A osteonecrose dos maxilares associada ao uso dos Bisfosfonatos é uma exposição óssea que persiste por mais de 8 semanas na cavidade oral em pacientes sob tratamento com bisfosfonatos e que não foram submetidos a radioterapia de cabeça e pescoço. Explicar o motivo do desenvolvimento destas lesões ósseas principalmente nos maxilares e desvendar a sua patofisiologia ainda é necessário. Por isso, o reparo ósseo em um modelo animal de osteonecrose dos maxilares associada ao uso de bisfosfonatos em ratos Wistar (Rattus norvegicus, albinus) foi avaliado através de análise microscópica e molecular. A amostra foi composta por 48 ratos machos, com 12 semanas de vida e peso aproximado de 300 gramas, que foram submetidos a administração de Ácido Zoledrônico, 0,6 mg/kg a cada 28 dias com um total de 5 doses. Os animais foram dividos em quatro grupos, cada um composto por 12 animais; 2 grupos de tratamento AZ e AZ-experimental (AZ-exp) e 2 grupos controles CO e CO-experimental (CO-exp) com administração de cloreto de sódio 0,9% no mesmo volume e frequencia do Ácido Zoledrônico. As soluções foram administradas por via intraperitoneal. O grupo AZ-exp e o CO-exp foram submetidos a exodontias dos molares superiores direitos e a realização de defeito ósseo no fêmur esquerdo 45 dias após a primeira aplicação das soluções. A eutanásia dos animais ocorreu após 150 dias do início do experimento. A avaliação histológica foi realizada através de análises qualitativa e quantitativa, verificou a presença de sequestros ósseos, áreas de osteonecrose e áreas de osso total por meio de estudos de microscopia óptica pela coloração Hematoxilina e Eosina. Análise quantitativa da expressão do RNAm de proteínas envolvidas no processo de reparo ósseo pelo método de reação em cadeia da polimerase em tempo real (RealTimePCR) também foi realizada para avaliação dos osteoclastos (RANK, RANKL e OPG), osteoblastos (ALPL e OCN) osteócitos (DMP-1 e PHEX) e vascularização (VEGF). Apenas o grupo com administração de AZ e realização de exodontias apresentou sequestros ósseos e áreas significativamente maiores de osteonecrose em comparação aos animais com exodontia e sem a administração de AZ. Não foram encontradas áreas de osteonecrose na análise do fêmur. A avaliação do osso total apresentou maior quantidade de osso nos animais submetidos a procedimentos cirúrgicos e administração de AZ, porém não houve significância estatística. Na análise molecular, a administração de AZ e a realização das exodontias e do defeito ósseo, provocou alteração na expressão de marcadores para osteoclastos. A administração do AZ associada à realização dos procedimentos cirúrgicos aumentou a expressão de marcadores iniciais da osteoblastogênese e diminuiu a expressão tardia dos osteoblastos. A expressão de marcadores de osteócitos foi menor após a administração de AZ e exodontias na maxila, no entanto, no fêmur, houve aumento na expressão destes marcadores. A realização das exodontias e de defeitos ósseos no fêmur aumentou a expressão de VEGF nos grupos com e sem administração de AZ. Estes resultados evidenciam a interferência do Ácido Zoledrônico no reparo ósseo dos alvéolos e dos defeitos ósseos nos fêmures, causando incidência de osteonecrose na maxila, atraso na remodelação óssea, e ausência de lesões de osteonecrose no fêmur. A presença de lesões de osteonecrose na maxila pode ser consequência do atraso na remodelação óssea, evidenciado pela alteração na expressão de marcadores para osteclastos e osteoblastos, e as mesmas ocasionam a diminuição da expressão de proteínas pelos osteócitos. / Bisphosphonate-related osteonecrosis of the jaws is a bone exposure persisting for more than 8 weeks in the oral cavity in patients receiving bisphosphonates and who did not undergo radiation therapy for head and neck. Explain the reason for these development bone lesions mainly in the jaws and unveil its pathophysiology is still needed. Therefore, the bone repair in an animal model of Bisphosphonate-related osteonecrosis of the jaws in Wistar rats (Rattus norvegicus, Albinus) was evaluated by microscopic and molecular analysis. The sample was composed of 48 male rats, with 12 weeks old and weighing approximately of 300 grams, who underwent the administration of zoledronic acid, 0.6 mg / kg every 28 days with a total of 5 doses. The animals were divided into four groups, each consisting of 12 animals; 2 treatment groups AZ and AZ-experimental (AZ-exp) and 2 control groups CO and CO-experimental (CO-exp) with sodium chloride administration 0.9% in the same volume and frequency of zoledronic acid. All solutions were administered intraperitoneally. The AZ-exp group and the CO-exp group underwent extractions rights upper molars and the bone defect was performed in the left femur 45 days after the first application of the solutions. Euthanasia of animals occurred after 150 days from the beginning of the experiment. Histological evaluation was performed through quantitative and qualitative analysis checked the presence of bone sequestration, osteonecrosis area and whole bone areas by means of optical microscopy by hematoxylin-eosin staining. Quantitative analysis of mRNA expression of proteins involved in bone repair by polymerase chain reaction method in real time (RealTimePCR) was also performed: osteoclasts (RANK, RANKL and OPG), osteoblasts (ALPL and OCN) osteocytes (PMD-1 and PHEX) and vascularization (VEGF). Only the group with administration of AZ and performing tooth extractions presented bone sequestration and significantly larger areas of osteonecrosis when compared to animals with extraction and without AZ administration. There were no areas of osteonecrosis of the femoral analysis. The evaluation of the whole bone showed more bone in animals undergoing surgeries and AZ administration but there was no statistical significance. Molecular analysis, AZ administration and the performance of tooth extractions and bone defect changed the expression of markers for osteoclasts. The AZ administration associated with surgical procedures increased the expression of early markers of osteoblastogenesis and decreased the late expression of osteoblasts. The expression of osteocytes markers was lower after AZ administration and maxillary tooth extractions; however, there was an increase in the expression of these markers in the femur. The performance of tooth extractions and bone defects in the femur increased VEGF expression in animals with and without administration of AZ. These results show the interference of zoledronic acid on bone repair of the sockets and bone defects in the femurs, causing incidence of osteonecrosis of the upper jaw, delayed bone remodeling, and no femoral osteonecrosis injuries. The presence of osteonecrosis injuries in the upper jaw can be the consequence of the delay on bone remodeling, evidenced by the change in the markers expression for osteoclasts and osteoblasts, and they cause the decreased in protein expression by osteocytes.

Estudos recentes sobre disfunção ventricular demonstram o potencial terapêutico da modulação da matriz extracelular. Isso se dá pela influência que a referida matriz tem sobre a função sistólica e a diastólica do coração. Outros estudos demonstram a influência do digital sobre os sistemas neurohormonais desbalanceados no cenário de disfunção ventricular levantando uma questão acerca do potencial do digital como modulador da deposição do colágeno intersticial e perivascular miocárdico. Sabendo-se da importância prognóstica que a concentração do colágeno no referido cenário tem, a literatura apresenta uma lacuna de conhecimento. Objetivo: Avaliar o papel do digital no remodelamento miocárdico em um modelo experimental. Material e Métodos: 60 ratos Wistar foram separados em 3 grupos de 20. Um grupo controle (GC); outro grupo submetido ao modelo experimental de uninefrectomia, administração de água de beber com 1% de NaCl e de aldosterona subcutânea (GA); e outro grupo submetido ao mesmo modelo experimental, mas também recebendo digitoxina na ração de comer na dose de 100 g/Kg/dia (GAD). Resultados: A fração de volume de colágeno intersticial e perivascular mostrou-se maior no GA comparado aos outros dois grupos (GC e GAD). O índice de desempenho miocárdico mostrou diferença estatisticamente significativa entre o GA (0,49 ± 0,08) e o GC (0,32 ± 0,06) e o GAD (0,4 ± 0,13) (p=0,001). Os níveis séricos de BNP mostraram diferença estatisticamente significativa entre o GA (1,07 ± 0,32 ng/ml) e o GC (0,75 ± 0,19 ng/ml) e o GAD (0,84 ± 0,21 ng/ml) (p=0,01). Os níveis de metaloproteinases não diferiram entre os grupos. Houve uma correlação positiva entre uma maior fração de encurtamento e menores níveis séricos de BNP no GAD. Conclusões: Esses dados demonstram que a digitoxina teve efeito reduzindo a deposição de colágeno intersticial e perivascular e melhorando a função cardíaca avaliada pelo BNP e IDM nesse modelo experimental / Recent studies on myocardial dysfunction are highlighting the therapeutic potential of the myocardial extracellular matrix management. Its interesting to highlight the importance of the dynamics of the cardiac extracellular matrix, because even the systolic and diastolic functions are implicated on it. Other studies showed that digital compounds may regulates the neuroendocrin misbalance due to myocardial impairment and influencing these systems the digital compounds may regulates the interstitial collagen deposition. Objective: To evaluate the role of the digital on a myocardial fibrosis in an experimental model, examining if the digital is able to prevent the collagen deposition. Methods: The sample was divided in 20 rats from the control group (CG); 20 rats submitted to a fibrosis experimental model in which the rats are uninefrectomized, drink water with 1% NaCl during the protocol and receive aldosterone through an osmotic minipump (AG); and 20 rats submitted to the same experimental model treated with digitoxin in a daily dose of 100 g/Kg (DAG). Results: The interstitial and perivascular collagen volume fraction showed a significant difference between the AG and the other 2 groups (CG and DAG). The myocardial performance index showed a significant difference between the AG (0.49 ± 0.08) and the CG (0.32 ± 0.06) and the DAG (0.40 ± 0.13) (p=0.001). The BNP levels showed a significant difference between the AG (1.07 ± 0.32 ng/ml) and the CG (0.75 ± 0.19 ng/ml) and the DAG (0.84 ± 0.21 ng/ml) (p=0.01). The metalloproteinases levels did not differ among the groups and there was a positive correlation between the shortening fraction and the BNP levels among the GAD animals. Conclusion: These data demonstrate the digitoxin positive effect on the myocardial collagen deposition in this experimental model of interstitial fibrosis and could have a new therapeutic target previously unexplored

Orientador: Wilson Nadruz Júnior / Dissertação (mestrado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-25T05:29:30Z (GMT). No. of bitstreams: 1 Mendes_PauloRobertoAraujo_M.pdf: 976609 bytes, checksum: d5b069ade658cb23f9494d04dbbb255d (MD5) Previous issue date: 2014 / Resumo: Pacientes hipertensos estão predispostos ao remodelamento do ventrículo esquerdo (VE) e frequentemente apresentam queda na função pulmonar quando comparados à população geral. Neste estudo investigamos a associação entre dados espirométricos e ecocardiográficos em indivíduos hipertensos não fumantes. Num estudo transversal, 107 pacientes hipertensos (60 mulheres) foram avaliados por análises clínicas, hemodinâmicas, laboratoriais e ecocardiográficos. A capacidade vital (CV), a capacidade vital forçada (CVF), o volume expiratório forçado no primeiro segundo (VEF1), o volume expiratório forçado no sexto segundo (VEF6) e a relação VEF1\CVF foram medidos através de espirometria. Nas mulheres, o índice de massa do VE e a relação E\Em correlacionaram-se com variáveis espirométricas, enquanto que a espessura relativa da parede somente se correlacionou com o percentual de CVF previsto. Nos homens, somente o índice de massa do VE se correlacionou com variáveis espirométricas. Análise de regressão tipo stepwise mostrou que o índice de massa do VE não esteve associado com nemhum parâmetro espirométrico após ajuste para os potenciais confundidores nos homens, enquanto que CVF e VEF6 se associaram de maneira significativa com a massa do VE e a relação E\Em no sexo feminino. Entretanto, marcadores inflamatórios tais como Proteína C reativa plasmática e os níveis séricos de metaloproteinases 2 e 9 não influenciaram estas associações. Em conclusão, o declínio na função pulmonar está independentemente associado com maior massa e pior função diastólica do VE em mulheres hipertensas / Abstract: Hypertensive patients are predisposed to left ventricular (LV) remodeling and frequently exhibit decline in lung function as compared to the general population. Here, we investigated the association between spirometric and echocardiographic data in non-smoking hypertensive subjects. In a cross-sectional study, 107 hypertensive patients (60 women) were evaluated by clinical, hemodynamic, laboratory and echocardiographic analysis. Vital capacity, forced vital capacity (FVC), forced expired volume in 1s (FEV1) and in 6s (FEV6) and FEV1/FVC ratio were estimated by spirometry. In women, LV mass index and E/Em ratio correlated with spirometric variables, while relative wall thickness only correlated with the percentage of predicted FVC. In men, only LV mass index correlated with spirometric variables. Stepwise regression analysis showed that LV mass index did not associate with any spirometric parameter after adjustment for potential confounders in men, while markers of restrictive and obstructive lung dysfunction, such as reduced FVC and FEV6, were significantly associated with LV mass and E/Em ratio in women. Furthermore, inflammatory markers such plasma C-reactive protein and matrix-metalloproteinases-2 and -9 levels did not influence these associations. In conclusion, decline in lung function is independently associated with higher LV mass and worse LV diastolic function in hypertensive women / Mestrado / Clinica Medica / Mestre em Clinica Medica

Orientador: Hernandes Faustino de Carvalho / Dissertação (mestrado) - Universidade Estadual de Campinas, Instituto de Biologia / Made available in DSpace on 2018-08-21T07:59:39Z (GMT). No. of bitstreams: 1 Rosa-Ribeiro_Rafaela_M.pdf: 2991640 bytes, checksum: 1c3d98622022906218832c47353b3c63 (MD5) Previous issue date: 2012 / Resumo: O desenvolvimento, a fisiologia e o câncer de próstata dependem de balanços entre os níveis de andrógenos e estrógenos, que agem via receptor de andrógenos (AR) e receptores de estrógeno (ER'alfa' e ER'beta'), respectivamente. Sabe-se que a cinética de morte das células epiteliais da próstata ventral de ratos (PV) após castração (grupo Cas), administração de alta dose de 17'beta'-estradiol (grupo E2) e combinação de ambos (grupo Cas+E2) é diferenciada, com um nítido efeito aditivo nesta última situação (Garcia-Florez et al, 2005). Neste trabalho, procuramos investigar elementos comuns e exclusivos a estas diferentes condições hormonais, empregando análises morfológicas, estudo de componentes da via AKT/PTEN, e análise da expressão diferencial de genes (utilizando microarranjos de DNA) combinada com identificação de fatores de transcrição (FT) a ela relacionados. O peso relativo da PV foi significativamente diminuído nos grupos Cas e Cas+E2. A castração promove um padrão de descamação das células epiteliais que deve contribuir para redução do número de células epiteliais. No grupo E2, foi marcante a presença agregados protéicos citoplasmáticos e proliferação das células epiteliais com freqüente estratificação do epitélio. Outro aspecto importante foi o descolamento das células musculares lisas do epitélio quando este apresentava dobras epiteliais. No grupo Cas+E2, foram observados os diferentes aspectos observados em cada grupo individual. A análise bioquímica mostrou redução significativa na fosforilação de 4EBP2 nos grupos E2 e Cas+E2 (tendência similar foi observada no grupo Cas). Há uma grande quantidade de genes que são diferencialmente expressos em relação ao controle e que são comuns aos diferentes tratamentos. Surpreendentemente, não houve redução da expressão de probasina no grupo E2. A análise de ontologias e de termos enriquecidos mostrou a ausência de termos enriquecidos no conjunto de genes exclusivos do grupo Cas+E2, o que se concilia bem com a idéia de que os eventos observados neste grupo são aqueles observados nos dois grupos individuais. Em conclusão, a busca por sítios de ligação de FT na região promotora dos genes mais regulados em cada grupo e a identificação de FT nas rede de dos termos enriquecidos indicou os FT Evi-1, NF-Y, HNF-4, Elk-1, GATA-2, c-Rel, v-Myb e NFkB como candidatos a atuarem na regulação dos eventos associados à regressão prostática / Abstract: Prostate development, physiology and cancer depend on a fine balance between the levels of androgens and estrogens acting via the androgen receptor (AR) and the estrogen receptors (ER'alpha' e ER'beta'), respectively. It is known that the kinetics of apoptosis are different in the rat ventral prostate (VP) of castrated rats (Cas group) and in rats subjected to 17'beta'-estradiol high dose (group E2) or their combination (group Cas+E2), with an evident additive effect in the latter situation (Garcia-Florez et al, 2005). In this work, we investigated elements in common and exclusive to each of these hormonal conditions, employing morphological analysis, components of the AKT/PTEN pathway and analyses of differential gene expression using DNA microarrays combined with a search for transcription factors (TF). The VP weight was significantly reduced in the Cas and Cas+E2 groups. In the E2 group, the remarkable effects were the presence of protein aggregates in the cytoplasm, and cell proliferation and layering of the epithelium. Another important aspect was the detachment of the smooth muscle cells from the epithelium when it showed infolds. In the Cas+E2 group, the different aspects observed in the individual groups were observed, except by less frequent epithelial layering. The biochemical analysis showed a significant reduction in 4EBP phosphorylation in the groups E2 and Cas+E2 (similar tendency was observed in the Cas group). There is a large number of differentially expressed genes as compared to the controls and shared by the different treatments, Surprisingly, there was no reduction in the expression of the probasin gene in the E2 group. The recovered gene onthologies and enrichment terms revealed the absence of enrichment terms among the genes exclusive to the Cas+E2 group, what conciliates with the Idea that the observed changes in this group are identical or at least very similar to those occurring in the individual treatments. In conclusion, the search for TF binding sites in the promoter region of the regulated genes and the identification of TF in the regulatory pathways obtained for the enrichement terms indicated the TF Evi-1, NF-Y, HNF-4, Elk-1, GATA-2, c-Rel, v-Myb and NFkB as candidates to regulate the events associated with prostate regression / Mestrado / Biologia Celular / Mestre em Biologia Celular e Estrutural

The abdominal aorta (AA) in man is a vulnerable artery prone to atherosclerosis as well as aneurysmatic dilation. The underlying aortic composition, mechanical properties as well as the mechanisms responsible for age-related changes and vascular disease are however largely unknown. The aims of this study were 1) to characterize the age- and gender-related changes of the aortic wall components in vivo, using a mechanical model based on ultrasound measurements of pulsatile aortic diameter changes combined with intra-arterial pressure; 2) to validate ultrasound measurements of diameter and intima-media thickness (IMT) of the AA in order to calculate wall stress; 3) to study the stress driven remodeling response of the aortic wall in healthy individuals and the influence of age and gender; and 4) to study wall stress and remodeling of the AA in diabetic patients in order to elucidate the protective influence of diabetes on abdominal aortic aneurysm formation. The stiffness of the isotropic material (mainly elastin) increased in males despite the known decrease in elastin content with age. Further, an exponential increase in stiffness of the anisotropic material (mainly collagen) in males at high physiological pressure was found. This might be due to changed isoforms of collagen and increased glycation with age. Females were less affected than males. The reproducibility of the ultrasound measurements of diameter and IMT in the AA was acceptable (CV; 4% and 11% respectively), making it possible to calculate circumferential aortic wall stress in vivo. The age-related remodeling of the arterial wall led to increased diameter, and compensatory thickening of the wall preventing the circumferential wall stress from increasing in the common carotid artery of males and females, and the AA of females. However, the compensatory increase in wall thickness was defect in the male AA, where stress increased with age. Pulsatile stress influenced the material parameters of the AA, leading to increased stiffness of anisotropic material (mainly collagen), whereas stiffness of isotropic material (mainly elastin) was unaffected. Patients with diabetes mellitus had increased aortic wall thickness than controls, generating less circumferential stress. This coincides with the known reduction of abdominal aortic aneurysms in diabetic patients and may act as a protective factor.

Le remodelage bronchique regroupe des entités physiopaphologiques commel’hypertrophie musculaire lisse dans l’asthme ou l’augmentation d’épaisseur bronchique surl’infiltration de cellules inflammatoires et l’accumulation de fibrose dans la BPCO. Cesremodelages sont corrélés à l’obstruction fonctionnelle et donc à la sévèrité de ces maladies.L’analyse de biopsies bronchique ou pulmonaire permet d’étudier ce phénoméne qui, aprèsune meilleure compréhension, est une cible thérapeutique intérressante. Le premier articleest une revue d’indications de bronchoscopie chez les patients de réanimation. La deuxièmeétude a montré une augmentation des fibrocytes sanguins au cours d’exacerbation sévèrede patient BPCO et une corrélation entre leur taux et le risque de décès du patient. La voiede signalisation du CXCR4 semble impliquée dans ce recrutement. La troisième étudecherche à explorer la localisation et les caractéristiques intra-pulmonaires des fibrocyteschez le patient BPCO à l’état stable. La quatrième étude a montré, in vivo, que le gallopamil,un inhibiteur calcique, pouvait diminuer la taille de muscle lisse bronchique de patientasthmatique sévère en ciblant la biogenèse mitochondriale. Ceci pourrait en faire une armethérapeutique interressante et totalement novatrice. La dernière étude a permis d’isoler unphénotype de patient asthmatique non sévère à « muscle lisse bronchique augmenté » quiprésente un risque accru d’exacerbation et de contrôle non optimal de leur asthme. Lesmitochondries semblent jouer un rôle clé comme dans l’asthme sévère. / Airway remodeling groups pathophysiological entities such as smooth musclehypertrophy in asthma or increase bronchial thickness due to infiltration of inflammatory cellsand fibrosis in COPD. These remodeling is correlated with the functional obstruction andtherefore with the severity of these diseases. The bronchial or lung biopsies analysis allowsto study this phenomenon which, after understanding, is an interesting therapeutic target.The first article is a review of indications of bronchoscopy in critically ill patients. The secondstudy showed an increase in blood fibrocytes during severe exacerbation of COPD patientand a correlation between their rate and the risk of patient death. CXCR4 signaling pathwayseems to be involved in the fibrocyte recruitment. The third study seeks to explore thelocation and characteristics of intra-pulmonary fibrocytes in stable COPD patients. The fourthstudy has shown, in vivo, that gallopamil, a calcium channel blocker, could reduce airwaysmooth muscle size in severe asthmatic patient by targeting mitochondrial biogenesis. Thiscould make it an interesting therapeutic weapon and totally innovative. The last study hasisolated a non-severe asthma phenotype with "increased bronchial smooth muscle," whichpresents an increased risk of exacerbation and a suboptimal control of their asthma. Themitochondria appear to play a key role as in severe asthma.

L’action du système nerveux périphérique sympathique sur le métabolisme osseux, via la sécrétion de neurotransmetteurs, comme le VIP, est clairement établie. Réciproquement, les cellules osseuses expriment des molécules possédant des propriétés chémo-attractives (NGF) ou répulsives (Sémaphorine 3a) suggérant que l’os est capable de contrôler sa propre innervation. Afin de mieux comprendre les relations entre système nerveux et cellules osseuses, notre travail s’est déroulé en deux étapes.Dans un premier temps, nous avons montré que la sympathectomie chimique à la guanéthidine monosulfatée, au niveau du périoste mandibulaire, modulait le ratio OPG/RANKL par l’intermédiaire de la voie cholinergique du système nerveux sympathique. Le traitement par du VIP des rats sympathectomisés a permis de rétablir le potentiel résorbant du système sympathique et ainsi confirmé le rôle prépondérant du VIP dans la modulation du métabolisme osseux au niveau du périoste mandibulaire. Dans un deuxième temps, nous avons évalué les variations d’expression du NGF et de la Séma3a, en fonction de la disponibilité locale en VIP. La sympathectomie, dans le compartiment ostéogène du périoste mandibulaire, a épuisé les réserves en proNGF et en Sema 3a et provoqué une migration de fibres sensorielles immunoréactives au CGRP où physiologiquement elles sont absentes. Dans compartiment non-ostéogène, la sympathectomie a induit une dégranulation des mastocytes et la libération de βNGF (forme mature) et le développement de fibres CGRP-IR.. Le traitement par le VIP10-28, un antagoniste des récepteurs du VIP, a provoqué des effets similaires à la sympathectomie. In vitro, le VIP n’a pas modifié l’expression relative des ARNm codant pour le NGF et la Séma 3a, augmenté celle de RANKL et diminué celle d’OPG. Le VIP10-28 a permis d’augmenter l’expression d’OPG, et de diminuer celle de Séma3a et de CGRP. L’ensemble de ce travail a permis de montrer que le système sympathique cholinergique, via le VIP, module à la fois le rapport OPG/RANKL et l’expression de NGF et de Sema3a au niveau des ostéoblastes et du périoste mandibulaire et renforce l’hypothèse d’une communication bidirectionnelle entre les cellules nerveuses et osseuses. / The action of the sympathetic nervous system on bone metabolism, via the secretion of neurotransmitters such as VIP, is clearly established. Conversely, bone cells express molecules with chemo-attractive properties (NGF) or -repulsive (Semaphorin3a), suggesting that bone can control its own innervation. To better understand the relationship between the nervous system and the bone cells, our work takes place in two stages. As a first step, we have shown that chemical monosulfate guanethidine sympathectomy, modulates in the mandible periosteum the OPG/RANKL ratio through the cholinergic nervous system. VIP treatment of sympathectomized rats restored to controls the resorption potential of the sympathetic system and thus confirmed the leading role of VIP in the modulation of bone metabolism in this bone envelope. In a second step, we evaluated, the variations in NGF and Sema3a expressions, according to the local availability in VIP. Sympathectomy, exhausted proNGF and Sema 3a stores in the osteogenic compartment of the periosteum and caused its invasion by CGRP immunoreactive sensory fibers, where they are physiologically absent. In the non-osteogenic compartment, sympathectomy induced mast cell degranulation and release of βNGF (the mature form) and sprouting of CGRP-IR fibers Treatment with VIP10-28, a VIP receptors antagonist, had effects similar to sympathectomy. In vitro, VIP did not alter the relative expression of mRNA encoding NGF and Sema 3a, increased RANKL and decreased OPG mRNAs. VIP10-28 increased OPG mRNA expression and decreased that of Sema3a and CGRP.In conclusion, this work showed that the cholinergic sympathetic system, via VIP, modulates the OPG/RANKL ratio and NGF and Sema3a expression in periosteal osteoblasts and strengthens the hypothesis of a bi-directional communication between nerve and bone cells.

The use of nanoparticles for disease treatment is an increasingly popular area of research. The potential for multi-functionality allows nanoparticles to be used as transport and delivery vehicles for drugs and as diagnostic aides, among other applications, to address the unmet needs of many disease treatments. One such class of disease is osteoporosis including severe disorders, like Paget’s disease, Osteogenesis Imperfecta and Legg Calve Perthes disease. In this dissertation, we discuss a nanoparticle system consisting of gold nanoparticles surface functionalized with primary amine bisphosphonates, which is a classification of pharmaceuticals that is common in the treatment of osteoporosis. Functionalized nanoparticles allow for greater intracellular concentrations of pharmaceutical, while the properties of the gold nanoparticles provide the ability to track the pharmaceutical and enhance imaging. We have synthesized and characterized bisphosphonate functionalized gold nanoparticles of controlled size of approximately 15 nm, which are suitable for cellular uptake, and functionalized the surface using self-assembly with pamidronate and alendronate. In one major finding of this study, inductively coupled plasma mass spectrometry was used to estimate approximate surface density of the bisphosphonates on the gold nanoparticles. This resulted in concentrations of approximately 0.65 molecules per nm2 (approximately 154 Å2/molecule) for pamidronate functionalized on gold, and approximately 2.6 molecules per nm2 (approximately 39 Å2/molecule) for alendronate functionalized on gold. This allows for more accurate estimates of pharmaceutical concentrations, during in vitro and in vivo studies. Additionally, we investigated the effects of bisphosphonate functionalized gold nanoparticles on the viability and morphology of osteoclast and osteoblast cells in vitro. We found that attaching the bisphosphonates to the surface of the nanoparticles leads to increased apoptotic effects of the bisphosphonates on the osteoclast cells compared to free bisphosphonates. Further, we showed bisphosphonate functionalized gold nanoparticles may have an effect on nuclei morphology that may provide an additional means of modulating bone resorption rather than just through influencing viability. Further we showed that it may be possible to target concentrations that are safe for osteoblasts, which is critical in determining potential treatment concentrations. These viability results bring to light a number of potential considerations into the optimization of potential treatments, such as dosing concentrations. Finally, detailed results are given on effects of bisphosphonate functionalized gold nanoparticles on important behavior and activity of osteoclast and osteoblast cells in vitro. We showed that while using concentrations below the toxicity threshold, some of the normal activity of the cells could be maintained. RANKL and ALP expression in osteoblasts were maintained when removing viability as a variable. Additionally, bone nodule formation was also maintained for osteoblasts and co-cultured in vitro systems. Finally, we showed that the introduction of bone in the in vitro studies adds a new degree of consideration as to the interaction of the bisphosphonates with the hydroxyapatite surface. This strong interaction with bone is an important consideration in further developing potential treatments for osteoporotic disease. This dissertation provides insights into the use of bisphosphonate functionalized gold nanoparticles as a potential treatment and means of study for bone remodeling disorders.

Parmi les processus fondamentaux qui ont lieu pendant le développement d'un organisme, la morphogenèse est un des plus complexes. De nombreuses études expérimentales ont contribué à mieux comprendre les mécanismes morphogénétiques dans les organismes vivants. Cependant peu de modèles mathématiques ont été proposés afin d'étudier la morphogenèse dans les tissus vivants. Dans ce contexte, la thèse se propose de développer de nouveaux modèles mathématiques pour étudier les changements de forme dans les tissus mous tubulaires. L'approche adoptée est macroscopique où le tissu biologique est considéré comme un milieu continu déformable. L'hypothèse principale sur laquelle se basent les modèles proposés est la suivante: pendant les processus de croissance et remodelage, des contraintes résiduelles peuvent s'accumuler dans le tissu et, une fois une valeur critique dépassée, le mener à un changement morphologique sous la forme d'une instabilité élastique. Pour cela, les modèles développés intègrent les théories modernes de croissance et remodelage, dans le cadre de la thermomécanique des systèmes ouverts. Ensuite, l'analyse de stabilité linéaire permet de calculer les seuils et modes de l'instabilité élastique en utilisant la méthode des déformations incrémentales superposées aux déformations finies. L'ensemble de ces techniques (théorie morpho-élastique) est utilisé dans cette thèse afin de modéliser deux différents processus morphogénétiques ayant lieu dans les tissus mous tubulaires : la formation d'une variété des formes dans le système gastro-intestinal et le flambage hélicoïdal dans les organes tubulaires avec précontraintes. / Among the fundamental processes involved in the development of an organism, morphogenesis is one of the most complex. During the past centuries, an amount of experimental studies have improved our actual knowledge of the mechanisms which drive many morphogenetic processes in living organisms. Only recently, experiments have been complemented with mathematical modeling as a tool for proving novel insights on morphogenesis in soft tissues. In this context, this thesis aims at developing new mathematical models for the formation of patterns and forms in soft tubular organs. A macroscopic approach is adopted, where the tissue is considered as a continuum body undergoing growth and remodeling. The main idea behind the proposed models is that during growth and remodeling, residual stresses can arise and once they exceed a critical value, an elastic instability can occur in the tissue and lead to a morphological change. Therefore, the morphoelastic models are developed integrating the modern theories of growth and remodeling within the framework of the thermo-mechanics of open systems. The occurrence of the elastic instability is investigated using the method of incremental deformations superposed on finite deformations. The critical thresholds for the onset of the instability are determined together with the modes of the associated instability pattern. The morphoelastic theory is applied to the modeling of different morphogenetic processes occurring in soft tubular organs and gives useful insights in two interesting problems: the formation of the wide range of patterns in the gastro-intestinal system and the occurrence of torsional instabilities in pre-stressed tubular organs.

Nous avons cherché à discerner les rôles respectifs de l’âge et de l’hypertension sur l’anatomie et la fonction des grosses et des petites artères en utilisant de nouvelles techniques non invasive. Au niveau macrovasculaire : En utilisant l’IRM aortique nous avons montré le rôle proéminent de l’âge et observé l’influence parallèle et combinée de l’âge et de l’hypertension sur la diminution de la rigidité. En utilisant la tonométrie de l’artère radiale nous avons montré des liens entre anatomie, fonction et athérosclérose dans une large population suggérant de nouvelles hypothèses pour relier fonction artérielle et risque cardiovasculaire Au niveau microvasculaire rétinien: Nous avons validé une nouvelle technologie d’imagerie de haute résolution: l’optique adaptative. Nous avons confirmé dans une large cohorte d’hypertendus la présence d’un remodelage eutrophique. Nous avons observé des variations à court terme de la lumière sans modification de la surface sectionnelle en cas de diminution de pression. Conjugués à la corrélation que nous avons établie entre le remodelage rétinien et les résistances périphériques totales, nos résultats plaident pour une part importante de remodelage fonctionnel. Avec le vieillissement, nous avons décrit un épaississement pariétal avec augmentation de la surface sectionnelle sans modification de la lumière. Nous avons confirmé la présence d’un remodelage hypertrophique chez les patients diabétiques. En conclusion : Nous avons validé et montré l’apport de nouvelles techniques d’imagerie artérielle non invasives sur de larges populations pour étudier le remodelage artériel dû à l’âge et aux facteurs de risque cardiovasculaires. / We’ve tried to decipher the respective roles of age and hypertension on anatomy and function of large and small arteries using new non invasive imaging techniques. Large arteries: Using Aortic MRI, we’ve shown the proeminent role of ageing on arterial stiffening as well as the parallel and combined influence of age and hypertension . Using radial artery tonometry, we’ve shown links between anatomy, function and atherosclerosis in a large population. This generates new hypothesis to link arterial function and cardiovascular risk. Retinal microcirculation: We’ve validated a new high resolution imaging technique : adaptive optics. We’ve confirmed in a large cohort of hypertensives the existence of an inward eutrophic remodeling. We’ve observed Blood pressure drop related short term vasodilatation without changes in wall cross section. Together with the relationship we also established between arteriolar remodeling and total peripheral resistances, our results advocate for an important part of functional remodeling in hypertension. With ageing, we’ve observed a parietal thickening with wall cross section increase without lumen modifications. We’ve also confirmed the presence of an hypertrophic remodeling in diabetic patients. In conclusion: We have validated and confirmed the added value of new arterial non invasive imaging techniques on large populations to study cardiovascular risk factors induced arterial remodeling