Introduction: the ErbB-Neuregulin1 (Nrg1) axis is essential for both cardiac development and maintenance of adult heart function. We previously demonstrated that, in adult rat ventricular cardiomyocytes (ARVCMs), acute treatment with Nrg1ß1 rapidly improves intracellular calcium recovery by activating PI3K/Akt pathway, which in turn increases endothelial nitric oxide synthase (eNOS) phosphorylation, NO production and cyclic GMP (cGMP)/protein kinase G (PKG) activation. Besides known effects in protection against oxidative stress and cytotoxic agents, recently Nrg1 has also been proposed as an important cardioprotective factor in the setting of myocardial ischemia-reperfusion (I/R) injury. PI3K/Akt and cGMP/PKG cascades are known to improve cardiomyocyte survival, calcium homeostasis and mitochondrial function during I/R events; in this perspective, the aim of this study was to assess the role of Nrg1/ErbB axis in acute cardiac protection against I/R injury, in terms of control of both calcium handling and mitochondrial activity. Methods: the role of Nrg1ß1 (12.5 nmol/L) was studied on isolated ARVCMs subjected to simulated I/R experiments; the peptide was administered during the last phase of ischemia and reperfusion to mimic pharmacological Post Conditioning. Cell survival and hypercontracture were evaluated with propidium iodide nuclear staining; Indo-1 probe was used to evaluate intracellular calcium transients in cardiomyocytes subjected to electrical field stimulation; mitochondrial membrane potential was analyzed through JC-1 fluorescent probe; Akt (Ser473), GSK-3ß (Ser9) and phospholamban (PLN, Ser16 and Thr17) phosphorylation levels were detected through immunofluorescence experiments. Specific PI3K and PKG inhibition elucidated their contribution in the effects produced by exogenous Nrg1ß1 administration. Results and conclusions: respect to I/R alone Nrg1ß1 treatment leads to increased ARVCMs survival and reduced hypercontracture by both improving intracellular calcium handling and maintaining mitochondrial membrane potential. These effects seem to be mediated by increased Akt, PLN and GSK3ß phosphorylation. Taken together, present data support the idea that manipulation of Nrg1ß1/ErbB signaling pathway may provide important clinical benefits in protection against myocardial I/R injury.
Rapid Neuregulin-1ß1 dependent control of calcium handling and mitochondrial function in a model of myocardial ischemia/reperfusion injury
GALLINA, CLARA;BRERO, Alessia;BASSINO, ELEONORA;FORNERO, SARA;ALLOATTI, Giuseppe;GALLO, Maria Pia;LEVI, Renzo
2012-01-01
Abstract
Introduction: the ErbB-Neuregulin1 (Nrg1) axis is essential for both cardiac development and maintenance of adult heart function. We previously demonstrated that, in adult rat ventricular cardiomyocytes (ARVCMs), acute treatment with Nrg1ß1 rapidly improves intracellular calcium recovery by activating PI3K/Akt pathway, which in turn increases endothelial nitric oxide synthase (eNOS) phosphorylation, NO production and cyclic GMP (cGMP)/protein kinase G (PKG) activation. Besides known effects in protection against oxidative stress and cytotoxic agents, recently Nrg1 has also been proposed as an important cardioprotective factor in the setting of myocardial ischemia-reperfusion (I/R) injury. PI3K/Akt and cGMP/PKG cascades are known to improve cardiomyocyte survival, calcium homeostasis and mitochondrial function during I/R events; in this perspective, the aim of this study was to assess the role of Nrg1/ErbB axis in acute cardiac protection against I/R injury, in terms of control of both calcium handling and mitochondrial activity. Methods: the role of Nrg1ß1 (12.5 nmol/L) was studied on isolated ARVCMs subjected to simulated I/R experiments; the peptide was administered during the last phase of ischemia and reperfusion to mimic pharmacological Post Conditioning. Cell survival and hypercontracture were evaluated with propidium iodide nuclear staining; Indo-1 probe was used to evaluate intracellular calcium transients in cardiomyocytes subjected to electrical field stimulation; mitochondrial membrane potential was analyzed through JC-1 fluorescent probe; Akt (Ser473), GSK-3ß (Ser9) and phospholamban (PLN, Ser16 and Thr17) phosphorylation levels were detected through immunofluorescence experiments. Specific PI3K and PKG inhibition elucidated their contribution in the effects produced by exogenous Nrg1ß1 administration. Results and conclusions: respect to I/R alone Nrg1ß1 treatment leads to increased ARVCMs survival and reduced hypercontracture by both improving intracellular calcium handling and maintaining mitochondrial membrane potential. These effects seem to be mediated by increased Akt, PLN and GSK3ß phosphorylation. Taken together, present data support the idea that manipulation of Nrg1ß1/ErbB signaling pathway may provide important clinical benefits in protection against myocardial I/R injury.
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