A ocorrência de disfunção renal em pacientes com insuficiência cardíaca descompensada é evento comum e associado a pior prognóstico. Entretanto, intervenções direcionadas a prevenção e tratamento da disfunção renal nestas circunstâncias revelaram resultados desapontadores. O objetivo deste estudo foi investigar o efeito da solução salina hipertônica (SSH) para a prevenção de disfunção renal em pacientes com insuficiência cardíaca descompensada. Trata-se de ensaio clínico duplo-cego e randomizado no qual pacientes com insuficiência cardíaca descompensada foram alocados para receber 100 ml de SSH (NaCl 7,5%) ou placebo (NaCl 0,9%) duas vezes ao dia por três dias. O desfecho primário foi a elevação igual ou superior a 0,3 mg/dl da creatinina sérica; o desfecho secundário foi a elevação de biomarcadores da função renal, e incluiu creatinina, cistatina C, lipocalina de neutrófilos associada a gelatinase (NGAL), excreção urinária da isoforma A1 do transportador de uréia, da isoforma 3 do trocador de Na+/H+ e de aquaporina 2 . Ao todo, 22 pacientes foram alocados no grupo SSH e 12 no grupo placebo. O desfecho primário ocorreu em 2 (10%) pacientes no grupo SSH e em 6 (50%) pacientes no grupo placebo (risco relativo 0,3; intervalo de confiança 95% de 0,09 a 0,98; P=0,01). Ao longo da intervenção, a concentração sérica tanto de creatinina como de cistatina C mantiveram-se estáveis no grupo HSS e elevaram-se no grupo placebo (p=0,004 e p=0,03, respectivamente). Não houve diferença estatisticamente significativa entre os grupos em relação à concentração sérica de NGAL. A expressão urinária dos transportadores de membrana de células tubulares esteve aumentada no grupo SSH em comparação ao grupo placebo, tanto para a isoforma A1 do transportador de uréia, quanto para a isoforma 3 do trocador de Na+/H+ e da isoforma 2 da aquaporina .Nossos dados permitem concluir que a administração de SSH pode atenuar a ocorrência de disfunção renal em pacientes com insuficiência cardíaca descompensada, por atuar na função tubular e na glomerular, achado com importantes implicações clínicas. / The occurrence of renal dysfunction is frequent among patients with decompensated heart failure and is associated with increased mortality. However, interventions targeted to prevention of renal dysfunction in this setting have been disappointing. The objective of this study was to investigate the effects of hypertonic saline solution for prevention of renal dysfunction in patients admitted for decompensated heart failure. This study is a double-blind randomized trial, in which patients with decompensated heart failure were assigned to receive a three-day course of 100ml of hypertonic saline solution (HSS - NaCl 7.5%) twice daily or placebo. The primary end point was an increase in serum creatinine of 0.3mg/dL or more. Main secondary end points were changes in biomarkers of renal function, including serum levels of creatinine, cystatine C, neutrophil gelatinase-associated lipocalin (NGAL) and the urinary excretion of aquaporin 2, urea transporter A1, and sodium/hydrogen exchanger 3. Altogether, 22 patients were assigned to HSS and 12 to placebo. The primary end point occurred in two (10%) patients in HSS group and six (50%) in placebo group (relative risk 0.3; 95% CI 0.09 - 0.98; P=0.01). Relative to baseline, serum creatinine and cystatin C levels were lower in HSS as compared to placebo (P=0.004 and 0.03, respectively). NGAL level was not statistically different between groups, however the urinary expression of aquaporin 2, urea transporter A1, and sodium/hydrogen exchanger 3 were significantly higher in HSS than in placebo. Therefore, the present data suggest that the administration of hypertonic saline solution to patients with decompensated heart failure may attenuate heart failure-induced acute kidney injury as indicated by improvement in both glomerular and tubular defects, a finding with important clinical implications.

Adicionalmente às alterações estruturais e funcionais observadas na insuficiência cardíaca (IC), as alterações na dinâmica do Ca2+ intracelular apresentam uma relação negativa com a contratilidade e função cardíaca. Para evitar a progressão da IC, o tratamento desta síndrome, conta atualmente com intervenções multifatoriais, incluindo a terapia farmacológica, com destaque aos -bloqueadores, e a terapia não medicamentosa incluindo o treinamento físico aeróbico, que quando realizado em intensidade adequada melhora a tolerância ao esforço e a qualidade de vida do portador de insuficiência cardíaca. É bem conhecido que os -bloqueadores melhoram a função cardíaca e dinâmica do Ca2+ intracelular na IC, além de minimizar a remodelação cardíaca, no entanto, ainda existem controvérsias sobre qual o melhor -bloqueador a ser utilizado e seus efeitos específicos na regulação de Ca2+ intracelular. Quanto aos efeitos do treinamento físico aeróbico na IC, seu efeito é reconhecido na melhora da tolerância aos esforços e qualidade de vida do paciente, o que é atribuído principalmente, a ganhos vasculares e músculos-esqueléticos. Em relação à função cardíaca, em trabalho anterior do nosso laboratório, o treinamento físico aeróbico melhorou a função cardíaca associado à melhora nos transientes de cálcio do miócito cardíaco. Como tanto o treinamento físico como os -bloqueadores apresentaram seus respectivos efeitos positivos, mas diferenciados, na presente tese estudou-se o efeito associado do treinamento físico ao tratamento com -bloqueadores sobre a função cardíaca e o fluxo de cálcio no cardiomiócito. Além disso, comparou-se qual -bloqueador (metoprolol ou carvedilol) seria mais efetivo na associação com o treinamento físico. Dividimos nossa amostra em 6 grupos, todos com IC. Dentre eles, 3 foram mantidos sedentários; salina (S), metoprolol (M) e carvedilol (C) e 3 foram treinados; treinados (T), treinados e tratados com metoprolol (MT) e treinados e tratados com carvedilol (CT). Verificamos que apenas os camundongos com IC treinados melhoraram a tolerância ao esforço, medida por teste máximo em esteira rolante, o que não foi observado no tratamento farmacológico isoladamente. Quanto à função cardíaca, observamos uma melhora da função nos grupos que realizaram treinamento físico ou foram tratados com metoprolol ou carvedilol isoladamente. Nos grupos que associaram as terapias farmacológicas e nãofarmacológica, observamos uma melhora da função apenas no grupo treinado carvedilol associada a um aumento da expressão da SERCA 2, proteína esta, envolvida na recaptação do Ca2+ para o reticulo sarcoplasmático, e, portanto, relacionada ao relaxamento da bomba cardíaca, apesar de não se observar aumento no pico do transiente de cálcio em cardiomiócitos isolados. Em conjunto, os resultados sugerem uma associação preferencial entre o treinamento físico e o carvedilol o que pode trazer implicações clínicas futuras importantes para o tratamento da IC. / In heart failure (HF), structural and functional alterations in myocardium are often paralleled by, defects in intracellular Ca2+ transients.. Within therapeutical strategies for heart failure, it is worth mentioning -blockers and aerobic exercise training. It is known that -blockers improve cardiac function and Ca2+ handling in heart failure. Indeed, -blockers present cardiac antiremodeling effects, but there are many controversies concerning which - blocker is more efficient in HF treatment and which would be its specific effects in Ca2+ regulation. In relation to aerobic exercise training effects in HF, it is important to highlight its positive effects in exercise tolerance and life quality improvement associated with vascular and skeletal muscle gains. We also observed previously that exercise training improves cardiac function associated with improved calcium transients in cardiac myocytes. Taking into consideration the positive impact of both exercise training and -blockers, presently we studied the associative effect of exercise training and -blockers on cardiac function and Ca2+ handling in cardiomyocytes in sympathetic hyperactivity induced HF in mice. We further compared which -blocker (metoprolol or carvedilol) would be more effective to be associated with exercise training. HF mice were assigned into 6 different groups, being 3 of them sedentary: saline (S), metoprolol (M) and carvedilol (C) and 3 exercisetrained: trained (T), trained and treated with metoprolol (MT), and trained and treated with carvedilol (CT). We observed that only HF mice exercise-trained improved exercise tolerance evaluated by maximum exercise test on a treadmill, which was not observed in -blocker treatment alone. Both exercise training and -blockers improved cardiac function evaluated by echocardiography, respectively. When exercise training was associated with -blockers, only HF mice exercise-trained and carvedilol-treated improved cardiac function associated with increased expression of SERCA 2 protein levels, which is involved in sarcoplasmic Ca2+ reuptake and cardiac relaxation. This response was not paralleled by changes in peak Ca2+ transients in isolated cardiac myocytes. Our results provide evidence for a superior effect of exercise training associated with carvedilol for improving cardiac function in HF mice.

Recentemente foi demonstrado que na insuficiência cardíaca (IC), a via final das doenças circulatórias e a maior causa de internação em idosos no Brasil, os danos morfo-funcionais da musculatura esquelética representam um preditor independente de mortalidade. Por outro lado, é conhecido que o treinamento físico aeróbico previne o aparecimento desses prejuízos, que potencialmente podem ter relação com alterações no transporte intracelular de Ca2+. Nesse sentido, o objetivo principal do presente estudo foi avaliar o efeito da IC e do treinamento físico aeróbico na IC em relação à função da musculatura esquelética, à expressão de proteínas que transportam Ca2+ no sóleo e no plantar (DHPRα1, DHPR α2, DHPR β1, RYR, NCX, SERCA 1, SERCA 2, parvalbumina) e à atividade da via proteolítica dependente deste íon nestes músculos (calpaína e calpastatina). Foram utilizados camundongos machos C57B7/6J controle e com inativação dos genes para os receptores α2A e α2C adrenérgicos com 7 meses de idade, quando estes apresentam IC induzida por hiperatividade simpática e 50% de mortalidade. A função muscular foi avaliada pelos testes de deambulação e resistência à inclinação. Tanto a expressão protéica como a atividade proteolítica foram avaliadas por Western blot. Os animais com IC apresentaram disfunção muscular, prejuízos nas proteínas relacionadas ao transiente de Ca2+ tanto no sóleo como no plantar, além de alterações na via proteolítica dependente deste íon em relação aos controle. O treinamento físico, por sua vez, preveniu o aparecimento dessas alterações funcionais e moleculares nos animais com IC. Em conclusão, o treinamento físico aeróbico mostrou-se uma terapia efetiva para a síndrome / Heart failure (HF) is a clinical syndrome with poor prognosis characterized by exercise intolerance, early fatigue and skeletal muscle myopathy, which has been considered an independent predictor of mortality. Conversely, aerobic exercise training prevents skeletal muscle dysfunction, which might be related to altered intracellular Ca2+ handling. Therefore, we tested whether HF would lead to alterations in skeletal musculature function related to changes in Ca2+ handling proteins expression (DHPRα1, DHPR α2, DHPR β1, RYR, NCX, SERCA 1, SERCA 2, parvalbumin) and activity of the Ca2+-dependent proteolysis (calpain and calpastatin) in soleus and plantaris muscles. The potential role of exercise training in preventing Ca2+ handling alterations was also studied. Male wild type and α2A e α2C adrenoceptor knockout (KO) mice on a C56BL/6J genetic background were studied at 7 months of age, when KO mice display HF and skeletal muscle myopathy associated with sympathetic hyperactivity and 50% of mortality. KO mice displayed skeletal muscle dysfunction paralleled by altered Ca2+ handling protein expression and Ca2+- induced proteolysis in both soleus and plantaris. Interestingly, exercise training prevented skeletal muscle dysfunction and Ca2+-induced proteolysis in both soleus and plantaris. Collectively, we provide evidence that improved net balance of Ca2+ handling proteins and decreased Ca2+-induced proteolysis upon exercise training is, at least in part, a compensatory mechanism against skeletal muscle myopathy of sympathetic hyperactivity-induced HF

A ocorrência de disfunção renal em pacientes com insuficiência cardíaca descompensada é evento comum e associado a pior prognóstico. Entretanto, intervenções direcionadas a prevenção e tratamento da disfunção renal nestas circunstâncias revelaram resultados desapontadores. O objetivo deste estudo foi investigar o efeito da solução salina hipertônica (SSH) para a prevenção de disfunção renal em pacientes com insuficiência cardíaca descompensada. Trata-se de ensaio clínico duplo-cego e randomizado no qual pacientes com insuficiência cardíaca descompensada foram alocados para receber 100 ml de SSH (NaCl 7,5%) ou placebo (NaCl 0,9%) duas vezes ao dia por três dias. O desfecho primário foi a elevação igual ou superior a 0,3 mg/dl da creatinina sérica; o desfecho secundário foi a elevação de biomarcadores da função renal, e incluiu creatinina, cistatina C, lipocalina de neutrófilos associada a gelatinase (NGAL), excreção urinária da isoforma A1 do transportador de uréia, da isoforma 3 do trocador de Na+/H+ e de aquaporina 2 . Ao todo, 22 pacientes foram alocados no grupo SSH e 12 no grupo placebo. O desfecho primário ocorreu em 2 (10%) pacientes no grupo SSH e em 6 (50%) pacientes no grupo placebo (risco relativo 0,3; intervalo de confiança 95% de 0,09 a 0,98; P=0,01). Ao longo da intervenção, a concentração sérica tanto de creatinina como de cistatina C mantiveram-se estáveis no grupo HSS e elevaram-se no grupo placebo (p=0,004 e p=0,03, respectivamente). Não houve diferença estatisticamente significativa entre os grupos em relação à concentração sérica de NGAL. A expressão urinária dos transportadores de membrana de células tubulares esteve aumentada no grupo SSH em comparação ao grupo placebo, tanto para a isoforma A1 do transportador de uréia, quanto para a isoforma 3 do trocador de Na+/H+ e da isoforma 2 da aquaporina .Nossos dados permitem concluir que a administração de SSH pode atenuar a ocorrência de disfunção renal em pacientes com insuficiência cardíaca descompensada, por atuar na função tubular e na glomerular, achado com importantes implicações clínicas. / The occurrence of renal dysfunction is frequent among patients with decompensated heart failure and is associated with increased mortality. However, interventions targeted to prevention of renal dysfunction in this setting have been disappointing. The objective of this study was to investigate the effects of hypertonic saline solution for prevention of renal dysfunction in patients admitted for decompensated heart failure. This study is a double-blind randomized trial, in which patients with decompensated heart failure were assigned to receive a three-day course of 100ml of hypertonic saline solution (HSS - NaCl 7.5%) twice daily or placebo. The primary end point was an increase in serum creatinine of 0.3mg/dL or more. Main secondary end points were changes in biomarkers of renal function, including serum levels of creatinine, cystatine C, neutrophil gelatinase-associated lipocalin (NGAL) and the urinary excretion of aquaporin 2, urea transporter A1, and sodium/hydrogen exchanger 3. Altogether, 22 patients were assigned to HSS and 12 to placebo. The primary end point occurred in two (10%) patients in HSS group and six (50%) in placebo group (relative risk 0.3; 95% CI 0.09 - 0.98; P=0.01). Relative to baseline, serum creatinine and cystatin C levels were lower in HSS as compared to placebo (P=0.004 and 0.03, respectively). NGAL level was not statistically different between groups, however the urinary expression of aquaporin 2, urea transporter A1, and sodium/hydrogen exchanger 3 were significantly higher in HSS than in placebo. Therefore, the present data suggest that the administration of hypertonic saline solution to patients with decompensated heart failure may attenuate heart failure-induced acute kidney injury as indicated by improvement in both glomerular and tubular defects, a finding with important clinical implications.

Bakgrund: Hjärtsvikt är en vanlig folksjukdom som idag ökar i prevalens, det är en av de vanligaste orsakerna till sjukhusvård, sjuklighet och dödlighet bland äldre. Sjukdomen går idag inte att bota utan kräver istället vård i form av lindring. Den kan påverka patienten på flera olika plan och innebära ett lidande. För att ge rätt stöd till rätt patient behöver dennes individuella behov och förmåga identifieras när det kommer till att handskas med påfrestningar i sårbara situationer. Syfte: Att beskriva patienters upplevelse av att leva med hjärtsvikt. Metod: En litteraturöversikt med kvalitativ design genomfördes och utgick från fjorton studier som beskrev upplevelsen ur ett patientperspektiv. Data samlades in med hjälp av databaserna CINAHL, MEDLINE och PsycInfo och analyserades med hjälp av Fribergs analysmetod.Resultat: I resultatet framkom tre huvudteman, begränsat liv i vardagen, att förlika sig med det nya livet och, behov av hjälp och stöd. Vidare sex subteman, att påverkas av andfåddhet och trötthet, emotionella och mentala reaktioner, acceptans och förändrad självbild, att behöva anpassa sig med nya strategier, stöd från nära relationer och bristande stöd från hälso- och sjukvård. Slutsats: Att söka förståelse kring upplevelsen av att leva med hjärtsvikt kan bidra till ökad kunskap kring hur patientens behov i praktiken kan tillgodoses. Särskilt då sjuksköterskan ansvarar för att främja hälsa, förebygga sjukdom, återupprätta hälsa och lindra lidande med ett holistiskt perspektiv. Sjuksköterskan kan stötta patienten genom att stärka ochstödja den individuella känslan av sammanhang. / Background: Heart failure is a common public health problem which increases in prevalence. It is a leading cause to hospitalizations, morbidity and mortality among the elderly. Since there is no cure availablethe disease requires a health care that focuses on alleviating. The patient can get affected in various ways and it can cause a suffering. To make sure proper support goes to the right patient, there is a need of identifying the individual needs and abilities when it comes to dealing with stressful events. Aim: To describe patients’ experience of living with heart failure Method: A literature review with a qualitative design was conducted including fourteen studies that focuses on the experience from a perspective of patients. Data was collected through the databasesCINAHL, MEDLINE and PsycInfo and applied analyses method was carried out through the method of Friberg. Result: The result revealed three main themes, living restricted in daily life, to reconcile with the new life, in need of help and support. Furthermore, six subthemes emerged, to be affected by shortness of breath and fatigue, emotional and mental reactions, acceptance and an altered self-image, having to adapt with new strategies, support by close relationships, and lack of support from health-care. Conclusion: Through an understanding of the experience of living with heart failure an increased knowledge can be gained about how the needs of the patients practically can be met. Especially since nurses has a responsibilityto promote health, prevent illness, restore health and alleviate suffering through a holistic perspective. Nurses can provide help and support for the patientby supporting the induvial sense of coherence.

Congestive heart failure (CHF) is associated with a significant economic burden that includes frequent emergency department visits, hospitalizations, and readmissions. The purpose of this study was to examine the differences, if any, between hospital readmission rates and insurance reimbursement rates for non-Hispanic Black and White CHF patients in California. The theoretical framework was Bandura's social cognitive theory. Secondary data for this quantitative study were obtained from the Office of Statewide Health Planning and Development and State Inpatient Databases from Healthcare Cost and Utilization for calendar year 2014-2016. A t-test and Levene's test for equality of variance were conducted on a sample of 11,905 patient records from 675 hospitals in California; the readmission discharge data and insurance reimbursement rates were analyzed by ethnicity and payer type. The results indicated that there was not a statistically significant difference between non-Hispanic Blacks as compared to non-Hispanic Whites in relation to readmission rates (M = 49.6, SD = 38.28) or insurance reimbursement rates (M = 50.88, SD = 36.52). Non-Hispanic Blacks had a higher readmission rate (36%) as compared to Whites (29%), and although these results are not significant, they support the need for healthcare professionals to develop programs that meet the needs of the community. The results of this study contribute to positive social change by providing information that healthcare professionals may be able to use to decrease CHF readmissions and improve access to care for non-Hispanic Blacks and other vulnerable patient groups.

Les nucléotides cycliques (NC) apparaissent comme des régulateurs majeurs du tonus vasculaire. Produits dans les cellules musculaires lisses (CMLs) par les cyclases, les NC agissent par l’intermédiaire des protéines effectrices qui modulent de nombreux mécanismes régulateurs de la vasomotricité. Les phoshodiestérases (PDEs) qui dégradent les NC assurent le contrôle spatiotemporel de leur réponse biologique. L’objectif de mon travail était de préciser les mécanismes par lesquels le tonus artériel est contrôlé par certaines PDEs. J’ai réalisé cette exploration en utilisant des artères coronaires ou mésentériques de rat, en situation physiologique puis dans un modèle d’insuffisance cardiaque (IC) chronique.Dans la première partie de mon travail, nous nous sommes intéressés au mécanisme par lequel les PDE3 et PDE4 régulent le tonus de l’artère coronaire.Par une approche expérimentale plus intégrée de réactivité vasculaire sur artère coronaires isolées, nous avons montré l’importance du canal potassique BK(Ca) dans l’effet relaxant des inhibiteurs de PDE3 et PDE4. De plus, nous avons constaté une contribution différentielle de ces canaux selon le mode de stimulation de la génération d’un NC, l’AMPc. A l’échelle de la CML, notre étude a permis de suggérer l’existence d’un signalosome impliquant les PDE3/PDE4 et du canal potassique BK(Ca), indiqué par la technique de Proximity Ligation Assay. En accord avec cette hypothèse, le contrôle des BK(Ca) par ces PDEs a pu être mis en évidence par la technique du Patch-Clamp. De manière intéressante, la contribution du couplage fonctionnel entre le canal BKCa et les PDE3 et PDE4 n’a pas été retrouvée dans les artères isolées de rats atteints d’IC. Ceci pourrait être expliqué par une diminution de l’expression des canaux BK(Ca) ainsi que, au moins partiellement, par une raréfaction de leur localisation à proximité de la PDE4B.Dans la deuxième partie de mon travail nous avons exploré les rôles des PDE3, PDE4 et de la PDE2 en particulier, dans la régulation du tonus de l’artère mésentérique de rats atteints d’IC, en comparaison aux rats contrôles. En effet, la contribution de la PDE2 est suggérée par la capacité d’un inhibiteur sélectif de cette enzyme à diminuer la réponse des artères isolées à une stimulation vasocontractante. Nous avons ensuite exploré les effets de cet inhibiteur dans d’autres types de réponses impliquant les NC.L’ensemble de ces travaux ajoute un niveau de complexité à la compréhension des mécanismes de la régulation de la vasomotricité des artères de résistance par les PDEs. Les altérations de cette voie de signalisation que nous avons mises en évidence dans l’IC pourraient contribuer à la dysfonction vasculaire qui accompagne la maladie. / Cyclic nucleotides (CN) appears to be important regulator of vascular tone. Produced in smooth muscle cells (SMCs) by cyclases, CN act through activation of key effectors involved in the regulation of vascular tone. Phoshodiesterases (PDEs), which degrade CN, provide spatiotemporal control of their biological responses. The aim of my work was to understand how vascular tone is controlled by some PDEs and how it is altered during heart failure (HF). For this, I used rat coronary and mesenteric arteries.In the first part of my work, we were interested in defining the contribution of PDE3 and PDE4 in the regulation of coronary tone. Using vascular reactivity technique, we demonstrate the importance of large-conductance Ca2+-activated potassium channel (BK(Ca)) channel in the relaxant effect of PDE3 and PDE4 inhibitor. Moreover, their contribution was different depending on the mode of cAMP production, a type of CN. At the cellular level, Proximity Ligation Assay experiments suggest the existence of a signalosome that involves PDE3/PDE4 andBK(Ca). According to this result, the control of BK(Ca) activity by these PDEs was demonstrated using the Patch-Clamp technique. Interestingly, the contribution of the functional coupling involving the BK(Ca) and PDE3/PDE4 is lost during HF. This may be explained by the decrease of BK(Ca) expression and [BK(Ca)-PDE4B] duplex signal in coronary arteries isolated from HF rat.In the second part of my work, we investigated the role of PDE3, PDE4 and PDE2 particularly, in the regulation of rat mesenteric artery tone during HF. In fact, PDE2 selective inhibitor, decreased the contractile response of isolated arteries. Then, we investigated the effect of PDE2 inhibition in other pathway that involve CN.Our findings underline the complexity of PDEs in the regulation of resistance artery tone. During HF, some CN signaling pathway are impaired leading to vascular dysfunction which can aggravate the pathology.

No description available.

Heart failure is a debilitating condition in which the heart cannot meet the metabolic demands of the body. Chronic β-adrenergic (β-AR) stimulation causes pathological myocardial remodeling that leads to heart failure, in part, by promoting apoptosis of cardiac myocytes. Work from our laboratory has shown that β-AR stimulated apoptosis is dependent on reactive oxygen species (ROS), but the molecular targets by which ROS mediate apoptosis is not known. One target of ROS that may contribute to activating the apoptosis pathway is the sarco-endoplasmic reticulum ATPase (SERCA2). SERCA2 is responsible for moving the large majority of intracellular calcium in the cardiac myocyte. We have identified that SERCA2 can undergo oxidative post-translational modification (OPTM) of cysteine C674: Low ROS increase activity while high ROS decreases. Since SERCA is the primary calcium transporter and is located in close proximity of the mitochondria, it is possible SERCA activity may affect the level of calcium in mitochondria, which in excess is a known activator of the intrinsic mitochondrial apoptosis pathway. Progressive loss of myocardial cells in ischemia and heart failure likely contributes to the pathogenesis of cardiomyopathy. We hypothesized that oxidation of SERCA2 at C674 increases mitochondrial calcium, thereby activating the mitochondrial apoptosis pathway. To address this thesis, we used a novel redox-insensitive SERCA2 mutation in which C674 is replaced by serine (C674S) to determine the role of oxidative inhibition of SERCA in H2O2-stimulated apoptosis in vitro. We tested our hypothesis using adult rat ventricular myocytes (ARVM) that overexpress wild type or SERCA C674 and assessed intra-organelle calcium content, mitochondrial function and activation of the apoptosis pathway. To measure mitochondrial calcium, we optimized the use of an ultrasensitive genetically-encoded calcium indicator (GECI) targeted to the mitochondria which was expressed in ARVM via adenovirus infection. Redox-insensitive SERCA C674S expressing ARVM displayed less sensitivity to H2O2-stimulated mitochondrial calcium uptake which was confirmed by measuring calcium sensitive pyruvate dehydrogenase phosphorylation status. Furthermore, SERCA C674S ARVM were protected from H2O2 -mediated apoptosis, indicated by a reduction in cytochrome c release and annexin V staining. Lastly, H2O2 treatment decreased the cytosolic ATP/ADP ratio and depolarized the mitochondrial membrane potential, however this was independent of SERCA C674 oxidation. Taken together, these experiments elucidate a novel role for SERCA2 activity in cardiac myocytes and provide a potential therapeutic target for reducing cardiac myocyte apoptosis, potentially improving cardiac function during heart failure.

No description available.