A sepse é uma síndrome clínica que ocasiona alterações hemodinâmicas promovendo hipoperfusão, disfunção orgânica e hipotensão responsiva ou não à ressuscitação volêmica. Os pacientes hipotensos não responsivos a expansão volêmica usualmente são tratados com medicações vasoativas. O emprego desses fármacos tais como noradrenalina e dopamina, nessa situação tornam-se imprescindíveis, porém não obstante o fato de restaurarem a pressão arterial podem comprometer a microcirculação. Assim, o presente estudo teve por objetivo avaliar comparativamente os efeitos da dopamina e noradrenalina para o tratamento da hipotensão decorrente de sepse em cães, por meio da imagem espectral obtida através da polarização ortogonal (OPS), correlacionando estes resultados com parâmetros hemodinâmicos metabólicos e convencionais de oxigenação. Para tanto, foram utilizados 14 cadelas em sepse grave decorrente de piometra apresentando no mínimo duas variáveis da resposta inflamatória sistêmica e no mínimo uma variável de disfunção orgânica na avaliação inicial e que foram submetidas a cirurgia de ovariosalpingohisterectomia. A microcirculação foi avaliada por meio da técnica de imagem obtida através da polarização ortogonal (OPS) da mucosa sublingual durante a cirurgia, e os animais cuja pressão arterial média (PAM) não atingisse valores superiores a 65 mmHg após administração de 15ml/kg em 15 minutos de solução de Ringer com lactato, foram randomizados em dois grupos distintos (Dopamina ou Noradrenalina) de acordo com o fármaco vasoativo a ser empregado para restabelecer a PAM para valores acima de 65 mmHg. Os agentes foram administrados em infusão contínua em dose crescente até a PAM alvo. Parâmetros hemodinâmicos, de ventilação e oxigenação bem como o índice de fluxo microvascular, densidade capilar e o escore DE BACKER, foram obtidos em diferentes tempos de avaliação. Não houve diferença estatística entre os grupos estudados nos parâmetros cardiovasculares, hemodinâmicos, ventilatórios, de oxigenação e da microcirculação encontrados com o OPS. Entretanto, no grupo Dopamina verificou-se incremento significativo dos parâmetros de densidade e fluxo capilar quando a PAM alvo foi alcançada. Por outro lado, todos os animais tratados com a noradrenalina alcançaram a PAM alvo sem outra intervenção terapêutica, enquanto que, dois animais do grupo Dopamina necessitaram de resgate de noradrenalina para o restabelecimento da PAM. Ademais a noradrenalina restabeleceu a pressão arterial mais rapidamente e com maior eficácia que a dopamina. Deste modo, conclui-se que tanto a dopamina quanto a noradrenalina quando empregadas para o tratamento de hipotensão decorrente da sepse grave/choque séptico, não prejudicam a microcirculação. / Sepsis is a clinical syndrome which promotes hemodinamics changes, resulting in hypoperfusion, organ dysfunction and hypotension, those being responsive or not to volemic resuscitation. Non-responsive patients must be treated with vasoactive drugs, such as dopamine and norepinephrine, in order to rescue the arterial pressure. However, the use of those drugs may also compromise the microvascular perfusion. The aim of this study was to evaluate the effects of dopamine and norepinephrine for the treatment of hypotension in septics dogs, using orthogonal polarization spectral imaging (OPS), correlating with metabolic, hemodynamic and oxygenation parameters. Fourteen animals with severe sepsis secondary to pyometra were included in the study. To assess the microcirculation perfusion we utilized the OPS technique in the sublingual mucosa during the surgery. The animals whose mean arterial pressure (MAP) were less than 65 mmHg after the fluid challenge Ringer lactate solution (15ml/kg in 15 minutes) were randomly allocated in two groups, according to the vasoactive drugs to be used (dopamine or norepinephrine). Those medications were administered through the infusion pump aiming to rescue the MAP to values greater than 65mmHg. Hemodynamic, ventilator and oxygenation parameters as well as the microvascular flow, capillary density and the DE BACKER score were obtained in different time points. No statistical difference between the groups regarding the following parameters was observed: cardiovascular, hemodynamic, ventilatory and microcirculation. In Dopamine group, there was a significant increase of capillary density and flow when the MAP goal was achieved. In the Norepinephrine group all animal reached the MAP goal without need of another intervention, while in the dopamine group two animals needed rescue medication with norepinephrine aiming to achieve the MAP goal. Norepinephrine recovered the pressure faster and more effectively when compared to dopamine. Finally, the use of dopamine and norepinephrine in hypotension treatment in severe sepsis/ septic shock do not impair the microcirculation.

Vascular cognitive impairment describes a heterogeneous condition in which cognitive decline is precipitated by underlying cerebrovascular dysfunction. Ageing, as well as vascular diseases such as hypertension, stroke, cerebral small vessel disease and cerebral amyloid angiopathy, are risk factors for vascular cognitive impairment. The precise mechanisms by which these conditions impact the cerebral vasculature to drive cognitive decline, however, are unknown. Previous research has indicated that vascular risk factors can lead to microvascular oxidative stress, inflammation and endothelial dysfunction that can lead to tissue hypoperfusion, the development of white and grey matter vascular lesions (microinfarcts and microbleeds) and cognitive impairment. It was hypothesised that ageing, a prominent risk factor for cognitive decline, would induce impairments on neurovascular coupling resulting from neurovascular unit disruption. It was further hypothesised that induction of chronic cerebral hypoperfusion would mediate neurovascular dysfunction and vascular lesion development through increased oxidative stress, resulting in cognitive decline. Finally, it was also hypothesised that neurovascular impairments resulting from ageing and chronic cerebral hypoperfusion would be exacerbated in the presence of amyloid deposition. Four studies were performed in order to test these hypotheses. Vascular risk factors can be reproduced using experimental mouse models and provide a valuable basis in which to test hypotheses and therapeutic interventions. As such, a primary aim of this thesis was to develop and validate sensitive MRI approaches that would allow the detection of vascular alterations in vivo. In the first series of studies, MRI techniques to assess resting cerebral blood flow, vessel number, vascular lesions and inflammation in experimental mice were validated using established in vivo and ex vivo techniques, so that these techniques could be used in subsequent studies for vascular assessments in vivo. Arterial spin labelling was developed to assess resting cerebral blood flow, and was able to detect reductions in blood flow following cerebral hypoperfusion that correlated well with those obtained from laser speckle imaging. Q-map imaging was able to detect reductions in vessel number in acute lesions, and in non-lesioned mice measures of vessel number correlated well with histopathological measures. Structural T2 imaging was performed in order to detect ischaemic and haemorrhagic lesions in chronically hypoperfused mice, and was validated using H&E and Perls’ staining. Finally, contrast-enhanced T2* imaging was used to detect iron oxide uptake by macrophages in the brains of hypoperfused mice, which was further validated by the identification of iron-containing macrophages in immunostained brain sections. The second study was conducted to test the hypothesis that ageing would impair neurovascular unit function and structure, and that these impairments would be exacerbated in the presence of amyloid pathology. The aim of the study was to incorporate previously developed in vivo imaging approaches in the assessment of vascular function and alterations in neurovascular unit structure in both wild type and TgSwDI mice. As predicted, ageing caused a pronounced deficit on measures of neurovascular coupling, however this was not exacerbated by accumulation of amyloid in TgSwDI mice and was not associated with alterations in baseline blood flow measured by arterial spin labelling. Structural assessment of the neurovascular unit revealed a loss of contact between astrocytic endfeet and vasculature, which was significantly associated with the impairment on neurovascular coupling, in addition to other markers of breakdown of the neurovascular unit such as loss of pericyte coverage and microglial activation. Age and thalamic vascular amyloid accumulation were also associated with an increase in the NADPH oxidase (NOX) subunit p47, indicative of increased oxidative stress. Data from this experiment indicate that ageing can profoundly impair neurovascular coupling, mediated by gliosis and loss of astrocytic contacts with vasculature. The third study aimed to test the hypothesis that chronic cerebral hypoperfusion (a prominent early feature of vascular cognitive impairment) would impair vascular function and induce the development of vascular lesions and cognitive decline. The impact of hypoperfusion on neurovascular coupling, ischaemic and haemorrhagic lesion burden and cognition was investigated in wild type and TgSwDI mice. Hypoperfusion induced deficits on neurovascular coupling, increased lesion burden and inflammation assessed with T2 and contrast-enhanced T2* imaging, and caused impairment on measures of learning and memory. Hypoperfusion was also associated with an increase in the levels of NOX2, NOX4 and 3-NT at 3 months following surgery, indicating persistent reactive oxygen species production and oxidative damage in hypoperfused mice. The findings from this study indicate that vascular dysfunction and cognitive impairment following hypoperfusion may be mediated by increased NADPH oxidase activity and resulting oxidative stress. The previous studies indicated that markers of oxidative stress were induced in response to ageing, vascular amyloid accumulation and cerebral hypoperfusion. The final study sought to determine whether increased NOX activity mediates downstream pathological effects on vascular function, vascular lesion development and cognitive decline following hypoperfusion. NOX activity was inhibited pharmacologically by administration of apocynin to hypoperfused TgSwDI mice for 3 months following surgery. Treatment with apocynin significantly restored neurovascular coupling to a level similar to sham-operated mice, and there was a trend toward reduction of ischaemic vascular lesions. However, it was unable to rescue the prominent inflammatory response or decline in cognitive ability, as apocynin-treated mice were no different on these measures to non-treated hypoperfused mice. The data indicate that whilst inhibiting NOX may have potential therapeutic value in improving vascular function, additional interventions, for example to reduce inflammation, may also be required in order to prevent cognitive decline. Overall, the work outlined within the thesis indicate that vascular risk factors of ageing, cerebral amyloid angiopathy and cerebral hypoperfusion may converge on common pathways involving oxidative stress and increased inflammation in order to drive vascular dysfunction and lead to cognitive decline. Inhibition of NOX activity was able to rescue vascular function, however the results indicate that this was not sufficient to protect against cognitive impairment, suggesting additional therapeutic targets may need to be sought in order to fully preserve vascular health and prevent cognitive decline.

A hipoperfusão cerebral crônica (HCC) é um importante fator de risco para o declínio cognitivo e outras disfunções cerebrais, tais como Doença de Alzheimer e Demência Vascular. O objetivo deste estudo foi investigar o efeito neuroprotetor do Resveratrol (RSV), avaliando parâmetros comportamentais, bioquímicos e morfológicos, em um modelo experimental de Demência Vascular. Ratos Wistar adultos foram submetidos ao modelo modificado da HCC através da oclusão permanente de 2 vasos (2VO) e tratamento diário, iniciado uma hora após oclusão permanente dos vasos, com injeções intraperitoneais (20 mg/kg) de RSV durante 7 dias. Os testes comportamentais foram realizados entre o 35° e 45° dias após a cirurgia - 2VO, através da tarefa do labirinto aquático de Morris, na qual os animais foram avaliados quanto ao desempenho da memória espacial. Ao final dos testes comportamentais, um grupo dos animais foi perfundido transcardiacamente para análise histológica, outro grupo foi submetido à eutanásia em 3 tempos (3, 14 e 45 dias após a lesão isquêmica) para avaliação da expressão de NGF (fator de crescimento do nervo). Os resultados demonstraram que o tratamento com RSV atenuou significativamente a morte das células piramidais na região CA1 do hipocampo e preveniu o déficit da memória espacial. O aumento da expressão de NGF foi evidenciado no 3° dia após indução da HCC em todos os animais isquêmicos e no 45° dia após indução da HCC nos animais tratados com RSV. Com base nesses dados, hipotetizamos que o aumento, em longo prazo, na expressão de NGF no hipocampo após a HCC pode caracterizar uma das vias envolvidas nos mecanismos neuroprotetores do RSV. / Chronic cerebral hypoperfusion (CCH) is an important risk factor for cognitive decline and other brain dysfunctions, such as Alzheimer’s Disease and Vascular Dementia. The aim of the present study was to investigate the neuroprotective effect of Resveratrol (RSV) on behavioral, biochemical and morphological parameters in an experimental model of Vascular Dementia. Adult Wistar rats were submitted to the CCH modified model by means of permanent 2-vessel occlusion (2VO) and daily treatment, initiated one hour after permanent vessel occlusion, with intraperitoneal injections (20 mg/kg) of RSV for 7 days. Behavioral testing was performed between the 35th and the 45th day after 2VO surgery in the Morris Water Maze task, allowing for the evaluation of spatial memory function. At the end of the behavioral assessment, half of the animals were transcardially perfused for histological analysis and the remaining were euthanized in 3 times (3. 14 and 45 days after ischemic injury) for NGF expression evaluation (neural growth factor). Results demonstrate that the treatment with RSV significantly attenuated pyramidal cell death in CA1 hippocampal field and prevented spatial memory impairment. The increase of NGF expression was evidenced on the 3rd day after CCH induction in all ischemic animals and on the 45th day after CCH induction in animals treated with RSV. On the basis of these data, we hypothesize that the long term increase in NGF expression in the hippocampus after CCH may characterize one pathway involved in the neuroprotective mechanisms of RSV.

No paciente séptico, utiliza-se como tratamento inicial a reposição volêmica com o objetivo de restabelecer a pressão arterial e consequentemente a perfusão tecidual. Os pacientes não responsivos a expansão volêmica usualmente são tratados com medicações vasoativas. O emprego desses fármacos tais como noradrenalina, nessa situação, torna-se imprescindível, porém a hiporresponsividade do sistema adrenérgico é um obstáculo rotineiro em pacientes sépticos. A vasopressina aparece como uma alternativa, tanto como fármaco de primeira escolha como resgate quando o tratamento com vasoativos adrenérgicos falha. A avaliação da microcirculação é imprescindível visto a sua importância na patogênese da sepse, e no acompanhamento das diferentes terapias. Assim sendo, o presente projeto tem por objetivo avaliar o uso da vasopressina e da noradrenalina no tratamento da hipotensão de cães em sepse decorrente de piometra por meio imagem espectral obtida através da polarização ortogonal (OPS) e sobre variáveis hemodinâmicas, bem como sobre parâmetros de oxigenação e ventilação. Foram utilizados 13 cães em sepse grave apresentando no mínimo duas variáveis da resposta inflamatória sistêmica e no mínimo uma variável de disfunção orgânica na avaliação inicial. Em todos os animais foi realizada ressuscitação volêmica inicial com 15ml/kg em 15 minutos de solução de Ringer com lactato. Caso durante a anestesia a pressão arterial média não assumir valores superiores a 65 mmHg e a pressão venosa central não variasse 2mmHg ou apresentasse valores superiores a 8 mmHg, os animais foram distribuídos em dois grupos. O Grupo VASO recebeu inicialmente 0,0002UI/kg/min de vasopressina e o Grupo NORA 0,05 mcg/kg/min noradrenalina, podendo ter o incremento de 0,0002U/kg/min e 0,02 mcg/kg/min da dose inicial, respectivamente, com o objetivo até se atingir a PAM acima de 65mmHg. Foram confrontados os parâmetros de valores de densidade e fluxo encontrados com o OPS nos dois grupos, bem como dados hemodinâmicos e de ventilação. As imagens coletadas utilizando o OPS foram processadas e analisadas por software especifico. Nao houve diferenca estatistica entre os grupos estudados nos parametros, hemodinamicos, ventilatorios, de oxigenacao e da microcirculacao encontrados com o OPS. A frequência cardíaca foi menor no grupo VASO no momento TG quando comparada ao grupo NORA. Os parametros de densidade e fluxo capilar não diferiram do basal em nenhum dos grupos. Deste modo, conclui-se que tanto a vasopressina quanto a noradrenalina quando empregadas para o tratamento de hipotensao decorrente da sepse grave/choque septico, nao prejudicam a microcirculacao / In septic patients, volume replacement is used as initial treatment in order to restore blood pressure and consequently the tissue perfusion. Nonresponders patients to the increase in preload are usually treated with vasoactive medications. Those agents such as norepinephrine, in this situation, it is essential, but the hyporesponsiveness of the adrenergic system is a common obstacle in septic patients. Vasopressin is an alternative, both like the drug of choice as rescue when treatment of adrenergic hyporesponsiveness. The evaluation of microcirculation is essential for its importance in the pathogenesis of sepsis, and to guide the different therapies. The aim of this project is to evaluate the use of vasopressin and norepinephrine in the treatment of hypotension in sepsis in dogs due to pyometra through spectral image obtained by orthogonal polarization (OPS) and on hemodynamic variables, as well as oxygenation and ventilation parameters. Thirteen dogs in severe sepsis were used, presenting at least two variables of systemic inflammatory response and at least one organ dysfunction variable at baseline. In all animals was performed initial volume resuscitation with 15ml / kg in 15 minutes of Ringer\'s lactate solution. If during anesthesia mean arterial pressure not assume values greater than 65 mmHg and central venous pressure did not vary 2 mmHg or present values greater than 8 mmHg, the animals were divided into two groups. The Group VASO received 0,0002UI / kg / min of vasopressin and Group NORA 0.1 mcg / kg / min of noradrenaline, may have increment 0,0002U / kg / min and 0. 1mcg / kg / min initial dose, respectively, in order to achieve MAP above of 65 mmHg. The density values of parameters were compared and found flow with OPS in both groups, and hemodynamic data and ventilation. The images collected using OPS were processed and analyzed by specific software. There was no statistical difference between the groups studied in the parameters, hemodynamic, ventilation, oxygenation and microcirculation found with OPS. The heart rate was lower in group VASO in TG moment compared to NORA group. The density and capillary flow parameters from baseline were similar in all groups. Thus, it is concluded that both noradrenaline and vasopressin when used to treat hypertension caused by severe / sepsis, septic shock, do not impair the microcirculation

A hipoperfusão cerebral crônica (HCC) é um importante fator de risco para o declínio cognitivo e outras disfunções cerebrais, tais como Doença de Alzheimer e Demência Vascular. O objetivo deste estudo foi investigar o efeito neuroprotetor do Resveratrol (RSV), avaliando parâmetros comportamentais, bioquímicos e morfológicos, em um modelo experimental de Demência Vascular. Ratos Wistar adultos foram submetidos ao modelo modificado da HCC através da oclusão permanente de 2 vasos (2VO) e tratamento diário, iniciado uma hora após oclusão permanente dos vasos, com injeções intraperitoneais (20 mg/kg) de RSV durante 7 dias. Os testes comportamentais foram realizados entre o 35° e 45° dias após a cirurgia - 2VO, através da tarefa do labirinto aquático de Morris, na qual os animais foram avaliados quanto ao desempenho da memória espacial. Ao final dos testes comportamentais, um grupo dos animais foi perfundido transcardiacamente para análise histológica, outro grupo foi submetido à eutanásia em 3 tempos (3, 14 e 45 dias após a lesão isquêmica) para avaliação da expressão de NGF (fator de crescimento do nervo). Os resultados demonstraram que o tratamento com RSV atenuou significativamente a morte das células piramidais na região CA1 do hipocampo e preveniu o déficit da memória espacial. O aumento da expressão de NGF foi evidenciado no 3° dia após indução da HCC em todos os animais isquêmicos e no 45° dia após indução da HCC nos animais tratados com RSV. Com base nesses dados, hipotetizamos que o aumento, em longo prazo, na expressão de NGF no hipocampo após a HCC pode caracterizar uma das vias envolvidas nos mecanismos neuroprotetores do RSV. / Chronic cerebral hypoperfusion (CCH) is an important risk factor for cognitive decline and other brain dysfunctions, such as Alzheimer’s Disease and Vascular Dementia. The aim of the present study was to investigate the neuroprotective effect of Resveratrol (RSV) on behavioral, biochemical and morphological parameters in an experimental model of Vascular Dementia. Adult Wistar rats were submitted to the CCH modified model by means of permanent 2-vessel occlusion (2VO) and daily treatment, initiated one hour after permanent vessel occlusion, with intraperitoneal injections (20 mg/kg) of RSV for 7 days. Behavioral testing was performed between the 35th and the 45th day after 2VO surgery in the Morris Water Maze task, allowing for the evaluation of spatial memory function. At the end of the behavioral assessment, half of the animals were transcardially perfused for histological analysis and the remaining were euthanized in 3 times (3. 14 and 45 days after ischemic injury) for NGF expression evaluation (neural growth factor). Results demonstrate that the treatment with RSV significantly attenuated pyramidal cell death in CA1 hippocampal field and prevented spatial memory impairment. The increase of NGF expression was evidenced on the 3rd day after CCH induction in all ischemic animals and on the 45th day after CCH induction in animals treated with RSV. On the basis of these data, we hypothesize that the long term increase in NGF expression in the hippocampus after CCH may characterize one pathway involved in the neuroprotective mechanisms of RSV.

A hipoperfusão cerebral crônica (HCC) é um importante fator de risco para o declínio cognitivo e outras disfunções cerebrais, tais como Doença de Alzheimer e Demência Vascular. O objetivo deste estudo foi investigar o efeito neuroprotetor do Resveratrol (RSV), avaliando parâmetros comportamentais, bioquímicos e morfológicos, em um modelo experimental de Demência Vascular. Ratos Wistar adultos foram submetidos ao modelo modificado da HCC através da oclusão permanente de 2 vasos (2VO) e tratamento diário, iniciado uma hora após oclusão permanente dos vasos, com injeções intraperitoneais (20 mg/kg) de RSV durante 7 dias. Os testes comportamentais foram realizados entre o 35° e 45° dias após a cirurgia - 2VO, através da tarefa do labirinto aquático de Morris, na qual os animais foram avaliados quanto ao desempenho da memória espacial. Ao final dos testes comportamentais, um grupo dos animais foi perfundido transcardiacamente para análise histológica, outro grupo foi submetido à eutanásia em 3 tempos (3, 14 e 45 dias após a lesão isquêmica) para avaliação da expressão de NGF (fator de crescimento do nervo). Os resultados demonstraram que o tratamento com RSV atenuou significativamente a morte das células piramidais na região CA1 do hipocampo e preveniu o déficit da memória espacial. O aumento da expressão de NGF foi evidenciado no 3° dia após indução da HCC em todos os animais isquêmicos e no 45° dia após indução da HCC nos animais tratados com RSV. Com base nesses dados, hipotetizamos que o aumento, em longo prazo, na expressão de NGF no hipocampo após a HCC pode caracterizar uma das vias envolvidas nos mecanismos neuroprotetores do RSV. / Chronic cerebral hypoperfusion (CCH) is an important risk factor for cognitive decline and other brain dysfunctions, such as Alzheimer’s Disease and Vascular Dementia. The aim of the present study was to investigate the neuroprotective effect of Resveratrol (RSV) on behavioral, biochemical and morphological parameters in an experimental model of Vascular Dementia. Adult Wistar rats were submitted to the CCH modified model by means of permanent 2-vessel occlusion (2VO) and daily treatment, initiated one hour after permanent vessel occlusion, with intraperitoneal injections (20 mg/kg) of RSV for 7 days. Behavioral testing was performed between the 35th and the 45th day after 2VO surgery in the Morris Water Maze task, allowing for the evaluation of spatial memory function. At the end of the behavioral assessment, half of the animals were transcardially perfused for histological analysis and the remaining were euthanized in 3 times (3. 14 and 45 days after ischemic injury) for NGF expression evaluation (neural growth factor). Results demonstrate that the treatment with RSV significantly attenuated pyramidal cell death in CA1 hippocampal field and prevented spatial memory impairment. The increase of NGF expression was evidenced on the 3rd day after CCH induction in all ischemic animals and on the 45th day after CCH induction in animals treated with RSV. On the basis of these data, we hypothesize that the long term increase in NGF expression in the hippocampus after CCH may characterize one pathway involved in the neuroprotective mechanisms of RSV.

A sepse é uma síndrome clínica que ocasiona alterações hemodinâmicas promovendo hipoperfusão, disfunção orgânica e hipotensão responsiva ou não à ressuscitação volêmica. Os pacientes hipotensos não responsivos a expansão volêmica usualmente são tratados com medicações vasoativas. O emprego desses fármacos tais como noradrenalina e dopamina, nessa situação tornam-se imprescindíveis, porém não obstante o fato de restaurarem a pressão arterial podem comprometer a microcirculação. Assim, o presente estudo teve por objetivo avaliar comparativamente os efeitos da dopamina e noradrenalina para o tratamento da hipotensão decorrente de sepse em cães, por meio da imagem espectral obtida através da polarização ortogonal (OPS), correlacionando estes resultados com parâmetros hemodinâmicos metabólicos e convencionais de oxigenação. Para tanto, foram utilizados 14 cadelas em sepse grave decorrente de piometra apresentando no mínimo duas variáveis da resposta inflamatória sistêmica e no mínimo uma variável de disfunção orgânica na avaliação inicial e que foram submetidas a cirurgia de ovariosalpingohisterectomia. A microcirculação foi avaliada por meio da técnica de imagem obtida através da polarização ortogonal (OPS) da mucosa sublingual durante a cirurgia, e os animais cuja pressão arterial média (PAM) não atingisse valores superiores a 65 mmHg após administração de 15ml/kg em 15 minutos de solução de Ringer com lactato, foram randomizados em dois grupos distintos (Dopamina ou Noradrenalina) de acordo com o fármaco vasoativo a ser empregado para restabelecer a PAM para valores acima de 65 mmHg. Os agentes foram administrados em infusão contínua em dose crescente até a PAM alvo. Parâmetros hemodinâmicos, de ventilação e oxigenação bem como o índice de fluxo microvascular, densidade capilar e o escore DE BACKER, foram obtidos em diferentes tempos de avaliação. Não houve diferença estatística entre os grupos estudados nos parâmetros cardiovasculares, hemodinâmicos, ventilatórios, de oxigenação e da microcirculação encontrados com o OPS. Entretanto, no grupo Dopamina verificou-se incremento significativo dos parâmetros de densidade e fluxo capilar quando a PAM alvo foi alcançada. Por outro lado, todos os animais tratados com a noradrenalina alcançaram a PAM alvo sem outra intervenção terapêutica, enquanto que, dois animais do grupo Dopamina necessitaram de resgate de noradrenalina para o restabelecimento da PAM. Ademais a noradrenalina restabeleceu a pressão arterial mais rapidamente e com maior eficácia que a dopamina. Deste modo, conclui-se que tanto a dopamina quanto a noradrenalina quando empregadas para o tratamento de hipotensão decorrente da sepse grave/choque séptico, não prejudicam a microcirculação. / Sepsis is a clinical syndrome which promotes hemodinamics changes, resulting in hypoperfusion, organ dysfunction and hypotension, those being responsive or not to volemic resuscitation. Non-responsive patients must be treated with vasoactive drugs, such as dopamine and norepinephrine, in order to rescue the arterial pressure. However, the use of those drugs may also compromise the microvascular perfusion. The aim of this study was to evaluate the effects of dopamine and norepinephrine for the treatment of hypotension in septics dogs, using orthogonal polarization spectral imaging (OPS), correlating with metabolic, hemodynamic and oxygenation parameters. Fourteen animals with severe sepsis secondary to pyometra were included in the study. To assess the microcirculation perfusion we utilized the OPS technique in the sublingual mucosa during the surgery. The animals whose mean arterial pressure (MAP) were less than 65 mmHg after the fluid challenge Ringer lactate solution (15ml/kg in 15 minutes) were randomly allocated in two groups, according to the vasoactive drugs to be used (dopamine or norepinephrine). Those medications were administered through the infusion pump aiming to rescue the MAP to values greater than 65mmHg. Hemodynamic, ventilator and oxygenation parameters as well as the microvascular flow, capillary density and the DE BACKER score were obtained in different time points. No statistical difference between the groups regarding the following parameters was observed: cardiovascular, hemodynamic, ventilatory and microcirculation. In Dopamine group, there was a significant increase of capillary density and flow when the MAP goal was achieved. In the Norepinephrine group all animal reached the MAP goal without need of another intervention, while in the dopamine group two animals needed rescue medication with norepinephrine aiming to achieve the MAP goal. Norepinephrine recovered the pressure faster and more effectively when compared to dopamine. Finally, the use of dopamine and norepinephrine in hypotension treatment in severe sepsis/ septic shock do not impair the microcirculation.

Cerebral small vessel pathology is now known to be associated with the development of cognitive impairment and mild motor impairments such as gait disturbance in a variety of neurodegenerative diseases. This dissertation explores the hypothesis that blood brain barrier dysfunction is an early event in cerebral ischemia and contributes to the development of cerebral small vessel disease (CSVD). A common rodent model of CSVD is permanent bilateral common carotid artery occlusion in the rat. This model was used to study several aspects of the progression of CSVD including the timecourse of blood brain barrier permeability changes following the onset of ischemia, gait disturbance, the expression of tight junction proteins and cytokine expression. It was determined that BBB permeability was elevated for 2 weeks following BCCAO and ischemic rats displayed lower gait velocity. There was no change in expression of TJ proteins. However, ischemic rats had higher levels of some proinflammatory cytokines and chemokines in brain tissue with no obvious changes in plasma levels. The mechanisms underlying the increase in BBB permeability were studied in vitro using artificial barriers made of confluent rat brain microvascular endothelial cells. Cerebral ischemia has been reported to cause an increase in plasma toxicity, likely by elevating the numbers of circulating microparticles (MPs). MPs isolated from the plasma of ischemic rats were applied to artificial barriers where it was found that they act mainly as vectors of TNF-α signaling. MPs induce activation of caspase-3 and the Rho/Rho kinase pathways. It is concluded that most of the increase in barrier permeability is due to apoptosis and disassembly of actin cytoskeleton and disruption of adherens junctions IV and not an increase in transcellular transport. The effects of treatment with the type III phosphodiesterase inhibitor cilostazol on dye extravasation in the brain, glial activation, white matter damage and motor performance were evaluated. It was determined that cilostazol could improve the increased BBB permeability and gait disturbance and microglial activation in optic tract following BCCAO. Also, the effects of treatment with cilostazol on plasma toxicity in vivo (24h and 14d following BCCAO) and artificial barriers (in vitro) were assessed. It was found that cilostazol could reduce plasma toxicity at 24h and improve increased endothelial barrier permeability that is induced by MP treatment respectively. In summary BBB dysfunction occurs in the rat model of chronic cerebral hypoperfusion with no differences in expression of TJ proteins. There is a mild motor disturbance in the form of lower gait velocity following BCCAO. Cytokines released in brain tissue may be associated with pathological consequences following BCCAO while there is no significant difference in plasma levels and circulating MPs may play a role in BBB dysfunction.

京都大学 / 0048 / 新制・課程博士 / 博士(医学) / 甲第22336号 / 医博第4577号 / 新制||医||1041(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 高橋 淳, 教授 渡邉 大, 教授 伊佐 正 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Arterial spin labeling is a completely non-invasive method for blood-flow measurement techniques. Alzheimer's disease pathology includes microvascular abnormalities in addition to practically all risk factors having a vascular component that reduces cerebral perfusion. Hemodynamic parameters of cerebral blood flow and arterial transit time were estimated via single-compartment modeling of pseudo continuous arterial spin labeling data and neurocognitive test scores (Alzheimer's disease assessment scale and mini-mental state examination) were compared between a group of healthy (N=20) and early Alzheimer's disease (N=25) subjects before and six months after the Alzheimer's subjects began treatment with cholinesterase inhibitors. The early Alzheimer's group showed improved CBF after 6 months' treatment in every Alzheimer's-prone region except the medial and lateral temporal lobes. No difference in arterial transit time was found between groups, indicating that the pathophysiological process causing hypoperfusion in Alzheimer's disease may differ from vascular dementia.