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Background The recent development of accurate methods to measure two-dimensional strain during dobutamine stress echocardiography has reactivated the debate as to the respective value of myocardial perfusion versus myocardial function assessment in detecting coronary stenoses. The aim of our study was to compare the effects of progressive coronary constrictions on two-dimensional strain and myocardial contrast echocardiography parameters during stress conditions. Methods Nine open-chest pigs were studied in the setting of various degrees of coronary constrictions. Two-dimensional strain and myocardial contrast echocardiography with Flash refilling sequence acquisitions were obtained at rest and during dobutamine infusion. Values of longitudinal strain (LS), circumferential strain (CS), radial strain (RS), and wall thickening, as well as myocardial perfusion parameter (A.b), were then calculated. Results At rest, accuracy for detecting coronary stenosis was higher for CS, LS, and A.b (74%, 67%, and 69%, respectively) than for RS or wall thickening (62% and 64%, respectively). Dobutamine stress echocardiography increased the accuracy of A.b and LS to 77% and to 73%, respectively. Sensitivity during stress was higher for CS (93%) and A.b (77%), whereas specificity was higher for LS (89%) than for other parameters. Combined evaluations (CS+A.b, CS+LS, and LS+A.b) during dobutamine stress echocardiography improved both sensitivity and accuracy for detecting coronary stenosis. Conclusion Quantitative evaluation of contraction by LS and CS analysis and perfusion (A.b) during stress echocardiography resulted in similar accuracy levels, whereas the radial component was less accurate. Maximal sensitivities and accuracies were obtained by combined evaluations during stress.

This study analyzed the oxidant generation during ischemia-reperfusion protocols of Langendorff-perfused rat hearts, preconditioned with a mitochondrial ATP-sensitive potassium channel (mitoKATP) opener (i.e., diazoxide). The autofluorescence of mitochondrial flavoproteins, and that of the total NAD(P)H pool on the one hand and the fluorescence of dyes sensitive to H2O2or O2•−[i.e., the dihydrodichlorofluoroscein (H2DCF) and dihydroethidine (DHE), respectively] on the other, were noninvasively measured at the surface of the left ventricular wall by means of optic fibers. Isolated perfused rat hearts were subjected to an ischemia-reperfusion protocol. Opening mitoKATPwith diazoxide (100 μM) 1) improved the recovery of the rate-pressure product after reperfusion (72 ± 2 vs. 16.8 ± 2.5% of baseline value in control group, P < 0.01), and 2) attenuated the oxidant generation during both ischemic (−46 ± 5% H2DCF oxidation and −40 ± 3% DHE oxidation vs. control group, P < 0.01) and reperfusion (−26 ± 2% H2DCF oxidation and −23 ± 2% DHE oxidation vs. control group, P < 0.01) periods. All of these effects were abolished by coperfusion of 5-hydroxydecanoic acid (500 μM), a mitoKATPblocker. During the preconditioning phase, diazoxide induced a transient, reversible, and 5-hydroxydecanoic acid-sensitive flavoprotein and H2DCF (but not DHE) oxidation. In conclusion, the diazoxide-mediated cardioprotection is supported by a moderate H2O2production during the preconditioning phase and a strong decrease in oxidant generation during the subsequent ischemic and reperfusion phases.

Cardiotoxicity represents the major side-effect limiting the clinical use of anthracyclines, especially doxorubicin, in cancer chemotherapy. The use of non-toxic prodrugs, or of liposome-encapsulated drugs, allows a better targeting of the tumours and may, therefore, improve the tolerance to the treatment. Using the model of isolated perfused rat heart, we have evaluated the cardiotoxicity of a novel prodrug of doxorubicin, HMR-1826, which consists of the association of doxorubicin to glucuronic acid. We have compared the cardiac effects (developed pressure, contractility and relaxation of the left ventricle) induced by HMR-1826 to those induced by doxorubicin and Doxil, a liposomal form of doxorubicin. HMR-1826 was administered intravenously every other day for 11 days at doses of 50–200 mg kg−1 per injection while doxorubicin was administered according to the same protocol at doses of 1–3 mg kg−1 per injection. Doxorubicin strongly decreased the cardiac functional parameters at the doses of 2.5 and 3 mg kg−1 per injection. Doxil (3 mg kg−1) and HMR-1826 (50–150 mg kg−1) were largely devoid of cardiotoxicity. HMR-1826 only induced significant alterations of the cardiac function at the highest dose used (200 mg kg−1 per injection). These alterations were much lower than those of doxorubicin at 2.5 mg kg−1 per injection, despite similar general toxicity symptoms (weight loss, nose bleeding and diarrhoea) at these respective doses. Thus, HMR-1826 appeared about 100-fold less cardiotoxic than doxorubicin. © 1999 Cancer Research Campaign

Attempts to reduce the incidence of congestive heart failure following anthracycline therapy include the replacement of the parent compounds (especially doxorubicin) by less cardiotoxic analogs. Among these analogs, idarubicin (4-demethoxy-daunorubicin) was shown to be less cardiotoxic than doxorubicin in phase II clinical trials, but its actual cardiotoxicity has never been evaluated in large series and has never been compared to that of doxorubicin in relevant experimental models. Using the isolated perfused rat heart model, we compared the cardiac effects (developed pressure, contractility and relaxation of the left ventricle) induced by idarubicin to those induced by doxorubicin. Drugs were administered i.v. every other day for 11 days at doses of 1, 2, 2.5 and 3 mg/kg per injection for doxorubicin and 0.5, 0.75 and 1 mg/kg per injection for idarubicin. We confirmed that similar general toxicity symptoms were obtained for a dose ratio of 1:4 (idarubicin:doxorubicin). However, at the maximum tolerated doses of both drugs (3 mg/kg per injection for doxorubicin and 0.75 mg/kg per injection for idarubicin), the cardiac toxicity of idarubicin remained significantly lower than that of doxorubicin. Anthracycline cardiac accumulation was evaluated in parallel and revealed a lower cardiac accumulation of idarubicin, which could explain the reduced cardiac toxicity of this analog. Direct perfusion of the drugs in the isolated hearts of untreated animals revealed that idarubicin was taken up more readily than doxorubicin in the cardiac tissue, despite the fact that it had less deleterious effects on cardiac function. This indicates that idarubicin also had less intrinsic cardiotoxicity than doxorubicin in this model.

The aim of this study was to investigate the effects of HMR1098, a selective blocker of sarcolemmal ATP-sensitive potassium channel (sarcK(ATP)), in Langendorff-perfused rat hearts submitted to ischemia and reperfusion. The recovery of heart hemodynamic and mitochondrial function, studied on skinned fibers, was analyzed after 30-min global ischemia followed by 20-min reperfusion. Infarct size was quantified on a regional ischemia model after 2-h reperfusion. We report that the perfusion of 10 microM HMR1098 before ischemia, delays the onset of ischemic contracture, improves recovery of cardiac function upon reperfusion, preserves the mitochondrial architecture, and finally decreases infarct size. This HMR1098-induced cardioprotection is prevented by 1 mM 2-mercaptopropionylglycine, an antioxidant, and by 100 nM nifedipine, an L-type calcium channel blocker. Concomitantly, it is shown that HMR1098 perfusion induces (i) a transient and specific inhibition of the respiratory chain complex I and, (ii) an increase in the averaged intracellular calcium concentration probed by the in situ measurement of indo-1 fluorescence. Finally, all the beneficial effects of HMR1098 were strongly inhibited by 5-hydroxydecanoate and abolished by glibenclamide, two mitoK(ATP) blockers. This study demonstrates that the HMR1098-induced cardioprotection occurs indirectly through extracellular calcium influx, respiratory chain complex inhibition, reactive oxygen species production and mitoK(ATP) opening. Taken together, these data suggest that a functional interaction between sarcK(ATP) and mitoK(ATP) exists in isolated rat heart ischemia model, which is mediated by extracellular calcium influx.

It has been proposed that activation of the mitochondrial ATP-sensitive potassium channel (mitoK(ATP)) is part of signaling pathways triggering the cardioprotection afforded by ischemic preconditioning of the heart. This work was to analyze the mitochondrial function profile of Langendorff-perfused rat hearts during the different phases of various ischemia-reperfusion protocols. Specifically, skinned fibers of ischemic preconditioned hearts exhibit a decline in the succinate-supported respiration and complex II activity during ischemia, followed by a recovery during reperfusion. Meanwhile, the apparent affinity of respiration for ADP (which reflects the matrix volume expansion) is increased during preconditioning stimulus and, to a larger extent, during prolonged ischemia. This evolution pattern is mimicked by diazoxide and abolished by 5-hydroxydecanoate. It is concluded that opening the mitoK(ATP) channel mediates the preservation of mitochondrial structure-function via a mitochondrial matrix shrinkage and a reversible inactivation of complex II during prolonged ischemic insult.

International audience; The aim of this study was to investigate mitochondrial alterations in an animal model of chronic myocardial ischemia in rats obtained by surgical constriction of the left coronary artery. Resting coronary blood flow was measured using the fluorescent microsphere technique. Contractile function, defined by rate-pressure product, and myocardial oxygen consumption were measured in a Langendorff preparation. The mitochondrial function was evaluated on permeabilized skinned fibers. Three weeks after surgery, ischemic hearts showed a significant decrease in coronary blood flow compared with sham. Hemodynamic measurements showed a significant systolic and diastolic dysfunction. Alterations in mitochondrial function in ischemic hearts were mainly characterized by a significant decrease in the maximal velocity and apparent half-saturation constant for ADP, loss of the stimulatory effect of creatine, and a stimulatory effect of exogenous cytochrome c. These functional alterations were supported by structural alterations characterized by mitochondrial clustering and swelling associated with membrane rupture. We conclude that the alterations in systolic function after chronic ischemia are supported by severe modifications of mitochondrial structure and function.

Cardiotoxicity is the major side-effect and limits the clinical use of the anthracyclines, doxorubicin and daunorubicin. A special problem is raised by the metabolites of these drugs, the amount of which may vary according to drug combinations and formulations. Doxorubicinol, the 13-dihydroderivative of doxorubicin, has been shown to be more cardiotoxic than unchanged doxorubicin. Daunorubicinol has been assumed also to be more cardiotoxic than unchanged daunorubicin but its direct effect on cardiac function has never been evaluated in preclinical models. We have compared the cardiac effects (developed pressure, contractility and relaxation of the left ventricle) induced by daunorubicinol to those induced by daunorubicin, using the model of the isolated perfused rat heart. After treatment of rats with 6 doses of 3 mg/kg intraperitoneally, daunorubicin strongly decreased the cardiac functional parameters, while daunorubicinol did not induce cardiotoxicity. Both treatments induced a similar accumulation of daunorubicinol in the myocardium, while daunorubicin was only found in the hearts of rats treated with this drug. Direct perfusion of the hearts of untreated rats with both drugs at 10 microM induced a depression in heart function, but daunorubicin induced a progressive increase in diastolic pressure, reflecting the difficulties encountered by the heart to maintain its activity in the presence of this drug, while daunorubicinol did not. We conclude that daunorubicinol is not responsible for the important cardiac toxicity of daunorubicin.

P. Dos Santos, M. K. Aliev, P. Diolez, F. Duclos, P. Besse, S. Bonoron-Adele, P. Sikk, P. Canioni and V. A. Saks. Metabolic Control of Contractile Performance in Isolated Perfused Rat Heart. Analysis of Experimental Data by Reaction:Diffusion Mathematical Model. Journal of Molecular and Cellular Cardiology (2000) 32, 1703–1734. The intracellular mechanisms of regulation of energy fluxes and respiration in contracting heart cells were studied. For this, we investigated the workload dependencies of the rate of oxygen consumption and metabolic parameters in Langendorff-perfused isolated rat hearts.31P NMR spectroscopy was used to study the metabolic changes during transition from perfusion with glucose to that with pyruvate with and without active creatine kinase system. The experimental results showed that transition from perfusion with glucose to that with pyruvate increased the phosphocreatine content and stability of its level at increased workloads. Inhibition of creatine kinase reaction by 15-min infusion of iodoacetamide decreased the maximal developed tension and respiration rates by a factor of two.31P NMR data were analyzed by a mathematical model of compartmentalized energy transfer, which is independent from the restrictions of the classical concept of creatine kinase equilibrium. The analysis of experimental data by this model shows that metabolic stability—constant levels of phosphocreatine, ATP and inorganic phosphate—at increased energy fluxes is an inherent property of the compartmentalized system. This explains the observed substrate specificity by changes in mitochondrial membrane potential. The decreased maximal respiration rate and maximal work output of the heart with inhibited creatine kinase is well explained by the rise in myoplasmic ADP concentration. This activates the adenylate kinase reaction in the myofibrillar space and in the mitochondria to fulfil the energy transfer and signal transmission functions, usually performed by creatine kinase. The activity of this system, however, is not sufficient to maintain high enough energy fluxes. Therefore, there is a kinetic explanation for the decreased maximal respiration rate of the heart with inhibited creatine kinase: i.e. a kinetically induced switch from an efficient energy transfer pathway (PCr–CK system) to a non-efficient one (myokinase pathway) within the energy transfer network of the cell under conditions of low apparent affinity of mitochondria to ADP in vivo. This may result in a significant decrease in the thermodynamic affinity of compartmentalized ATPase systems and finally in heart failure.

Paclitaxel strongly potentiates the cardiotoxicity of doxorubicin in the clinical setting. In this study, we aimed (1) to determine whether this potentiation could be reproduced in an ex vivo model and, if so, (2) to select drugs and protocols that did not cause this potentiation. The effect of paclitaxel and docetaxel on the cardiotoxicity induced by doxorubicin and epirubicin was studied using the model of isolated perfused rat heart. Cardiac performances were evaluated after several combination protocols administered every 2 days over a period of 12 days, and anthracycline concentrations in the heart and liver were determined on Day 12. When administered simultaneously, paclitaxel strongly potentiated the cardiotoxicity of doxorubicin ex vivo, and this effect was not due to Cremophor EL, the solvent used in the formulation of paclitaxel. The potentiation of anthracycline cardiotoxicity could be avoided by the replacement of doxorubicin by epirubicin, and/or of paclitaxel by docetaxel. Cardiotoxic potentiation was also avoided by the introduction of a 24-h lag time between the repetitive injections of doxorubicin and docetaxel. The concentration of doxorubicin and its cardiotoxic metabolite, doxorubicinol, in the heart and liver was not significantly altered by the taxanes, but that of epirubicin was increased twofold both in the heart and the liver. These results show that the potentiation of doxorubicin-induced cardiotoxicity by paclitaxel can be reproduced with an ex vivo model, and that it is not related to an increase in tissue concentration of the drug or active metabolite. Our model, therefore, may be useful for the selection of anthracycline-containing protocols with no increased risk of cardiotoxicity for the patients.

1. In order to develop a predictive model for the preclinical evaluation of anthracycline cardiotoxicity and the means of preventing it, we have studied the functional parameters of perfused hearts isolated from rats receiving repeated doses of several anthracyclines. 2. The anthracyclines studied were doxorubicin, epirubicin, pirarubicin and daunorubicin, and we also studied a liposomal formulation of daunorubicin (DaunoXome) and the co-administration of dexrazoxane (ICRF-187) and doxorubicin. 3. Anthracyclines were administered i.p. at equimolar doses corresponding to 3 mg kg-1 per injection of doxorubicin, every other day for a total of six doses. Dexrazoxane was used at the dose of 30 mg kg-1 per injection and was administered either 30 min before or 30 min after doxorubicin. We evaluated any general toxicity towards the animals as well as alterations of left ventricular contractility and relaxation ex vivo. 4. Epirubicin and daunorubicin were significantly less cardiotoxic than doxorubicin, and neither pirarubicin nor DaunoXome caused significant alterations in cardiac function. There was a direct relationship between the decrease in cardiac contractility or relaxation and anthracycline accumulation in the heart, evaluated after the same treatment schedule. 5. Dexrazoxane induced a significant protection against doxorubicin-induced cardiac toxicity when administered 30 min before doxorubicin, whereas this protection was ineffective when administered 30 min after doxorubicin. Direct perfusion of DaunoXome in isolated hearts of untreated animals resulted in a 12-fold reduction of the accumulation of daunorubicin in heart tissue as compared to the perfusion of free daunorubicin, and did not cause alterations in cardiac function at a dosage for which free daunorubicin induced major alterations. 6. The isolated perfused rat heart appears to be a valuable model for screening of new anthracyclines and of strategies for circumventing anthracycline cardiotoxicity.

We have compared the cardiotoxicity of 3 anthracyclines in a model of isolated perfused rat heart using the Langendorff technique. The contractile state and ventricular compliance were studied. Doxorubicin, epirubicin and pirarubicin were perfused at concentrations of 10(-6) and 10(-5) M during 70 min. The cardiac accumulation of the drugs was studied by HPLC. No significant alteration of cardiac functional parameters was observed at 10(-6) M. At 10(-5) M, epirubicin produced a significantly greater alteration of cardiac contractility than doxorubicin, whereas pirarubicin exerted first an inotropic effect followed by a recovery to initial values at the 60th min. Anthracycline accumulation in the heart was dose-dependent; epirubicin accumulated to a 30% greater extent than doxorubicin and pirarubicin heart concentrations were 4-5 times higher than those of doxorubicin at the end of the perfusion. These results suggest that doxorubicin and epirubicin have the same intrinsic cardiac toxicity, and that their distinct clinical cardiotoxicity must be explained by pharmacokinetic differences, whereas pirarubicin is much less cardiotoxic than the other anthracyclines because of different pharmacodynamic properties.

The aim of this study was to investigate the mechanisms of the cardiodepressive action of ionic and non-ionic contrast media currently used in coronary arteriography. Experiments carried out on isolated preparations of cat papillary muscle, treated with increasing concentrations of Telebrix 39, Radioselectan, Hexabrix and Iopamidol 370 and 300, on the one hand and, on the other hand, a pre-determined dose of Telebrix and Iopamidol 300 during hypoxia and subsequent reoxygenation, showed that, at constant Ca2+ concentrations: (1) The cardiodepressive effects of contrast media are correlated with the hyperosmolality that they induce. When osmolality was higher than 400 mOsm, all the products caused a reduction of the peak force (PF: 40.49 +/- 5.15%), the maximum velocity of contraction (Vmax: 39.85 +/- 3.66%) and of the peak velocity of relaxation (Vrelax: 23.30 +/- 2.20%) (P less than 0.01). The time to peak force (TPF), on the other hand, remained constant, whereas the half-relaxation time (THR) was increased. No significant differences were observed between these effects and those induced by control iso-osmolar solutions when the same osmolality was induced. In practice, however, the critical hyperosmolality value of 400 mOsm is never reached when using non-ionic contrast media such as Hexabrix and Iopamidol 300. This could explain the excellent tolerance to these substances. (2) During hypoxia and reoxygenation, the effect of hyperosmolality is more marked. Thus, the non-ionic contrast medium, Iopamidol 300 (340 mOsm), reduces the hypoxic contractility depression (PF: 53.10 +/- 2.60% compared to the control values of 47.60 +/- 5.00%, P less than 0.01), whereas, at the same dose, the ionic medium Telebrix is hyperosmolar (440 mOsm) and induces a more pronounced hypoxic depression of contractility (P less than 0.01). The critical hyperosmolality is never reached during ventriculography (320 mOsm), whatever the medium used, but it can be observed during coronary arteriography. It is, therefore, important to use non-ionic contrast media in the investigation of unstable angina and of acute myocardium infarction.

The purpose of the present study was to investigate the effects of low concentrations of nifedipine on the cat myocardium under normal conditions, and during hypoxia with subsequent reoxygenation, and, thus, to find out whether the protective effect on hypoxic myocardium described by different investigators existed at these levels. Experiments carried out on five series of each 10 papillary muscles showed that: (1) Nifedipine (0.02 μg/ml) exerted a negative ino tropic effect on maximum unloaded velocity of shortening and total force (Vmax: 92.91 ± 2.92%; PF: 92.10 ± 2.60%), whereas depression of peak velocity of relaxation became marked only at 0.2 μg/ml (Vrelax: 88.87 ± 3.91%). This dose-dependent effect is also Ca$ $-dependent. (2) Nifedipine (0.02 μ/ml) protected against the depression of Vmax, PF and Vrelax during hypoxia (PO2 = 25 mm Hg) Vmax: 82.60 ± 3.45% v. 78.45 ± 1.35% control; PF: 54.65 ± 4.41% v. 48.74 ±3.51% and Vrelax: 92.55 ± 4.50% v. 68.47 ± 1.30%). (3) The association of nifedipine (0.02 μ/ml) and nitroglycerin (0.3 μ/ml) accelerated the recovery of the depression of peak velocity of relaxation during reoxygenation (Vrelax: 99.90 ± 5.63% v. 89.96 ± 5.15% nifedipine alone).