Stanley R. Jolly

Canine myocardial reperfusion injury. Its reduction by the combined administration of superoxide dismutase and catalase.

Therapy directed against the toxic effects of reactive oxygen species may reduce the final extent of ischemic injury in otherwise viable tissue irreversibly injured by the abrupt reoxygenation of reperfusion. In four groups of dogs, superoxide dismutase plus catalase (groups I—III) or saline (controls) (group IV) was infused into the left atrium. Group I received the infusion for 2 hours, beginning 15 minutes before occlusion of the left circumflex coronary artery (90 minutes) and ending 15 minutes after reperfusion. Group II received the infusion for 1 hour starring 15 minutes before reperfusion. Group III received the infusion for 1 hour beginning 40 minutes after reperfusion. Dogs were killed the next day, and infarct size was determined by dissection and weighing, and confirmed histologically. Infarct size expressed as percent of the anatomic area at risk was: group I, 19.4 ± 5.0; group II, 21.8 ± 3.3; group III, 47.6 ± 10.3; group IV, 43.6 ± 3.5 (mean ± sem). Analysis of variance followed by Duncans multiple range test showed that ultimate infarct size as assessed in groups I and II differed significantly (P < 0.05) from that observed in the control animals in group IV, whereas infarct size between groups III and IV did not differ significantly (P > 0.05). The percent of left ventricle at risk did not differ between the four groups. The beneficial effects of superoxide dismutase plus catalase could not be explained by hemodynamic differences. Similar protection of jeopardized myocardium in groups I and II suggest that potentially viable tissue is salvaged by scavenging free radicals during early reperfusion. Lack of protection in group III suggests that injury has occurred within the first 40 minutes of reperfusion. The results of this investigation demonstrate that the ‘primary’ myqcardial cellular damage due to ischemia is additive to the cardiac cell damage during the phase of reperfusion, and that the “secondary” effects are mediated by toxic metabolites of oxygen.

Reduction of myocardial infarct size by neutrophil depletion: Effect of duration of occlusion

Experiments were performed in the dog to examine the effects of neutropenia on ultimate infarct size resulting from short (90 minutes) or prolonged (4 hours) circumflex coronary artery occlusion. Sheep antiserum to canine neutrophils was used to produce neutropenia. Control animals received nonimmune serum. Neutrophil infiltration into myocardial infarcts was examined using histopathologic techniques and a semiquantitative scoring system. In 90-minute occlusions with 24-hour reperfusion, neutropenia was associated with the development of significantly smaller infarcts: normopenic group, 43.2% +/- 3.3% (n = 7) vs. neutropenic group, 26.6% +/- 3.7% (n = 10) of the area at risk, means +/- SEM. However, in 4-hour occlusion with 6-hour reperfusion experiments, the tendency of neutrophil depletion to reduce infarct size did not reach statistical significance (46.4% +/- 7.2% vs. 31.5% +/- 6.0% of the area at risk, normopenic vs. neutropenic) despite differences in neutrophil infiltration into the reperfused region. The observed differences in ultimate infarct size could not be attributed to differences in myocardial oxygen consumption. The results suggest that a significant amount of myocardial infarction induced by a limited duration of coronary artery occlusion followed by reperfusion is neutrophil dependent and appears to be less important in determining the fate of myocardium subjected to more prolonged periods of ischemia followed by reperfusion.

Effect of BW755C in an occlusion-reperfusion model of ischemic myocardial injury

BW755C is a new antiinflammatory agent which predominantly inhibits lipoxygenase over cyclooxygenase. Effects of BW755C have been examined in a canine, occlusion-reperfusion, model of ischemic myocardial injury. In pentobarbital anesthetized open-chest dogs, the proximal left circumflex coronary artery (LCX) was occluded for 90 minutes and slowly reperfused using a micrometer-driven occluder. Thirty minutes before occlusion, animals randomly received BW755C, 3 mg/kg (n = 7), or 10 mg/kg (n = 8), or saline (n = 16) by intravenous infusion. The thoracotomy was closed and the animals subsequently were killed at 24 hours. Infarct size and anatomic area dependent on the occluded LCX were determined by a dual staining technique using triphenyltetrazolium and Evans blue. Both doses of BW755C significantly reduced the ultimate extent of irreversible myocardial ischemic injury, whether results were expressed as grams of infarcted tissue or as percent of risk region infarcted. No difference in risk region size was observed between groups. No effects of BW755C on heart rate, arterial pressure, or left circumflex flow were observed. BW755C (10 mg/kg) did not significantly inhibit ex vivo platelet aggregation in response to collagen, adenosine diphosphate, or arachidonic acid. These results suggest that inhibition of lipoxygenase may reduce the extent of ischemic damage to the heart.

Cardioprotective effects of ibuprofen in experimental regional and global myocardial ischemia

Summary The effects of the nonsteroidal antiinflammatory agent, ibuprofen, were evaluated in two animal models of myocardial ischemia: regional myocardial ischemia produced by temporary occlusion followed by reperfusion of the canine left circumflex coronary artery (LCX), and global ischemia in the isolated, blood-perfused feline heart. In control dogs (n = 12), 60 min of LCX occlusion followed by 24 h of reperfusion resulted in 51 ± 4% of the area at risk, and 20 ± 2% of the total left ventricle becoming infarcted. The average infarct size of the group of dogs receiving oral ibuprofen (n = 12: dose of 12.5 mg/kg, q4h) was 33 ± 5% of the area at risk, and 12 ± 2% of the total left ventricle, a reduction in infarct size as compared to the controls of 35% (p > 0.02) and 39% (p < 0.02), respectively. The mechanism by which ibuprofen provided these beneficial effects could not be attributed to a reduction in myocardial oxygen consumption (M&OV0312;O2), estimated by rate–pressure products, or redistribution of regional myocardial blood flow, assessed by radioactive microspheres. The direct effect of ibuprofen on ischemic myocardium was assessed in an isolated, blood-perfused cat heart model of global ischemia. After 60 min of normothermic global ischemia, left ventricular pulse pressure of control hearts (n = 8) at 5 and 10 mm Hg end-diastolic pressure was reduced by 56 ± 97% and 60 ± 12%, respectively, relative to preischemic values. Administration of ibuprofen (12.5 mg/kg, i.v., to the blood donor cat, n = 6) did not significantly prevent the ischemia-induced deterioration of left ventricular function. Normothermic ischemia for 60 min resulted in a significant reduction in M&OV0312;O2 in both the control and ibuprofen-treated hearts. However, after 1 h of reperfusion, the average M&OV0312;O2 for the ibuprofen-treated hearts was 96% greater (4.5 ± 0.4 versus 2.3 ± 0.5 ml/min/100 g) than the average M&OV0312;O2 of the control hearts. While conventional thinking contends that myocardial infarct size may be limited by a reduction in M&OV0312;O2 or a redistribution of blood flow, the data from this study indicate that these two mechanisms are not relevant in the case of ibuprofen.

Effects of lodoxamide on ischemic reperfused myocardium

The beneficial effects of lodoxamide tromethamine (U42585E) have been examined in a canine model of myocardial ischemic injury. Lodoxamide was infused 20 mg/kg/h i.v. starting 30 min before occlusion of the proximal left circumflex coronary artery (LCX) and continuing through 90 min of ischemia. Lodoxamide produced a significant reduction in ultimate infarct size measured at 24 h by postmortem tetrazolium perfusion staining. Infarct size expressed as a percent of the anatomical area at risk was 21.7 ± 2.7 in the treated group vs. 47.0 ± 3.1 in the control group (mean ± SEM). No significant difference in area at risk was observed between treated and control groups. Salvage occurred primarily in subepicardial and midmyocardial tissue without apparent lateral protection. Histological examination confirmed gross results of postmortem staining. The protection appeared to be unrelated to myocardial oxygen demand since no hemodynamic differences between groups were observed at the time of occlusion or throughout the 24-h experimental course. Concurrent studies of ex vivo platelet aggregation showed no effect of lodoxamide on adenosine diphosphate (ADP), collagen, and arachidonic acid-induced aggregation. In vivo antithrombotic effects were evaluated in four conscious dogs by inducing LCX thrombosis with low-amperage stimulation (50 μA for 24 h) of the intimal surface. Occlusive thrombi occurred in all four dogs and were similar to controls. These results suggest that lodoxamide reduces myocardial ischemic injury by a mechanism unrelated to oxygen demand or antiplatelet effects.

Cardiovascular Depression by Verapamil: Reversal by Glucagon and Interactions with Propranolol

Verapamil-induced cardiovascular depression has been examined in dial-urethane-anesthetized open chest dogs. Verapamil was administered slowly intravenously until the mean arterial pressure was decreased by approximately 45 mm Hg. The dose of verapamil required to reach the hemodynamic endpoint was 1,495 +/- (SE) 165 micrograms/kg. In a second group, interactions between beta-adrenergic blockade, propranolol 1 mg/kg (i.v.), and verapamil were examined. Although propranolol alone had only minor hemodynamic effects, the cardiac depressant dose of verapamil was reduced significantly to 450 +/- 105 micrograms/kg. After cardiovascular depression with verapamil or verapamil plus propranolol, glucagon was administered to assess its inotropic activity using cumulative doses of 5, 15, and 45 micrograms/kg over 20 min. Glucagon produced a dose-dependent recovery of heart rate, mean arterial pressure, and PR interval. Depressed contractility assessed by peak positive dP/dt and right ventricular isometric contractile force also recovered after glucagon. These results suggest a significant interaction between the potency of verapamil as a myocardial depressant and the state of the myocardium as affected by beta-blockade. Cardiac depression by verapamil or verapamil in combination with propranolol was reversible by glucagon.

Protective Effects of AICAriboside in the Globally Ischemic Isolated Cat Heart

The effects of a purine precursor, AICAriboside (5-aminoimidazole-4-carboxamide riboside), on postischemic recovery of myocardial function and adenine nucleotides have been studied in the isolated blood-perfused cat heart. The isolated hearts received either AICAriboside or saline prior to 60 min of global ischemia and during 60 min of subsequent reperfusion. After 60 min of global ischemia and reperfusion, left ventricular function of the AICAriboside-treated hearts approached preischemic values, whereas contractile function of the saline-treated hearts remained depressed. At 60 min postischemia, recovery of left ventricular developed pressure (LVDP) was 62 +/- 10% for the saline-treated hearts as compared to 93 +/- 8% for the AICAriboside-treated hearts. Left ventricular compliance of the saline-treated hearts was decreased slightly at 60 min postischemia. In contrast, left ventricular compliance was increased in the isolated hearts which received AICAriboside. Myocardial ATP concentrations were decreased significantly at 60 min postischemia in both the saline-treated hearts and in the AICAriboside-treated hearts relative to nonischemic hearts. Similarly, total adenine nucleotides (TAN = ATP + ADP + AMP) were decreased significantly in both the saline-treated hearts and AICAriboside-treated hearts relative to nonischemic hearts. The present study demonstrates that AICAriboside protected globally ischemic hearts from the mechanical dysfunction associated with an ischemic insult but did not restore ATP or the total adenine nucleotide pool of the postischemic myocardium.

Platelet depletion in experimental myocardial infarction

SummaryAccumulation of platelets in the microvasculature after acute myocardial ischemia may exacerbate tissue injury through the formation of microthrombi and by the release of vasoactive substances. To assess the role of platelets in myocardial ischemic injury and infarction, circulating platelets were reduced by 94±2% (mean±S.E.M.) with sheep antiserum to canine platelets. Regional myocardial ischemia was produced by occlusion of the left circumflex coronary artery (LCCA) for 90 min followed by reperfusion for 5 hours. Infarct size did not differ significantly between antiplatelet serum and nonimmune serum groups: 36±8vs. 43±4% of the area at risk, determined by a post-mortem dual staining technique (p>0.05). A second occlusion-reperfusion control group, sacrificed at 24 hours, did not differ from 5 hr reperfused groups with regard to infarct size. Coronary sinus thromboxane B2 (TXB2) concentrations were not altered significantly by platelet depletion. Histopathologic examination confirmed the presence of necrosis in the infarcted myocardium and revealed substantial leukocytic infiltration in both groups. The results suggest that circulating platelets are not required for the full expression of myocardial ischemic injury resulting from temporary coronary artery occlusion followed by reperfusion.

Beneficial effects of nafazatrom on ischemic reperfused myocardium

The effect of nafazatrom, a new antithrombotic agent, was studied in a canine model of regional myocardial ischemia. Nafazatrom was administered 1 mg/kg intravenously every 6 h for 48 h. After 24 h of drug or placebo administration, animals underwent 90 min of occlusion of the proximal left circumflex coronary artery followed by gradual reperfusion over a period of 30 min. Twenty-four hours later, the animals were sacrificed and infarct size was determined by histochemical staining with triphenyltetrazolium chloride. Nafazatrom-treated animals had a significant reduction in infarct size expressed as a percent of the anatomical area at risk for infarction: 21 +/- 5% in the treated group vs. 41 +/- 5% in the control group (X +/- S.E.M., P less than 0.05). Histological examination confirmed the gross results of postmortem histochemical staining. Salvage of ischemically jeopardized tissue appeared to be unrelated to myocardial oxygen demand as there were no hemodynamic differences between groups. The beneficial effects of nafazatrom are presumably related to a limitation of autolytic processes on the heart during and after ischemia as a result of the drugs ability to inhibit lipoxygenase and to prevent the enzymatic degradation of prostacyclin.

Experimental coronary artery thrombosis in the absence of thromboxane A2 synthesis: evidence for alternate pathways for coronary thrombosis

The actions of the thromboxane synthetase inhibitor, U-63557A. wore evaluated in vivo in anesthetized open-chest dogs by inducing left circumflex coronary artery (LCCA) thrombosis with low amperage electrical stimulation (100 μA for 6 h) of the intimal surface of the vessel, and ex vivo by assessing platelet aggregation and TXB2 production. U-63557A, 10 mg/kg × 5 mg/kg/h i. v., reduced ex vivo platelet aggregation in response to arachidonic acid (0.65 mM) by 93 + 2% (p < 0.05. ± SEM), whereas the concurrent formation of TXB2 was decreased by 78 + 8% (p < 0.05). TXB2 concentration also was reduced significantly in vivo as measured from coronary sinus blood samples; however. 6-keto-PGF concentration was unchanged from predrug values. Despite the significant inhibiton of platelet aggregation and TXB2 production, thrombus mass was not reduced: control, 32.0 + 5.9 mg (n 7): U-63557A. 30.8 × 12.0 mg (n 5, p NS). These results suggest that U-63557A effectively inhibits TXA2 synthetase. but lacks antithrombotic activity in our experimental model. Therefore, substances other than TXA2 may be capable of mediating occlusive coronary artery thrombosis.