Subcellular Creatine Kinase Alterations

Abstract

Abstract-We have tested the hypothesis that decreased functioning of creatine kinase (CK) at sites of energy production and utilization may contribute to alterations in energy fluxes and calcium homeostasis in congestive heart failure (CHF). Heart failure was induced by aortic banding in 3-week-old rats. Myofilaments, sarcoplasmic reticulum (SR), mitochondrial functions, and CK compartmentation were studied in situ using selective membrane permeabilization of left ventricular fibers with detergents (saponin for mitochondria and SR and Triton X-100 for myofibrils). Seven months after surgery, animals were in CHF. A decrease in total CK activity could be accounted for by a 4-fold decrease in activity and content (Western blots) of mitochondrial CK and a 30% decrease in M isoform of CK (MM-CK) activity. In myofibrils, maximal force, crossbridge kinetics, and ␣-myosin heavy-chain expression decreased, whereas calcium sensitivity of tension development remained unaltered. Myofibrillar CK efficacy was unchanged. Calcium uptake capacities of SR were estimated from the surface of caffeine-induced tension transient (S Ca ) after loading with different substrates. In CHF, S Ca decreased by 23%, and phosphocreatine was 2 times less efficient in enhancing calcium uptake.Oxidative capacities of the failing myocardium measured as oxygen consumption per gram of fiber dry weight decreased by 28%. Moreover, the control of respiration by creatine, ADP, and AMP was severely impaired. Our observations provide evidence that alterations in CK compartmentation may contribute to alterations of energy fluxes and calcium homeostasis in CHF. (Circ Res. 1999;85:68-76.)Key Words: mitochondrial respiration Ⅲ myofibril Ⅲ compartmentation Ⅲ sarcoplasmic reticulum Ⅲ skinned fiber T he mechanisms underlying the decline in cardiac pump function in heart failure are incompletely understood. They lead to a gradual increase in left ventricular (LV) end diastolic pressure and a decrease in systolic pressure. In the past decade attention has been focused on the alterations of the various steps in excitation-contraction coupling and intracellular calcium homeostasis, 1-3 whereas the possible involvement of a mismatch in energy supply and demand has received less attention. 4 -6 It has been shown recently that in human heart failure, there is a generalized alteration of the creatine kinase (CK) system with a decrease in total enzyme activity and velocity and alteration in the isoenzyme pattern, which could contribute to the pathogenesis of heart failure. 5 Moreover, in patients with dilated cardiomyopathy, the phosphocreatine (PCr)/ATP ratio, governed by CK activity, may be a predictor of both total and cardiovascular mortality. 7 However, the precise cellular mechanisms by which altered CK may compromise energy fluxes and contractility are not well understood.CK is an important enzyme involved in energy maintenance and energy transfer in muscle and brain cells. It catalyzes the reversible transfer of a phosphate moiety between ATP and creatine. Four d...

Oxidative capacity in failing hearts

“…Myocardial PCr/ATP is decreased in patients (22) and in large and small animal models of LVH secondary to pressure and volume overload and postinfarction LV remodeling (11,21,23,36,37,39). Furthermore, abnormalities in oxidative phosphorylation have been described in in vitro studies of failing myocardium (6,32). Consistent with these observations, we (18,24) recently reported that ATP synthase and adenine nucleotide translocator (ANT) protein expression is decreased in animals with CHF secondary to postinfarction remodeling or pacinginduced heart failure.…”

31P NMR Detection of Subcellular Creatine Kinase Fluxes in the Perfused Rat Heart

“…Indeed most human and animal cardiac pathologies (dilated hypertrophy, diabetes, heart failure, and genetic cardiomyopathy) are associated with alterations of mito-CK expression or activity (43)(44)(45). We believe that our strategy of analysis will prove to be valuable to explore the pathways of energy transfer in these models and could contribute to a better understanding of the implication of bioenergetics in the development of cardiac failure.…”

Voluntary physical activity alterations in endothelial nitric oxide synthase knockout mice

“…Tissue samples were weighed, homogenized in an ice-cold buffer (50 mg wet wt/ml) containing (in mM) 5 HEPES (pH 8.7), 1 EGTA, and 1 dithiothreitol with Triton X-100 (0.1%), and incubated for 60 min at 4°C for complete enzyme extraction (8). The total activities of CK, AK, lactate dehydrogenase (LDH), cytochrome-c oxidase (COX), and citrate synthase (CS) were assayed (30°C, pH 7.5) with coupled enzyme systems (7).…”