title: Impaired diastolic function after exchange of endogenous troponin I with C-terminal truncated troponin I in human cardiac muscle.
abstract: The specific and selective proteolysis of cardiac troponin I (cTnI) has been proposed to play a key role in human ischemic myocardial disease, including stunning and acute pressure overload. In this study, the functional implications of cTnI proteolysis were investigated in human cardiac tissue for the first time. The predominant human cTnI degradation product (cTnI(1-192)) and full-length cTnI were expressed in Escherichia coli, purified, reconstituted with the other cardiac troponin subunits, troponin T and C, and subsequently exchanged into human cardiac myofibrils and permeabilized cardiomyocytes isolated from healthy donor hearts. Maximal isometric force and kinetic parameters were measured in myofibrils, using rapid solution switching, whereas force development was measured in single cardiomyocytes at various calcium concentrations, at sarcomere lengths of 1.9 and 2.2 mum, and after treatment with the catalytic subunit of protein kinase A (PKA) to mimic beta-adrenergic stimulation. One-dimensional gel electrophoresis, Western immunoblotting, and 3D imaging revealed that approximately 50% of endogenous cTnI had been homogeneously replaced by cTnI(1-192) in both myofibrils and cardiomyocytes. Maximal tension was not affected, whereas the rates of force activation and redevelopment as well as relaxation kinetics were slowed down. Ca(2+) sensitivity of the contractile apparatus was increased in preparations containing cTnI(1-192) (pCa(50): 5.73+/-0.03 versus 5.52+/-0.03 for cTnI(1-192) and full-length cTnI, respectively). The sarcomere length dependency of force development and the desensitizing effect of PKA were preserved in cTnI(1-192)-exchanged cardiomyocytes. These results indicate that degradation of cTnI in human myocardium may impair diastolic function, whereas systolic function is largely preserved.
authors.affiliation: Laboratory for Physiology, Institute for Cardiovascular Research, VU Medical Center, Amsterdam, the Netherlands.
authors.forename: Nadiya A
authors.initials: NA
authors.lastname: Narolska
authors.forename: Nicoletta
authors.initials: N
authors.lastname: Piroddi
authors.forename: Alexandra
authors.initials: A
authors.lastname: Belus
authors.forename: Nicky M
authors.initials: NM
authors.lastname: Boontje
authors.forename: Beatrice
authors.initials: B
authors.lastname: Scellini
authors.forename: Sascha
authors.initials: S
authors.lastname: Deppermann
authors.forename: Ruud
authors.initials: R
authors.lastname: Zaremba
authors.forename: Rene J
authors.initials: RJ
authors.lastname: Musters
authors.forename: Cris
authors.initials: C
authors.lastname: dos Remedios
authors.forename: Kornelia
authors.initials: K
authors.lastname: Jaquet
authors.forename: D Brian
authors.initials: DB
authors.lastname: Foster
authors.forename: Anne M
authors.initials: AM
authors.lastname: Murphy
authors.forename: Jennifer E
authors.initials: JE
authors.lastname: van Eyk
authors.forename: Chiara
authors.initials: C
authors.lastname: Tesi
authors.forename: Corrado
authors.initials: C
authors.lastname: Poggesi
authors.forename: Jolanda
authors.initials: J
authors.lastname: van der Velden
authors.forename: Ger J M
authors.initials: GJ
authors.lastname: Stienen
mesh_terms: D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D003971:Diastole; D006801:Humans; D007537:Isometric Contraction; D007700:Kinetics; D009200:Myocardial Contraction; D032383:Myocytes, Cardiac / Q000502:physiology / Q000648:ultrastructure; D009210:Myofibrils / Q000502:physiology / Q000648:ultrastructure; D012518:Sarcomeres / Q000648:ultrastructure; D017384:Sequence Deletion; D014336:Troponin / Q000032:analysis / Q000378:metabolism; D019210:Troponin I / Q000737:chemistry / Q000378:metabolism*
chemical_list: D014336:Troponin; D019210:Troponin I; D017868:Cyclic AMP-Dependent Protein Kinases