Nitric oxide inhibits cardiac energy production via inhibition of mitochondrial creatine kinase.
Nitric oxide biosynthesis in cardiac muscle leads to a decreased oxygen consumption and lower ATP synthesis. It is suggested that this effect of nitric oxide is mainly due to the inhibition of the mitochondrial respiratory chain enzyme, cytochrome c oxidase. However, this work demonstrates that nitric oxide is able to inhibit soluble mitochondrial creatine kinase (CK), mitochondrial CK bound in purified mitochondria, CK in situ in skinned fibres as well as the functional activity of mitochondrial CK in situ in skinned fibres. Since mitochondrial isoenzyme is functionally coupled to oxidative phosphorylation, its inhibition also leads to decreased sensitivity of mitochondrial respiration to ADP and thus decreases ATP synthesis and oxygen consumption under physiological ADP concentrations. (+info)
Myogenic signaling of phosphatidylinositol 3-kinase requires the serine-threonine kinase Akt/protein kinase B.
The oncogene p3k, coding for a constitutively active form of phosphatidylinositol 3-kinase (PI 3-kinase), strongly activates myogenic differentiation. Inhibition of endogenous PI 3-kinase activity with the specific inhibitor LY294002, or with dominant-negative mutants of PI 3-kinase, interferes with myotube formation and with the expression of muscle-specific proteins. Here we demonstrate that a downstream target of PI 3-kinase, serine-threonine kinase Akt, plays an important role in myogenic differentiation. Expression of constitutively active forms of Akt dramatically enhances myotube formation and expression of the muscle-specific proteins MyoD, creatine kinase, myosin heavy chain, and desmin. Transdominant negative forms of Akt inhibit myotube formation and the expression of muscle-specific proteins. The inhibition of myotube formation and the reduced expression of muscle-specific proteins caused by the PI 3-kinase inhibitor LY294002 are completely reversed by constitutively active forms of Akt. Wild-type cellular Akt effects a partial reversal of LY294002-induced inhibition of myogenic differentiation. This result suggests that Akt can substitute for PI 3-kinase in the stimulation of myogenesis; Akt may be an essential downstream component of PI 3-kinase-induced muscle differentiation. (+info)
Myocardial creatine kinase kinetics in hearts with postinfarction left ventricular remodeling.
This study examined whether alterations in myocardial creatine kinase (CK) kinetics and high-energy phosphate (HEP) levels occur in postinfarction left ventricular remodeling (LVR). Myocardial HEP and CK kinetics were examined in 19 pigs 6 wk after myocardial infarction was produced by left circumflex coronary artery ligation, and the results were compared with those from 9 normal pigs. Blood flow (microspheres), oxygen consumption (MVO2), HEP levels [31P magnetic resonance spectroscopy (MRS)], and CK kinetics (31P MRS) were measured in myocardium remote from the infarct under basal conditions and during dobutamine infusion (20 micrograms. kg-1. min-1 iv). Six of the pigs with LVR had overt congestive heart failure (CHF) at the time of study. Under basal conditions, creatine phosphate (CrP)-to-ATP ratios were lower in all transmural layers of hearts with CHF and in the subendocardium of LVR hearts than in normal hearts (P < 0.05). Myocardial ATP (biopsy) was significantly decreased in hearts with CHF. The CK forward rate constant was lower (P < 0.05) in the CHF group (0.21 +/- 0.03 s-1) than in LVR (0.38 +/- 0.04 s-1) or normal groups (0.41 +/- 0.03 s-1); CK forward flux rates in CHF, LVR, and normal groups were 6.4 +/- 2.3, 14.3 +/- 2.1, and 20.3 +/- 2.4 micromol. g-1. s-1, respectively (P < 0.05, CHF vs. LVR and LVR vs. normal). Dobutamine caused doubling of the rate-pressure product in the LVR and normal groups, whereas CHF hearts failed to respond to dobutamine. CK flux rates did not change during dobutamine in any group. The ratios of CK flux to ATP synthesis (from MVO2) under baseline conditions were 10.9 +/- 1.2, 8. 03 +/- 0.9, and 3.86 +/- 0.5 for normal, LVR, and CHF hearts, respectively (each P < 0.05); during dobutamine, this ratio decreased to 3.73 +/- 0.5, 2.58 +/- 0.4, and 2.78 +/- 0.5, respectively (P = not significant among groups). These data demonstrate that CK flux rates are decreased in hearts with postinfarction LVR, but this change does not limit the response to dobutamine. In hearts with end-stage CHF, the changes in HEP and CK flux are more marked. These changes could contribute to the decreased responsiveness of these hearts to dobutamine. (+info)
Attenuation of myocardial injury due to oxygen free radicals (OFR) by pretreatment with OFR or calcitonin gene-related peptide.
AIM: To study the cardioprotective effects of oxygen free radicals (OFR) and calcitonin gene-related peptide (OGRP) pretreatment on myocardial damages due to OFR in isolated perfused rat heart. METHODS: The hearts were perfused in a Langendorff mode. OFR were generated by electrolysis of Krebs-Henseleit (K-H) solution. RESULTS: OFR pretreatment reduced the impairment of cardiac contractile function, the decrease of coronary flow and the increase of creatinine kinase (CK) release due to OFR, and the effect exhibited period dependence and cycle-dependence. 1-(5-isoquinolinylsulfonyl)-2-methylpiperazine (H-7), an inhibitor of protein kinase C, abolished the protection of OFR pretreatment (CK release = 110 +/- 7, 215 +/- 23, 169 +/- 14, 240 +/- 30, and 113 +/- 19 kU.L-1 for control, OFR, OFR pretreatment, OFR pretreatment plus H-7, and H-7, respectively). CGRP pretreatment also protected the myocardium damages elicited by OFR in isolated perfused rat heart. CONCLUSIONS: OFR or CGRP pretreatment protected myocardium against injury elicited by OFR, and the effect of OFR pretreatment was related to the activation of PKC. (+info)
The relationship between periprocedural myocardial infarction and subsequent target vessel revascularization following percutaneous coronary revascularization: insights from the EPIC trial. Evaluation of IIb/IIIa platelet receptor antagonist 7E3 in Preventing Ischemic Complications.
OBJECTIVES: We sought to determine whether periprocedural myocardial infarction complicating percutaneous coronary revascularization is associated with subsequent clinical restenosis, as judged by the need for target vessel revascularization. BACKGROUND: Although myocardial enzyme elevation following angioplasty is associated with increased late mortality, its effect on subsequent clinical restenosis, as assessed by the need for late target vessel revascularization (TVR), is unknown. METHODS: Serial myocardial enzyme determinations were performed on 2,099 patients who underwent angioplasty or atherectomy in the Evaluation of IIb/IIIa platelet receptor antagonist 7E3 in Preventing Ischemic Complications (EPIC) trial. Thirty-day survivors were prospectively followed for three years for adverse clinical events including death and need for TVR. RESULTS: Within the study population, periprocedural creatine kinase (CK) elevation was a predictor of late mortality. Among patients with elevated CK, however, a paradoxical decrease in the need for late TVR was present. This relationship became progressively more profound as the magnitude of CK release increased. Late TVR occurred in 29.8% of patients with no CK elevation, 24.8% with CK elevation to >3 times normal, and 16.9% with >10 times elevation (hazard ratio 0.51, 95% CI 0.29, 0.91). CONCLUSIONS: In the EPIC study, patients with periprocedural MI were less likely to develop clinical restenosis as measured by the need for TVR. Mechanistically, although it is unlikely that CK elevation prevents vascular renarrowing per se, myocardial necrosis impairs the clinical manifestation of restenosis, thereby reducing the need for ischemia-driven TVR. This novel finding 1) highlights the potential discordance between angiographic and clinical measures of restenosis, and 2) has implications for clinical trials, as therapies that reduce periprocedural MI may be associated with a perceived excess of restenosis when measured by the need for TVR. (+info)
Protective effect of quinaprilat, an active metabolite of quinapril, on Ca2+-overload induced by lysophosphatidylcholine in isolated rat cardiomyocytes.
We examined the effects of quinaprilat, an active metabolite of quinapril (an angiotensin converting enzyme (ACE) inhibitor) on the increase in intracellular concentration of Ca2+ ([Ca2+]i) (Ca2+-overload) induced by lysophosphatidylcholine (LPC) in isolated rat cardiomyocytes. LPC (15 microM) produced Ca2+-overload with a change in cell-shape from rod to round. Quinaprilat but not quinapril at 20 or 50 microM attenuated the LPC-induced increase in [Ca2+]i and the change in cell-shape in a concentration-dependent manner. Since quinaprilat has an inhibitory action on ACE and quinapril has practically no inhibitory action on ACE, it is likely that the inhibitory action of quinaprilat on ACE is necessary for the protective effect of the drug against LPC-induced changes. We therefore examined the effects of enalapril (another ACE inhibitor with the weak inhibitory action on ACE) and enalaprilat (an active metabolite of enalapril with an inhibitory action on ACE) on the LPC-induced changes. Both enalapril and enalaprilat attenuated the LPC-induced Ca2+-overload, suggesting that the inhibitory action on ACE may not mainly contribute to the protective effect of ACE inhibitors against LPC-induced Ca2+-overload. This suggestion was supported by the fact that neither ACE (0.2 U/ml) nor angiotensin II (0.1-100 microM) increased [Ca2+]i in isolated cardiomyocytes. Furthermore, application of bradykinin (0.01-10 microM) did not enhance the protective effect of quinaprilat against LPC-induced changes. LPC also increased release of creatine kinase (CK) from the myocyte markedly, and quinaprilat but not quinapril attenuated the LPC-induced CK release. Unexpectedly, both enalapril and enalaprilat did not attenuate the LPC-induced CK release. Neither quinapril nor quinaprilat changed the critical micelle concentration of LPC, suggesting that these drugs do not directly bind to LPC. We conclude that quinaprilat attenuates the LPC-induced increase in [Ca2+]i, and that the protective effect of quinaprilat on the LPC-induced change may not be related to a decrease in angiotensin II production or an increase in bradykinin production. (+info)
Maturation of the myogenic program is induced by postmitotic expression of insulin-like growth factor I.
The molecular mechanisms underlying myogenic induction by insulin-like growth factor I (IGF-I) are distinct from its proliferative effects on myoblasts. To determine the postmitotic role of IGF-I on muscle cell differentiation, we derived L6E9 muscle cell lines carrying a stably transfected rat IGF-I gene under the control of a myosin light chain (MLC) promoter-enhancer cassette. Expression of MLC-IGF-I exclusively in differentiated L6E9 myotubes, which express the embryonic form of myosin heavy chain (MyHC) and no endogenous IGF-I, resulted in pronounced myotube hypertrophy, accompanied by activation of the neonatal MyHC isoform. The hypertrophic myotubes dramatically increased expression of myogenin, muscle creatine kinase, beta-enolase, and IGF binding protein 5 and activated the myocyte enhancer factor 2C gene which is normally silent in this cell line. MLC-IGF-I induction in differentiated L6E9 cells also increased the expression of a transiently transfected LacZ reporter driven by the myogenin promoter, demonstrating activation of the differentiation program at the transcriptional level. Nuclear reorganization, accumulation of skeletal actin protein, and an increased expression of beta1D integrin were also observed. Inhibition of the phosphatidyl inositol (PI) 3-kinase intermediate in IGF-I-mediated signal transduction confirmed that the PI 3-kinase pathway is required only at early stages for IGF-I-mediated hypertrophy and neonatal MyHC induction in these cells. Expression of IGF-I in postmitotic muscle may therefore play an important role in the maturation of the myogenic program. (+info)
Kinetics of lactate and pyruvate transport in cultured rat myotubes.
Skeletal muscle transport of lactate and pyruvate was studied in primary cultures of rat myotubes, applying the pH-sensitive fluorescent indicator 2', 7'-bis(carboxyethyl)-5(6)-carboxyfluorescein. The initial rate of decrease in intracellular pH (pHi) upon lactate or pyruvate incubation was used to determine total transport (carrier mediated and diffusion). Both lactate and pyruvate transport could be inhibited by a combination of 0.5 mM 4,4'-diisothiocyanostilbene-2, 2'-disulfonic acid, 5 mM mersalyl and 10 mM alpha-cyano-4-hydroxycinnamate. The kinetic parameters, Km and Vmax, for carrier-mediated transport of lactate were 9.9+/-1.1 mM and 0. 69+/-0.02 mmol l-1 s-1, respectively. For pyruvate, Km and Vmax were 4.4+/-1.3 mM and 0.30+/-0.05 mmol l-1 s-1, respectively. The diffusion component of the total transport was 0.0040+/-0.0005[S] (n=4) and 0.0048+/-0.0003[S] (n=4) for lactate and pyruvate, respectively. Furthermore, it was observed that the two monocarboxylate transporter isoforms present in mature skeletal muscles, MCT1 and MCT4 (formerly called MCT3 (M.C. Wilson, V.N. Jackson, C. Heddle, N.T. Price, H. Pilegaard, C. Juel, A. Bonen, I. Montgomery, O.F. Hutter, A.P. Halestrap, Lactic acid efflux from white skeletal muscle is catalyzed by the monocarboxylate transporter isoform MCT3, J. Biol. Chem. 273 (1998) 15920-15926)), were also expressed in primary culture of myotubes. (+info)