Submitted by Luciana Ferreira (lucgeral@gmail.com) on 2014-10-29T14:34:23Z No. of bitstreams: 2 Dissertação - Iury Valentim Jorge Zoghaib - 2013.pdf: 1607539 bytes, checksum: d92b3313edee2773935cb667be5b908d (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Approved for entry into archive by Luciana Ferreira (lucgeral@gmail.com) on 2014-10-30T10:48:45Z (GMT) No. of bitstreams: 2 Dissertação - Iury Valentim Jorge Zoghaib - 2013.pdf: 1607539 bytes, checksum: d92b3313edee2773935cb667be5b908d (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) / Made available in DSpace on 2014-10-30T10:48:45Z (GMT). No. of bitstreams: 2 Dissertação - Iury Valentim Jorge Zoghaib - 2013.pdf: 1607539 bytes, checksum: d92b3313edee2773935cb667be5b908d (MD5) license_rdf: 23148 bytes, checksum: 9da0b6dfac957114c6a7714714b86306 (MD5) Previous issue date: 2013-10-18 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / The LQFM030 was obtained by molecular simplification of nutlins (inhibitors of p53-MDM2 interaction) and, having demonstrated excellent antineoplastic activity in vitro (cytotoxicity against K-562 cell line with IC50 = 23 M) and in vivo against Ehrlich ascitic tumor with increased survival in treated animals developed the pharmacokinetic study of p.o. in rats. It was used for LC-MS/MS analytical method (Applied Biosystems MDS Sciex API 3200) previously validated together. O LQFM030 was administered to 3 rats (mean weight 186g) in predetermined dose of 100 mg/kg by gavage. After administration, samples were collected from 1.0 mL of blood by cannulation of the left jugular vein with heparinized syringe, the intervals 0-8 h. The blood samples were identified and centrifuged to obtain plasma which was frozen at -20°C until analysis. The kinetic parameters were calculated in the software WinNonlin 5.0 (Pharsight™). Results (mean ± SD) half-life (t1/2) 3.61 ± 0.68 h, total clearance (CLT) 36.49 ± 2.23 mL/min/kg, volume of distribution (Vd) 11,40 ± 1.58 L/kg. The LQFM030 had low value of t1/2, Vd high and high value of CLT. These values allow us to understand that the prototype study demonstrated a good safety profile of tissue distribution and/or has been extensively eliminated. When compared with the kinetic parameters obtained in other studies, it was observed difference in results is justified by the high interspecies variability, mainly in basal metabolic rate and body weight, once the route of administration, orally and intraperitoneally, are kinetically similar. / O LQFM030 foi obtido por simplificação molecular dos nutlins (inibidores da interação MDM2-p53) e, tendo demonstrado excelente atividade antineoplásica in vitro (citotoxicidade contra a linhagem celular K-562 com IC50 = 23 M) e in vivo contra tumor ascítico de Ehrlich, com aumento de sobrevida em animais tratados, desenvolveu-se o estudo do perfil farmacocinético, p.o., em ratos. Empregou-se método analítico em LC-MS/MS (Applied Biosystems MDS Sciex API 3200) previamente validado em colaboração. O LQFM030 foi administrado a 3 ratos (peso médio de 186g), em dose preestabelecida de 100 mg/kg, por gavagem. Após a administração, foram coletadas amostras de 1,0 mL de sangue, por canulação da veia jugular esquerda, com seringa heparinizada, nos intervalos de tempo de 0 a 8 h. As amostras sanguíneas foram identificadas e centrifugadas para obtenção do plasma, que foi congelado a -20ºC até o momento da análise. Os parâmetros cinéticos foram calculados no software Winnonlin 5.0 (Pharsight™). Resultados (média ± DP): meia-vida (t1/2) 3,61 ± 0,68 h; clearance total (CLT) 36,49 ± 2,23 mL/min/kg; volume de distribuição (Vd) 11,40 ± 1,58 L/kg. O LQFM030 apresentou baixo valor de t1/2, elevado Vd e elevado valor de CLT. Estes valores permitem entender que o protótipo estudado demonstrou um bom perfil de distribuição tecidual e/ou foi extensivamente eliminado. Quando comparados com parâmetros cinéticos obtidos em outros estudos, observou-se diferença nos resultados, justificada pela elevada variabilidade interespécies, principalmente na taxa de metabolismo basal e peso corporal, uma vez que as vias de administração, oral e intraperitoneal, são cineticamente semelhantes.

L’imagerie préclinique est devenue une ressource incontournable pour l’évaluation de paramètres physiopathologiques, pour le suivi du développement tumoral ainsi que pour le développement de thérapies anticancéreuses. Les évolutions technologiques apparues ces dernières années ont conduit au développement de nouvelles modalités d’imagerie ayant un fort potentiel de translation vers la clinique. Ce manuscrit présente diverses approches par imagerie échographique, photoacoustique et de fluorescence dans le proche infrarouge pour le suivi de la pathologie cancéreuse. Dans un premier temps, nous nous sommes intéressés à la caractérisation de l’hypoxie et son suivi au cours du temps dans différents modèles de cancers humains. Différentes stratégies d’imagerie multimodale ont ensuite été mises en oeuvre pour évaluer l’efficacité d’une nouvelle prodrogue thérapeutique permettant la libération d’une molécule active dans le proche environnement tumoral sur des modèles humains de tumeurs pancréatiques, mammaires, pulmonaires. Enfin, dans un contexte de recherche translationnelle, nous avons exploré le potentiel de l’imagerie photoacoustique et de la fluorescence infrarouge pour la mise en évidence de l’invasion ganglionnaire tumorale en mettant en oeuvre des modèles de ganglions sentinelles minimalement envahis. Au cours de ce travail, nous avons montré l’intérêt du suivi de l’hypoxie tumorale en onco-pharmacologie et mis en évidence le fort potentiel de l’imagerie PA pour les approches translationnelles en oncologie. La principale limitation correspond à la profondeur relativement faible des régions explorables, mais ce point suscite actuellement de nombreux développements technologiques. Les études de faisabilité réalisées ainsi que la validation de protocoles de preuves de concept permettront l’exploitation en routine de ces nouvelles modalités d’imagerie. / Preclinical imaging has become an unavoidable step for pathophysiological parameters assessments, for the follow up of tumor growth and for the anticancer therapies development. Technological improvements have emerged in recent years, allowing the emergence of new imaging modalities with a high potential for translation into clinical practice. This manuscript presents several approaches by ultrasound imaging, photoacoustics and near infrared fluorescence in order to monitor the cancer pathology. Initially, we focused on the characterization of hypoxia and its longitudinal assessment in various preclinical models of human cancers. Various multimodal imaging strategies were implemented to assess the efficacy of a new therapeutic prodrug allowing the release of an active molecule in the tumor microenvironment on human models of pancreatic, breast and lung tumors. Finally, in a context of translational research, we explored the potential of photoacoustic and near infrared fluorescence imaging to highlight the lymph node invasion by cancer cells implementing minimally invaded sentinel lymph node models. In this work, we have shown the interest in monitoring the tumor hypoxia in onco-pharmacology and highlighted the high potential of photoacoustic imaging for oncology translational approaches. The main limitation is the relatively shallow depth of regions that we can explore, but this point is currently subject to many technological developments. Feasibility studies performed and validation of proof of concept protocols will enable routine exploitation of these new imaging modalities.

The progress of cell therapy can be greatly facilitated by using suitable experimental models. It is essential to verify the clinical usefulness of new healing procedures obtained in studies on laboratory animals by using a large animal model. One of suitable models well acceptable in medical community is undoubtedly the miniature pig, which resembles humans in terms of physiology and body proportions. This PhD thesis presents the summary of our experimental studies relating to possible exploitation of mesenchymal and neural stem cells in the healing of locomotive apparatus and neural tissue disorders in humans or animals. The first part of the thesis briefly describes the current issue of cell therapy and animal models, mesenchymal cells and/or their combination with new types of scaffolds, neurogenesis, neural stem cells and their potential application in therapy of spinal cord injury. The second part is focused on the goals and methodology, the individual publications being listed in the third part. Our experiments with iatrogenic physeal defect in rabbits, which served as a model of the occurrence of valgous deformation in the clinical practice, showed the positive preventive and therapeutical effects of a new type of scaffolds seeded with allogeneic mesenchymal stem cells in animals without...

<div>Osteoporosis is the most common bone disease in older adults and is characterized by low bone mass and increased fragility. Women are at a higher risk for osteoporosis because of the rapid loss of bone during menopause. The decline of estrogen is accompanied by an increased bone resorption and a decreased bone formation which results in negative bone balance. Due to adverse effects on the uterus, breast and cardiovascular system, hormone replacement therapy has been discouraged. Nutritional strategies for osteoporosis prevention are being sought. It has been suggested that (poly)phenol-rich fruits may have bone protective effects. Blueberries are one of the richest sources of (poly)phenols, thus the aim of this dissertation was to determine whether a blueberry-enriched diet could aid in bone loss prevention in the ovariectomized rat model.</div><div><br></div><div>There are hundreds of blueberry varieties which differ in (poly)phenol profiles and content. Five blueberry varieties (Ira, Montgomery, SHF2B1-21:3, Onslow and Wild Blueberry) were chosen to assess the bioavailability of its individual (poly)phenols. Bioavailability of individual phenolic metabolites was determined through a pharmacokinetic study in ovariectomized rats. The results showed that Montgomery blueberry had significantly higher bioavailability of malvidin, cyanidin and myricetin metabolites, while Ira had significantly higher bioavailability of quercetin metabolites, thus suggesting that the absorption of blueberry polyphenols and their potential to reach target tissues differed between blueberry varieties.</div><div><br></div><div>It is important to assess what is the most appropriate dose of blueberry necessary to exert beneficial effects on bone. To determine the most adequate dose of wild blueberry to prevent bone loss in ovariectomized rats, a randomized crossover study was carried out to assess the effects of four different blueberry doses on net bone calcium retention over a 10-day treatment period. The results showed that the only dose to significantly increase net bone calcium retention by 25.6% (p = 0.0426) was the 5% blueberry diet (% w/w), while the higher doses of 10% and 15% had no effect on net bone calcium retention. This informed the last study where Montgomery blueberry and wild blueberry at a 5% dose (% w/w) were chosen to investigate the effects of an 8-week chronic feeding study on calcium metabolism, kinetics, bone microarchitecture and strength and polyphenol metabolism and distribution. A chronic consumption of the wild blueberry resulted in a trend towards minimal trabecular bone loss protection in comparison to the control diet (p=0.08). Kinetic modeling of calcium showed that the Montgomery blueberry had anabolic effects on bone through significantly increasing calcium absorption and bone deposition. The phenolic metabolism differed among blueberry varieties due to each berry’s polyphenol content and profiles and a chronic consumption of blueberry resulted in significant changes in absorption and metabolism of polyphenols. The bone marrow was investigated to determine whether there was any accumulation of phenolic acids in the tissue. Hippuric acid accumulation was significantly higher with the Montgomery blueberry treatments in comparison to control diet. Interestingly, hippuric acid content in the bone marrow was significantly and positively correlated with bone deposition calculated from kinetic modeling. Although no differences were observed on bone mineral density, strength, and microarchitecture, previous studies with a duration of 12-14 weeks have shown significant protection of a blueberry-enriched diet on bone mineral density. Because our study showed a trend for increased trabecular bone (p = 0.08) with the blueberry treatments, we conclude that an 8-week treatment was insufficient time to detect significant differences between the control and blueberry treatments. Since previous researchers before us have reported significant attenuation to bone loss immediately after OVX, it is possible that blueberry that in our study, blueberry was unable to rescue bone once lost after ovariectomy.<br></div><div><br></div><div>A blueberry-enriched diet resulted in a minimal protection to bone after stabilized to OVX, but showed significant increases in calcium absorption and bone turnover in ovariectomized rats. Colonic metabolite profiles from the chronic consumption of blueberry significantly changed over time, thus providing an insight into the effects of blueberry consumption on polyphenol metabolism.<br></div><div><br></div>

Anxiety disorders are the most common class of mental disorders in the United States, and they both promote and exacerbate disorders of substance abuse. Mounting evidence of sex differences in the relationship between anxiety disorders and alcoholism supports the potential existence of an anxiety-dependent vulnerability to alcohol abuse in women compared with men. One potential point of overlap in the physiological systems involved in anxiety response and reward processing is the amygdala. Here, a model of chronic stress in rodents was employed to probe changes in the electrophysiological and biochemical properties of the amygdala at a post-stress baseline and during a post-stress first exposure to alcohol. Electrophysiological data revealed that neurons in the centromedial amygdala were more responsive to stimulation in the basolateral amygdala in females compared with males, but a history of chronic stress altered the female response to match that of males with or without a history of chronic stress. Protein analysis of postsynaptic glutamatergic receptor expression and phosphorylation in the amygdala did not indicate any differences based on sex or exposure to stress or alcohol. These data demonstrate a sex difference in stress-induced alterations in amygdala circuitry and indicate a potential role for this circuitry in the comorbidity of anxiety disorders and alcoholism.

京都大学 / 新制・課程博士 / 博士(医学) / 甲第23601号 / 医博第4788号 / 新制||医||1055(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 羽賀 博典, 教授 戸井 雅和, 教授 伊藤 貴浩 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

<p>Preclinical rodent models were used to investigate the neural signatures of strategy change during the delay discounting decision making task. Neural signatures were assessed using advanced statistical techniques (de-mixed principal component analysis). </p>

This paper presents an intraoperative MRI-guided, patient-mounted robotic system for shoulder arthrography procedures in pediatric patients. The robot is designed to be compact and lightweight and is constructed with nonmagnetic materials for MRI safety. Our goal is to transform the current two-step arthrography procedure (CT/x-ray-guided needle insertion followed by diagnostic MRI) into a streamlined single-step ionizing radiation-free procedure under MRI guidance. The MR-conditional robot was evaluated in a Thiel embalmed cadaver study and healthy volunteer studies. The robot was attached to the shoulder using straps and ten locations in the shoulder joint space were selected as targets. For the first target, contrast agent (saline) was injected to complete the clinical workflow. After each targeting attempt, a confirmation scan was acquired to analyze the needle placement accuracy. During the volunteer studies, a more comfortable and ergonomic shoulder brace was used, and the complete clinical workflow was followed to measure the total procedure time. In the cadaver study, the needle was successfully placed in the shoulder joint space in all the targeting attempts with translational and rotational accuracy of 2.07 ± 1.22mm and 1.46 ± 1.06 degrees, respectively. The total time for the entire procedure was 94 min and the average time for each targeting attempt was 20 min in the cadaver study, while the average time for the entire workflow for the volunteer studies was 36 min. No image quality degradation due to the presence of the robot was detected. This Thiel-embalmed cadaver study along with the clinical workflow studies on human volunteers demonstrated the feasibility of using an MR-conditional, patient-mounted robotic system for MRI-guided shoulder arthrography procedure. Future work will be focused on moving the technology to clinical practice.

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Yes / Non-invasive methods to monitor tumour growth are an important goal in cancer drug development. Thermographic imaging systems offer potential in this area, since a change in temperature is known to be induced due to changes within the tumour microenvironment. This study demonstrates that this imaging modality can be applied to a broad range of tumour xenografts and also, for the first time, the methodology’s suitability to assess anti-cancer agent efficacy. Mice bearing subcutaneously implanted H460 lung cancer xenografts were treated with a novel vascular disrupting agent, ICT-2552, and the cytotoxin doxorubicin. The effects on tumour temperature were assessed using thermographic imaging over the first 6 hours post-administration and subsequently a further 7 days. For ICT-2552 a significant initial temperature drop was observed, whilst for both agents a significant temperature drop was seen compared to controls over the longer time period. Thus thermographic imaging can detect functional differences (manifesting as temperature reductions) in the tumour response to these anti-cancer agents compared to controls. Importantly, these effects can be detected in the first few hours following treatment and therefore the tumour is observable non-invasively. As discussed, this technique will have considerable 3Rs benefits in terms of reduction and refinement of animal use. / University of Bradford