Submitted by Neusa Fagundes (neusa.fagundes@unioeste.br) on 2018-07-10T19:46:34Z No. of bitstreams: 2 Joziana_Pastro2018.pdf: 1649555 bytes, checksum: 769cd69a9e879a09907152dd2e64987c (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) / Made available in DSpace on 2018-07-10T19:46:34Z (GMT). No. of bitstreams: 2 Joziana_Pastro2018.pdf: 1649555 bytes, checksum: 769cd69a9e879a09907152dd2e64987c (MD5) license_rdf: 0 bytes, checksum: d41d8cd98f00b204e9800998ecf8427e (MD5) Previous issue date: 2018-03-13 / Conselho Nacional de Pesquisa e Desenvolvimento Científico e Tecnológico - CNPq / Retinopathy of prematurity (ROP) is a vasoproliferative disease, related to blindness, in Premature Newborns (PN) who, within other factors, used oxygen during hospitalization. Early visual follow-up of preterm newborns by ophthalmologists makes it possible to identify the stage and grade of the disease and the indication of treatment or attendance of its evolution. Nursing orientation is essential to the PN relatives, since ROP is not an easily seen disease, so, parents being aware that the ophthalmologic follow-up is of vital importance for the visual recovery of the baby will make a difference in the child future visual health. Therefore, this study aims to evaluate the prevalence and evolution of ROP in PN, hospitalized in a Neonatal Intensive Care Unit (NICU) and attended at a referral ophthalmologic follow-up service. This is a quantitative, descriptive and exploratory study, carried out in the city of Cascavel. Data collection was done in medical records of PN, born between January 2014 and June 2016, hospitalized in the NICU of the West Paraná University hospital, and followed by the outpatient clinic of Cascavel Eyes Hospital, totaling 181 charts. A quantitative analysis was performed through descriptive and inferential statistic. The prevalence of ROP was 11.31% (n = 50). Relating its characteristics to evolution, the diagnoses prevalent in the PN of the study were respiratory diseases (41.99%) and among comorbidities, sepsis prevailed (63.54%). 80 PN required blood transfusion (44,20%) and 152 (83.98%) used oxygen therapy, with a predominance of mask device (n = 141; 77,90%) and orotracheal tube - TOT (n = 100; 55.25%), on average for 15 days. The highest concentration recorded was hood (45.0%). The time of use and the O2 concentration of TOT, time of mask use and time and concentration of O2 of Continuous Positive Airway Pressure (CPAP), were significant to trigger ROP. Among ROP cases, ten (5.5%) children had severe ROP and required laser treatment. All the PN diagnosed with ROP used oxygen. ROP prevailed in moderate preterm newborns (44%), with ROP grade 3 being the most severe, affecting both extreme and moderate PN. Respiratory diseases, sepsis and procedures such as blood transfusion and oxygen therapy influenced the presence of ROP, with prevalence in moderate preterm newborns. The adoption of ophthalmologic screening for ROP detection, with background examinations of the eyes in the NICU contributed to the early treatment and prevention of severe visual impairment and blindness caused by ROP. Thus, nurses are fundamentally important, from the prevention, the diagnosis until the course of the treatment and for discharge counselling. / A Retinopatia da Prematuridade (ROP) é uma enfermidade vasoproliferativa, relacionada à cegueira, em recém-nascidos prematuros (RNPT) que, dentre outros fatores, utilizaram oxigênio durante a hospitalização. O acompanhamento visual precoce de prematuros por oftalmologistas possibilita a identificação do estágio e do grau da doença, assim como a indicação de tratamento ou de acompanhamento da sua evolução. A orientação da enfermagem é primordial aos familiares dos RNPT, visto que a ROP não é uma doença de fácil visualização; portanto, os pais estarem cientes de que o acompanhamento oftalmológico é de vital importância para a recuperação visual do bebê fará diferença na vida futura da criança. Assim, este estudo objetiva avaliar a prevalência e a evolução da ROP em RNPT internados em uma Unidade de Terapia Intensiva Neonatal (UTIN) e acompanhados em serviço de seguimento oftalmológico de referência. Trata-se de uma pesquisa quantitativa, descritiva e exploratória, realizada no município de Cascavel. A coleta de dados ocorreu em prontuários de RNPT, nascidos entre janeiro de 2014 e junho de 2016, hospitalizados na UTIN do Hospital Universitário do Oeste do Paraná – HUOP, e em seguimento pelo ambulatório do Hospital de Olhos de Cascavel, totalizando 181 prontuários. Realizou-se análise quantitativa, por meio de estatística descritiva e inferencial. A prevalência de ROP foi de 11,31% (n=50). Relacionados às suas características e evolução, os diagnósticos prevalentes nos RNPT do estudo foram as doenças respiratórias (41,99%) e, entre as comorbidades, a sepse prevaleceu (63,54%). Necessitaram de transfusão de sangue 80 RNPT (44,20%), e 152 (83,98%) fizeram uso de oxigenioterapia, com predomínio do dispositivo máscara (n=141; 77,90%) e tubo orotraqueal - TOT (n=100; 55,25%), em média por 15 dias. A maior concentração registrada foi por halo (45%). O tempo de uso e a concentração de O2 de TOT, tempo de uso de máscara e tempo e concentração de O2 do Continuous Positive Airway Pressure (CPAP) foram significativos para desencadear a ROP. Dentre os casos de ROP, dez (5,5%) crianças tiveram ROP grave e necessitaram de tratamento com laser. Todos os RNPT diagnosticados com ROP fizeram uso de oxigênio. A ROP prevaleceu nos prematuros moderados (44%), sendo o grau 3 de ROP o mais grave encontrado, acometendo tanto RNPT extremos quanto moderados. Doenças respiratórias, sepse e procedimentos como a transfusão de sangue e a oxigenioterapia influenciaram na presença da ROP, com prevalência em prematuros moderados. A adoção da triagem oftalmológica para a detecção da ROP, com exames de fundo de olho sistematicamente realizados na UTIN, contribuíram para o tratamento precoce e a prevenção de deficiências visuais graves e cegueira causados pela ROP. Nesse sentido, o enfermeiro é de fundamental importância, desde a prevenção, o diagnóstico até o decorrer do tratamento e a alta.

Submitted by Renata Lopes (renatasil82@gmail.com) on 2016-02-26T11:57:05Z No. of bitstreams: 1 jomaraoliveiradossantosyogui.pdf: 569717 bytes, checksum: be9f77d66f6c940acf5caed0b7a11a98 (MD5) / Approved for entry into archive by Adriana Oliveira (adriana.oliveira@ufjf.edu.br) on 2016-03-03T13:37:39Z (GMT) No. of bitstreams: 1 jomaraoliveiradossantosyogui.pdf: 569717 bytes, checksum: be9f77d66f6c940acf5caed0b7a11a98 (MD5) / Made available in DSpace on 2016-03-03T13:37:39Z (GMT). No. of bitstreams: 1 jomaraoliveiradossantosyogui.pdf: 569717 bytes, checksum: be9f77d66f6c940acf5caed0b7a11a98 (MD5) Previous issue date: 2015-03-11 / A Retinopatia da Prematuridade(ROP) é uma das principais causas de cegueira na infância. A ROP pode ser evitável ou minimizada, desde que diagnosticada em tempo hábil. Objetivos: Determinar a ocorrência de retinopatia da prematuridade e avaliar os principais fatores associados e implicados no seu desenvolvimento. Métodos: Trata-se de um estudo transversal de base hospitalar, realizado no período de 2002 a 2012, no Hospital Universitário, HU/CAES/UFJF, de Juiz de Fora/MG-Brasil. A população de estudo é composta por 392 recém-nascidos, com idade gestacional < 37 semanas e qualquer peso ao nascimento. Resultados:A ocorrência da ROP no HU/CAS/UFJF foi de 29,9%. Os fatores associados à ROP, após a regressão logística, foram: baixo peso ao nascer, oxigenoterapia (cateter nasal e CPAP) e sepse neonatal. Conclusão: O presente estudo demonstrou significantes fatores associados à ROP após realizar a análise multivariada: peso ao nascer (foi demonstrado que quanto mais baixo o peso ao nascer maior é a ocorrência da ROP), oxigenioterapia (cateter nasal e CPAP) e sepse neonatal. / The retinopathy of prematurity(ROP) is one of the main causes of childhood blindness. The ROP can be avoided or minimized as long as it is diagnosed in time. Purposes: To determine the occurrence of retinopathy of prematurity and assess the main associated factors involved in its development. Methods: Seccional study carried out from 2002 at 2012, at the University Hospital HU/CAES/UFJF, Juiz de Fora (MG) – Brazil. The sample was composed of 392 newborns, with less than 37 weeks of gestational age and any birth weight. Results: The occurrence of ROP in HU/CAS/UFJF was of 29, 9%. The associated factors of ROP after the logistic regression were birth weight, oxygen Therapy (nasal catheter, CPAP), neonatal sepsis. Conclusion: The present study showed significant associated factors to ROP: birth weight (It was noticed that the lower the birth weight, the higher is the occurrence of ROP), oxygen Therapy (nasal catheter, CPAP) and neonatal sepsis.

Les rétinopathies ischémiques et la forme humide de la dégénérescence maculaire liée à l’âge (DMLA) sont la principale cause de malvoyance respectivement chez les personnes en âge de travailler et les personnes agées. Les néovascularisations choroïdiennes (NVC) et rétiniennes et l’œdème maculaire associés à ces pathologies sont traités à l’aide de biomolécules qui ciblent uniquement la voie de signalisation des VEGF. Le développement de thérapies indépendantes de cette voie permettrait d’offrir aux patients résistants aux anti-VEGF une alternative thérapeutique pour préserver leur vision. Les intégrines αvβ3 et αvβ5, impliquées dans la néovascularisation oculaire, apparaissent ainsi comme une cible alternative intéressante. La Lébécetine (LCT), une lectine de type-C, de 30 kDa et de structure hétérodimèrique issue de venin de serpent Macrovipera lebetina interagit spécifiquement avec les intégrines α5β1 et αvβ3, αvβ5. La LCT a une activité anti-angiogénique in vitro sur des cellules endothéliales microvasculaire du cerveau humain (HBMEC) et in vivo sur le modèle de la membrane chorioallantoïde du poulet (CAM). Afin d’étudier son effet sur la néovascularisation oculaire, nous avons eu recours aux modèles d’angiogenèse ex vivo utilisant des explants aortique ou choroïdien cultivés en présence de LCT, puis son effet a été évalué in vivo dans un modèle de NVC chez la souris et également sur la néovascularisation rétinienne dans le modèle de rétinopathie induite par l’oxygène (RIO). Nos données démontrent qu’une injection unique de LCT est capable de réduire la NVC et rétinienne dans ces modèles sans affecter les vaisseaux quiescents matures indiquant un bon profil d’innocuité. / Ischemic retinopathies and the wet form of age-related macular degeneration (AMD) are characterized by devastating angiogenesis responsible for the majority of irreversible blindness. Current therapies include use of anti-VEGF agents to reduce choroidal neovascularization and edema. These treatments are effective in most cases, but spontaneous or acquired resistance to anti-VEGF highlight a need for additional alternative therapies. In recent years, pharmacological inhibition of αvβ3 and αvβ5, which regulate endothelial cell proliferation and stabilization, have emerged as new therapeutic tools for the treatment of these diseases. Lebecetin (LCT), a 30-kDa heterodimeric C-type lectin that is isolated from Macrovipera lebetina venom, interacts with α5β1 and αv-containing integrins (αvβ3, αvβ5). We previously showed that LCT has an anti-angiogenic effect in vitro on human brain microvascular endothelial cells (HBMEC) and in vivo in a chick chorioallontoic membrane assay (CAM). To evaluate the inhibitory effect of LCT on ocular angiogenesis, we cultured aortic and choroidal explants in the presence of LCT and analyzed the effect of LCT on choroidal neovascularization in the mouse CNV model and on retinal neovascularization in the oxygen induced retinopathy (OIR) model. Our data demonstrated that a single injection of LCT efficiently reduced choroidal and retinal neovascularization in these models with no significant effect on mature blood vessels predicting a good safety profile.

In my thesis I dealt with the theme of very serious and widespread diseases of civilization posed by diabetes mellitus (diabetes), type 2. Incidence of the disease is considered a pandemic of the 21st century. I focused on macro- and microvascular complications of type 2 diabetes mellitus. The work is divided into two parts. The theoretical part deals with the description of type 2 diabetes in terms of its origin, course, diagnosis and treatment, including komplkací. In the practical part I am devoted to a survey of diabetic patients in the waiting rooms of several diabetics. Through the questionnaire, I investigated how respondents have access to their disease. Whether they know their blood glucose levels as they are treated, whether they realize the seriousness of their illness if they have already expressed complications of diabetes and how to approach the treatment, if observe regime measures. The respondents' answers, I analyzed and processed using graphs.

Abstract The incidence of childhood onset type 1 diabetes in Finland has been the highest in the world for several decades. Optimal management of the disease presents a lifelong challenge to the affected individuals. Related complications are common and include an ocular pathology called diabetic retinopathy. Type 1 diabetes with its ramifications can impact on several facets of a patient’s physical and psychological well-being. This study aimed to assess the ophthalmic findings and to evaluate the characteristics of general well-being of a population-based cohort of paediatric patients with type 1 diabetes and a population-based cohort of young adults with type 1 diabetes since childhood. The prevalence and risk factors of diabetic retinopathy were assessed of the population-based paediatric cohort in the catchment area of the Northern Ostrobothnia Hospital District and these were compared to a similar paediatric cohort studied 18 years previously. There was no significant change in the overall prevalence of diabetic retinopathy (12%) during the study period. Furthermore, glycaemic balance and other risk factors of diabetic retinopathy had remained almost unchanged. A population-based cohort of young adults was evaluated in 2007 for the prevalence and severity of diabetic retinopathy. Most of the cohort subjects (94%) had developed diabetic retinopathy and in every third subject there was evidence of proliferative retinopathy. Health related quality of life was the same as that in the age- and gender-standardised control population. For the most part, the young adults with a long duration of type 1 diabetes fared equally well as the general population in the measured social aspects. However, proliferative diabetic retinopathy was associated with lower educational achievements and poorer health related quality of life as well as with a higher probability of unemployment or being pensioned. Glycaemic balance and prevalence of diabetic retinopathy have remained unchanged in paediatric cohorts for nearly two decades despite concurrent advances in care. Social well-being was mainly restricted in young adults exhibiting signs of proliferative diabetic retinopathy. The negative impact of advanced complications of type 1 diabetes already in these young adults highlights the importance of strict metabolic control to maintain overall well-being. / Tiivistelmä Lapsuusiässä alkavan tyypin 1 diabeteksen ilmaantuvuus on ollut Suomessa maailman korkein usean vuosikymmenen ajan. Hyvän hoitotasapainon ylläpitäminen on elinikäinen haaste sairastuneelle. Diabeteksen liitännäissairaudet ovat yleisiä, kuten myös silmänpohjissa todettava diabeettinen retinopatia. Tyypin 1 diabetes voi komplikaatioineen vaikuttaa laajasti potilaan fyysiseen ja psyykkiseen hyvinvointiin. Tässä tutkimuksessa pyrittiin arvioimaan silmien terveydentilaa ja yleiseen hyvinvointiin liittyviä tekijöitä tyypin 1 diabetesta sairastavien lasten sekä lapsena diabetekseen sairastuneiden nuorten aikuisten väestöpohjaisissa potilasaineistoissa. Diabeettisen retinopatian esiintyvyys ja riskitekijät tutkittiin väestöpohjaisessa lapsipotilasaineistossa Pohjois-Pohjanmaan sairaanhoitopiirin alueella ja tuloksia verrattiin vastaavaan 18 vuotta aiemmin tutkittuun potilasaineistoon. Diabeettisen retinopatian esiintyvyys (12 %) ei ollut merkittävästi muuttunut tutkimusaikana. Glykeeminen tasapaino ja muut diabeettisen retinopatian riskitekijät olivat pysyneet kohorttien välillä oleellisilta osin ennallaan. Lapsena sairastuneiden nuorten aikuisten väestöpohjaisesta potilasaineistosta arvioitiin diabeettisen retinopatian esiintyvyys ja vaikeusaste vuonna 2007. Enemmistölle potilaista (94 %) oli kehittynyt diabeettinen retinopatia ja kolmanneksella todettiin proliferatiivinen retinopatia. Terveyteen liittyvä elämänlaatu oli verrattavissa ikä- ja sukupuolivakioituun verrokkiväestöön. Sosiaalista hyvinvointia mittaavat tulokset olivat pääosin yhtäläiset muuhun väestöön verrattuna. Proliferatiivisella diabeettisella retinopatialla havaittiin kuitenkin yhteys huonompaan terveyteen liittyvään elämänlaatuun ja koulutustasoon sekä korkeampiin työttömyys- ja eläköitymislukuihin. Tutkitussa aineistossa lapsipotilaiden glykeeminen tasapaino sekä diabeettisen retinopatian esiintyvyys pysyivät ennallaan lähes kahden vuosikymmenen ajan hoitojen kehittymisestä huolimatta. Nuorten aikuisten sosiaalisessa hyvinvoinnissa esiintyi poikkeavuuksia lähinnä proliferatiivista diabeettista retinopatiaa sairastavilla. Tyypin 1 diabeteksen pitkälle edenneiden komplikaatioiden negatiivinen vaikutus jo nuorella aikuisiällä korostaa hyvän hoitotasapainon tärkeyttä yleisen elämänlaadun ylläpitämisessä.

Les complications vasculaires rétiniennes constituent des évènements qui peuvent être observés au cours de rétinopathies pouvant être à l’origine d’une cécité à tous les stades de la vie. Ces complications concernent particulièrement la rétinopathie du prématuré, la rétinopathie diabétique et la dégénérescence maculaire liée à l’âge. Les lipides offrent de nombreuses possibilités pour prévenir et éventuellement freiner le développement de ces rétinopathies. Parmi eux, la classe des plasmalogènes est particulièrement riche en acides gras poly-insaturés (AGPI), qui sont libérés par une phospholipase indépendante du calcium (iPLA2) et qui sont précurseurs de métabolites biologiquement actifs. Certains de ces métabolites sont connus pour être impliqués dans la modulation de l’angiogenèse rétinienne. L’objectif de ce travail de thèse a été d’évaluer l’implication des plasmalogènes dans le développement vasculaire rétinien par l’intermédiaire de la libération des AGPI par la iPLA2. Pour vérifier cette hypothèse, nous avons caractérisé les évènements cellulaires et moléculaires du développement vasculaire rétinien postnatal chez un modèle animal d’inhibition de la iPLA2 rétinienne que nous avons préalablement développé, ceci de manière comparative avec un modèle de déficience totale en plasmalogènes. Nous avons également tenté de mettre en évidence de potentielles altérations du métabolisme des plasmalogènes chez au cours d’une rétinopathie à composante vasculaire chez l’homme, la rétinopathie diabétique. Nos résultats ont suggéré que les plasmalogènes sont indispensables pour le développement physiologique des vaisseaux rétiniens. Ils seraient impliqués dans le contrôle de la formation de la trame astrocytaire et la mise en place du réseau endothélial par l’intermédiaire des AGPI libérés par la iPLA2. Les mécanismes moléculaires impliqueraient la voie des Angiopoïétines-Tie sans affecter celle du VEGF. Chez l’homme, nous avons noté une réduction des AGPI circulants, en particulier l’acide docosaexanéïque et l’acide arachidonique, sur les phosphatidyl-éthanolamines chez tous les patients diabétiques avec ou sans rétinopathie diabétique, sans implication des formes plasmalogènes. Nos résultats suggèrent une implication du métabolisme des plasmalogènes dans le contrôle du développement vasculaire en période péri-natale mais pas au cours de la rétinopathie diabétique. Ce contrôle serait exercé par l’intermédiaire des AGPI libérés par la iPLA2. / Retinal vascular complications are secondary events of several retinopathies that result in blindness at all ages. Such complications can be observed in retinopathy of prematurity, diabetic retinopathy and age-related macular degeneration. Lipids, and particularly polyunsaturated fatty acids (PUFAs), display beneficial properties in the prevention of such retinopathies. Among the different lipid classes, the plasmalogen subclass is particularly interesting since it is known to be rich in PUFAs. These PUFAs are known to be released by a calcium-independent phospholipase (iPLA2) and further converted into biologically active metabolites. Some of these metabolites are known to be involved in the modulation of retinal angiogenesis. The aim of this work was to evaluate the involvement of plasmalogens in retinal vascular development through PUFA release by iPLA2. To check this hypothesis, we have comparatively characterized cellular and molecular mechanisms of postnatal retinal vascular development in an animal model of retinal iPLA2 inhibition as well as in a model of plasmalogens deficiency. On the other hand, we have attempted to identify potential alterations in plasmalogen metabolism in diabetic retinopathy. Our results suggest that plasmalogens are essential for the physiological development of retinal vessels. They are involved in the control of astrocyte template formation and the development of the primary vascular network through PUFA released by iPLA2. Molecular mechanisms by which PUFAs from plasmalogens control retinal vascular development involve Angiopoietin-Tie pathways, without affecting those involving VEGF. In the human study, we have observed a decrease in the bioavailability of circulating PUFAs, and especially docosaexaneic acid and arachidonic acid binded to phosphatidyl-ethanolamine in all diabetic patients with or without diabetic retinopathy. Plasmalogens were not involved in these modifications. Our results suggest that plasmalogen metabolism is involved in the control of primary vascular growth during retinal development but not in diabetic retinopathy. Plasmalogens may control early steps of retinal vascular development through the release of PUFAs by iPLA2.

Orientadores: Jacqueline Mendonça Lopes de Faria, José Butori Lopes de Faria / Tese (doutorado) - Universidade Estadual de Campinas, Faculdade de Ciências Médicas / Made available in DSpace on 2018-08-26T10:54:36Z (GMT). No. of bitstreams: 1 Duarte_DiegoAndreazzi_D.pdf: 28811769 bytes, checksum: 726e2f631efd3a14fedaf1f8148784aa (MD5) Previous issue date: 2014 / Resumo: A retinopatia diabética (RD) é uma doença devastadora que está entre as maiores causas de cegueira entre pessoas na idade adulta em todo o mundo. Considerada multifatorial e progressiva, a RD afeta células neurais e gliais, e também elementos vasculares da retina. Sabe-se que diversas vias estão envolvidas na patogênese da RD, no entanto, os mecanismos que levam a exacerbação da inflamação e morte de células gliais/neurais o que caracteriza a neurodegeneração da retina, ainda permanecem desconhecidos. Diante disso, a redução desses fatores tem sido extensivamente estudada como alvo no combate a RD. As Sirtuínas, histonas desacetilases dependentes de nicotinamida adenina dinucleotídeo (NAD+), atuam em resposta a vários estresses e atualmente tem sido relacionadas às importantes funções moleculares na regulação de várias doenças. Considerado um redox sensível, a SIRT1 pode estar reduzida em condição de doença, o que agravar ainda mais a situação patológica. No entanto, não se sabe ao certo o mecanismo de modulação e/ou atuação da SIRT1 frete às doenças neurodegenerativas, tais como a RD. No Artigo I, foram avaliados os possíveis efeitos protetores do cacau rico em polifenóis na retina diabética. As células Müller da retina (rMC-1) foram expostos por 24h à glicose normal (NG), alta glicose (HG) ou peróxido de hidrogênio (H2O2) e submetidas ao tratamento com cacau na presença ou não de um inibidor da SIRT1 ou siRNA. O estudo animal foi desenvolvido em ratos experimentalmente diabéticos induzidos por estreptozotocina e randomizado para receber tratamentos com cacau com baixa, intermediária, ou elevada dose de polifenol (0,12 mg; 2,9 mg; 22,9 mg/kg/dia) por gavagem durante 16 semanas. As células expostas a H2O2 ou HG apresentaram aumento de proteína acídica fibrilar glial (GFAP) e acetil-RelA/p65 e diminuição da atividade/expressão da SIRT1. Estes efeitos foram anulados pelo cacau, que diminuiu a produção de espécies reativas de oxigênio e reduziu a ativação da poli(ADP-ribose) polimerase-1 (PARP-1); melhorou os níveis intracelulares de NAD+ e consequentemente aumentou da atividade da SIRT1. As retinas dos ratos diabéticos exibiram os primeiros marcadores de retinopatia acompanhada pela eletrorretinografia prejudicada. A presença de diabetes levou a ativação da PARP-1 e diminuição dos níveis de NAD+, resultando em comprometimento da SIRT1. O aumento na acetilação do RelA/p65 levou na hiperexpressão do GFAP. A administração oral de cacau polifenol restaurou as alterações acima referidas. Este estudo revelou que o cacau enriquecido com polifenóis teve efeito protetor da retina diabética restabelecendo a via da SIRT-1. No Artigo II, foi investigado o possível efeito terapêutico de células derivadas de animais saudáveis (Dock7 m +/+ Leprdb db/m) e diabéticos (BKS.Cg-Dock7 m +/+ Leprdb/J, db/db) na retinopatia diabética (RD). Os camundongos db/db (espontaneamente diabéticos) com 8 semanas de idade foram randomizados para receber uma única injeção intravenosa de PBS ou células early outgrowth (EOCs) de doadores db/m ou db/db. Quatro semanas mais tarde, os animais foram sacrificados e os olhos enucleados. Para estudo in vitro, o meio condicionado das EOCs (EOC-CM) foi gerado a partir do cultivo de EOCs de animais db/m e db/db. As células rMC-1 foram expostas por 24h a NG ou HG e submetidas ao tratamento com db/m ou db/db EOC-CM, em presença ou não de um inibidor farmacológico (EX527) ou gênico (siRNA) da SIRT1. Nos ratos diabéticos, houve um aumento de marcadores de RD e do dano oxidativo, acompanhado por uma diminuição da proteína SIRT1 e seguido pelo aumento da acetilação da lisina-310 do complexo p65-NFkB. A terapia celular com EOCs reduziu significativamente todas as alterações mencionadas acima. As rMC-1 expostas a HG apresentaram aumento da expressão de GFAP, fator de crescimento do endotélio vascular e Nox4, acompanhado pelo aumento dos níveis de espécies reativas de oxigênio e acetil-lisina-310-p65-NFkB. Além disso, a expressão/atividade da SIRT1 foram reduzidas em ambiente diabético. O tratamento com EOC-CM impediu todas estas alterações. Este estudo demonstra que a capacidade parácrina das EOCs, na secreção de fatores, é eficaz no restabelecimento da via de SIRT1 retina, e assim, proteger a retina dos insultos diabéticos. Em resumo, a presente tese fornece evidências que tanto a administração oral do cacau enriquecido com polifenóis quanto à terapia celular com EOCs, conferem neuroproteção da retina aos insultos do diabetes. Portanto, intervenções que modulem a atividade das sirtuínas são promissoras no tratamento farmacológico da retinopatia diabética / Abstract: The diabetic retinopathy (RD) is a devastating disease and the principal cause of blindness among people in adulthood worldwide. The RD is considered a multifactorial and progressive disease, affecting neuronal and glial cells, and also vascular elements of the retina. It is known that several pathways are involved in the pathogenesis of RD, however, the mechanisms that lead to exacerbation of inflammation and death of glial/neuronal cell, characterizing retinal neurodegeneration, remain unknown. Therefore, the reduction of these factors have been extensively studied as a therapeutic target against RD. Sirtuin 1 (SIRT1), a family of histone deacetylase enzyme, acts in response to various stresses and, currently, has been related to important molecular functions in the regulation of various diseases. Considered a redox-sensitive, SIRT1 may be reduced under disease condition, whereby aggravate the pathological situation. However, is not known the mechanism of modulation/activity of SIRT1 in neurodegenerative diseases, such as RD. In the article I, were studies the possible protective effects of cocoa in the diabetic retina were assessed. rMCs exposed to NG, HG or H2O2 were submitted to cocoa treatment in the presence or absence of SIRT-1 inhibitor and siRNA. The experimental animal study was conducted in streptozotocin-induced diabetic rats and randomized to receive low, intermediate, or high polyphenol cocoa treatments via daily gavage for 16 weeks (i.e., 0.12 mg/kg/day, 2.9 mg/kg/day, or 22.9 mg/kg/day of polyphenols). The rMCs exposed to HG or H2O2 exhibited increased GFAP and acetyl-RelA/p65 and decreased SIRT1 activity/expression. These effects were cancelled out by cocoa, which decreased ROS production and PARP-1 activity, augmented the intracellular pool of NAD+, and improved SIRT1 activity. The rat diabetic retinas displayed the early markers of retinopathy accompanied by markedly impaired electroretinogram. The presence of diabetes activated PARP-1 and lowered NAD+ levels, resulting in SIRT1 impairment. This augmented acetyl RelA/p65 had the effect of upregulated GFAP. Oral administration of polyphenol cocoa restored the above alterations in a dose-dependent manner. This study reveals that cocoa enriched with polyphenol improves the retinal SIRT-1 pathway, thereby protecting the retina from diabetic milieu insult. In the article II, were investigated the possible therapeutic effect of cells derived from control (db/m) and spontaneously diabetic (db/db) mice on diabetic retinopathy. The db/db mice with 8 weeks of age were randomized to receive a unique intravenous injection of PBS or 0,5x105 db/m EOCs or 0,5x105 db/db EOCs. Four weeks later, the animals were euthanized and the eyes enucleated. For in vitro study, EOC-CM was generated from db/m and db/db EOCs cultures. rMCs were exposed for 24h to NG or HG combined or not with db/m or db/db EOC-CMs. In diabetic rats, there was an increase of DR and oxidative damage markers, accompanied by decrease in SIRT1 protein followed by lysine-310-p65-NF?B acetylation. The treatment with cells from db/m significantly reduced all the above-mentioned, but interestingly the treatment with cells from db/db mice fully restored the above alterations to normal levels. rMCs exposed to HG displayed GFAP and VEGF expression up regulated, accompanied by increase in Nox4 expression and ROS levels, and acetyl-lysine-310-p65-NF?B. SIRT1 protein expression and activity were markedly reduced in diabetic milieu conditions. The treatment with both EOC-CMs prevented all these abnormalities, but db/db EOC-CM fully restored to NG conditions. This study demonstrates that endocrine capacity of EOCs is effective in improving retinal SIRT1 pathway thus protecting the retina from diabetic milieu insult. In summary, compelling novel evidence is provided herein that either through oral administration of polyphenol enriched cocoa or cell therapy with EOCs, conferred retinal neuroprotection against diabetic insults in animal models. The identification of SIRT-1 as a potential therapeutic target in the treatment of diabetic retinopathy may provide new perspective in the pharmacological treatment of this diabetic complication / Doutorado / Clinica Medica / Doutor em Clínica Médica

<p>Stopa prematuriteta konstantno raste, a sve vi&scaron;e prevremeno rođenih beba koje prežive neonatalni i perinatalni period zahteva adekvatno sistemsko i oftalmolo&scaron;ko praćenje i tretman. Prematuritet sam po sebi, predstavlja rizik za razvoj vida. Taj rizik dodatno povećava prisustvo prematurne retinopatije i refraktivnih mana. Skrining i tretman promena na retini u sklopu prematuriteta, kao i optička korekcija refraktivnih mana obezbeđuju uslove za razvoj vida. Cilj ovog istraživanja je bio utvrditi promene biometrijskih karakteristika oka prematurusa, sa i bez prematurne retinopatije, tokom &scaron;estogodi&scaron;njeg praćenja, utvrditi refraktivni status ove dve grupe dece sa &scaron;est godina, te povezati promene biometrijskih karakteristika oka sa refrakcijom. U ispitivanje je uključeno 192 prevremeno rođena deteta (384 oka). Vr&scaron;ena su tri pregleda, u uzrastu od 3 meseca, 12 meseci i 6 godina starosti. Nakon prvog pregleda ispitivani uzorak je, u zavisnosti od nalaza na očnom dnu, podeljen na bebe sa prematurnom retinopatijom i bebe bez ovog oboljenja, a kako bi se uporedile proučavane karakteristike. U sva tri navedena uzrasta je pregledano očno dno i merene su tri glavne biometrijske karakteristike koje utiču na refraktivni status: aksijalna dužina oka, dubina prednje očne komore i debljina očnog sočiva. Sa &scaron;est godina je određena refrakcija, uključujući zakrivljenost rožnjače odnosno kornealni astigmatizam, vidna o&scaron;trina bez korekcije refraktivne mane, a zatim i nakon korekcije. Ostali neophodni podaci su dobijeni iz medicinske dokumentacije. Pokazano je da prematurna retinopatija utiče na refraktivni status i vidni ishod sa &scaron;est godina, pre svega uticajem na zakrivljenost rožnjače, aksijalnu dužinu oka i dubinu prednje očne komore, dok su promene debljine očnog sočiva bez posebnog značaja. Vidne o&scaron;trine oba oka u pred&scaron;kolskom uzrastu prevremeno rođene dece sa prematurnom retinopatijom su statistički značajno manje kada se uporede sa vidnim o&scaron;trinama prevremeno rođene dece bez retinopatije, ali razlika u ovim dvema vrednostima gubi statističku značajnost nakon optičke korekcije. Učestalost astigmatizma u prematurnoj populaciji sa 6 godina je visoka, a od onih koji imaju astigmatizam u tom uzrastu, vi&scaron;e od dve trećine je imalo prematurnu retinopatiju. Ukupna učestalost miopije, hipermetropije i emetropije u prematurnoj populaciji sa 6 godina je 18,9%, 54,7% i 13,2%, a preostalih 13,2% su anizometropi. U vi&scaron;e od trećine dece, sa prematurnom retinopatijom u najranijem uzrastu, sa &scaron;est godina postoji gubitak vidne sposobnosti, koji je najče&scaron;će blag, no može biti i potpun.</p> / <p>The number of babies born prematurely is constantly growing, so more and more of them need appropriate systemic and ophthalmologic monitoring and treatament. Prematurity itself, represents a risk for the development of vision. This risk further reinforces the presence of retinopathy of prematurity and refractive errors. Screening and treatment of retinopathy, as well as optical correction of refractive errors provide adequate conditions for the development of vision. The aim of this study was to determine changes of biometric features of eyes of the prematurely born infants, with and without retinopathy of prematurity, during the six-year follow-up. We also want to determine the refractive status of these two groups of children in the six year, and to link changes of biometric features with their eye refraction. The study included 192 prematurely born children (384 eyes). The examinations were performed at 3 months, 12 months and 6 years and they included fundus examination and measurements of the ocular axial length, anterior chamber depth and lens thickness. After dividing the subjects into two groups, based on the results of the fundus examination at 3 months, the results of the children with and without retinopathy were compared. At the age of six we also determine the refraction of eyes, including the curvature of the cornea and corneal astigmatism, visual acuity without correction of refractive error, and then after it. Other necessary data were obtained from medical documentation. It is shown that retinopathy of prematurity affects the refractive status and visual outcome at sixth year, primarily by the changes of curvature of the cornea, the axial length of the eye and the depth of the anterior chamber, while the change in the lens thickness has no significance. Visual acuity in both eyes in the preschool age preterm born children with retinopathy of prematurity are significantly less, when compared to visual acuity in preterm infants without retinopathy, but the difference in these two values lost statistical significance after optical correction. The rate of astigmatism in preterm population of six years is high, and of those who have astigmatism in this age group, more than two-thirds had retinopathy of prematurity. The overall incidence of myopia, hypermetropia and emetropia in the prematurely born, six years old children is 18.9%, 54.7% and 13.2%, and the remaining 13.2% have anisometropia. In more than a third of children with retinopathy of prematurity at an early age, with six years, there has been loss of visual acuity, which is usually mild, but it can be complete.</p>

Contexte : Le diabète de type 2 (DT2) constitue une pathologie majeure, au lourd fardeau, associ ée à de nombreuses complications, comme la rétinopathie diabétique (RD). Des facteurs modifiables, comme l’alimentation, ont déjà été identifiés pour le DT2 et la RD mais certains aspects de leurs rôles restent à préciser. Objectifs : Les objectifs de cette thèse étaient d’examiner le rôle de l’alimentation, en particulier la consommation d’acides gras (AGs), et des autres facteurs modifiables liés au mode de vie sur le risque de DT2 et de synthétiser, interpréter et analyser la relation entre l’alimentation et la RD. Résultats : Les résultats suggèrent que le rôle des AGs sur le risque de DT2 et de la RD pourrait être différent selon leur type, et même varier au sein d’un groupe comme les AG polyinsaturés (AGPI). Les résultats suggèrent aussi qu’une forte adhésion aux recommandations alimentaires n’est pas associée avec le développement d’un DT2, mais en revanche une forte adhérence aux autres recommandations de santé (concernant le tour de taille, l’activité physique et le statut tabagique) est fortement associée avec un moindre risque de DT2. On a montré qu’avoir un mode de vie sain aurait pu empêcher la survenue de plus de la moitié des cas de DT2. Conclusions : Cette thèse a permis de préciser l’importance et la complexité du rôle de l’alimentation dans le développement du DT2 et de la RD. Elle montre aussi l’impact des comportements sains dans la pathologie de DT2 et confirme que le DT2 est en grande partie, une maladie évitable. Les efforts devraient se focaliser sur la modification des comportements de santé à la fois dans la population générale et atteinte de DT2 et notamment encourager une alimentation modérée et variée. / Background : Type 2 diabetes (T2D) presents a significant health burden that is associated with many complications, such as diabetic retinopathy (DR), that further burden people with diabetes. Modifiable risk factors, such as the diet, have been identified for both T2D and DR; yet certain aspects of the role of the diet remain unclear. Objectives : The main objectives of this thesis were therefore to examine the role and impact of the diet, and in particular, the consumption of fatty acids (FAs), and other modifiable behaviours on the risk of T2D and to summarize, interpret and analyze the relationship between the diet and DR using data from both the E3N and AusDiab cohort studies. Results : The results suggest that the role of FAs on the risk of T2D and DR may differ between and within subgroups, and by individual polyunsaturated fatty acids (PUFAs). The findings also suggest that strongly adhering to national dietary guidelines is not associated with the development of T2D, but strongly adhering to other recommendations for healthy behaviours (for waist circumference, physical activity and smoking) is strongly inversely associated with T2D. Modifiable behaviour could have prevented more than half of the cases of T2D. Conclusions : This work underlines the importance and the complexity of the role of the diet in the development of T2D and DR. It also illustrates the impact of healthy behaviour in the etiology of T2D and confirms that T2D is largely preventable. Efforts should focus on the modification of multiple healthy behaviours in populations, and promote diets that are moderate and widely varied.

<p>Transforming growth factor β-activated kinase 1 (TAK1), a hub kinase at the convergence of multiple signaling pathways, is critical to the development of the central nervous system and has been found to play a role in cell death and apoptosis. TAK1 may have the potential to elucidate mechanisms of cell cycle and neurodegeneration. The Belecky-Adams laboratory has aimed to study TAK1 and its potential roles in cell cycle by studying its role in chick retinal development as well as its possible implication in the progression of diabetic retinopathy (DR). Chapter 3 includes studies that explore TAK1 in a study in chick retinal development and TAK1 in in vitro studies in retinal microglia. Using the embryonic chick, immunohistochemistry for the activated form of TAK1 (pTAK1) showed localization of pTAK1 in differentiated and progenitor cells of the retina. Using an inhibitor or TAK1 activite, (5Z)-7-Oxozeaenol, in chick eye development showed an increase in progenitor cells and a decrease in differentiated cells. This study in chick suggests TAK1 may be a critical player in the regulation of the cell cycle during retinal development. Results from experimentation in chick led to studying the potential role of TAK1 in inflammation and neurodegeneration. TAK1 has previously been implicated in cell death and apoptosis suggesting that TAK1 may be a critical player in inflammatory pathways. TAK1 has been implicated in the regulation of inflammatory factors in different parts of the CNS but has not yet been studied specifically in retina or in specific retinal cells [3, 4]. Chapter 2 includes studies from the Belecky-Adams laboratory of in vitro work with retinal microglia. Retinal microglia were treated with activators and the translocation to the nucleus of a downstream factor of TAK1 was determined: NF-kB. Treatment of retinal microglia in the presence of activators with TAKinib, an inhibitor of TAK1 activation, revealed that TAK1 inhibition reduces the activation of downstream NF-kB. Together this data suggests that TAK1 may be implicated in various systems of the body and further studies on its mechanisms may help elucidate potential therapeutic roles of the kinase.</p>