Bypass surgery in patients with coronary artery disease. Indications based on the multicenter randomized trials.
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Angina, Unstable↗
Biomedical subjects
R Bolli
Publications and source records attributed to R Bolli.
The monocarboxylate carrier from bovine heart mitochondria: partial purification and its substrate-transporting properties in a reconstituted system.
The monocarboxylate (pyruvate) carrier from bovine heart mitochondria was extracted from submitochondrial particles with Triton X-114 in the presence of cardiolipin. By a single hydroxylapatite chromatography step a 125-fold purification of the carrier protein could be achieved. High pyruvate/pyruvate-exchange activity was recovered, when the protein was reconstituted into phospholipid vesicles. No transport activity was observed, when the isolation occurred in the absence of phospholipids. The 2-cyano-4-hydroxycinnamate sensitive pyruvate exchange reaction was strongly temperature sensitive and dependent on the amount of protein reconstituted. Other 2-ketoacids caused competitive inhibition of the pyruvate uptake. Inhibitors of other mitochondrial carries, however, had very low or no effect on the monocarboxylate exchange. The influence of different -SH group reagents on the measured pyruvate/pyruvate-exchange in the reconstituted system was similar to the one observed with intact mitochondria. It is concluded that the described procedures for extraction, purification and reconstitution of the mitochondrial monocarboxylate carrier conserved the functional properties of the protein.
Animals↗
Extraction, partial purification and functional reconstitution of two mitochondrial carriers transporting keto acids: 2-oxoglutarate and pyruvate.
Bovine heart submitochondrial particles were treated with a medium containing Triton X-114 and cardiolipin. The extract was subjected to hydroxyapatite chromatography. Only a few major polypeptides of similar molecular masses were found in the eluate, as shown by electrophoresis in an SDS-polyacrylamide gel stained with silver. The eluate was reconstituted into liposomes and was shown to catalyse two different transport activities: 2-oxoglutarate-2-oxoglutarate exchange sensitive to phthalonate and phenylsuccinate and pyruvate-pyruvate exchange sensitive to 2-cyano-4-hydroxycinnamate. Since both activities were found to have characteristics similar to those described for intact mitochondria, it was concluded that at least two of the polypeptides found in the hydroxyapatite eluate correspond to the two mitochondrial carriers.
Animals↗
Cytochrome c oxidase from Paracoccus denitrificans in Triton X-100: aggregation state and kinetics.
Cytochrome c oxidase from Paracoccus denitrificans was homogeneously dispersed in Triton X-100. Using gel exclusion chromatography and sucrose gradient centrifugation analysis a molecular weight of the detergent-protein complex of 155,000 was determined. After subtraction of the bound detergent (111 mol/mol heme aa3) a molecular weight of 85,000 resulted, which agreed well with the model of a monomer containing two subunits. This monomer showed high cytochrome c oxidase activity when measured spectrophotometrically in the presence of Triton X-100 (Vmax = 85 s-1). The molecular activity, plotted according to Eadie-Hofstee, was monophasic as a function of the cytochrome c concentration. A Km of 3.6 X 10(-6) M was evaluated, similar to the Km observed in the presence of dodecyl maltoside [Nałecz et al. (1985).
Electron Transport↗
Factors that determine the occurrence of arrhythmias during acute myocardial ischemia.
To elucidate the determinants of arrhythmias during acute myocardial ischemia, a 25-minute coronary artery occlusion was performed in 101 open-chest dogs. Heart rate correlated positively with the number of premature ventricular complexes (PVCs) and the occurrence of ventricular tachycardia (VT) and fibrillation (VF). The size of the occluded coronary bed was positively correlated with the occurrence of VF, but exhibited only a weak association with PVCs and VT. Coronary collateral flow exhibited a strong negative correlation with all forms of arrhythmias. Importantly, a sensitive coupling was present, whereby small differences in flow were associated with large differences in rhythm disorders. VT occurred in all ranges of occluded bed size examined, but within any given range, its incidence was inversely related to collateral flow. Thus, VT is relatively independent of occluded bed size, and is determined primarily by the degree of myocardial hypoperfusion. In contrast, even in the presence of low flows, VF did not develop with occluded beds less than 28% of left ventricular mass; above this critical occluded bed size, the incidence of VF was inversely related to collateral perfusion. VF is therefore determined by the association of a large occluded bed with a poor collateral function. This study identifies and systematically analyzes three major determinants of arrhythmias during acute myocardial ischemia: the heart rate, the amount of ischemic myocardium, and the severity of myocardial flow reduction. The finding that these factors account for most of the variability in the occurrence of malignant tachyarrhythmias has important methodological and clinical implications.
Animals↗
Techniques for the study of bovine cytochrome-c oxidase monomer-dimer association.
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Animals↗
Disparity of reperfusion arrhythmias after reversible myocardial ischemia in open chest and conscious dogs.
Myocardial reperfusion after brief, reversible ischemia is frequently associated with malignant arrhythmias in experimental animals. These observations have been extrapolated to humans despite being restricted to anesthetized, open chest preparations. No data are available regarding the incidence of reperfusion arrhythmias after reversible (less than 20 minutes) ischemia in the conscious state. Thus, reperfusion arrhythmias after a 15 minute occlusion of the left anterior descending coronary artery were compared in 24 open chest dogs (17 anesthetized with pentobarbital and 7 with chloralose plus urethane) and 25 conscious, unsedated, trained dogs. The incidence of all rhythm disorders (single premature ventricular complexes, pairs, ventricular tachycardia and fibrillation) was markedly and significantly lower in conscious than in either pentobarbital- or chloralose-anesthetized dogs. The disparity was not accounted for by differences in coronary collateral flow, coronary reactive hyperemia or occluded bed size. The conscious animals, however, exhibited lower heart rates and arterial pressures during reperfusion than did the open chest dogs, suggesting a lower level of adrenergic stimulation, which might have contributed to the reduced incidence of reperfusion arrhythmias. Coronary reperfusion after 15 minutes of occlusion is unlikely to precipitate ventricular tachyarrhythmias in the conscious, trained dog, even after severe ischemia. The occurrence of these rhythm disorders in anesthetized models may reflect the influence of surgical trauma or excessive adrenergic activity, or both. Reperfusion arrhythmias after reversible ischemia may be considerably less common in the clinical setting than previously postulated on the basis of open chest animal experiments.
Anesthesia↗
N-2-mercaptopropionylglycine improves recovery of myocardial function after reversible regional ischemia.
Myocardial reperfusion after reversible regional ischemia is known to result in delayed recovery of contractile function, but the mechanism responsible for this phenomenon remains unclear. We examined the ability of N-2-mercaptopropionylglycine, a synthetic thiol compound with oxygen free radical scavenging properties, to attenuate postischemic dysfunction in open chest dogs undergoing a 15 minute occlusion of the left anterior descending coronary artery followed by 4 hours of reperfusion. Treated animals received an infusion of N-2-mercaptopropionylglycine (50 mg/kg per h) for 4 hours starting 15 minutes before coronary occlusion. Collateral flow, as determined with radioactive microspheres after 10 minutes of ischemia, was 0.07 +/- 0.01 ml/min per g (mean +/- SE) in both control (n = 20) and treated (n = 13) groups. The occluded vascular bed, as determined by postmortem perfusion, averaged 26.1 +/- 1.2% of the weight of the left ventricle in control and 29.6 +/- 1.3% in treated animals. Systolic wall thickening (an index of regional function) was assessed with an epicardial pulsed Doppler probe. The two groups exhibited comparable systolic thickening under baseline conditions and similar degrees of dyskinesia during ischemia. Nevertheless, recovery of function (expressed as percent of baseline) was considerably greater in the treated dogs at 1 hour (44.6 versus 12.8%, p = 0.05), 2 hours (64.0 versus 31.6%, p less than 0.02), 3 hours (77.1 versus 36.7%, p less than 0.01) and 4 hours of reperfusion (75.0 versus 40.0%, p less than 0.05). Thus, N-2-mercaptopropionylglycine produced a significant and sustained improvement in recovery of contractile function after a brief episode of regional myocardial ischemia.(ABSTRACT TRUNCATED AT 250 WORDS)
Amino Acids, Sulfur↗
Myocardial infarct size quantification by MR imaging early after coronary artery occlusion in dogs.
The feasibility of using magnetic resonance (MR) imaging to estimate myocardial infarct size was explored in an in vitro model using only the inherent differences in contrast between infarcted and noninfarcted myocardium. Eight dogs underwent coronary occlusion; their hearts were removed 6 hours later. Estimates of T2 for normal and infarcted myocardium were derived from MR images. Infarct size was quantified anatomically using triphenyltetrazolium-chloride (TTC) staining and compared with MR estimates. The T2 values derived from the images clearly discriminated between infarcted (126 +/- 22 msec) and normal myocardium (88 +/- 10 msec, P less than .05), providing images with good contrast between normal and infarcted myocardium. Comparable differences in T2 values were also noted from spectrometric determinations. Estimates of infarct size by MR imaging compared well with TTC estimates (r = 0.98) over a wide range of infarct sizes from 3% to 29% of the left ventricular mass. These results suggest the potential for in vivo quantification of infarct size based on the inherent contrast difference between infarcted and normal myocardium.
Animals↗
Validation of a single crystal for measurement of transmural and epicardial thickening.
Conventional measurement of wall thickening with two transit-time crystals requires considerable skill, is associated with myocardial trauma, and does not discern the function of different layers of the left ventricular (LV) wall. To overcome these limitations, we have developed a 10 MHz pulsed Doppler technique that measures thickening at any depth of the LV wall from a single crystal sutured to the epicardium. To verify its accuracy, we compared measurements of thickening fraction (TF) by pulsed Doppler and transit-time methods in 25 open-chest dogs. The epicardial Doppler crystal was placed over an intramyocardial crystal positioned either in the subendocardium or midwall. The epicardial crystal acted as both the Doppler transducer and the transit-time transmitter, so that TF was measured by each technique at the same site. A wide range of regional function (transmural TF:-28-42%, epicardial TF:-20-28%) was produced by coronary occlusion followed by reperfusion, and by isoproterenol and phenylephrine infusions. There was a good correlation between the two methods, both for transmural TF (r = 0.98, 107 paired measurements) and epicardial TF (r = 0.99, 70 paired measurements). Despite marked changes in function, the two techniques yielded similar measurements under all of the conditions tested (base line, ischemia, 5, 15, 30, 60, 120, and 180 min of reperfusion, isoproterenol and phenylephrine). One-millimeter errors in selecting the depth of the Doppler sample volume did not significantly affect the accuracy of TF measurements. Thus the single pulsed Doppler crystal provides a simple, atraumatic, and accurate means for measuring myocardial function, both transmurally and in selected layers of the LV wall.
Animals↗
Beta-adrenergic stimulation reverses postischemic myocardial dysfunction without producing subsequent functional deterioration.
The prolonged myocardial dysfunction observed after reversible ischemia (stunned myocardium) has been postulated to result from an inability of the myocytes to replenish ATP stores. Accordingly, one would expect inotropic stimulation to result in minimal increase in contractile function, or possibly even further deterioration. To test this hypothesis, studies were performed in open-chest dogs undergoing a 15-minute occlusion of the left anterior descending coronary artery (LAD) followed by 4 hours of reperfusion. Systolic wall thickening, an index of regional myocardial function, was measured in the LAD-dependent territory with ultrasonic crystals. Thickening fraction was 20.8 +/- 3.0% (mean +/- standard error of the mean) under baseline conditions, decreased to -18.6 +/- 1.6% during LAD occlusion, and was still severely depressed after 3 hours of reperfusion (2.6 +/- 3.4%). Thickening fraction remained stable between 3 and 4 hours of reperfusion in 5 untreated control dogs. In 9 treated dogs, isoproterenol (0.1 microgram/kg/min intravenously for 30 minutes starting 3 hours after reperfusion) increased thickening fraction to values (24.8 +/- 4.5%) that were similar to those at baseline. Thirty minutes after discontinuation of isoproterenol administration, thickening fraction had returned to pre-isoproterenol levels. Thus, reperfused, severely depressed myocardium responds dramatically to beta-adrenergic stimulation without subsequent adverse effects on function in the short-term. These findings imply that the stunned myocardium can generate ATP, and therefore do not support the view that an inability to replenish ATP stores is the cause of postischemic dysfunction. More important, this study suggests that postischemic dysfunction in humans may be effectively reversed with inotropic therapy without short-term deleterious sequelae.
Adenosine Triphosphate↗
The role of subunit III in bovine cytochrome c oxidase. Comparison between native, subunit III-depleted and Paracoccus denitrificans enzymes.
In order to obtain information on the role of subunit III in the function and aggregation state of cytochrome c oxidase, the kinetics of ferrocytochrome c oxidation by the bovine cytochrome c oxidase depleted of its subunit III were studied and compared with those of the oxidase isolated from P. denitrificans which contains only two subunits. The aggregation state of both enzymes dispersed in dodecyl maltoside was also compared. The two-subunit oxidase from P. denitrificans gave linear Eadie-Hofstee plots and the enzyme resulted to be monomeric (Mr = 82 000) both, in gel filtration and sucrose gradient centrifugation studies. The bovine heart subunit III depleted enzyme, under conditions when the P. denitrificans cytochrome c oxidase was in the form of monomers, was found to be dimeric by sucrose gradient centrifugation analysis. At lower enzyme concentrations monomers were, however, detected by gel filtration. Depletion of subunit III was accompanied by the loss of small polypeptides (VIa, VIb and VIIa) and of almost all phospholipid (1-2 molecules were left per molecule of enzyme). The electron-transfer activity of the subunit III-depleted enzyme showed a monophasic Eadie-Hofstee plot, which upon addition of phospholipids became non-linear, similar to that of the control bovine cytochrome c oxidase. One of the roles of subunit III may be that of stabilising the dimers of cytochrome c oxidase. Lack of this subunit and loss of phospholipid is accompanied by a change in the kinetics of electron transfer, which might be the consequence of enzyme monomerisation.
Animals↗
Effect of ibuprofen on the healing phase of experimental myocardial infarction in the rat.
The healing phase of acute myocardial infarction (AMI) is initiated by proteolysis of necrotic myocardium, followed by infiltration of fibroblasts and deposition of collagen. To assess whether ibuprofen, a potent antiinflammatory agent, preserves existing collagen and enhances deposition of new collagen during infarct healing, biochemical and morphologic studies were made of experimentally induced myocardial infarcts in untreated rats and in rats treated with ibuprofen. All treated rats received 12.5 mg/kg of ibuprofen at 1, 6 and 18 hours after AMI. Group 1 rats underwent measurement of myocardial hydroxyproline (HP) content at 24 hours after AMI. Group 2 rats received ibuprofen, 12.5 mg/kg, twice a day for 2 additional days, with measurement of myocardial HP at 3 days (group 2a) or 21 days (group 2b) after AMI. Group 3 rats received ibuprofen, 12.5 mg/kg, twice a day for 6 additional days with measurement of HP content, or infarct size and degree of thinning at 21 days after AMI. Compared with untreated rats, ibuprofen-treated rats had significantly greater amounts of HP in the infarct at 24 hours (group 1, 8.9 +/- 2.2 nmol/mg dry weight vs untreated, 7.1 +/- 2.8 nmol/mg dry weight, p less than 0.04) and at 21 days (group 2b, 112 +/- 37 nmol/mg dry weight vs untreated, 91 +/- 39 nmol/mg dry weight, p less than 0.05, and group 3, 125 +/- 51 nmol/mg dry weight vs untreated, 91 +/- 39 nmol/mg dry weight, p less than 0.003). Substantial scar thinning was noted in all rats; no difference in scar thinning was noted between treated and untreated rats at 21 days after AMI.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
Molecular conversion between monomeric and dimeric states of the mitochondrial cytochrome b-c1 complex: isolation of active monomers.
Bovine heart cytochrome b-c1 complex dispersed in 0.1% dodecylmaltoside, 10 mM Tris-HCl (pH 7.4), was subjected to filtration on Ultrogel AcA 34 columns. Apparent Mr values of about 400,000 and 170,000 were estimated for the enzyme-detergent complex in the presence and absence of 50 mM KCl, respectively. Similar Mr values (about 390,000 and 160,000) were obtained after sucrose gradient centrifugation of the b-c1 complex species isolated using Ultrogel filtration. Both species contained eight polypeptides, as in the original cytochrome b-c1 complex. The experiments suggest that the two species represent a dimer and a monomer of the b-c1 complex. The molecular conversion between the monomeric and dimeric state of the enzyme was found to be reversible. Both monomers and dimers of the b-c1 complex were competent to catalyze QH2:cytochrome c reductase activity with approximately the same maximal velocity. The finding that both molecular forms of the enzyme appear equally active does not support functional models based exclusively on a dimeric b-c1 complex.
Animals↗
The aggregation state of bovine heart cytochrome c oxidase and its kinetics in monomeric and dimeric form.
The monomeric and dimeric forms of bovine cytochrome c oxidase (EC 1.9.3.1) were obtained from gel filtration chromatography on Ultrogel AcA 34 and analyzed. Both species contained all 12-13 subunits described for this enzyme. In the dimer 320 molecules [3H]dodecyl-beta-D-maltoside were bound per heme aa3 and in the monomer 360 molecules per heme aa3. The monomers contained 10 mol of tightly bound phospholipid/mol heme aa3 and the dimers 14. Sedimentation coefficients of 15.5-18 S for the dimer and 9.6 S for the monomer were calculated from sucrose density centrifugation analysis and analytical centrifugation. By the laser beam light-scattering technique a Stokes radius of 70 A for the dimeric detergent-lipid-protein complex was measured. From those parameters and the densitometric determined partial specific volumes of the detergent and the enzyme, the molecular weights of 400,000 for the protein moiety of the dimer and 170,000-200,000 for the monomer were calculated. Under very low ionic strength conditions the monomer/dimer equilibrium was found to be dependent on the protein concentration. At low enzyme concentrations (10(-9) M) monomers were predominant, whereas at concentrations above 5 X 10(-6) M the amounts of dimers and higher aggregates were more represented. The cytochrome c oxidase activity, measured spectrophotometrically and analyzed by Eadie-Hofstee plot, was biphasic as a function of cytochrome c concentration for the dimeric enzyme. Pure monomers gave monophasic kinetics. The data, fitting with a homotropic negative cooperative mechanism for the dimer of cytochrome c oxidase, are discussed and compared with other described mechanisms.
Animals↗
The interconversion between monomeric and dimeric bovine heart cytochrome c oxidase.
Monomers and dimers of bovine heart cytochrome c oxidase (EC 1.9.3.1.) were separated by gel filtration chromatography on Ultrogel AcA 34 or by sucrose gradient centrifugation. Factors influencing the interconversion of the two aggregation states of this enzyme were analyzed. At very low ionic strength, in the presence of dodecyl maltoside, monomers were the main species. Salts appeared to stabilize the dimeric form, divalent cations being more efficient than monovalent. High enzyme concentrations favoured the formation of dimers, also at low ionic strength. The type of detergent had a strong influence on the monomer-dimer interconversion; in Triton X-100 and dodecyl maltoside (at high ionic strength) cytochrome c oxidase was homogenously dispersed in its dimeric form, while in Tween-80 gel filtration showed only large particles eluting in the void volume. In cholate monomers and aggregates were observed but no dimers. The aggregation state had an influence on the steady state kinetics of the ferrocytochrome c oxidase activity. Monomers showed linear Eadie-Hofstee plots, whilst the dimeric and aggregated enzyme gave nonlinear Eadie-Hofstee plots. Ionic strength, enzyme concentration and type of detergent were affecting the enzyme's kinetics in a way consistent with the molecular form obtained by the gel filtration or sedimentation analysis. The data support a negative cooperative mechanism for the interaction of cytochrome c with the dimeric enzyme, as proposed earlier (K.A. Nałecz et al., (1983) Biochem. Biophys. Res. Commun., 114, 822-828).
Animals↗
Influence of alpha-adrenergic blockade on platelet-mediated thrombosis in stenosed canine coronary arteries.
The functional significance of platelet alpha-adrenergic receptors in vivo is uncertain. The aim of this study was to elucidate their role in experimental coronary thrombosis. In 46 open-chest dogs with a critical coronary stenosis produced by plicating the coronary artery wall with a suture, blood flow showed cyclical reductions followed by an abrupt return to control levels. The flow reduction were previously shown to be caused by platelet aggregation. In the present study they were unaffected by heparin (1,000 U . kg-1 . h-1) at the start of the experiment but consistently abolished by aspirin (30 mg . kg-1) at the end of the protocol, further supporting platelet aggregation as the primary mechanism. After 1 h of observation, dogs were assigned to one of the following groups: control (no intervention, n = 9), phentolamine (a nonselective alpha-blocker) "low dose" (0.07 mg . kg-1 bolus followed by 0.1 mg . kg-1 . min-1, n = 4), phentolamine "high-dose" (3 mg . kg-1 bolus followed by 0.07 mg . kg-1 . min-1, n = 6), prazosin (an alpha 1-blocker, 2 mg . kg-1, n = 8), yohimbine (an alpha 2-blocker, 3 mg . kg-1, n = 11) or prazosin + yohimbine (same doses, n = 8). In controls, cyclical reductions in flow continued unchanged for another hour. Phentolamine effected a partial dose-related inhibition of flow reductions; however, prazosin and yohimbine, given separately or in combination, failed to produce any significant effect despite an alpha-blocking action equivalent to or greater than that of phentolamine (alpha-agonist dose-response studies).(ABSTRACT TRUNCATED AT 250 WORDS)
Adrenergic alpha-Antagonists↗
Enhancement of recovery of myocardial function by oxygen free-radical scavengers after reversible regional ischemia.
Reperfusion after reversible regional ischemia has been shown to result in delayed recovery of myocardial function, but the mechanism responsible for this phenomenon remains unknown. We explored the potential role of oxygen-free radicals as mediators of postischemic dysfunction in open-chest dogs undergoing a 15 min occlusion of the left anterior descending coronary artery (LAD) followed by 2 hr of reperfusion. Treated animals (n = 19) received an infusion of the oxygen free-radical scavengers superoxide dismutase (SOD; 15,000 U/kg) and catalase (CAT; 55,000 U/kg) for 1 hr starting 15 min before LAD occlusion, while control animals (n = 20) received an equal volume of saline. SOD and CAT produced no discernible effect on heart rate, aortic pressure, or left atrial pressure. Collateral flow to the ischemic zone (radioactive microspheres) was 0.07 +/- 0.01 ml/min/g in both groups. The size of the occluded bed as determined by postmortem perfusion was 26.1 +/- 1.2% of the left ventricle in the control group and 26.5 +/- 0.9% in the treated group. Systolic wall thickening (an index of regional function) was assessed with an epicardial pulsed-Doppler probe. The two groups exhibited comparable systolic thickening under baseline conditions and similar degrees of dyskinesia during ischemia. Nevertheless, recovery of function (expressed as percent of baseline) was considerably greater in the treated dogs, both at 1 hr (43.8 +/- 14.3 vs 12.8 +/- 11.6) and 2 hr of reperfusion (74.2 +/- 8.4 vs 31.6 +/- 9.8, p less than .005). This improved recovery of function obtained with SOD and CAT suggests that oxygen-free radicals play an important role in the genesis of myocardial dysfunction after a brief episode of regional ischemia.
Animals↗