RESUMO
Loss of function or/and death of cardiomyocytes is one of the major contributing factors in the development of heart failure. Cytosolic Hsp60 can directly interact and regulate activation of some kinases and sequestrate certain proapoptotic molecules to avoid the cardiomyocyte apoptosis. We assumed that Akt1 kinase, a downstream effector of PI3 kinase, can interact with Hsp60. Our aim was to clarify the interaction of Akt1 and Hsp60 and to investigate the Akt1 expression in normal and failing hearts in acute and chronic stress. The experimental mouse models of inducible myocarditis and DCM-like pathology were developed in our laboratory. Akt1 and phospho-Akt1 (pS473) expression were studied by Western blot analysis. Co-immunoprecipitation method was used to test complex formation of Akt1 and Hsp60. The interaction of Hsp60 and Akt1 was detected for the first time by co-immunoprecipitation method in normal myocardium and under pathology as well. There were no significant changes in the level of Akt1 expression in both myocardia. At the same time we observed significant decrease in Akt1 phosphorylation at the final stage of DCM-like pathology but not at experimental myocarditis. The final stage of heart failure in mouse model of DCM-like pathology was characterized by reduced level of phospho-Akt1/Akt1 (pS473; -26%; P<0.05), whereas no differences were found in total Akt1 protein content. We suggest a possible involvement of cytoplasmic Hsp60 in regulation of Akt1 activity at heart failure progression.
RESUMO
Physiological stresses (heat, hemodynamics, genetic mutations, oxidative injury and myocardial ischemia) produce pathological states in which protein damage and misfolded protein structures are a common denominator. The specialized proteins family of antistress proteins - molecular chaperons (HSPs) - are responsible for correct protein folding, dissociating protein aggregates and transport of newly synthesized polypeptides to the target organelles for final packaging, degradation or repair. They are inducible at different cell processes such as cell division, apoptosis, signal transduction, cell differentiation and hormonal stimulation. HSPs are involved in numerous diseases including cardiovascular pathologies, revealing changes of expression and cell localization. We studied the possible changes in expression level of abundant mitochondrial chaperon Hsp60 and main human cytochrome P450 monooxygenase (2E1 isoform) at dilated cardiomyopathy (DCM) progression at the end stage of heart failure using Western blot analysis. The ischemic and normal humans' hearts were studied as control samples. We observed the decrease of Hsp60 level in cytoplasmic fraction of DCM- and ischemia-affected hearts' left ventricular and significant increase of Hsp60 in mitochondrial fractions of all hearts investigated. At the same time we detected the increase of P450 2E1 expression level in ischemic and dilated hearts' cytoplasmic fractions in comparison with normal myocardium and no detectable changes in microsomal fractions of hearts investigated which could be linked with increased level of oxidative injury for DCM heart muscle. In addition, all the changes described are accompanied by significant decrease of ATPase activity of myosin purified from DCM-affected heart in comparison with normal and ischemic myocardia as well. The data obtained allow us to propose a working hypothesis of functional link between antistress (HSPs) and antioxidative (cytochromes) systems at DCM progression.
