analyzing-oxygen-consumption-rate-primary-cultured-mouse-neonatal

Analyzing Oxygen Consumption Rate in Primary Cultured Mouse Neonatal Cardiomyocytes Using an Extracellular Flux Analyzer

Mitochondria and oxidative metabolism are critical for maintaining cardiac muscle function. Research has shown that mitochondrial dysfunction is an ...

University of California San Diego

Veterans Administration Healthcare, San Diego

β1 integrins (β1) transduce mechanical signals in many cells, including cardiac myocytes (CM). Given their close localization, as well as their role in mechanotransduction and signaling, we hypothesized that caveolin (Cav) proteins might regulate integrins in the CM. β1 localization, complex formation, activation state, and signaling were analyzed using wild-type, Cav3 knockout, and Cav3 CM-specific transgenic heart and myocyte samples. Studies were performed under basal and mechanically loaded conditions. We found that: (1) β1 and Cav3 colocalize in CM and coimmunoprecipitate from CM protein lysates; (2) β1 is detected in a subset of caveolae; (3) loss of Cav3 caused reduction of β1D integrin isoform and active β1 integrin from the buoyant domains in the heart; (4) increased expression of myocyte Cav3 correlates with increased active β1 integrin in adult CM; (5) in vivo pressure overload of the wild-type heart results in increased activated integrin in buoyant membrane domains along with increased association between active integrin and Cav3; and (6) Cav3-deficient myocytes have perturbed basal and stretch mediated signaling responses. Thus, Cav3 protein can modify integrin function and mechanotransduction in the CM and intact heart.

Caveolin modulates integrin function and mechanical activation in the cardiomyocyte.

Cardiac fibrosis is one of the major components of the healing mechanism following any injury of the heart and as such may contribute to both systolic and diastolic dysfunction in a range of pathophysiologic conditions. Canonically, it can occur as part of the remodeling process that occurs following myocardial infarction or that follows as a response to pressure overload. Integrins are cell surface receptors which act in both cellular adhesion and signaling. Most importantly, in the context of the continuously contracting myocardium, they are recognized as mechanotransducers. They have been implicated in the development of fibrosis in several organs, including the heart. This review will focus on the involvement of integrins and integrin-related proteins, in cardiac fibrosis, outlining the roles of these proteins in the fibrotic responses in specific cardiac pathologies, discuss some of the common end effectors (angiotensin II, transforming growth factor beta 1 and mechanical stress) through which integrins function and finally discuss how manipulation of this set of proteins may lead to new treatments which could prove useful to alter the deleterious effects of cardiac fibrosis.

Integrins and integrin-related proteins in cardiac fibrosis.

Continuous contraction-relaxation cycles of the heart require strong and stable connections of cardiac myocytes (CMs) with the extracellular matrix (ECM) to preserve sarcolemmal integrity. CM attachment to the ECM is mediated by integrin complexes localized at the muscle adhesion sites termed costameres. The ubiquitously expressed cytoskeletal protein talin (Tln) is a component of muscle costameres that links integrins ultimately to the sarcomere. There are two talin genes, Tln1 and Tln2. Here, we tested the function of these two Tln forms in myocardium where Tln2 is the dominant isoform in postnatal CMs. Surprisingly, global deletion of Tln2 in mice caused no structural or functional changes in heart, presumably because CM Tln1 became up-regulated. Tln2 loss increased integrin activation, although levels of the muscle-specific β1D-integrin isoform were reduced by 50%. With this result, we produced mice that had simultaneous loss of both CM Tln1 and Tln2 and found that cardiac dysfunction occurred by 4 wk with 100% mortality by 6 mo. β1D integrin and other costameric proteins were lost from the CMs, and membrane integrity was compromised. Given that integrin protein reduction occurred with Tln loss, rescue of the phenotype was attempted through transgenic integrin overexpression, but this could not restore WT CM integrin levels nor improve heart function. Our results show that CM Tln2 is essential for proper β1D-integrin expression and that Tln1 can substitute for Tln2 in preserving heart function, but that loss of all Tln forms from the heart-muscle cell leads to myocyte instability and a dilated cardiomyopathy.

Robert S. Ross

Division of Cardiology,
Department of Medicine,
Division of Cardiology, Department of Medicine

University of California San Diego

Analyzing Oxygen Consumption Rate in Primary Cultured Mouse Neonatal Cardiomyocytes Using an Extracellular Flux Analyzer

Robert S. Ross

.css-o250i3{font-size:18px;font-family:Open Sans,sans-serif;line-height:21.6px;}Division of Cardiology,Department of Medicine,Division of Cardiology, Department of Medicine

University of California San Diego

Analyzing Oxygen Consumption Rate in Primary Cultured Mouse Neonatal Cardiomyocytes Using an Extracellular Flux Analyzer

Division of Cardiology,
Department of Medicine,
Division of Cardiology, Department of Medicine