Introdução: A simpatectomia torácica é o único tratamento, definitivo e eficaz, para a hiperidrose primária. A ramicotomia é um procedimento cirúrgico tão eficaz, mais conservador e com menos efeitos adversos que a simpatectomia convencional, contudo foi abandonada pela alta taxa de recidiva, atribuída, até então, à secção incompleta dos ramos comunicantes, ao desenvolvimento de outras vias de condução para o estímulo central e à regeneração neural. A avaliação histológica dos ramos comunicantes simpáticos após a ramicotomia, pode ajudar a entender o processo de recidiva dos sintomas da hiperidrose e, dessa forma contribuir para o desenvolvimento de estratégias para evitá-la. MATERIAL E MÉTODOS: 28 suínos foram submetidos à ramicotomia por videotoracospia e divididos randomicamente em 5 grupos, sacrificados com 15, 45, 90, 135 e 180 dias de pós-operatório (DPO). Os segmentos operados foram removidos cirurgicamente e submetidos à avaliação macroscópica da regeneração assim como análise histológica dos ramos comunicantes brancos e cinzentos para quantificação da reação inflamatória, deposição de fibras de colágeno grossas e finas, fibras reticulares e células de Schwann por imuno-histoquímica. Os dados foram comparados ao grupo controle, composto por segmentos intactos, não operados. RESULTADOS: Não houve regeneração macroscópica no grupo de 15 DPO sendo presente em 41,6% dos casos no grupo 180 DPO (p < 0,05). A reação inflamatória foi determinante no processo de degeneração Walleriana, com presença importante das células de Schwann nos ramos pré-ganglionares (p < 0,05), as células de Schwann apresentaram evolução semelhante nos dois ramos a partir do grupo de 45DPO, mantendo-se em menor número nos ramos cinzentos. As fibras de colágeno foram cruciais na cicatrização e as fibras reticulares importantes na regeneração neural, com correlação negativa entre elas (r = - 0,414; p < 0,01). A deposição de fibras de colágeno foi maior nos ramos cinzentos, apresentando pico de deposição no grupo 135 DPO e declínio importante no grupo 180 DPO (p < 0,05). CONCLUSÕES: A ramicotomia permite a secção completa de todos os ramos comunicantes do gânglio simpático. A taxa de regeneração histológica deve ser maior que a taxa de recidiva dos sintomas no humano, devido a regenerações não funcionais. O processo regenerativo é similar nos ramos brancos e cinzentos, com tendências menores para os últimos. A regeneração dos ramos comunicantes deve ser um dos principais fatores de recidiva da hiperidrose após a ramicotomia / INTRODUCTION: Thoracic sympathectomy is the only definitive and efficient treatment for primary hyperhidrosis. The ramicotomy is a surgical procedure that is as efficient as conventional sympathectomy but more conservative, having less adverse effects then conventional sympathectomy. This procedure was abandoned on account of the high recurrence rate, attributed to the incomplete section of the rami communicantes and to the development of new pathways of conduction to the central stimuli. MATHERIAL AND METHODS: Twenty-eight swine underwent bilateral videothoracoscopic ramicotomy and were randomly divided into 5 groups. The animals were sacrificed at 15, 45, 90, 135 and 180 days post-operative POD. The segments were removed and evaluated for macroscopic regeneration and histological analysis of the white and gray rami communicantes analyzing the inflammatory reaction, deposition of thin and thick collagen fibers, reticular fibers and Schwann cells. The data was compared to intact segments of sympathetic trunk as a positive control. RESULTS: There was neither macroscopic nor microscopic regeneration at the 15 POD group. The remaining groups had an average of 41,6% of regeneration, more significant at the 180 POD group (p<0.05). The inflammatory reaction was crucial in the process of Wallerian degeneration, with an important participation of the Schwann cells in the pre-ganglionic rami (p<0.05). The Schwann cells presented a similar evolution in both rami beginning at the 45 POD group, with a smaller count in the gray rami. The collagen fibers were significant in the cicatrization and the reticular fibers were important in neural regeneration, with a meaningful negative correlation between them (r = - 0,414; p < 0,01). The rate of deposition of the collagen fibers was greater in the white rami when compared to the gray rami in the first trimester and less important in the second trimester (p<0.05). CONCLUSIONS: Ramicotomy allows complete section of all rami communicantes of the sympathetic ganglia. The histological regeneration might be greater than the recurrence rates of clinical symptoms as seen in the human being due to non-functional regenerations. The restoration process is similar in both white and gray rami, with smaller tendencies in the last one. The regeneration of the could be one of the main factors for recurrence of hyperhidrosis following ramicotomy

Uma abordagem mais moderna sobre o binômio estresse térmico-degeneração foi realizada, visando monitorar a integridade de acrossomo, mitocôndrias e cromatina. Raros são os trabalhos sobre os efeitos da somatotropina bovina exógena (bST) sobre a qualidade seminal de touros que tenham passado por períodos de estresse térmico e conseqüente degeneração testicular. Foram objetivos do trabalho: 1) Comparar as características seminais e espermáticas de touros normais e touros submetidos a estresse térmico testicular (tratados ou não com bST); 2) Avaliar a integridade da membrana plasmática e acrossomal, função mitocondrial e defeitos cromossômicos de espermatozóides de touros normais e touros submetidos a estresse térmico testicular (tratados ou não com bST). Para tanto, o experimento foi dividido em 4 fases: Fase Pré-Insulação: período do dia 1 ao dia 35 (dia 1 = dia do início do experimento), Fase Pré-Tratamento: período do dia 36 ao dia 63 (colocação da bolsa insuladora no dia 36 por 96 horas), Fase Tratamento: período do dia 64 ao dia 119 (aplicações de bST), Fase Pós-tratamento: período do dia 120 ao dia 154. O sêmen de dezesseis touros Simental (Bos taurus taurus) foi coletado semanalmente ao longo de 22 semanas (154 dias). Os touros foram divididos em quatro grupos: o Grupo CONT foi o controle, não foi submetido a qualquer tratamento; o Grupo INSUL sofreu insulação testicular; o Grupo bST recebeu aplicações de somatotropina bovina (Lactotropin® 1 mg/kg PV) a cada quatorze dias a partir do dia 64; o Grupo IbST sofreu insulação e recebeu aplicações de somatotropina. Os animais foram avaliados quanto às características físicas e seminais. O estresse térmico testicular influenciou negativamente o perímetro escrotal, a motilidade espermática, o vigor, o movimento de massa, mas aumentou o total de defeitos espermáticos. A somatotropina bovina influenciou a quantidade de cabeças piriformes nos ejaculados. Os defeitos de cromatina foram influenciados pelo estresse térmico testicular. A população de espermatozóides com membrana plasmática íntegra, acrossomo intacto e potencial mitocondrial foi reduzida em função do estresse térmico testicular / Modern approach concerning binomial thermal stress-testicular degeneration was done in order to study chromatin, acrosomal and mitochondrial integrity. Furthermore, there are few relates about effects of exogenous bovine somatotropine (bST) on seminal quality of bulls which were submitted to thermal stress and consequent testicular degeneration. The objectives of this study were: 1) Comparing seminal and spermatic features of bulls which were submitted to testicular thermal stress (treated or not with bST); 2) Evaluating of the plasmatic and acrosomal membrane integrity, mitochondrial function, and chromosomal defects from the spermatozoa of these bulls. The experiment was divided in 4 phases. Pre-insulation phase: from day 1 to 35 (day 1 = first day of experiment), Pre-treatment phase: from day 36 to 63, Treatment phase: from day 64 to 119 (treatment with bST) and Post-treatment phase: from day 120 to 154. Semen of sixteen Simmental bulls (Bos taurus taurus) was collected once a week, during 22 weeks (154 days). Bulls were split in four groups: Group CONT was the control, and it was not treated or insulated; Group INSUL was insulated; Group bST was treated with bovine somatotropine (Lactotropin® 1.2 mg/kg BW) each 14 days since day 64. Group IbST was insulated and treated with bST. Physical and seminal features of bulls were evaluated. Thermal stress decreased scrotal circumference, sperm motility, vigour and gross motility, and increased total sperm defects. The bST increased pyriform heads in ejaculated. Testicular thermal stress increased chromatin defects. Sperm population with intact plasmatic membranes, intact acrosome and mitochondrial function were reduced by testicular thermal stress

Nesta tese explora-se um conjunto de requisitos especiais de concepção e desenvolvimento de software que asseguram um alto grau de flexibilidade e eficiência para o controle de sistemas produtivos. Desenvolve-se assim, um método que inclui a técnica de degeneração (redução gradual do nível de serviços de um sistema) no projeto do software de controle de sistemas produtivos. Apresentam-se inicialmente os conceitos fundamentais considerados no projeto do software de sistemas de controle como: sistemas produtivos, sistemas de controle, sistemas distribuídos, arquitetura em n-camadas (“middleware") e sistemas multi-agentes. A seguir, introduz-se a aplicação de requisitos padrões para o desenvolvimento do software de controle com qualidade e com a característica de orientação a objetos. O trabalho apresenta também exemplos específicos relacionados com o controle em Edifícios Inteligentes adotados como estudo de casos, que ilustram a aplicação do método desenvolvido. Os respectivos artefatos resultantes da aplicação de cada etapa do método também são descritos e comprovam o potencial desta abordagem. / This thesis explores a set of special requirements for software development that assure high degree of flexibility and efficiency for control of productive systems. Thus, it investigates a method that includes the degeneration technique (i.e., a gradual reduction of the service level of a system) in the development of control software for productive systems. The text presents initially the basic concepts considered in the development of control software such as productive systems, distributed control systems, middleware architecture and multi-agent systems. Following, it introduces the application of standard requests for the development of control software with quality and object orientation features. The work also presents specific examples related with the control in Intelligent Buildings which have been adopted as case study and that illustrate the application of the proposed method. The artifacts generated from the application of each step of the method are also described and confirm the potential of the proposed approach.

Diabetic retinopathy is an ischemic retinal neovascular disease causing vision loss among adults. The studies presented involve the design and testing of a gene therapy vector to inhibit retinal revascularization, similar to that found in diabetic retinopathy. Gene therapy has proven to be an effective method to introduce therapeutic proteins to treat retinal diseases. Targeting a specific cell type and expression of therapeutic proteins according to the tissue microenvironment should have an advantage over traditional gene therapy by avoiding unwanted transgene expression. Hypoxia plays a significant role in the pathophysiology of many retinal ischemic diseases. Retinal Mèuller cells provide structural and functional support to retinal neurons, as well as playing a significant role in retinal neovascularization. Targeting Mèuller cells may be an effective strategy to prevent retinal neovascularization under pathological conditions. ... The hypoxia regulated, glial specific vector successfully reduced the abnormal neovascularization in the periphery by 93% and reduced the central vasobliterated area by 90%. A substantial amount of exogenous endostatin was produced in the retinas of P17 OIR mice. A significant increase in human endostatin protein and reduced vascular endothelial growth factor (VEGF) were identified by Western blot and ELISA, respectively. These findings suggest hypoxia-regulated, glial cell-specific scAAV mediated gene expression may be useful to prevent blindness found in devastating retinal diseases involving neovascularization. / by Manas Ranjan Biswal. / Thesis (Ph.D.)--Florida Atlantic University, 2012. / Includes bibliography. / Mode of access: World Wide Web. / System requirements: Adobe Reader.

INTRODUÇÃO: O aumento da expectativa de vida tem contribuído para o crescimento da população idosa em todo o mundo. O envelhecimento é uma condição de risco para o desenvolvimento de doenças, entre as quais as demências, que responderão nas próximas décadas por um número significativo de idosos com comprometimento cognitivo, comportamental e funcional. A ampliação de cuidados a esses pacientes está associada ao aumento de sua expectativa de vida; por isso é crescente o número de indivíduos dementes em estágios avançados. Os quadros demenciais comprometem gradualmente o comportamento e a cognição, e observa-se uma progressiva deterioração também na comunicação. As características da linguagem em fases mais avançadas têm sido pouco detalhadas nas demências. Daí a necessidade de se disponibilizar instrumento, em língua portuguesa brasileira, para avaliação da linguagem na demência em fases moderada e grave, o FLCI (Inventário de Comunicação Lingüístico Funcional). O FLCI gera dados para auxiliar o diagnóstico, acompanhamento e evolução da doença e, além disso, para orientar familiares e cuidadores. O FLCI foi aplicado a população com doença de Alzheimer (DA). Desconhece-se sua aplicabilidade em outros quadros demenciais como doença de Alzheimer associada a demência vascular (DA+DV), demências do espectro lobar frontotemporal (DLFT) em fases avançadas. OBJETIVOS: 1.realizar a tradução e adaptação transcultural do FLCI para uso na população brasileira; 2.comparar o desempenho de pacientes com DA, DA+DV e DLFT em fase moderada e grave e 3.correlacionar as habilidades lingüístico-cognitivas com a funcionalidade na vida cotidiana. MÉTODOS: A amostra foi composta por 57 sujeitos, sendo 24 com DA, 24 com DA+DV e 09 com DLFT, com idade a partir de 60 anos e CDR (Clinical Dementia Rating) em níveis 2 e 3, moderado e grave respectivamente. Foi realizada a tradução e adaptação transcultural do FLCI e verificada a consistência interna, confiabilidade inter e intra-examinadores, assim como a validade de critério do instrumento pela correlação com o Mini-Exame do Estado Mental (MEEM). O desempenho lingüístico-cognitivo dos sujeitos agrupados segundo diagnóstico foi analisado por meio da aplicação do FLCI, para comparação de médias de desempenho nos diferentes sub-testes. Foi possível correlacionar os aspectos cognitivos com a funcionalidade por meio da escala FAST (Functional Assessment Staging) e analisar a correlação desta com o FLCI. Verificou-se também o efeito da idade e da escolaridade no desempenho comunicativo dos pacientes. RESULTADOS: As análises estatísticas indicaram que o FLCI apresenta consistência interna satisfatória (alfa de Cronbach=0,890), ótima confiabilidade intra e interexaminador (coeficiente de correlação interclasse - ICC=0,999 e 0,100 respectivamente) e ótima validade de critério ao ser correlacionado com o MEEM. Todos os sub-testes que compõem o FLCI apresentaram diferenças significativas para a amostra total classificada de acordo com a gravidade da demência após ser correlacionada a pontuação total do teste com a escala FAST. De acordo com a classificação nosológica da demência, somente um sub-teste do FLCI apresentou diferenças estatisticamente significativas: \"compreensão de sinais e emparelhamento objeto-figura\". As variáveis idade e a escolaridade não apresentaram influência sobre o desempenho comunicativo da amostra. Comparando-se o perfil de desempenho, a partir da pontuação média em cada sub-teste do FLCI, observou-se melhor desempenho na maioria dos sub-testes para o grupo DLFT, em seguida o grupo DA e por último o grupo de DA+DV. A partir da comparação da pontuação total do FLCI com a FAST modificada, foi possível correlacionar a funcionalidade com as habilidades de comunicação. Verificou-se correlação significativa nas análises entre a escala FAST e a pontuação total do FLCI e escala FAST entre os itens dicotômicos (pontuados como sim/não) e itens pontuáveis (pontuação aberta) que compõem o FLCI. CONCLUSÃO: O FLCI, versão em português, é instrumento válido e confiável para avaliação de pacientes com demência avançada, útil para identificar as habilidades de comunicação de dementes em fases moderada e grave. O FLCI vem preencher importante lacuna de carência de instrumentos eficazes para intervenções de linguagem e comunicação em pacientes com demência em fase avançada. / BACKGROUND: The increase in life expectancy has contributed to the growth of elderly population all over the world. Aging is a risk factor for many diseases - including dementia - that, in the next decades, will answer for a significant number of elderly with cognitive, behavioral and functional deficits. The expansion of care for these patients is associated to the increase in their life expectancy; therefore there is also an increase in the number of demented individuals in advanced stages. Dementia gradually undertakes behavior and cognition, and a progressive deterioration in communication is also observed. Language characteristics in advanced stages of dementia have been little detailed in literature, hence the need to provide a Brazilian Portuguese version of an instrument to evaluate language in moderate and severe dementia, the Functional Linguistic Communication Inventory (FLCI). The FLCI generates data to help diagnosis, monitoring and evolution of the disease and, moreover, for the orientation of family and caregivers. The FLCI have been used in population with Alzheimer\"s disease (AD), but its applicability in other types of dementia, such as Alzheimer\"s associated to vascular dementia (AD+VD) and frontotemporal lobar degeneration spectrum dementia (FTLD) in advanced stages, is unknown. PURPOSE: 1. to translate and culturally adapt the FLCI for use with the Brazilian population; 2. to compare the performances of patients with moderate and severe AD, AD+VD, and FTLD; and 3. to correlate cognitive-linguistic abilities and functionality in daily life. METHODS: Participants were 57 subjects (24 with AD, 24 with AD+VD, and 09 with FTLD) with ages 60 years and up and levels 2 or 3 in the Clinical Dementia Rating (CDR), moderate or severe, respectively. It was carried out the translation and cultural adaptation of the FLCI, and internal consistency, intra- and inter-examiners reliability were verified, as well as the criterion validity of the instrument through its correlation with the Mini-Mental State Examination (MMSE). The cognitive-linguistic performance of the subjects grouped according to diagnosis was analyzed by comparing mean scores on different subtests of the FLCI. It was possible to correlate cognitive aspects and functionality through the FAST scale (Functional Assessment Staging) and to analyze its correlation with the FLCI. It was also verified the effects of age and education level on the communicative performance of these patients. RESULTS: Statistical analysis indicated that the FLCI presents satisfactory internal consistency (Cronbach\"s ?=.890), great intra- and inter-examiner reliability (interclass correlation coefficient - ICC=.999 and .100, respectively), and great criterion validity when correlated to the MMSE. All subtests that compose the FLCI presented significant differences for the total sample classified according to the severity of dementia, after total score on the test was correlated to the FAST scale. According to the nosological classification of dementia, only one FLCI subtest showed significant differences: \"sign comprehension and object-to-picture matching\". The variables age and education level did not influence the communicative performance of the sample. When performance profiles based on the average score in each FLCI subtest were compared, it was observed better performance of the FTLD group in most subtests, followed by the AD group and, last, by the AD+VD group. Functionality and communication abilities were correlated based on the comparison between total score on the FLCI and the modified FAST scale. It was verified a correlation between total, dichotomous (scored yes/no) and scored (open scored) items of FLCI and the FAST scale. CONCLUSION: The Brazilian Portuguese version of the FLCI is a valid and reliable instrument to evaluate patients with advanced dementia, useful to identify communication abilities of demented in moderate and severe stages. The FLCI fulfills an important lack of efficient instruments for language and communication intervention in patients with dementia in advanced stages.

Dentre os diversos fatores que provocam diminuição no desempenho reprodutivo, a degeneração testicular térmica é o motivo mais freqüente de baixa fertilidade em Bos taurus no Brasil. Baseados nos efeitos sobre a secreção de hormônios hipofisários e gonadais, o GH vem sendo estudado para o tratamento da infertilidade masculina. Um delineamento experimental tipo blocos ao acaso utilizou dezesseis touros adultos submetidos a 4 tratamentos em esquema fatorial 2x2 (0 e 96 horas de insulação testicular, 0 e 1,2 mg bST/kg PV), com o objetivo de testar os efeitos da bST no tratamento de touros submetidos a insulação testicular. Motilidade, alterações de acrossoma, defeitos de cauda e cabeça, gota protoplasmática proximal e defeitos espermáticos totais aumentaram em conseqüência da insulação testicular. As concentrações seminais de Testosterona foram temporariamente diminuídas em resposta a insulação testicular. A ocorrência de gota protoplasmática distal, anomalias de peça intermediária e concentrações seminais de IGF1 não foram afetadas pela insulação testicular. A somatotropina recombinante bovina não afetou as características espermáticas ou concentrações seminais de Testosterona e IGF1. / Testicular heat degeneration is the most common cause of poor fertility of Bos taurus bulls in the tropics. The Growth Hormone has been studied in man infertility treatment with some progress. A randomly blocks experimental design used 16 mature bulls allotted in 4 treatments in a 2x2 factorial arrangement (0 e 96 hours of scrotal insulation, 0 e 1,2 mg bST/kg BW) was performed to asses the effects ob bulls submitted to scrotal insulation. Motility, abnormal acrosome, tail and head defects, proximal droplet, and abnormal sperm increased, and seminal plasma Testosterone was temporally increased in response to scrotal insulation. Distal droplet, midpiece and seminal plasma IGF1 were not affected by bST. The bST did not affect sperm characteristics or seminal Testosterone and IGF1.

Hepcidin is a well-known iron-regulatory hormone that plays a key role in maintaining peripheral iron homeostasis. The presence and wide-spread distribution of hepcidin in the brain suggests that this peptide may also be an important player in brain iron homeostasis. In this study, we tested the hypothesis that hepcidin exerts an important role in the regulation of brain iron content, which might benefit iron-associated NDs such as PD. We also examined the hypothesis that hepcidin could control iron transport processes via regulating iron transport proteins in the brain cells, thus maintaining brain iron homeostasis. / In conclusion, the results of the present study implied that hepcidin plays an important role in maintaining brain iron homeostasis. Hepcidin is beneficial for PD and this effect is related to its iron-regulatory effect via inhibiting iron accumulation in the substantia nigra. Hepcidin effectively controls iron uptake and release through regulating iron transport proteins expressions in the brain, which would contribute to brain iron homeostasis. Therefore, manipulation of hepcidin level in the brain has a potential to be developed into a novel preventive approach for the iron-associated NDs such as PD. / In the second part, we investigated the effect of hepcidin on the processes of iron uptake and release in the cultured brain cells including neurons, astrocytes and brain vascular endothelial cells (BVECs). The expressions of iron uptake proteins such as transferrin receptor 1 (TfR1) and divalent metal transporter 1 (DMT1) as well as the iron exporter ferroportin 1 (Fpn1) were also observed. We found that hepcidin reduced both iron uptake and release via decreasing iron transport proteins expressions in these brain cells, which would contribute to its iron regulatory effect. Finally, we further explored the mechanisms underlying the regulatory effect of hepcidin on the iron transporters in the last part, and found that the action of hepcidin in reducing TfR1 expression is a direct and cAMP-PKA (Cyclic Adenosine 3', 5'-monophosphate/ Protein Kinase-A) pathway-dependent event. / Iron is a transition trace metal essential for mammalian cellular and tissue viability. It also plays important roles in the central nervous system (CNS), including embryonic brain development, myelination, and neurotransmitters synthesis. However, abnormal iron accumulation has been demonstrated in a number of neurodegenerative diseases (NDs) such as Parkinson's (PD), Alzheimer's (AD) and Huntington's diseases (HD). Currently very little is known about the mechanisms involved in brain iron homeostasis and therefore it is not known why and how iron is abnormally increased in the brain. However, given the essential role that excess iron plays in the pathological processes in the NDs, to suppress the accumulated iron is expected to be an effective strategy to prevent and treat these NDs. / To investigate whether hepcidin could benefit iron-associated NDs including PD and whether this beneficial role is related to its iron-regulatory function in the brain, in the first part of study, we investigated the effects of hepcidin on the 6-hydroxydopamine (6-OHDA) in vitro and in vivo PD models. We found that in primary cultured mesencephalic (MES) neurons, hepcidin overexpression via adenovirus-hepcidin (Ad-hepcidin) infection prevented 6-OHDA-induced increase in cellular iron content and protected the MES neurons. In the 6-OHDA model of PD in vivo, overexpression of hepcidin in the substantia nigra via Ad-hepcidin intranigral injection significantly prevented iron accumulation and dopaminergic neurons loss in the pars compacta of substantia nigra (SNc). These effects were accompanied by a marked improvement in motor performance of the PD animals. These findings indicate that hepcidin could benefit iron-associated NDs such as PD and via its iron-regulatory role in the brain. / Du, Fang. / Adviser: Ya Ke. / Source: Dissertation Abstracts International, Volume: 73-06, Section: B, page: . / Thesis (Ph.D.)--Chinese University of Hong Kong, 2011. / Includes bibliographical references (leaves 152-173). / Electronic reproduction. Hong Kong : Chinese University of Hong Kong, [2012] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Electronic reproduction. [Ann Arbor, MI] : ProQuest Information and Learning, [201-] System requirements: Adobe Acrobat Reader. Available via World Wide Web. / Abstract also in Chinese.

Retinal hypoxia is associated with many retinal diseases, such as diabetic retinopathy. Current retinal research suggests that retinal hypoxia appears prior to the onset of diabetic retinopathy. The preliminary association of retinal hypoxia to the early stages of diabetic retinopathy is stimulating the development of new technologies to measure the oxygen content of retinal tissue. Frequency domain phosphoresence lifetime imaging (PLI) is a promising technology that enables the mapping of the oxygen content across the entire retina in the form of two-dimensional images. The two-dimensional images generated from the PLI process are a spatial mapping of the retinal tissue's oxygen tension. Currently, the phosphorescent based oxygen tension PLI measurements contain contaminating auto-fluorescent signals in addition to the desired phosphorescent signals. These auto-fluorescent signals artificially inflate the oxygen tension readings due to the nature of fluorescent signals in phosphorescent imaging. Additionally, the maps generated through PLI appear to contain oxygen tension information from both the retinal vasculature and the choroidal vasculature. The choroidal vasculature is situated directly behind the retina and can have a different oxygen tension value than the retinal vasculature. This research enhanced the PLI system by mathematically eliminating the contaminating auto-fluorescent signals and investigated the methods aimed at separating the PO2s of the retinal and choroidal vasculature beds. In addition, the application of the enhanced PLI technology to the investigation of retinal oxygen changes in a rat model of type I diabetes yielded results that suggest a hyperoxic to hypoxic trend prior to the onset of diabetic retinopathy.

Lam Wun. / Thesis (M.Phil.)--Chinese University of Hong Kong, 2005. / Includes bibliographical references (leaves 140-153). / Abstracts in English and Chinese. / ABSTRACT --- p.i / ACKNOWLDGEMENT --- p.iii / TABLE OF CONTENT --- p.iv / ABBREVIATIONS --- p.x / LISTS OF TABLES --- p.xi / LISTS OF FIGURES --- p.xii / Chapter 1. --- INTRODUCTION / Chapter 1.1 --- Neurodegeneration and triplet repeat diseases --- p.1 / Chapter 1.2 --- Polyglutamine diseases --- p.2 / Chapter 1.3 --- Polyglutamine nuclear inclusions --- p.4 / Chapter 1.3.1 --- Kinetics of polyglutamine nuclear inclusion formation --- p.4 / Chapter 1.3.2 --- Roles of protein inclusions in neurodegeneration --- p.7 / Chapter 1.4 --- Polyglutamine pathogenic pathways --- p.8 / Chapter 1.4.1 --- Protein depletion theory --- p.9 / Chapter 1.4.2 --- Induction of apoptotic pathways --- p.13 / Chapter 1.5 --- Previous study on NI proteins --- p.14 / Chapter 1.6 --- Drosophila model for studying polyglutamine diseases --- p.15 / Chapter 1.6.1 --- Drosophila model for studying human diseases --- p.15 / Chapter 1.6.2 --- GAL4/UAS gene expression system --- p.15 / Chapter 1.6.3 --- Drosophila polyglutamine models --- p.17 / Chapter 1.7 --- Objectives of the study --- p.21 / Chapter 2. --- MATERIALS AND METHODS / Chapter 2.1 --- Drosophila genetics --- p.22 / Chapter 2.1.1 --- Drosophila culture --- p.22 / Chapter 2.1.2 --- GAL4/UAS gene expression system --- p.22 / Chapter 2.1.3 --- Eye phenotypic analysis --- p.25 / Chapter 2.1.4 --- Polyglutamine fly models --- p.25 / Chapter 2.1.5 --- Generation and characterization of GFP-polyglutamine transgenic fly models --- p.25 / Chapter 2.2 --- Proteomic identification of nuclear inclusion proteins --- p.26 / Chapter 2.2.1 --- Proteomic identification of NI proteins by SDS-insolubility of NIs --- p.26 / Chapter 2.2.2 --- Proteomic identification of NI proteins by FA-solubility of NIs --- p.27 / Chapter 2.2.2.1 --- Approach overview --- p.27 / Chapter 2.2.2.2 --- Sample preparation for two-dimensional gel electrophoresis --- p.27 / Chapter 2.2.2.3 --- Two-dimensional gel electrophoresis --- p.29 / Chapter 2.2.2.4 --- Polyacrylamide gel staining --- p.31 / Chapter 2.2.2.5 --- Computer analysis of 2D patterns --- p.31 / Chapter 2.2.2.6 --- In-gel trypsin digestion --- p.32 / Chapter 2.2.2.7 --- Mass spectrometric analysis --- p.33 / Chapter 2.2.3 --- Detection of NIs by flow cytometry --- p.34 / Chapter 2.3 --- SDS-polyacrylamide gel electrophoresis (SDS-PAGE) --- p.34 / Chapter 2.3.1 --- Sample preparation for SDS-PAGE --- p.34 / Chapter 2.3.2 --- SDS-PAGE --- p.35 / Chapter 2.4 --- Immunodetection --- p.36 / Chapter 2.4.1 --- Electroblotting --- p.36 / Chapter 2.4.2 --- Western blotting --- p.36 / Chapter 2.4.3 --- Filter trap assay --- p.37 / Chapter 2.5 --- Sav antibody production --- p.38 / Chapter 2.5.1 --- Sav peptide synthesis --- p.38 / Chapter 2.5.2 --- Rabbit immunization --- p.38 / Chapter 2.6 --- Cryosectioning and immunostaining of adult fly heads --- p.39 / Chapter 2.7 --- Alcohol dehydrogenase assay --- p.40 / Chapter 2.8 --- Semi-quantitative reverse transcription- Polymerase Chain Reaction --- p.41 / Chapter 2.8.1 --- Total RNA preparation from fly heads --- p.41 / Chapter 2.8.2 --- Reverse transcription- Polymerase Chain Reaction (RT-PCR) --- p.41 / Chapter 2.9 --- Reagents and buffers --- p.42 / Chapter 3. --- RESULTS / Chapter 3.1 --- Transgenic polyglutamine fly models --- p.48 / Chapter 3.1.1 --- Characteristics of MJD polyglutamine fly model --- p.48 / Chapter 3.1.1.1 --- Overexpression of expanded truncated human MJD proteins in Drosophila causes eye degeneration --- p.49 / Chapter 3.1.1.2 --- Overexpression of expanded truncated human MJD proteins in Drosophila results in nuclear inclusion formation --- p.49 / Chapter 3.1.1.3 --- Formic acid dissolves fly polyglutamine nuclear inclusions --- p.51 / Chapter 3.1.1.3.1 --- Formic acid dissolves fly polyglutamine NIs as shown by Western blot analysis --- p.51 / Chapter 3.1.1.3.2 --- Formic acid dissolves fly polyglutamine NIs as shown by filter trap assay --- p.53 / Chapter 3.1.2 --- Summary --- p.55 / Chapter 3.2 --- Proteomic identification of nuclear inclusion (NI) proteins --- p.56 / Chapter 3.2.1 --- Proteomic identification of NI proteins by SDS-insolubility of NIs --- p.56 / Chapter 3.2.2 --- Proteomic identification of NI proteins by FA-solubility of NIs --- p.63 / Chapter 3.2.2.1 --- Two-dimensional gels showing differential protein spots as potential NI proteins --- p.63 / Chapter 3.2.2.2 --- NI protein candidates identified by the 2D approach --- p.75 / Chapter 3.2.3 --- Study of polyglutamine NI proteins by flow cytometry analysis --- p.90 / Chapter 3.2.3.1 --- Detection of fly polyglutamine NIs by flow cytometry --- p.90 / Chapter 3.2.3.2 --- Characterization of a new GFP-polyglutamine fly model --- p.92 / Chapter 3.3 --- Characterization of the nuclear inclusion protein candidates --- p.96 / Chapter 3.3.1 --- Glyceraldehyde-3-phosphate dehydrogenase (GAPDH) --- p.96 / Chapter 3.3.1.1 --- Confirmation of GAPDH as a NI protein --- p.97 / Chapter 3.3.1.2 --- Discussion --- p.97 / Chapter 3.3.2 --- Receptor of activated protein kinase C (RACK1) --- p.99 / Chapter 3.3.2.1 --- Confirmation of RACK1 as a NI protein --- p.99 / Chapter 3.3.2.1.1 --- Colocalization of RACK1 with NIs --- p.99 / Chapter 3.3.2.1.2 --- Formic Acid extracts RACK1 from NIs --- p.101 / Chapter 3.3.2.2 --- Reduction of soluble RACK1 protein level in polyglutamine fly --- p.101 / Chapter 3.3.2.2.1 --- Soluble RACK1 protein level reduced in polyglutamine fly --- p.101 / Chapter 3.3.2.2.2 --- RACK1 transcript level remains unchanged in polyglutamine fly --- p.103 / Chapter 3.3.2.3 --- Overexpression of RACK 1 partially suppresses polyglutamine degeneration --- p.105 / Chapter 3.3.2.4 --- Discussion --- p.107 / Chapter 3.3.3 --- Warts (Wts) --- p.111 / Chapter 3.3.3.1 --- Overexpression of Wts partially suppresses polyglutamine degeneration --- p.111 / Chapter 3.3.3.2 --- Wts mutant slightly enhances polyglutamine degeneration --- p.113 / Chapter 3.3.3.3 --- Genetic analysis of Warts pathway in polyglutamine pathogenesis --- p.113 / Chapter 3.3.3.3.1 --- Overexpression of Salvador partially suppresses polyglutamine degeneration --- p.116 / Chapter 3.3.3.3.2 --- Hpo mutant slightly enhances polyglutamine degeneration --- p.119 / Chapter 3.3.3.3.3 --- Overexpression of DIAP1 partially suppresses polyglutamine degeneration --- p.119 / Chapter 3.3.3.4 --- Discussion --- p.121 / Chapter 3.3.4 --- Alcohol dehydrogenase (Adh) --- p.122 / Chapter 3.3.4.1 --- Adh activity is reduced in polyglutamine flies --- p.122 / Chapter 3.3.4.2 --- Overexpression of Hsp70 partially restores the reduced Adh activity in polyglutamine flies --- p.122 / Chapter 3.3.4.3 --- Discussion --- p.125 / Chapter 3.3.5 --- Genetic analysis of other NI protein candidates --- p.127 / Chapter 3.3.5.1 --- Overexpression of CG7920 protein partially suppresses polyglutamine degeneration --- p.127 / Chapter 3.3.5.2 --- Pten dsRNA slightly enhances polyglutamine degeneration --- p.129 / Chapter 3.3.6 --- Summary --- p.131 / Chapter 4. --- DISSCUSSION / Chapter 4.1 --- Protein depletion theory --- p.133 / Chapter 4.2 --- Comparison of different approaches for identification of NI proteins --- p.134 / Chapter 4.3 --- Long-term significance --- p.136 / Chapter 4.4 --- Future studies --- p.137 / Chapter 4.4.1 --- Characterization of other NI protein candidates --- p.137 / Chapter 4.4.2 --- Study of NI proteins by an alternative approach --- p.137 / Chapter 4.4.3 --- Study of NI proteins using other polyglutamine fly models --- p.137 / Chapter 5. --- CONCLUSION --- p.139 / Chapter 6. --- REFERENCES --- p.140

La scoliose de l’adulte est une pathologie dont la prévalence augmente avec de le vieillissement de la population. De plus, la demande fonctionnelle est de plus en plus importante chez ces patients. De nombreuses études ont montré un bénéfice du traitement chirurgical de la déformation de l’adulte par rapport au traitement médical en termes d’amélioration des scores fonctionnels, de qualité de vie et de satisfaction. Néanmoins, cette chirurgie est associée à de nombreuses complications (jusque 50% dans certaines séries), pour la plupart mécaniques telle la pseudarthrose, la rupture d’implant, la dégradation des étages adjacents. Ainsi, si l’analyse radiographique de la scoliose a bien été explorée, certaines questions demeurent comme les phénomènes responsables d’une aggravation brutale de la déformation chez certains patients ou les causes d’échecs. L’objectif de ce travail était d’analyser la scoliose de l’adulte, en 3D à l’aide de la stéréoradiographie et d’évaluer le système musculaire de ces patients afin de mettre en évidence des relations entre déformations rachidiennes notamment par l’analyse du plan horizontal, et dégénérescence musculaire.La 1e partie de ce travail était consacrée à l’analyse 3D de la scoliose : tout d’abord avec l’analyse de reproductibilité chez l’adulte des mesures 3D effectuées par stéréoradiographie, puis par l’analyse de l’alignement global de ces patients avec une déformation rachidienne, à l’aide d’un nouveau paramètre prenant en compte la position de la jonction cervico-céphalique. Apres avoir analysé l’alignement postural, le système musculaire pelvi-rachidien a été étudié dans la 2e partie. En effet, en plus de l’alignement du squelette, c’est l’activation du système musculaire qui est responsable du maintien d’une posture érigée. Nous avons décrit les caractéristiques musculaires des patients avec une scoliose lombaire et analysé les relations avec les paramètres radiographiques de la déformation, montrant que selon le type de déformation certains groupes musculaires étaient plus touchés par l’atrophie et l’infiltration graisseuse. En les comparant à des sujets jeunes et âgés sans déformation, nous avons observé que les patients avec une scoliose avaient une dégénérescence musculaire à la fois liée à la déformation et au vieillissement. Dans une 3e partie, en faisant le lien entre les données de la posture par la stéréoradiographie et les données musculaires de l’IRM, nous avons utilisé un modèle musculosquelettique personnalisé pour mieux comprendre les contraintes exercées sur les segments vertébraux et donc pour essayer d’expliquer les faillites mécaniques.Ainsi, l’association de mesures 3D radiographiques et de l’analyse musculaire pourrait permettre en comprenant mieux les phénomènes dégénératifs, de mieux prédire l’aggravation de la déformation et donc de la prévenir par une rééducation ciblée. Par exemple, un renforcement des érecteurs du rachis mais aussi des fléchisseurs pourrait permettre de mieux maintenir la posture. Et, un renforcement des érecteurs et fléchisseurs de hanche pourrait permettre d’activer de manière plus efficace, les mécanismes de compensation telle la rétroversion pelvienne. De plus, la mise en évidence de facteur de risque musculosquelettique d’aggravation de la déformation entrainerait une prise en charge plus précoce de ces patients. Une analyse longitudinale serait donc intéressante. / Adult degenerative scoliosis is a pathology whose prevalence increases with the aging of the population. Moreover, the functional demand is more and more important in these patients. Many studies have shown a benefit of surgical treatment of adult spinal deformity compared to medical treatment in terms of improved functional scores, quality of life and satisfaction. Nevertheless, this surgery is associated with many complications (up to 50% in some series), mostly mechanical such as nonunion, implant rupture, degradation of adjacent levels. Thus, if the radiographic analysis of scoliosis has been well explored, some questions remain like the phenomena responsible for a sudden worsening of the deformation in certain patients or the causes of failures. The aim of this work was to analyze adult scoliosis in 3D using stereoradiography and to evaluate the muscular system of these patients in order to highlight the relationships between spinal deformities, particularly by the horizontal plane analysis, and muscle degeneration.The first part of this work was dedicated to the 3D analysis of scoliosis: first, with the analysis of reproducibility in the adult of 3D stereoradiographic measurements, then by the analysis of the global alignment of these patients with spinal deformity, using a new parameter taking into account the position of the cervico-cephalic junction. After analyzing the postural alignment, the spino-pelvic muscular system was studied in the second part. Indeed, in addition to the alignment of the skeleton, it is the activation of the muscular system that is responsible for maintaining an erect posture. We described the muscular features of patients with lumbar scoliosis and analyzed the relationships with the radiographic parameters of the deformity, showing that depending on the type of deformity some muscle groups were more affected by atrophy and fatty infiltration. Comparing them to young and elderly subjects without deformity, we observed that patients with scoliosis had muscle degeneration that was both related to deformity and aging. In the third part, by linking stereoradiographic posture data with muscular MRI data, we used a personalized musculoskeletal model to better understand the constraints on vertebral segments and therefore to try to explain the mechanical failures.Thus, the combination of 3D radiographic measurements and muscle analysis could better predict muscle degeneration and worsening of deformity and thus prevent it by targeted rehabilitation. For example, a strengthening of the erectors of the spine but also of the flexors could allow better maintaining the posture. And, a strengthening of the erectors and hip flexors could allow activating more effectively, compensation mechanisms such pelvic retroversion. In addition, the demonstration of a musculoskeletal risk factor worsening the deformity would lead to an earlier management of these patients. A longitudinal analysis would be interesting.