[pt] MÓDULO II - SLIDES COMPLEMENTARES - ADM1668

Abstract

Aims: Nitroxyl (HNO) interacts with thiols to act as a redox-sensitive modulator of protein function. It enhances sarcoplasmic reticular Ca2+ uptake and myofilament Ca2+ sensitivity, improving cardiac contractility. This activity has led to clinical testing of HNO donors for heart failure. Here we tested whether HNO alters the inhibitory interaction between phospholamban (PLN) and the sarcoplasmic reticulum Ca2+-ATPase (SERCA2a) in a redox-dependent manner, improving Ca2+ handling in isolated myocytes/hearts. Results: Ventriculocytes, sarcoplasmic reticulum (SR) vesicles, and whole hearts were isolated from control (wildtype [WT]) or PLN knockout (pln(-/-)) mice. Compared to WT, pln(-/-) myocytes displayed enhanced resting sarcomere shortening, peak Ca2+ transient, and blunted -adrenergic responsiveness. HNO stimulated shortening, relaxation, and Ca2+ transient in WT cardiomyocytes, and evoked positive inotropy/lusitropy in intact hearts. These changes were markedly blunted in pln(-/-) cells/hearts. HNO enhanced SR Ca2+ uptake in WT but not pln(-/-) SR-vesicles. Spectroscopic studies in insect cell microsomes expressing SERCA2a +/- PLN showed that HNO increased Ca2+-dependent SERCA2a conformational flexibility but only when PLN was present. in cardiomyocytes, HNO achieved this effect by stabilizing PLN in an oligomeric disulfide bond-dependent configuration, decreasing the amount of free inhibitory monomeric PLN available. Innovation: HNO-dependent redox changes in myocyte PLN oligomerization relieve PLN inhibition of SERCA2a. Conclusions: PLN plays a central role in HNO-induced enhancement of SERCA2a activity, leading to increased inotropy/lusitropy in intact myocytes and hearts. PLN remains physically associated with SERCA2a; however, less monomeric PLN is available resulting in decreased inhibition of the enzyme. These findings offer new avenues to improve Ca2+ handling in failing hearts. Antioxid. Redox Signal. 19, 1185-1197.American Heart AssociationT32 NIH Training GrantItalian Society of CardiologyISHR-ES/ServierCoordenação de Aperfeiçoamento de Pessoal de Nível Superior (CAPES)NIHNational Science FoundationAHA Scientist Development GrantFondation LeducqPeter Belfer LaboratoryAbraham and Virginia Weiss ProfessorshipAmerican Heart Association GIAJohns Hopkins Med Inst, Div Cardiol, Baltimore, MD 21205 USAVirginia Polytech Inst & State Univ, Dept Biochem, Blacksburg, VA 24061 USAClin Montevergine, Mercogliano, ItalyUS Army Edgewood Chem Biol Ctr, Aberdeen, MD USAExcet Inc, Springfield, VA USAUniv Maryland, Sch Med, Dept Biochem & Mol Biol, Baltimore, MD 21201 USAUniversidade Federal de São Paulo UNIFESP EPM, Dept Farmacol, São Paulo, BrazilUniv Alabama Birmingham, Dept Med, Birmingham, AL 35294 USAJohns Hopkins Univ, Dept Chem, Baltimore, MD 21218 USAMed Univ Graz, Div Cardiol, Graz, AustriaVanderbilt Univ, Dept Clin Pharmacol, Nashville, TN 37235 USAOhio State Univ, Davis Heart & Lung Res Inst, Dept Physiol & Cell Biol, Columbus, OH 43210 USAUniv Cincinnati, Cincinnati, OH USAEdward Via Coll Osteopath Med, Blacksburg, VA 24060 USAUniv Perugia, Dept Clin Med, I-06100 Perugia, ItalyUniversidade Federal de São Paulo UNIFESP EPM, Dept Farmacol, São Paulo, BrazilAmerican Heart Association: 0815217EAmerican Heart Association: 10POST4140001NIH: R01CA102428NIH: K02HL094692NIH: R01HL079283NIH: R01 HL26057NIH: R01 HL64018NIH: R01 HL101235NIH: 1 R15HL091410NIH: R01 HL075265NIH: R01 HL091923National Science Foundation: CHE-1213438AHA Scientist Development Grant: 10SDG2640109Web of Scienc