular y del tejido lamelar utilizando un estereomicroscopio y el FESEM. Al nivel del tejido lamelar, se ha podido observar la disposición de las lamelas en distintas localizaciones, como en las trabéculas y sus intersecciones. Además, se ha podido evaluar la porosidad tisular natural y la producida por enfermedades óseas. Esta información se ha introducido en los modelos EF para definir las propiedades en las muestras. Las ecuaciones elásticas y resistentes estimadas consideran un comportamiento ortótropo y es posible cuantificar cómo afectan la DMO y la porosidad a estas propiedades. Se ha estudiado el comportamiento mecánico a compresión de muestras humanas de hueso trabecular procedente de cabezas femorales. Estas muestras se clasifican en tres grupos: sano, artrósico y osteoporótico. Estas muestras se han estudiado mediante ensayos experimentales, imagen médica y modelos de EF, permitiendo obtener gran información sobre el comportamiento mecánico del hueso. La respuesta mecánica está muy influenciada por la microarquitectura trabecular. Los resultados obtenidos más relevantes son el módulo aparente, la tensión de fallo y la deformación de fallo. Se han podido generar modelos de EF y estudiar la morfometría de las muestras. Los modelos de EF han permitido estimar las propiedades a nivel de tejido: módulo de Young a nivel de tejido y las deformaciones de fluencia y fallo. Los resultados muestran que a nivel de tejido no se aprecian variaciones en el módulo de Young aunque el hueso presente una patología ósea. Un comportamiento de fluencia similar se ha observado para todos los grupos, con diferencias únicamente en la deformación de fallo. El estudio morfométrico permite analizar los parámetros morfométricos que diferencian las muestras sanas de las enfermas. Se han correlacionado dichos parámetros con la respuesta mecánica para analizar los que tienen mayor influencia sobre ella. Las correlaciones permiten estimar la respuesta mecánica del hueso utilizando imagen médica sin realizar ensayos experimentales. Los parámetros obtenidos de la respuesta mecánica también se han relacionado entre sí, permitiendo conocer el comportamiento de cada una de las muestras que es diferente según la enfermedad que padecen. También, se han estudiado muestras bioinspiradas y estructuras TPMS obtenidas mediante fabricación aditiva. Estas muestras se han fabricado con PLA como material de impresión. Las muestras muestran un comportamiento ortótropo como las muestras trabeculares. Los simulantes pueden ser utilizados para hacer estudios previos cuando no se disponga de las muestras óseas. El tratamiento de fracturas óseas de gran tamaño requiere de técnicas no convencionales para la fijación del hueso. Suelen utilizarse andamios óseos con triple periodicidad que tienen como resultado propiedades ortótropas iguales en las tres direcciones ortogonales. Estos sistemas deben disponer de rigideces similares al hueso alrededor del defecto para evitar el aflojamiento del implante. Por ello, se propone una metodología que permite diseñar andamios óseos paciente-específicos que permiten ajustar las propiedades mecánicas ortótropas en función del hueso. / [CA] En esta tesi, el principal objectiu és analitzar l'efecte de l'osteoporosi en el comportament mecànic de l'os trabecular. El comportament de l'os trabecular s'ha abordat des de diferents plantejaments, com són els assajos experimentals, els models d'elements finits (EF) i la imatge mèdica obtinguda per micro-CT. L'impacte de l'osteoporosi s'ha estudiat en diferents escales. En primer lloc, s'ha abordat un estudi a nivell de teixit lamellar de l'os trabecular per a estimar les equacions que ens permeten definir les propietats elàstiques i resistents en funció de la densitat mineral òssia i la porositat del teixit. Estes equacions inferides en els models d'EF permetran estudiar el comportament del teixit ossi lamel·lar abastant un ampli rang de porositats i DMOs. S'ha realitzat un estudi de l'estructura trabecular i del teixit lamel·lar utilitzant un estereomicroscopi i el FESEM. Al nivell del teixit lamel·lar, s'ha pogut observar la disposició de les lamel·les en diferents localitzacions, com en les trabècules i les seues interseccions. A més, s'ha pogut avaluar la porositat tissular natural i la produïda per malalties òssies. Tota esta informació ha sigut incorporada als models numèrics per a poder definir les propietats en les mostres trabeculars. Les equacions elàstiques i resistents estimades consideren un comportament ortòtrop i és possible quantificar com afecten la DMO i la porositat a estes propietats. S'ha estudiat el comportament mecànic a compressió de mostres humanes d'os trabecular procedent de caps femorals i s'han classificat en tres grups: sa, artròsic i osteoporòtic. S'han estudiat mitjançant assajos experimentals, imatge médica i models d'EF, obtenint gran informació sobre el comportament mecànic de l'os. La resposta mecànica està fortament influenciada per la microarquitectura trabecular. Els resultats obtinguts més importants són el mòdul aparent, la tensió de fallada i la deformació de fallada. S'han generat models d'EF i estudiat la morfometria. Els models d'EF han permés estimar les propietats a nivell de teixit: el mòdul de Young a nivell de teixit i les deformacions de fluència i fallada. A nivell de teixit no s'aprecien variacions en el mòdul de Young encara que l'os present una patologia òssia. Un comportament de fluència similar s'ha observat per a tots els grups, amb diferències únicament en la deformació de fallada. L'estudi de la morfometria ha permés analitzar els paràmetres morfomètrics que diferencien les mostres sanes de les malaltes, i fins i tot a diferenciar entre diferents patologies. S'han correlacionat estos paràmetres amb la resposta mecànica per a analitzar aquells que tenen major influència sobre ella. Les correlacions obtingudes permeten estimar la resposta mecànica de l'os trabecular utilitzant les imatges micro-CT sense necessitat de realitzar assajos experimentals. Els paràmetres obtinguts de la resposta mecànica també s'han relacionat entre si. Estes relacions permeten conéixer el comportament de cadascuna de les mostres que és diferent segons la malaltia que patixen. També, s'han estudiat mostres bioinspirades i estructures TPMS obtingudes mitjançant fabricació additiva. Estes mostres bioinspirades s'han fabricat amb PLA com a material d'impressió. Les mostres mostren un comportament ortòtrop com les trabeculars. Estos simulants poden ser utilitzats per a fer estudis previs quan no es dispose de les mostres òssies. El tractament de fractures òssies de gran grandària requerix de tècniques no convencionals per a la fixació de l'os. Solen utilitzar-se bastides òssies amb triple periodicitat que tenen com a resultat propietats ortòtropes iguals en les tres direccions ortogonals. Estos sistemes han de disposar de rigideses similars a l'os en el veïnatge del defecte per a evitar l'afluixament de l'implant. S'ha proposat una metodologia que permet dissenyar bastides òssies pacient-específics que permeten ajustar les propietats mecàniques ortòtropes en cada direcció en funció de l'os. / [EN] The main objective of this thesis is to analyse the effect of osteoporosis on the mechanical behaviour of cancellous bone. The behaviour of cancellous bone has been addressed through different approaches: experimental tests, finite element (FE) models and medical imaging obtained by micro-CT. The impact of osteoporosis has been studied on different scales. Firstly, a study has been undertaken at the lamellar tissue level of trabecular bone to estimate the equations that define the elastic and strength properties as a function of bone mineral density and tissue porosity. These equations are inferred from FE models and allow us to characterise the behaviour of lamellar bone tissue covering a wide range of porosities and BMDs. A study of the cancellous structure and lamellar tissue has been carried out using a stereomicroscope and FESEM. At lamellar tissue level, it was possible to observe the arrangement of lamellae in different locations, such as trabeculae and at their intersections. In addition, it has been possible to evaluate the natural tissue porosity and that produced by bone diseases. All this information has been incorporated to the numerical models in order to define the properties of cancellous bone samples. The estimated elastic and strength equations consider an orthotropic behaviour and it is possible to quantify how BMD and porosity affect these properties. On the other hand, the mechanical behaviour under compression of human trabecular bone samples from femoral heads has been studied. These samples are classified into three study groups: healthy, osteoarthrosic and osteoporotic. The samples have been studied using experimental tests, medical imaging obtained and FE models, enabling to obtain a great deal of information regarding the mechanical behaviour of the bone. The trabecular microarchitecture strongly influences the mechanical response. The most important results obtained are the apparent modulus, the failure stress and the failure strain. It was possible to generate FE models and study the morphometry of the specimens. The FE models have made it possible to estimate the properties at the tissue level. The results revealed that at the tissue level, Young's modulus does not vary appreciably, even if the bone shows bone pathology. A similar yield behaviour is observed for all groups, with differences only in the final failure strain. The morphometry study has enabled to analyse the morphometric parameters that differentiate between healthy and diseased specimens and even to differentiate between different pathologies. The morphometric parameters have been correlated with the mechanical response to analyse those that have the most significant influence on the mechanical response. The correlations obtained make it possible to estimate the mechanical response of trabecular bone using micro-CT images without the need for experimental tests. The parameters obtained from the mechanical response have also been related. These relationships permit to know the behaviour of each of the samples, which is different depending on the disease they suffer. Finally, bioinspired samples and minimal surface area triple periodic structures obtained by additive manufacturing have been studied. These samples are made of PLA as the printing material. The samples show a similar orthotropic behaviour to the trabecular samples. These simulants can be used for pre-studies when bone samples are not available. The treatment of large bone fractures requires unconventional techniques for bone fixation. Bone scaffolds with triple periodicity that result in equal orthotropic properties in all three orthogonal directions are often used. These systems must have bone-like stiffness in the vicinity of the defect to avoid implant loosening. To this end, a methodology has been proposed to design patient-specific bone scaffolds that allow the orthotropic mechanical properties in each direction to be calibrated according to a specific bone. / The author and the supervisors of this PhD thesis acknowledge the Ministerio de Ciencia e Innovación y Universidades and the European Regional Development Fund (FEDER) for the financial support received through the projects PID2020-118920RB-I00 and PID2020-118480RB-C21 and C22 funded by MCIN/AEI/10.13039/501100011033, and the Generalitat Valenciana for Plan FDEGENT 2018 and Programa PROMETEO/2021/046. / Megías Díaz, R. (2024). Analysis of osteoporosis effect on the mechanical behaviour and morphometry of human cancellous bone [Tesis doctoral]. Universitat Politècnica de València. https://doi.org/10.4995/Thesis/10251/206355

Orientador: Guilherme de Paula Nogueira / Banca: Yuri Tani Utsunomiya / Banca: Marco Antonio Rodrigues Fernandes / Resumo: O objetivo desse estudo foi inferir a Densidade Mineral óssea (DMO) a partir da imagem radiográfica (Raios-X) usando como referência a DMO de uma escada de alumínio mensurada por Absorciometria por de Duas-Energias (DXA). Para isso foram utilizadas 30 amostras de tecido ósseo cortical e 30 amostras de tecido ósseo trabecular de osso bovino "in vitro". Foram mensurados neste estudo a Densidade Mineral Óssea (DMO), o Conteúdo Mineral Ósseo (CMO) ambos obtidos pelo densitômetro LUNAR®-DPX ALPHA; a Quantidade Mineral Óssea (QMO) massa das cinzas após a calcinação das amostras; a densidade real das amostras (dReal=massa/volume). Foram feitas 10 tomadas de RX com as 60 amostras, entremeadas pela escada de alumínio (referencial densitométrico). A correlação entre as técnicas (DR-tons de cinza e DXA g/cm2) gerou equações de regressão para cada uma das dez radiografias e permitiu inferir a densidade mineral óssea (DMODR), obtida através da conversão dos tons de cinza em densidade radiográfica pelo software ImageJ® para cada uma das 10 tomadas radiográficas; calculou-se então a média da densidade mineral óssea calculada pela densitometria radiográfica (XDMODR) de todas as dez radiografias. Foi observado que a média da densidade real das amostras foi de 2,2±0,23g/cm2 e 1,05±0,09g/cm2 enquanto que a média da densidade mensurada pelo DXA foi de 0,73±0,11g/cm2 e 0,22±0,11g/cm2, observou que a media da DMODR ficou em 0,93±0,11g/cm2 e 0,34±0,14 g/cm2 para o osso cortical e trabecular respect... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: The aim of this study was to infer the bone mineral density (BMD) from the radiographic image (X-X) with reference to the BMD of an aluminum step wedge measured by absorptiometry Dual-energy (DXA). For this we used 30 samples of cortical bone 30 and cancellous bone tissue samples of bovine bone "in vitro". Were measured in this study Bone Mineral Density (BMD), bone mineral content (BMC) both obtained by densitometer LUNAR® DPX-ALPHA; Volume Bone Mineral (QMO) mass of ash after calcination of the samples; the real density of the samples (dReal = mass / volume). 10 taken RX were made with the samples 60, interspersed by aluminum step wedge (densitometric reference). The correlation between the techniques (gray DR-tones and DXA g / cm 2) generated regression equations for each of the ten X-rays and allowed to infer bone mineral density (DMODR) obtained by converting grayscale radiographic density by ImageJ® for each of the 10 radiographic taken. It was then calculated average bone mineral density calculated by X-ray densitometry (XDMODR) radiographs of all ten. Was observed that the average true density of the samples was 2.2±0.23g/cm2 and 1.05±0.09g/cm2 while the average density measured by DXA was 0.73±0.11g/cm2 and 0.22±0.11g/cm2 interesting that the average DMODR was 0.93±0.11g/cm2 and 0.34±0.14g/cm2 for cortical and cancellous bone respectively. The average of the BMC was 0.19±0.08g and 0.9±0.02g, and the average QMO 0.14±0.04g and 0.05±0 03g for cortical and cancellous bo... (Complete abstract click electronic access below) / Mestre

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No. of bitstreams: 1 fernandes_ra_me_araca_int.pdf: 2718642 bytes, checksum: 4dea3e0dc607519bbdbdb41a172e9277 (MD5) Previous issue date: 2019-01-18 / Coordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES) / O objetivo desse estudo foi inferir a Densidade Mineral óssea (DMO) a partir da imagem radiográfica (Raios-X) usando como referência a DMO de uma escada de alumínio mensurada por Absorciometria por de Duas-Energias (DXA). Para isso foram utilizadas 30 amostras de tecido ósseo cortical e 30 amostras de tecido ósseo trabecular de osso bovino “in vitro”. Foram mensurados neste estudo a Densidade Mineral Óssea (DMO), o Conteúdo Mineral Ósseo (CMO) ambos obtidos pelo densitômetro LUNAR®–DPX ALPHA; a Quantidade Mineral Óssea (QMO) massa das cinzas após a calcinação das amostras; a densidade real das amostras (dReal=massa/volume). Foram feitas 10 tomadas de RX com as 60 amostras, entremeadas pela escada de alumínio (referencial densitométrico). A correlação entre as técnicas (DR-tons de cinza e DXA g/cm2) gerou equações de regressão para cada uma das dez radiografias e permitiu inferir a densidade mineral óssea (DMODR), obtida através da conversão dos tons de cinza em densidade radiográfica pelo software ImageJ® para cada uma das 10 tomadas radiográficas; calculou-se então a média da densidade mineral óssea calculada pela densitometria radiográfica (XDMODR) de todas as dez radiografias. Foi observado que a média da densidade real das amostras foi de 2,2±0,23g/cm2 e 1,05±0,09g/cm2 enquanto que a média da densidade mensurada pelo DXA foi de 0,73±0,11g/cm2 e 0,22±0,11g/cm2, observou que a media da DMODR ficou em 0,93±0,11g/cm2 e 0,34±0,14 g/cm2 para o osso cortical e trabecular respectivamente. A média da CMO foi de 0,19±0,9g e 0,08±0,02g e a media da QMO 0,14±0,04g e 0,05±0,03g para o osso cortical e trabecular respectivamente. Foi possível obter bons coeficientes de determinação entre todas as variáveis estudadas: CMO e QMO, R2 =0,747; DMO e dReal com R2 = 0,765; dReal e DMODR; DMO e XDMODR, respectivamente 0,764 e 0,856. Para as correlações foram usadas amostras de dois tecidos ósseos cortical e trabecular, o que sugere distinção entre as amostras com as metodologias utilizadas / The aim of this study was to infer the bone mineral density (BMD) from the radiographic image (X-X) with reference to the BMD of an aluminum step wedge measured by absorptiometry Dual-energy (DXA). For this we used 30 samples of cortical bone 30 and cancellous bone tissue samples of bovine bone "in vitro". Were measured in this study Bone Mineral Density (BMD), bone mineral content (BMC) both obtained by densitometer LUNAR® DPX-ALPHA; Volume Bone Mineral (QMO) mass of ash after calcination of the samples; the real density of the samples (dReal = mass / volume). 10 taken RX were made with the samples 60, interspersed by aluminum step wedge (densitometric reference). The correlation between the techniques (gray DR-tones and DXA g / cm 2) generated regression equations for each of the ten X-rays and allowed to infer bone mineral density (DMODR) obtained by converting grayscale radiographic density by ImageJ® for each of the 10 radiographic taken. It was then calculated average bone mineral density calculated by X-ray densitometry (XDMODR) radiographs of all ten. Was observed that the average true density of the samples was 2.2±0.23g/cm2 and 1.05±0.09g/cm2 while the average density measured by DXA was 0.73±0.11g/cm2 and 0.22±0.11g/cm2 interesting that the average DMODR was 0.93±0.11g/cm2 and 0.34±0.14g/cm2 for cortical and cancellous bone respectively. The average of the BMC was 0.19±0.08g and 0.9±0.02g, and the average QMO 0.14±0.04g and 0.05±0 03g for cortical and cancellous bone respectively. It was possible to obtain good determination coefficients between all variables: BMC and QMO, R2 = 0.747; BMD and dReal with R2 = 0.765; dReal and DMODR; BMD and XDMODR respectively 0.764 and 0.856. For correlations were used two samples of cortical and cancellous bone tissues, suggesting a difference between the samples with the methodologies used.

Si l’obésité a longtemps été considérée comme protectrice pour le squelette, sa prise en charge nutritionnelle s’accompagne d’une perte osseuse. Dans le but d’étudier les mécanismes liant le tissu adipeux et le tissu osseux, des rats ont été nourris avec un régime riche en graisse et en sucre afin d’induire une obésité et ses complications métaboliques. Puis, ces rats ont suivi un programme de prise en charge de l’obésité associant un rééquilibrage nutritionnel à de l’activité physique. Des investigations de la densité, de la micro architecture trabéculaire, de la qualité corticale, des propriétés biomécaniques et des paramètres cellulaires du tissu osseux ont été réalisées sur le squelette des rats. L’obésité induite a amélioré les paramètres densitométriques et corticaux des rats. La prise en charge de l’obésité a entraîné des effets contrastés. Le rééquilibrage de l’alimentation n’a pas altéré la densité ni les paramètres biomécaniques et corticaux du tissu osseux des rats en dépit d’altérations de la micro architecture et des perturbations l’activité cellulaire osseuse. L’exercice a augmenté la densité minérale osseuse du tibia des rats. Nos résultats montrent que l’inclusion de l’exercice dans un programme de prise en charge de l’obésité permet de potentialiser la perte de masse grasse et de maintenir l’intégrité du squelette. En étudiant l’impact d’un régime obésogène et de la prise en charge de l’obésité sur le tissu osseux chez le rat, ce travail de thèse apporte sa contribution dans la compréhension des mécanismes liant le tissu adipeux et le tissu osseux. / While obesity has long been considered protective of bone tissue, its nutritional management is linked to a concomitant bone loss. In order to access these mechanisms, male rats were given a high fat / high sucrose diet to induce obesity and its resulting metabolic disorders. Then, these rats participated in a well balanced nutritional program combined, or not, with physical activity in order to treat their obesity. Investigations on density, trabecular micro-architecture, cortical quality, biomechanical properties and cellular parameters of bone tissue were performed on rat’s skeleton. Obesity had improved the bone density and cortical parameters of rats. Obesity management had induced mixed effects. The well balanced diet did alter neither the bone density nor biomechanical and cortical parameters despite the presence of alterations in the trabecular micro architecture and some disturbances of bone cellular activity. Exercise by itself had increased the tibia bone mineral density. Our results show that including exercise in obesity management allows increasing the fat mass loss and maintaining the skeleton integrity. By studying the impact of an obesogenic diet and obesity treatment on bone tissue in rats, this study brings its contribution to the understanding of the mechanisms linking adipose tissue and bone tissue.

[Truncated abstract] In this work a computer simulation program, similar to a finite element program, is used to study the relationship between skeletal tissue structure and function. Though other factors affect the shape of bone (e.g., genetics, hormones, blood supply), the skeleton adapts its shape mainly in response to the mechanical environment to which it is exposed throughout life. The specific relationship between the mechanical environment and the mechanical adaptation response of the skeleton is reviewed. Theories of mechanical adaptation are applied to the sites of tendon attachment to bone (entheses), the adaptation of generalised trabecular bone (i.e., Wolff’s Law of trabecular architecture), sesamoid bones that are often found where a tendon wraps around a bony pulley, and the internal trabecular structure of a whole bony sesamoid such as the patella. The relative importance of compression rather than tension in bone adaptation theories is still not fully understood. Some mechanical adaptation theories suggest that an overwhelming tensile stress at a skeletal location does not stimulate bone deposition, but would instead lead to bone resorption. The skeletal locations studied in this work were chosen because they have been proposed to be in tension. Computer simulations involving models are an ideal method to analyse the mechanical environment of a skeletal location. They are able to determine the mechanical stresses at, and the stress patterns around, complex biological situations. This study uses a two dimensional computer simulation program, Fast Lagrangian Analysis of Continua (Flac), to analyse the stress at the skeletal locations, and to test theories of mechanical adaptation over time by simulating physiological adaptation. The initial purpose of this study is to examine the stress in the skeletal tissue in generalised trabeculae, anatomical sites where a tendon wraps around a bony pulley, in the trabecular networks that fill the patella, and at tendon attachments. A secondary purpose, that follows directly from the first, is to relate the results of these initial stress analyses to existing and hypothetical skeletal tissue remodelling theories, to suggest how the complex skeletal structures might be generated solely in response to their mechanical environment. The term “remodelling” is used throughout this work to refer to mechanical adaptation of bone, usually at a surface of bone, rather than the internal regeneration of osteons (Haversion systems)

Afin de mieux comprendre le comportement mécanique de l’os trabéculaire et d’améliorer la prédiction du risque de fracture, l’objectif de cette thèse est de développer un modèle numérique « bio-fidèle » prenant en compte l’hétérogénéité tissulaire, et de déterminer les contraintes mécaniques au sein des travées osseuses, dans le domaine de déformation élastique. À l’échelle tissulaire, une étude par nanoindentation a permis de dissocier les comportements élastiques et plastiques de l’os en fonction de sa composition (minéral/collagène). Ainsi, le comportement élastique du tissu osseux serait principalement lié à sa quantité de minéral alors que ses propriétés plastiques seraient davantage liées à la phase organique. Une loi reliant le degré de minéralisation de l’os (DMB) au module élastique a été déterminée dans l’os humain. La création d’un modèle numérique reproduisant de manière rigoureuse le comportement élastique de l’os trabéculaire, nécessite la prise en compte de l’hétérogénéité de la quantité de minéral (DMB) et donc son acquisition en 3D. Grâce à une méthode de recalage d’image 2D/3D, les acquisitions de microtomographie ont été comparées aux valeurs obtenues par microradiographie quantitative, méthode de référence de mesure du DMB. Sous certaines conditions, la microtomographie permet une évaluation correcte de l’hétérogénéité minérale. La création et l’analyse d’un modèle numérique par éléments finis de l’os trabéculaire, à partir des images de tomographie, a montré l’importance des paramètres du modèle (taille et formulation des éléments) ainsi que le rôle de l’hétérogénéité minérale sur l’évaluation des contraintes locales appliquées aux travées osseuses. / Finite element modeling has become more and more suitable to estimate the mechanical properties of trabecular bone. Such models tend to be used to evaluate bone fracture risk. The main goal of this study was to create a bio-faithful model of trabecular bone to evaluate elastic stresses fields in trabeculae. In a first part, a nanoindentation study lead to a dissociation of elastic and plastic behavior of bone tissue, depending of its composition (mineral/collagen). At osteon level, bone elastic behavior mainly depends on mineral quantity whereas its plastic behavior mainly depends on collagen maturity. The relation between degree of mineralization of bone (DMB) and elastic modulus have been determined for human bone. The purpose of second part of the study was to evaluate DMB heterogeneity inside a 3D model of trabecular bone. Using 2D/3D registration, we compare the results obtain with high resolution microtomography to those from quantitative microradiography, the goldstandard method used to measure DMB. We prove that it was possible to obtain a good evaluation of mineral heterogeneity in trabecular bone by tomography. The last part of this study, is dedicated to the creation of a finite element model of trabecular bone. After analyzing the influence of finite element modeling parameter on the assessment of mechanical response (size and element formulation), we showed that the integration of mineral heterogeneity at the tissue level lead to strong modifications of stress fields in bone trabeculae. The results of this study prove that bone mineral heterogeneity is an important parameter and should be taken into account when evaluating trabecular bone mechanical properties.

L'analyse de la microstructure osseuse joue un rôle important pour étudier des maladies de l'os comme l'ostéoporose. Des nouveaux scanners périphériques haute résolution (HR-pQCT) permettent de faire des acquisitions de la micro-architecture osseuse in-vivo sur l'homme. Toutefois la résolution spatiale de ces appareils reste comparable à la taille des travées osseuses, ce qui limite leur analyse quantitative. L'objectif de cette thèse est de proposer de nouvelles approches jointes super-résolution/ segmentation pour une analyse quantitative plus fine des images HR-pQCT in-vivo de la structure osseuse trabéculaire. Dans une première étape nous nous sommes concentrés sur des méthodes 2D de super-résolution avec régularisation par variation totale (TV) puis par variation totale d'ordre plus élevé (Higher Degree TV), avec minimisation par un algorithme ADMM (Alternating Direction Method of Multipliers). Ensuite, nous avons proposé une méthode itérative combinant le principe de Morozov et la méthode de Newton pour estimer le paramètre de régularisation TV. Comparé à la méthode UPRE (Unbiased Predictive Risk Estimator), la méthode proposée est plus rapide et ne requiert pas un balayage exhaustif des valeurs des paramètres. Nous avons développé dans une deuxième étape une méthode de super-résolution/segmentation conjointe avec un a priori basé sur la Variation Totale et une relaxation convexe (Tvbox), qui permet d'améliorer les paramètres quantitatifs de l'os et de la connectivité 3D. La méthode a été validée sur des images expérimentales micro-CT déteriorées artificiellement. Finalement, en vue de l'application à des images réelles HR-pQCT, nous nous sommes intéressés à une approche conjointe semi-aveugle super-résolution/segmentation qui vise à estimer à la fois l'image binaire super-résolue et le noyau de convolution. Des résultats sur des images micro-CT et HR-pQCT sont présentés. En conclusion, notre travail montre que les méthodes d'optimisation basées sur la régularisation TV sont prometteurs pour améliorer la quantification de la micro-architecture osseuse sur des images HR-pQCT. / The investigation of trabecular bone micro-architecture provides relevant information to determine the bone strength, an important parameter in osteoporosis investigation. While the spatial resolution of clinical CT is not sufficient to resolve the trabecular structure, the High Resolution peripheral Quantitative CT (HR-pQCT) has been developed to investigate bone micro-architecture in-vivo at peripheral sites (tibia and radius). Despite this considerable progress, the quantification of 3D trabecular bone micro-architecture in-vivo remains limited due to a lack of spatial resolution compared to the trabeculae size. The objective of this thesis is to propose new joint super-resolution/segmentation approaches for improving the quantitative analysis of in-vivo HR-pQCT images of the trabecular bone structure. To begin with, we have investigated 2D super-resolution methods based on Total Variation (TV) and Higher Degree Total Variation (HDTV) and Alternating Direction Method of Multipliers (ADMM) minimization. Afterwards, an iterative method combining the Morozov principle and the Newton method was proposed in order to estimate the TV regularization parameter. The proposed method provides a very good regularization parameter only in few iterations compared with the UPRE method that requires an extensive scanning of parameter values. Furthermore, we have developed a 3D joint super-resolution/segmentation method based on a TV a prior with a convex relaxation (TVbox). The validation of the proposed methods was made on experimental micro-CT bone images artificially deteriorated. The results showed an improvement of the bone parameters and 3D connectivity with the TVbox method. Moreover, we have investigated a semi-blind joint super-resolution/ segmentation approach aiming to estimate both the binary super-resolved image and the assumed Gaussian blurring kernel that is not known for the real HR-pQCT images. Results on micro-CT and HR-pQCT experimental bone images were presented. In conclusion, our work has shown that TV based regularization methods promise to improve the quantification of bone micro-architecture from HR-pQCT images.