Einleitung: Die im Genom der Schweine integrierten porzinen endogenen Retroviren (PERV) gehören zu den potentiellen humanpathogenen Erregern, die eines der Risiken bei der Xenotransplantation darstellen. Für die Abschätzung des Infektionsrisikos von PERV sind drei Verfahrensweisen von Bedeutung: Erstens die Evaluierung der PERV-Freisetzung aus porzinen Zellen und Geweben; Zweitens die Etablierung eines In vivo-Infektionsmodells und Drittens ein retrospektives Screenen von Patienten. Methoden: Die PERV-Expression in Inselzellen von Schweinen der Deutschen Landrasse wurde in vitro und in vivo evaluiert. Anschließend wurde PERV auf nicht-humanen Primatenzellen passagiert. In einem zweiten Modellversuch wurde murinen Zellen in vitro und SCID-Mäusen in vivo zellfreies PERV appliziert. Schließlich wurden Seren von Patienten analysiert. Ergebnisse: Die untersuchten Inselzellen setzten keine Viruspartikel frei und konnten somit weder humane Zellen noch BALB/c-Mäuse infizieren. Die verwendeten Affenzellen produzierten infektiöses PERV mit geringer Replikation. Weder in den murinen Modellversuchen noch in den untersuchten Patienten wurde eine Übertragung von PERV beobachtet. Schlussfolgerung: Schweine der Deutschen Landrasse könnten als Ausgangsbasis für die Zucht sicherer Schweine für die Xenotransplantation dienen. Da keine Infektion verschiedener muriner Zellen mit PERV beobachtet wurde, muss angenommen werden, dass Publikationen anderer Arbeitsgruppen, die eine PERV-Infektion in SCID-Mäusen diagnostizierten, ein falsch-positives Ergebnis aufgrund von Mikrochimärismus oder aufgrund von Pseudotypisierungen mit murinen endogenen Retroviren wiedergeben. Unsere Befunde werden dadurch erhärtet, dass der Rezeptor für PERV-A auf murinen Zellen nicht exprimiert wird und diese auch in vitro nicht infiziert werden konnten. In Übereinstimmung mit den weltweit etwa 200 behandelten Patienten konnte auch in den beiden neuen Studien keine Übertragung von PERV festgestellt werden. / Objective: Porcine endogenous retroviruses (PERVs) are integrated in the porcine genome and are able to infect human cells in vitro. Therefore, PERVs are one of the possible pathogens which poses a risk for xenotransplantations. In this study three significant methods were used to evaluate the infectious risk of PERV: The release of PERV particles from porcine cells and tissues, the formation of an in vivo infection model and a retrospective screening of patients. Methods: Islet cells from german landrace pigs were co-cultivated with human cells in vitro and were transplanted in BALB/c mice in vivo. Serial passaging experiments were performed with nonhuman primate cells. Murine cells were incubated in vitro and SCID mice in vivo with PERV. Sera of patients who were treated ex vivo with porcine liver cells and who had received islet cells were investigated for antibodies against PERV. Results: No virus release were observed in german landrace islet cells, thus they were neither able to infect human cells nor BALB/c mice. The used nonhuman primate cells released low replicating PERVs. None of the murine cells could be infected by PERV and no provirus integration was observed in different SCID mice organs. PERV-specific antibodies were found in none of the investigated patients. Conclusion: German landrace pigs could be used as a source for breeding safe genetically modified pigs suitable for xenotransplantation. Since there were no detectable PERV infection of different murine cells and SCID mice, it have to be supposed, that previously reported PERV transmissions to SCID mice might be due to microchimerism or to pseudotyping of murine endogenous retroviruses. Our results were confirmed by the fact, that the receptor for PERV-A is not expressed on murine cells and that these cells could not be infected in vitro. The absence of a PERV transmission in the investigated patients, correspond to the results obtained from approximately twohundred treated patients worldwide.

A mucosite oral é um efeito colateral de quimioterapia e radioterapia de cabeça e pescoço e consiste de uma comorbidade severa com a presença de xerostomia, lesões ulcerativas, infecções secundárias e dor, passíveis de alterar e até interromper a terapia antineoplásica, diminuindo a possibilidade de controle da doença. A mucosite oral pode ser tratada com variados esquemas terapêuticos, incluindo a utilização de laser de baixa intensidade, que diferem em tipo de laser, densidade de energia, comprimento de onda utilizados, entre outros parâmetros de irradiação. Este trabalho utilizou mesmos parâmetros de irradiação laser (30 mW, 7,5 J/cm2, 0,04 mm diâmetro do feixe, 10 s por ponto, 3 pontos no dorso e 2 pontos no ventre da língua, com distância de 1 mm da superfície, 48/48 h por até 20 dias), diferindo apenas o comprimento de onda utilizado, &lambda; = 660 nm e &lambda; = 780 nm. Em seguida, os efeitos da irradiação foram comparados entre si e em relação ao grupo controle não tratado. Foi feita a padronização de modelo animal de mucosite oral em ratos utilizando radiação gama de fonte panorâmica, em dose única de 20 Gy, para avaliação uniforme dos grupos. Para tanto, foi elaborado escore clínico detalhado ponto a ponto, com correlação estatística, que se mostrou eficiente na avaliação clínica desta comorbidade. Após análise estatística dos dados coletados (p < 0,05), pode-se verificar que existem diferenças entre as terapias com laser, além de sua superioridade em relação ao grupo Controle. Os animais do grupo laser &lambda; = 660 nm apresentaram maior produção de colágeno em mucosa e fechamento de feridas mais rapidamente nos dias 14 e 20 quando comparados ao grupo laser &lambda; = 780 nm. Ambos os tratamentos foram superiores em relação ao grupo Controle. Os tratamentos laser também se mostraram superiores ao Controle na modulação da inflamação, formação de pseudomembrana e estimulação de angiogênese. Estes resultados indicam que os benefícios obtidos pelos dois comprimentos de onda, nas mesmas condições de irradiação, são diferentes, podendo ser utilizados de forma complementar em pacientes. / A well-known side effect of head and neck chemotherapy and radiotherapy is oral mucositis. This severe comorbidity presents xerostomya, ulcers, secondary infections and pain that can alter and even force to interrupt tumor-targeted therapies. Various treatments currently exist such as in particular low-level laser therapies (LLT) with different parameters including laser type, energy density, or wavelength. In the present study we propose to compare the effects of LLT at two different wavelengths, &lambda;= 660 nm and &lambda; = 780 nm, on an animal model. For this purpose we separated rats in three groups: one control group and one group for each wavelength. Apart from the wavelengths, the other laser irradiation parameters were identical: 30 mW, 7,5 J/cm2, 0,04 mm diameter, 10 s per point, 3 points on the upper side and 2 points on the lower side of the tongue , 1 mm away from the surface, 48/48 h until 20 days. An animal model of oral mucositis in rats was established by irradiation with a panoramic source of 60Co, single dose of 20 Gy, to ensure uniform evaluation of the groups. In addition, I elaborated a novel clinical point-to-point score scheme with statistical correlation that is proven to be efficient for clinical evaluation of this comorbidity. After statistical analysis using p < 0,05, we observed differences between each laser therapies and also in relation to the control group. The animals treated with &lambda; = 660 nm presented a higher amount of collagen in the mucosal region and faster wound healing when compared to the animals treated with &lambda; = 780 nm. Both laser treatments showed superior efficiency than the control group for collagen generation and wound healing speed, as well as in the inflammatory modulation, pseudomembrane production and angiogenesis stimulation. These results indicate that both laser treatments are beneficial in the wound healing process, with complementary effects depending on the wavelength. This suggests that laser therapies at various wavelengths could be simultaneously used for oral mucositis treatment.

A dengue é doença causada pelo vírus dengue (DENV), que acomete cerca de 390 milhões de pessoas no mundo, representando uma ameaça à saúde pública mundial. Até o momento não se dispõe de tratamento específico ou de vacinas para a maioria da população. Um dos maiores obstáculos para o desenvolvimento de vacinas ou para a compreensão da biologia do vírus é a falta de modelos animais que mimetizem a doença vista em humanos. Neste trabalho, o principal objetivo foi a busca de modelos alternativos de infecção com um isolado clínico (JHA1) de DENV sorotipo 2 (DENV2) capaz de infectar camundongos imonocompetentes após administração pela via intracraniana. Foram testadas duas vias alternativas de inoculação viral (intraperitoneal e intravenosa) em camundongos adultos imunocompetentes, além da via intracraniana em camundongos C57BL/6. Por fim, tentativas de mapeamento de mutações relacionadas à neurovirulência do JHA1 em camundongos imunocompetentes. Os resultados obtidos contribuem para a caracterização do JHA1 e para a busca de modelos experimentais alternativos de DENV. / Dengue fever is a disease caused by the dengue virus (DENV), which affects around 390 million people worldwide, posing a threat to global public health. To date, there is no specific treatment or vaccination for the majority of the population. One of the major obstacles to vaccine development or understanding the biology of the virus is the lack of animal models that mimic the disease seen in humans. In this work, the main objective was to search for alternative models of infection with a clinical isolate (JHA1) of DENV serotype 2 (DENV2) capable of infecting immunocompetent mice after administration by the intracranial route. Two alternative routes of viral inoculation (intraperitoneal and intravenous) were tested in immunocompetent adult mice, in addition to the intracranial route in C57BL/6 mice. Finally, attempts to map mutations related to JHA1 neurovirulence in immunocompetent mice. The results obtained contribute to the characterization of JHA1 and to the search for alternative experimental models of DENV.

O enfisema pulmonar é caracterizado pela destruição das paredes alveolares, sendo a fumaça de cigarro o principal agente etiológico. Pretendeu-se, neste estudo, comparar os efeitos terapêuticos do transplante de células mononucleares da medula óssea (BMMC) e células mesenquimais (CTM) em animais com enfisema pulmonar induzido por exposição à fumaça de cigarro. Camundongos fêmeas da linhagem C57Bl6/J foram expostos à fumaça de cigarro durante 90 dias e, 21 dias após o término do período de exposição, receberam BMMC ou CTM derivadas da medula óssea de camundongos machos da linhagem C57Bl6/J, expressando a proteína GFP. As análises morfométricas mostraram que os tratamentos com BMMC e CTM foram eficientes na recuperação do parênquima pulmonar dos animais expostos à fumaça de cigarro. Testes de fluorescência direta e PCR mostraram a migração de BMMC e CTM para o pulmão. Desta forma, pode-se concluir que, morfologicamente, a terapia celular com BMMC ou CTM é eficaz no tratamento do enfisema pulmonar resultante da exposição à fumaça de cigarro em modelo animal. / Pulmonary emphysema is characterized by destruction of alveolar walls, and the cigarette smoking is the main etiologic agent. It was intended in this study to compare the therapeutic effects of the transplantation of bone marrow mononuclear cells (BMMC) and mesenchymal stem cells (MSC) in animals with pulmonary emphysema induced by exposure to cigarette smoke. C57Bl6/J female mice were exposed to cigarette smoke for 90 days and 21 days after the end of the exposure time, received BMMC or MSC derived from bone marrow of C57Bl6/J male mice expressing the GFP protein. The morphometric analysis showed that treatments with BMMC and MSC were efficient in recovering the lung of animals exposed to cigarette smoke. Fluorescence and PCR tests showed the migration of BMMC and MSC to the lung. Thus, it is concluded the cell therapy with BMMC or CTM is morphologically effective in the treatment of pulmonary emphysema resulting from exposure to cigarette smoke in an animal model.

Objetivos: Verificar alterações na mecânica e parâmetros de morfometria pulmonar em um estudo temporal sobre enfisema em um modelo animal. Métodos:Setenta camundongos Balb / c receberam instilação nasal de solução de papaína ou salina e foram estudados no 1º, 3º, 15º, 28º e 40º dias após a instilação. Resistência das vias aéreas (Raw), Resistência do tecido (Gtis) e elastância tecidual (Htis), foram analisados. Intercepto Linear Médio (Lm), a proporção do volume de fibras elásticas e colágenas, macrófagos (MAC), o número de células que expressam MMP12 e a expressão de -isoprostano 8 no parênquima pulmonar, foram medidos. Resultados: Comparando os grupos papaína e solução salina ao longo do tempo, foi observado um aumento de Lm após o dia 28, associado a uma diminuição no Htis e Gtis. Houve um aumento na proporção do volume das fibras colágenas a partir do dia 15 até o dia 40, enquanto a proporção do volume de fibras elásticas foi aumentada somente no dia 40. Analisando o número de macrófagos, houve um aumento no dia 1 e manteve-se crescente até o dia 40. A expressão de MMP12 aumentou a partir do dia 3 até o dia 40. No entanto, a expressão de isoprostano 8 foi maior apenas no dias 1 e 3. Conclusão: Um aumento significativo no intercepto linear médio (Lm) após o dia 28 de instilação de papaína foi associado a uma piora na função pulmonar caracterizando assim o enfisema pulmonar. No entanto, no dia 40, as diferenças na morfometria foram mantidos, mas não houve diferenças na mecânica respiratória. O remodelamento da matriz extracelular observada no parênquima pulmonar no dia 40 poderia explicar estes resultados / Objectives: To verify how mechanical and morphometry parameters change in a temporal study of emphysema in an animal model. Methods: Balb/c mice received either a nasal drop of papain solution (Papa) or saline (Sal) and were studied on the 1st, 3rd, 15th, 28th and 40th days after instillation. We evaluated airway resistance (Raw), tissue damping (Gtis) and tissue elastance (Htis). Using morphometry, we measured mean linear intercept (Lm), volume proportion of elastic and collagen fibers, number of macrophages (MAC), number of cells expressing MMP12 and the expression of 8-isoprostane in parenchyma. Results: Comparing Papain and Saline groups in each time window, we observed an increase in Lm after the 28th day associated to a decrease in Htis and Gtis. The volume proportion of collagen fibers increased from the 15th to the 40th day, while the volume proportion of elastic fibers increased only on the 40th day. Analyzing the macrophages number, there was an increase on the 1st day, and it continued increasing until the 40th day. The expression of MMP12 increased from the 3rd until 40th day. However, the expression of 8-isoprostane increased only on the 1st and the 3rd day. Conclusions: A significant increase in mean linear intercept (Lm) after the 28th day of papain instillation was associated to a worsening in lung function. However, on the 40th day, differences in morphometry maintained but there were no differences in respiratory assessment. The extracellular matrix remodeling observed in lung parenchyma on the 40th day could explain these results

La fonction du ventricule droit est un déterminant majeur de la capacité fonctionnelle et du pronostic dans l’hypertension pulmonaire. La survenue dans ce contexte d’une insuffisance cardiaque droite requérant un support inotrope est associée à un taux de mortalité supérieur à 40%. Les déterminants cellulaires et moléculaires du découplage entre le coeur droit et la circulation artérielle pulmonaire sont méconnus, d’autant qu’il existe une grande hétérogénéité fonctionnelle parmi les patients soumis au même niveau de résistances vasculaires pulmonaires.Dans une première étude expérimentale, nous avons mis évidence à partir d’un modèle porcin d’hypertension pulmonaire chronique que les indices fonctionnels systoliques du ventricule droit mesurés en échocardiographie sont davantage corrélés au couplage ventriculo-artériel qu’à la performance contractile propre du ventricule droit. La réponse du ventricule droit à l’exercice ou à un stress pharmacologique a été peu documentée jusqu’à présent dans l’hypertension pulmonaire. Apartir de notre modèle porcin, nous avons montré que l’altération de la réserve contractile du ventricule droit est fortement associée au découplage ventriculo-artériel. La réserve contractile pourrait être un marqueur sensible et précoce de dysfonction ventriculaire droite. Dans une troisième étude, nous montrons la relation forte entre le couplage ventriculo-artériel et la plasticitéhémodynamique, fonctionnelle et moléculaire du ventricule droit dans un contexte de surcharge de pression chronique. Les variations d’expression de l’isoforme β de la chaîne légère de la myosine cardiaque pourraient déterminer l’efficacité du travail cardiaque droit. Nous avons par ailleurs constaté expérimentalement chez le rat que le remodelage géométrique et fonctionnel du ventricule droit en condition de surcharge barométrique chronique est associé à une infiltration macrophagique dumuscle cardiaque droit.Nos résultats physiopathologiques pourraient permettre une meilleure stratification des patients souffrant d’hypertension pulmonaire chronique. Ces mécanismes pourraient par ailleurs constituer autant de cibles thérapeutiques pour optimiser la fonction cardiaque droite lorsque la postcharge du ventricule droit n’est pas complètement corrigée, d’autant que les thérapies vaso-actives pulmonaires usuelles auraient des effets directs controversés sur le remodelage du ventricule droit. / Right ventricular function is a major determinant of functional capacity and prognosis in pulmonary hypertension. Right heart failure related to pulmonary hypertension is associated with a mortality rate up to 40% when inotrope support is necessary. Cellular and molecular determinants of right ventricular-pulmonary arterial coupling are misunderstood, while a wide functional range is remarkable among patients sharing the same degree of pulmonary vascular resistance.In a first experimental study, we showed from a porcine model of chronic pulmonary hypertension that usual non-invasive indices of right ventricular function are rather associated with ventricular-arterial coupling than with contractility. Right ventricular response to exercise or to pharmacological stress has been poorly reported in pulmonary hypertension. In our piglet model, we showed that impairment of right ventricular contractile reserve is strongly associated with ventricular-arterial uncoupling. Rightventricular reserve might be a sensitive marker of early ventricular dysfunction. In a third study, we highlighted that a strong relationship between ventricular-arterial coupling and functional and molecular plasticity of the pressure overloaded right ventricle. Gene expression of the beta-myosin heavy chain may be related to right heart efficiency. We also oberved experimentally in rats that structural and functional remodeling of the pressure overloaded right ventricle is associated withmacrophagic infiltration in the myocardium.Our pathophysiologic results could improve patient’s stratification in chronic pulmonary hypertension.These mechanisms may represent innovative targeted therapies to improve right ventricular function despite persistent elevated afterload.

La douleur neuropathique est un syndrome secondaire à une maladie ou à une lésion affectant le système nerveux somatosensoriel. Environ 30% des patients souffrant de douleurs neuropathiques présentent des troubles de l’humeur. Les causes biologiques de ces comorbidités ne sont pas clairement établies. Grâce à l’utilisation d’un modèle murin de douleur neuropathique, nous avons cherché à comprendre l’apparition des conséquences émotionnelles de cette douleur. Pour cela, nous avons cherché à identifier des régions cérébrales impliquées dans les différentes composantes et conséquences de la douleur ainsi que les modifications moléculaires y prenant place. Nous avons mis en évidence une ségrégation corticale de la douleur avec l’intégration de la composante sensorielle par le cortex insulaire postérieur d’une part et l’intégration de la composante aversive et des conséquences émotionnelles par le cortex cingulaire antérieur d’autre part. Nous avons ensuite montré l’implication de la protéine MKP-1 dans l’expression des comportements de type anxiodépressif dans notre modèle. / Neuropathic pain is defined as a pain caused by a lesion or disease of the somatosensory nervous system. Around 30% of neuropathic pain patients develop mood disorders. The biologic bases of these comorbidities are not clearly established. Using a murine model of neuropathic pain, we tried to understand the emotional consequences of neuropathic pain. Thus, we identified cerebral regions involved in the different components of pain and molecular modifications taking place in these regions. We showed a cortical separation of the pain experience with on one hand the integration of the sensory component of pain in the posterior insular cortex and on the other hand the integration of the aversive component and the emotional consequences of pain in the anterior cingulate cortex (ACC). Looking at the molecular modifications in the ACC, we showed that MKP-1, a protein able to dephosphorylate the MAPK, is involved in the development of pain-related mood disorders in our model of neuropathic pain.

La dépression majeure (DM) est une pathologie complexe et hétérogène associée avec des altérations du réseau cérébral, une dérégulation de l’axe hypothalamus-pituitaire-surrénales et avec des déficits de la neurogenèse adulte hippocampique (NHA). De nombreuses évidences pointent sur l’implication de la NHA dans les troubles de l’humeur et les troubles anxieux, ce qui a conduit à la formulation de l’ « hypothèse neurogénique », laquelle postule que des neurones néo-formés dans l’hippocampe du sujet adulte sont impliqués dans l’étiologie et dans l’efficacité du traitement de la DM. L’objectif de cette étude a été de déterminer le rôle des neurones formés à l’âge adulte après que les animaux aient été exposés à un stress ainsi que les mécanismes sous-jacents. Nos résultats suggèrent que l’augmentation de la neurogenèse est suffisante pour estomper les effets d’un stress chronique au niveau comportemental et hormonal, et donc pour induire un effet de type antidépresseur, comportementalement et physiologiquement. Les effets surviennent sans doute via le noyau du lit de la strie terminale antéro-dorsale. / Major depressive disorder (MDD) is a complex and heterogeneous disorder hypothesized to be associated with alterations in brain circuitry, dysregulations of the hypothalamic–pituitary–adrenal axis and impairments in adult hippocampal neurogenesis (AHN). Multiple lines of evidence point to the involvement of AHN in mood and anxiety disorders, leading to the formation of the “neurogenesis hypothesis”, which postulates that adult-born hippocampal neurons are involved in the etiology and treatment efficacy of MDD. The purpose of this study was to determine the role of adult-born neurons after the onset of stress exposure and the mechanism that underlies the observed results. Our results suggest that increasing neurogenesis is sufficient to buffer against the effects of chronic stress on certain behavioral and endocrine levels and thus to display antidepressant-like effects, both behaviorally and physiologically. Adult-born neurons might have exerted some of their effects via the anteromedial division of the bed nucleus of the stria terminalis (BSTMA).

Introdução: A esquizofrenia é um dos principais transtornos mentais e promove distúrbios comportamentais e cognitivos. Apesar do grande impacto social, pouco se conhece sobre a patofisiologia e etiologia da esquizofrenia. Pacientes com desordens psicóticas têm aumentado risco de desenvolver epilepsia, e pacientes com epilepsia do lobo temporal tem alta frequência de psicoses. A utilização de modelos experimentais animais de esquizofrenia e epilepsia pode ajudar a entender a neurobiologia dessas doenças e ter papel importante no desenvolvimento de novas estratégias terapêuticas. As interações complexas das crises epilépticas e quadros psicóticos tem sido aos poucos desvendadas. O óxido nítrico (NO) é um importante modulador da neurotransmissão e doadores de NO como o nitroprussiato de sódio (NPS) tem efeitos promissores em pacientes com esquizofrenia. Em modelos animais de epilepsia, NO pode ter efeito pró ou anticonvulsivante, sugerindo que o uso de NPS em pacientes com epilepsia e psicose pode não ser tão simples. Objetivos: Desenvolver um modelo animal de epilepsia e psicose comórbida, e compará-lo do ponto de vista comportamental e neuropatológico com modelos animais puros de epilepsia e esquizofrenia. Além disso, verificar a possível modulação desencadeada pelo tratamento com NPS nos modelos citados. Metodologia: Ratos Wistar machos (260-300g) foram divididos em grupos controle (SAL+SAL), epilepsia puro (PILO+SAL), esquizofrenia (SAL+QUET), epilepsia e psicose comórbida (PILO+QUET) e suas versões tratadas com NPS (SAL+SAL+NPS, PILO+SAL+NPS, SAL+QUET+NPS e PILO+QUET+NPS). Esses animais foram avaliados em diferentes aspectos comportamentais (campo aberto, teste de inibição pré- pulso, memória de reconhecimento de objeto e memória de trabalho), e filmados ao longo de 71 dias para o monitoramento de crises epilépticas espontâneas recorrentes. Também foram avaliadas a perda neuronal e a expressão de nNOS em diferentes regiões cerebrais. Resultados: NPS reduziu a frequência de crises nos animais com epilepsia e psicose comórbida quando comparados a animais não tratados. NPS também teve efeito ansiolítico no modelo animal de epilepsia com e sem psicose, e diminuiu os correlatos comportamentais de sintomas positivos dos animais no modelo de psicose e no de epilepsia + psicose, além de reverter o prejuízo no filtro sensório motor nos animais que receberam pilocarpina e quetamina. No reconhecimento de objeto o modelo de epilepsia e epilepsia+psicose o tratamento com NPS não interferiu no desempenho de memória de curto prazo, porém NPS melhorou a memória de longo prazo no modelo de psicose. NPS preservou regiões como o córtex entorrinal e subiculum nos animais epilépticos, mas a camada granular e córtex pré-límbico revelaram perda significativa nos animais controles. Alta expressão de nNOS em diferentes regiões cerebrais foram visualizadas nos animais com epilepsia, com destaque para aqueles que receberam tratamento com NPS. Conclusão: O tratamento com NPS foi efetivo na amenização de sintomas correlatos comportamentais de psicose no modelo puro de esquizofrenia e no comórbido VIII com epilepsia. Além disso, não exacerbou crises e contribuiu para diminuição delas nos animais do modelo de epilepsia e psicose, que apresentou grandes similaridades com o que é encontrado em casos clínicos. Nossos resultados sugerem que o uso do NPS pode ter resultados na melhoria dos sintomas da psicose interictal humana, assim como já observado para a esquizofrenia. / Introdution: Schizophrenia is a major mental disorder that affects 1% of young adults and has a great impact on quality of life, behavior, and cognition. Despite the high social burden, little is known about the pathophysiology and etiology of schizophrenia. Patients with psychotic disorders have increased risk of developing epilepsy, and patients with temporal lobe epilepsy have a high frequency of psychoses. The use of experimental animal models of epilepsy and schizophrenia may help to better understand the neurobiology of these diseases and play an important role in the development of potential new therapeutic strategies. The complex interactions of seizures and psychotic symptoms are being unveiled. Nitric oxide (NO) is an important modulator of neurotransmission and NO donors such as sodium nitroprusside (SNP) have shown promising effects in schizophrenic patients. In animal models of epilepsy, NO may exert pro- or anticonvulsant effects, suggesting that the use of SNP in patients with epilepsy and psychosis may not be so straightforward. Objectives: To develop an animal model of epilepsy and comorbid psychosis, and compare it from the behavioral and neuropathological point of view with pure animal models of epilepsy and schizophrenia. It was hoped, to verify the possible modulation triggered by the treatment with SNP. Metodology: Males Wistar rats weighing 260-300g were divided in controls group (SAL+SAL), pure epilepsy (PILO+SAL), schizophrenia (SAL+QUET), comorbid psychosis (PILO+KET) and their versions treated with SNP (SAL+SAL+SNP, PILO+SAL+SNP, SAL+KET+SNP and PILO+KET+SNP). These animals were evaluated in a variety of behavioral tests (open field, prepulse inhibition startle reflex, object recognition and working memory), and were filmed over 71 days to monitor spontaneous recurrent seizures. We also evaluated neuronal loss and nNOS expression in different brain regions. Results: SNP reduced seizure frequency in animals with epilepsy and psychosis when compared to those not treated with SNP. SNP also showed anxiolytic effects in the animal model of epilepsy with or without psychosis, decreased behavioral correlates of positive symptoms in the animal model of epilepsy + psychosis, and prevented sensorimotor gating deficits in epilepsy + psychosis IX model, The object recognition test showed that in epilepsy and epilepsy + psychosis models the treatment with SNP did not interfere with short-term memory performance, but SNP improved long-term memory in the psychosis model. SNP preserved brain regions like entorhinal cortex and subiculum in epileptic animals, but the granular layer and prelimbic cortex revealed significant loss in controls animals. High expression of NOS in same brain regions was observed in the animal model of epilepsy, especially in the animals receiving SNP. Conclusion: Treatment with SNP was effective in ameliorating symptoms of behavioral correlates of psychosis in the pure model of schizophrenia and the model comorbid with epilepsy. Moreover, SNP did not exacerbate seizures and reduced seizure frequency in the animal model of epilepsy + psychosis, which showed striking similarities to clinical cases. Our results suggest that the use of SNP could ameliorate symptoms of human interictal psychosis, such as those observed in schizophrenia.

With 9.9 million new dementia cases each year, Alzheimer’s and Parkinson’s disease (AD and PD) are the most prevalent form of neurodegenerative disorder (NDG) affecting the aging population. Despite years of pharmaceutical research, no cure is yet available. Most neuropathological aspects of these diseases are extremely complex but the study of the rare genetic cases allowed to model these diseases in animals and uncover key pathophysiological processes. Transgenic Drosophila NDG models have been used for in vivo studies for many years with a range of relevant phenotypes. The cellular and molecular biology of the Central Nervous System, as well as the mechanisms underlying neurodegeneration, are well conserved between Drosophila and Humans (with a 75% of human disease-related genes having homologs in flies). Most NDG studies are performed in the aging flies. However, there are reports of measurable phenotypes for a variety of AD and PD models in juvenile Drosophila melanogaster (larval stage) with an unexploited considerable potential for drug discovery and screening for this outstanding model. Here I sought to develop a new assay for research into NDGs that focus on the earliest phenotypes. During this Ph.D. project a customized crawling assay apparatus was developed, for the assessment of locomotor ability in humanised larval Drosophila (overexpressing human proteins/peptides linked to AD and PD). A locomotor phenotype was identified in larvae overexpressing different variation of Amyloid-β42, tau and α-Synuclein pan neutrally: these animals crawl on agarose surface at a reduced mean speed when compared to controls. The defect was proven partially rescuable by administration of Tacrine and Methylene Blue, renewing the importance of such models for future applications in drug discovery and screening. The motor impairment supports the hypothesis of a neurotoxic effect of the protein/peptide. Thus, to test this further, the overexpression of the human transgenes was restricted to neurons involved in larval olfaction (olfactory impairment is often the earliest symptom in PD and AD) and odour associated learning tasks (both PD and AD are characterized by severe cognitive dysfunction). Interestingly, larvae overexpressing the Amyloid-β42 ARC peptide in the Olfactory Sensory Neurons showed a subtle navigation defect during chemotaxis (in 1-Hexanol odour gradient) that could possibly be addressed to premature neural habituation to the olfactory stimulus. Furthermore, the overexpression of the peptide in the larval Mushroom Bodies influenced the performances of the animals in associative learning tasks. Lastly, using immunohistochemistry and confocal imaging techniques I showed that the gross morphology of neurons is not altered by the targeted overexpression of the Amyloid-β42 ARC. Even though physiological studies are required to characterize the chemosensory/learning defect shown by the Amyloid-β42 ARC larvae, this Ph.D. work further confirms that the effects of the overexpression of the human transgenes are robust and measurable already at larval stage. These findings may also be relevant to the development of new, fast, and cost-effective compound screening procedures, for applications in early stages of the drug discovery process.