Luc Hittinger

Role of vascular endothelium in exercise-induced dilation of large epicardial coronary arteries in conscious dogs.

BACKGROUNDnThe role of vascular endothelium in the control of epicardial coronary artery vasomotion during treadmill exercise remains unclear. Therefore, we examined the consequences of in vivo balloon endothelial denudation on external coronary diameter of the left circumflex artery during exercise in conscious dogs.nnnMETHODS AND RESULTSnSeven dogs instrumented for the measurement of arterial blood pressure, external coronary artery diameter, and coronary blood flow were studied during exercise before and up to 21 days after balloon endothelial denudation of the proximal left circumflex artery. Endothelial denudation was confirmed by abolition of the epicardial coronary artery dilation induced by acetylcholine (0.3 microgram/kg IV) and reactive hyperemia. Epicardial coronary vasodilation was observed in the control state during treadmill exercise (+5.2 +/- 1.0%). In contrast, a marked vasoconstriction was observed 3 (-4.6 +/- 0.6%) and up to 6 days after endothelial denudation. Complete epicardial coronary artery dilation in response to acetylcholine and exercise was restored 9 days after endothelial denudation. In addition, epicardial coronary artery vasomotor responses to acetylcholine and treadmill exercise were closely correlated (r = .82, P < .001). Reactive dilation was not completely restored 21 days after endothelial denudation, but reactive hyperemia and exercise vasomotor responses during the 21 days follow-up were correlated (r = .70, P < .001). Vasodilation induced by nitroglycerin (1 microgram/kg IV) was reduced by 25% (P < .01) 3 days after endothelial denudation and returned to its corresponding control level 3 days later. Prazosin (50 micrograms/kg IV) significantly attenuated the exercise-induced coronary artery constriction after endothelial denudation (+1.5 +/- 1.4% versus -4.6 +/- 1.0%).nnnCONCLUSIONSnThese data demonstrate that endothelium is essential for the mediation of epicardial coronary dilation during exercise and may protect these vessels against the vasoconstrictor effect of endogenous catecholamines.

Magnetic resonance imaging of targeted catheter-based implantation of myogenic precursor cells into infarcted left ventricular myocardium.

OBJECTIVESnThis study was designed to test the hypothesis that myocardial implantation of myogenic precursor cells (MPC) loaded with iron oxide can be reliably detected in vivo by cardiac magnetic resonance imaging (MRI).nnnBACKGROUNDnIn vivo imaging of targeted catheter-based implantation of MPC into infarcted left ventricular (LV) myocardium is unavailable.nnnMETHODSnThe study was conducted in seven farm pigs (four with anterior myocardial infarction), in which autologous MPC were injected through a percutaneous catheter allowing for LV electromechanical mapping and guided micro-injections into normal and infarcted myocardium. Cardiac MRI was used to detect implanted MPC previously loaded with iron oxide nanoparticles.nnnRESULTSnMagnetic resonance imaging data were compared with LV electromechanical mapping and cross-registered pathology. All 9 injections into normal and 12 injections into locally damaged myocardium were detected on T2-weighted spin echo and inversion-recovery true-fisp MRI (low signal areas) with good anatomical concordance with sites of implantation on electromechanical maps. All sites of injection were confirmed on pathology that showed in all infarct animals iron-loaded MPC at the center and periphery of the infarct as expected from MRI.nnnCONCLUSIONSnTargeted catheter-based implantation of iron-loaded MPC into locally infarcted LV myocardium is accurate and can be reliably demonstrated in vivo by cardiac MRI. The ability to identify noninvasively intramyocardial cell implantation may be determinant for future experimental studies designed to analyze subsequent effects of such therapy on detailed segmental LV function.

Stimulation of Bradykinin B1 Receptors Induces Vasodilation in Conductance and Resistance Coronary Vessels in Conscious Dogs Comparison With B2 Receptor Stimulation

Background—Constitutive bradykinin B1 receptors have been identified in dogs; however, their physiological implications involving the coronary circulation remain to be determined. This study examined, in conscious dogs, the coronary response to des-Arg9-bradykinin (a B1 receptor agonist) and the mechanisms involved. Methods and Results—Eleven dogs were instrumented with a left ventricular micromanometer, a circumflex coronary catheter, a cuff occluder, a Doppler flow probe, and ultrasonic crystals to measure coronary blood flow velocity (CBFv) and coronary diameter (CD). Intracoronary des-Arg9-bradykinin (3 to 100 ng/kg) and bradykinin (0.1 to 10 ng/kg) did not modify systemic hemodynamics but dose-dependently increased CBFv and CD. Des-Arg9-bradykinin was less potent than bradykinin. Hoe 140 (a B2 antagonist, 10 μg/kg) abolished the effects of bradykinin but did not influence the effects of des-Arg9-bradykinin. When CBFv increase was prevented by the cuff occluder, CD responses to bradykinin and des-Arg9...

Role of creatine kinase in cardiac excitation-contraction coupling: studies in creatine kinase-deficient mice

To understand the role of creatine kinase (CK) in cardiac excitation‐contraction coupling, CK‐deficient mice (CK–/–) were studied in vitro and in vivo. In skinned fibers, the kinetics of caffeine‐induced release of Ca2+ was markedly slowed in CK–/– mice with a partial restoration when glycolytic substrates were added. These abnormalities were almost compensated for at the cellular level: the responses of Ca2+ transient and cell shortening to an increased pacing rate from 1 Hz to 4 Hz were normal with a normal post‐rest potentiation of shortening. However, the post‐rest potentiation of the Ca2+ transient was absent and the cellular contractile response to isoprenaline was decreased in CK–/– mice. In vivo, echocardiographically determined cardiac function was normal at rest but the response to isoprenaline was blunted in CK–/– mice. Previously described compensatory pathways (glycolytic pathway and closer sarcoplasmic reticulum‐mitochondria interactions) allow a quasi‐normal SR function in isolated cells and a normal basal in vivo ventricular function, but are not sufficient to cope with a large and rapid increase in energy demand produced by β‐adrenergic stimulation. This shows the specific role of CK in excitation‐contraction coupling in cardiac muscle that cannot be compensated for by other pathways.

Macrophages improve survival, proliferation and migration of engrafted myogenic precursor cells into MDX skeletal muscle.

Transplantation of muscle precursor cells is of therapeutic interest for focal skeletal muscular diseases. However, major limitations of cell transplantation are the poor survival, expansion and migration of the injected cells. The massive and early death of transplanted myoblasts is not fully understood although several mechanisms have been suggested. Various attempts have been made to improve their survival or migration. Taking into account that muscle regeneration is associated with the presence of macrophages, which are helpful in repairing the muscle by both cleansing the debris and deliver trophic cues to myoblasts in a sequential way, we attempted in the present work to improve myoblast transplantation by coinjecting macrophages. The present data showed that in the 5 days following the transplantation, macrophages efficiently improved: i) myoblast survival by limiting their massive death, ii) myoblast expansion within the tissue and iii) myoblast migration in the dystrophic muscle. This was confirmed by in vitro analyses showing that macrophages stimulated myoblast adhesion and migration. As a result, myoblast contribution to regenerating host myofibres was increased by macrophages one month after transplantation. Altogether, these data demonstrate that macrophages are beneficial during the early steps of myoblast transplantation into skeletal muscle, showing that coinjecting these stromal cells may be used as a helper to improve the efficiency of parenchymal cell engraftment.

Preserved Vasodilator Effect of Bradykinin in Dogs With Heart Failure

BACKGROUNDnIn heart failure (HF), vasoconstrictor systems are activated and endothelium-derived vasodilation is blunted. Bradykinin, a potent vasodilator, may play an important role in this setting. However, it is not known whether its vasodilator effect is modified in HF.nnnMETHODS AND RESULTSnFourteen chronically instrumented dogs were studied in the control state and in pacing-induced HF (250 bpm for 3 weeks). The dose-dependent decrease in mean aortic pressure (MAP) induced by acetylcholine was significantly blunted in HF. In contrast, in both control and HF, bradykinin infusion caused similar dose-dependent decreases in MAP and increases in cardiac output (CO). This vasodilator effect of exogenous bradykinin was potentiated similarly in both states by enalaprilat, which blocks both angiotensin conversion and bradykinin degradation. For evaluating the role of endogenous bradykinin, the effects of enalaprilat were compared with those of ciprokiren, a pure renin inhibitor. In control, ciprokiren did not produce any effect. Enalaprilat, however, produced a significant decrease in MAP and a significant increase in CO, which were attributed to the inhibition of bradykinin degradation, because these effects were absent after pretreatment with Hoe 140 (a bradykinin B2 receptor antagonist). In contrast, in HF, vasodilator effects of ciprokiren were observed, but enalaprilat produced larger changes in MAP and CO, and after Hoe 140, the hemodynamic effects of enalaprilat were significantly decreased, showing the effects of endogenous bradykinin, which were similar to those measured in control.nnnCONCLUSIONSnIn this model of HF with a blunted endothelium-derived vasodilation, the vasodilator effects of exogenous and endogenous bradykinin are preserved. These results suggest that bradykinin may play an important role in HF, in which vasoconstriction is present and endothelium-dependent vasodilation is blunted.

Recovery of a normal coronary vascular reserve after rejection therapy in acute human cardiac allograft rejection.

During acute rejection, coronary vascular reserve is severely impaired in human orthotopic heart transplants. To evaluate the effects of rejection therapy on coronary vascular reserve, the ratio of peak-to-resting coronary flow velocity was assessed with a coronary Doppler catheter and a maximally vasodilating dose of intracoronary papaverine (12 mg) in nine allograft recipients without rejection (group 1) and in six recipients before and after treatment of an acute episode of rejection (group 2). All the patients had normal epicardial coronary arteries and were free of left ventricular hypertrophy. In group 2 during rejection, the coronary vascular reserve was significantly lower than in group 1, in which all the patients had a peak-to-resting coronary flow velocity ratio greater than 4 (2.3 +/- 0.5 vs. 5.4 +/- 0.8, respectively, p less than 0.001). In group 2 after treatment of rejection, the peak-to-resting coronary flow velocity ratio was similar to that of group 1 (4.7 +/- 0.8). Heart rate, left ventricular volumes and pressures, hemoglobin concentration, and arterial oxygen pressure were similar in the two groups. This study provides evidence that alterations of coronary vascular reserve because of acute rejection were reversible after treatment of the rejection episode.

Chronic Infusion of Bradykinin Delays the Progression of Heart Failure and Preserves Vascular Endothelium-Mediated Vasodilation in Conscious Dogs

Background—This study examined the effects of chronic bradykinin infusion on hemodynamics and myocardial and endothelial functions during the development of heart failure. Methods and Results—Sixteen instrumented dogs were randomized to receive through the left atria either vehicle or bradykinin (1 &mgr;g/min) during ventricular pacing (250 bpm, 5 weeks). Hemodynamic and left ventricular (LV) parameters and the vasodilator responses to intravenous acetylcholine (0.3 to 3 &mgr;g/kg) and nitroglycerin (1 to 10 &mgr;g/kg) were examined in the control and after 3 and 5 weeks of pacing. The expression of endothelial NOS in femoral, carotid, and renal arteries was determined by Western blot analysis. After 3 weeks of pacing, changes in LV diastolic and systolic parameters were significantly lower in bradykinin-treated than vehicle-treated dogs (LV end-diastolic pressure, +10±3 versus +19±2 mm Hg; time constant of LV isovolumic relaxation, +11±2 versus +17±1 ms; LV wall thickening, −33±18% versus −75±9%; and cardiac output, −16±6% versus −32±6%; all P <0.05). Compared with vehicle-treated dogs, bradykinin-treated dogs had a reduced rightward shift of the diastolic LV pressure-diameter relation and a reduced diastolic LV wall stress. Similar trends were observed after 5 weeks. The vasodilator response to nitroglycerin was preserved in both groups. The response to acetylcholine was blunted in vehicle-treated but preserved in bradykinin-treated dogs. Vascular endothelial NOS expression decreased in vehicle-treated but was preserved in bradykinin-treated dogs. Conclusions—In conscious dogs, chronic bradykinin infusion delays the heart failure progression by preserving LV diastolic and systolic functions and by preserving vascular endothelial function.

Endothelial Modulation of β-Adrenergic Dilation of Large Coronary Arteries in Conscious Dogs

BACKGROUNDnEndothelium-derived relaxing factors have been described as important intermediates in beta-adrenergic vasodilation of resistance coronary vessels, but their involvement at the level of large epicardial coronary arteries remains controversial. Therefore, we examined the role of vascular endothelium in the beta-adrenergic-mediated vasodilation of large epicardial coronary arteries in conscious dogs.nnnMETHODS AND RESULTSnNine dogs were instrumented for measurement of left circumflex coronary artery diameter (CD) by sonomicrometry and coronary blood flow velocity (CBFv) with a Doppler technique in response to graded doses of isoproterenol (0.001 to 0.1 microgram/kg IV bolus). Under control conditions, isoproterenol induced dose-dependent increases in CD and CBFv. When CBFv was kept constant at its baseline value by inflation of a cuff occluder, isoproterenol still induced dose-dependent increases in CD, but the latter were of lesser magnitude than those observed under normal CBFv conditions (110 +/- 20 versus 170 +/- 30 microns, respectively, ie, a reduction of 33% of the dilatory response at 0.1 microgram/kg, P < .01). In the same dogs, the coronary endothelium was then mechanically removed at the site of CD measurement by a balloon angioplasty technique. After this procedure, the dose-dependent increases in CD induced by isoproterenol under either normal or controlled CBFv conditions were overimposable, and their magnitude was similar to that of the increases observed in the presence of an intact endothelium when CBFv was kept constant. After beta 1-adrenergic receptor blockade by atenolol (1 mg/kg), isoproterenol-induced increases in CD were abolished either when CBFv was kept constant or after endothelium removal.nnnCONCLUSIONSnIn conscious dogs, the direct stimulating effect of isoproterenol on beta 1-adrenergic receptors is endothelium-independent at the level of large coronary arteries. The endothelium reinforces the dilatory response to isoproterenol through an indirect, flow-dependent mechanism.

Comparisons of the effects of nicorandil, pinacidil, nicardipine and nitroglycerin on coronary vessels in the conscious dog: role of the endothelium

1 The vasodilator properties of nicorandil on large and small coronary arteries were compared to those of nicardipine, pinacidil, nitroglycerin and acetylcholine in six conscious dogs. 2 Intravenous bolus injections of acetylcholine (0.1 μg kg−1), nitroglycerin (0.3–3 μg kg−1), pinacidil (10–100 μg kg−1), nicardipine (3–30 μg kg−1) and nicorandil (10–100 μg kg−1) dose‐dependently increased circumflex coronary artery diameter and decreased coronary vascular resistance, indicating vasodilator effects on both conduit and resistance coronary arteries. 3 Three days after removal of the endothelium of the circumflex coronary artery (balloon angioplasty), pinacidil‐ and nicardipine‐induced dilatation of large coronary arteries was greatly reduced (both − 76%, P<0.01) whereas that produced by nitroglycerin and nicorandil was decreased only slightly and to a similar extent for both drugs (− 19%, P<0.01 and −28%, P<0.05, respectively). 4 Thus in conscious dogs, nicardipine‐ and pinacidil‐induced dilatation of large coronary arteries is endothelium‐dependent. In contrast, the vasodilator effects of nitroglycerin and nicorandil on conduit vessels are endothelium‐independent. 5 Finally, our results demonstrate that nicorandil dilates the large coronary arteries through its nitrate‐like action and that the ATP‐potassium channel opening properties of the drug are not involved in this effect in the conscious dog.