Cardiac hypertrophy defines an adaptive process brought about in response to sustained increases in haemodynamic work. Cardiomyocytes undergo an initial compensatory phase in which enlargement and contractility alterations normalise wall stress and maintain adequate perfusion of organs. In pathological hypertrophy, this deteriorates to a decompensated state characterised by ventricular dysfunction and predisposition to heart failure. In contrast, physiological hypertrophy and associated enhanced cardiac functioning arising from chronic exercise training does not progress to heart failure. Determination of the molecular pathways underlying myocardial hypertrophy remains a challenge for cardiovascular research. The objective of the work presented in this thesis was to identify genes differentially expressed during pathological and physiological hypertrophy in order to enhance our knowledge of the mechanistic processes involved. A reverse Northern hybridisation method was applied to profile the expression of specifically selected genes in the hypertrophic models examined. Functional categories represented in the gene panel assembled included cardiac contractile and cytoskeletal markers, matrix metalloproteinases, vasoactive pathway factors, calcium handling genes, ion channels, cardiac regulatory factors, signalling pathway intermediates, apoptotic factors and histone deacetylases. In order to investigate pathological hypertrophy, a deoxycorticosterone acetate-salt (DOCA-salt) rat model was utilised. DOCA-salt treated rats used in this study demonstrated a 1.4-fold increase in heart weight to body weight ratio compared to controls. Impaired cardiac function indicative of a decompensated pathological phenotype in the DOCA-salt treated group was demonstrated by way of decreased chamber size, impaired myocardial compliance and significantly reduced cardiac output. Reverse Northern hybridisation analysis of 95 selected genes identified a number of candidates with differential expression in hearts of DOCA-salt treated rats. Increased gene expression was demonstrated for the collagenase MMP1 and stress-activated signal transduction factor Sin1. In contrast, the sarcoplasmic reticulum calcium ATPase SERCA-2 and anti-apoptotic factor BCL2l-10 genes exhibited decreased expression. To investigate changes in gene expression associated with physiological hypertrophy, use was made of an endurance run-trained rat model. The run-trained rats used in this study demonstrated a 24.1% increase in heart weight to body weight ratio and improvements in performance consistent with physiological cardiac adaptation. These performance indicators included improvements in systolic volume, cardiac output, myocardial compliance and bio-energetic function. Reverse Northern hybridisation expression analysis of 56 genes identified a number of differentially expressed mRNA transcripts in run-trained hypertrophied hearts. Four genes shown to demonstrate reduced expression in the run-trained rat model were interleukin-1 receptor associated kinase (IRAK1) and the developmentally expressed transcription factors Nkx-2.3, dHAND, and IRX-2. Based upon the reverse Northern hybridisation results, four genes were selected for Western blotting analysis of rat cardiac tissue. Of these, MMP1 and a putative isoform of Sin1 exhibited increased levels in DOCA-salt treated hypertrophic left ventricular tissue, results that correlate with the findings of increased mRNA expression for these two genes. Therefore, this study identified MMP1 and Sin1 as candidates involved in pathological but not physiological hypertrophy. This finding is in accord with other recent investigations demonstrating that pathological hypertrophy and physiological hypertrophy are associated with distinct molecular phenotypes. An aside to the major objective of identifying genes differentially regulated in left ventricular hypertrophy involved the application of the P19CL6 cell in vitro model of cardiomyogenesis to compare protein expression during hypertrophy and development. The Sin1 isoform, found to be up-regulated during DOCA-salt induced hypertrophy, was also shown to be more abundant in differentiating, than non-differentiating, P19CL6 cells. This result is consistent with the developing paradigm that implicates 'fetal' genes in the hypertrophic remodelling process.

Orientador: Antonio Carlos Cicogna / Resumo: Introdução: Diversos modelos experimentais têm avaliado o processo de remodelação cardíaca (RC); dentre eles, destaca-se a indução à estenose aórtica supravalvar (EAo). Os mecanismos fisiopatológicos responsáveis pela depressão da função cardíaca incluem alterações no trânsito de cálcio (Ca2+) e em suas proteínas regulatórias. O treinamento físico (TF) tem sido utilizado na terapêutica das cardiopatias. Na patologia cardíaca por sobrecarga pressórica, o TF restaura, total ou parcialmente, a atividade e/ou expressão das proteínas regulatórias do trânsito de Ca2+, otimizando o fluxo de Ca2+ intracelular e atenuando o prejuízo funcional cardíaco. Objetivo: Analisar a participação do trânsito de Ca2+ e suas proteínas reguladoras na melhoria da função cardíaca de ratos com EAo e disfunção ventricular pelo TF. Material e Métodos: Ratos Wistar machos (70-90 g), submetidos à cirurgia de EAo, foram divididos em dois grupos: controle operado (Sham) e EAo. Após 18 semanas da cirurgia, foi analisada função cardíaca para redistribuição dos grupos: não expostos ao TF (Sham, n= 36 e EAo, n= 29) e treinados (ShamTF, n= 33 e EAoTF, n= 32) durante 10 semanas. O treinamento físico aeróbio (TFa) em esteira foi realizado com velocidade equivalente ao limiar de lactato, obtida durante os testes de esforço (inicial, 4a e 7a semanas e final). A RC foi avaliada por ecocardiografia, músculo papilar e cardiomiócito isolados e macroscopia post mortem. O trânsito de cálcio miocárdico foi analisado pela e... (Resumo completo, clicar acesso eletrônico abaixo) / Abstract: Introduction: Several experimental models have been proposed for the study of cardiac remodeling (CR); among them, the induction of supravalvular aortic stenosis (AoS). The pathophysiological mechanisms responsible for the cardiac function depression include changes in calcium (Ca2+) and its regulatory proteins. Exercise training (ET) has been used in the management of cardiopathies. In cardiac pathology due to pressure overload, ET completely or partially restores the activity and/or expression of regulatory proteins of Ca2+ handling, optimizing intracellular Ca 2+ flow and attenuating cardiac functional impairment. Objective: To analyze the participation of Ca2+ handling and its regulatory proteins in the improvement of the cardiac function of rats with aortic stenosis and ventricular dysfunction by ET. Material and Methods: Male Wistar rats (70-90 g) submitted to supravalvular aortic stenosis (AoS) were divided into two groups: operated control (Sham) and aortic stenosis (AoS). After 18 weeks of the surgical procedure, cardiac function analysis was performed for redistribution of the groups: non-exposed to exercise training (Sham, n = 36 and AoS, n = 29) and trained (ShamET, n = 33 and AoSET, n = 32) for 10 weeks. The treadmill exercise training was performed with a velocity equivalent to the lactate threshold, obtained during effort tests (initial, 4th and 7th weeks, and final). CR was evaluated by echocardiography, papillary muscle and cardiomyocyte isolated and postmort... (Complete abstract click electronic access below) / Doutor

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