Search

Restoration of coronary flow after hyperkalemic cardioplegic arrest (HCA) is associated with a unique metabolic reperfusion injury (RI) manifested by declining nucleotide stores despite their end-ischemic preservation. Prevention of this RI by exogenous provision of adenosine with or without inhibition of adenosine’s major catabolic enzyme was assessed in 27 dogs subjected to 60 minutes of HCA. The effect of aortic root infusion of 40 mg/kg of adenosine in addition to adenosine deaminase inhibition by 10 mg/kg of EHNA (group 2) initiated during 60 minutes of reperfusion on myocardial adenosine triphosphate (ATP) and creatine phosphate (CP) stores and coronary blood flow (CBF) were compared to animals having adenosine infusion alone (group 3) or controls (group 1). Although ATP levels were preserved at the end of HCA in all groups, adenosine infusion with or without EHNA prevented the significant 23 percent decline in ATP stores incurred during unmodified reperfusion (p<0.01, group 1). The CP stores decreased (p<0.05, all groups) during arrest, but were restored to preischemic levels during reperfusion. When measured 60 minutes after aortic unclamping, CBF was 312 percent of preischemic flow in group 3 (p<0.01), only 170 percent in group 2 (p<0.05), and unchanged in controls (group 1). The data indicate that provision of adenosine as a nucleotide precursor prevents the metabolic RI following HCA. In addition, inhibition of adenosine catabolism is not necessary for this salutary effect, nor is adenosines efficacy solely mediated by augmentation of CBF.

Metabolic evidence of improved delivery of cardioplegic solutions by adjuvant use of nitroglycerin (NTG) and reinfusing these solutions distal to an obstructed coronary artery was sought in 40 dogs subjected to cold cardioplegic arrest. The left anterior descending coronary artery was occluded prior to initiating arrest by ultra-aortic root infusion. Cardioplegic solution was reinfused with the left anterior descending occluded throughout ischemia (Group I), or with this artery reopened, to simulate a completed distal anastomosis (Group II). Serial biopsy specimens of the left ventricular apex were assayed for adenosine triphosphate and creatine phosphate, while specimens from the posterior left ventricular wall served as controls. Regional myocardial temperatures were recorded throughout ischemia. Half of the hearts in each group received 300 μg of nitroglycerin in the cardioplegic solution. Adenosine triphosphate was preserved in myocardium distal to a patent coronary artery whether nitroglycerin was added to the cardioplegic solution or not (control, control + NTG). Moreover, nitroglycerin did not prevent the 26% to 34% (p < 0.05) decline in adenosine triphosphate levels when the left anterior descending remained obstructed throughout ischemia (Group I, I + NTG). However, opening the left anterior descending for reinfusion of cardioplegic solution allowed adenosine triphosphate to be preserved at end-ischemia (Group II, II + NTG). In addition, the metabolic reperfusion injury manifested by a 37% (p < 0.01) decline in adenosine triphosphate levels after aortic unclamping (Group II) was obviated when nitroglycerin was added to the cardioplegic solution delivered in this manner (II + NTG). The depletion of cardioplegic solution stores during ischemia was more severe in the experimental groups than in controls (p < 0.05). These metabolic changes did not correlate with regional myocardial temperature gradients. The data indicate that myocardium jeopardized by coronary stenoses can be preserved as well as myocardium supplied by a patent coronary artery by adjuvant use of nitroglycerin and varying the mode of delivery of the cardioplegic solution.

• Potassium-induced cardioplegia was studied in 38 mongrel dogs supported by normothermic cardiopulmonary bypass and subjected to 60 minutes of aortic cross clamping followed by 30 minutes of reperfusion. A study of preischemic and postischemic ventricular function and myocardial high-energy phosphate compounds, lactate, and glycogen showed substantial preservation of high-energy phosphates and ventricular performance when potassium cardioplegia was used. However, the substantial depression in contractility observed following ischemia and reperfusion suggests that potassium cardioplegia alone does not provide adequate intraoperative protection of the myocardium.(Arch Surg 113:976-980, 1978)

Intracellular sodium and calcium accumulation were measured after 30 and 40 minutes of ischemia and 30 minutes of reperfusion in newborn (0 to 7 day), neonatal, (14 to 21 day), and adult (4 to 6 month) New Zealand white rabbit heart preparations. Newborn hearts showed twofold and threefold elevations of calcium when exposed to 30 and 40 minutes of ischemia and reperfusion, respectively, whereas sodium increase was noted only in the 40 minute group. Conversely, adult hearts exhibited sodium elevation if exposed to ischemia for 30 minutes and calcium accumulation occurred only after 40 minutes of ischemia. There was no significant alteration in intracellular sodium and calcium levels after ischemic intervals of 30 and 40 minutes in the neonate. These studies demonstrate that the immature newborn heart is the most susceptible and the neonatal heart is the least susceptible to injury sustained by ischemia followed by reperfusion. These differences in susceptibility to ischemia may be due to age-dependent alterations of intramembrane ionic pumps and channels.

In an attempt to arbitrate the reputed clinical efficacy of pulsatile flow during reperfusion in minimizing ischemic injury, 32 mongrel dogs supported by normothermic cardiopulmonary bypass were subjected to 30 minutes (Groups IC and IP) or 60 minutes (Groups IIC and IIP) of global myocardial ischemia. The effect of pulsatile flow (P) initiated during 30 minutes of reperfusion on the recovery of myocardial adenosine triphosphate (ATP) and creatine phosphate (CP) stores, coronary blood flow, and myocardial water content (MWC) was compared to the effect of linear reperfusion (C) in another group of animals. ATP stores, which significantly decreased to 43% and 53% of preischemic levels (Groups IC and IP, respectively, p < 0.01) and 36% and 31% of control values (Groups IIC and IIP, respectively, p < 0.001), did not increase with either pulsatile or linear reperfusion. CP stores, depleted 97% during ischemia in all groups, returned to preischemic levels regardless of the mode of reperfusion flow. Coronary blood flow measured 30 minutes after aortic unclamping was not significantly different from control flow in any group. MWC significantly decreased during ischemia from 80.5% ± 0.8% to 76.5% ± 1.1% in Group IC and from 81.8% ± 1.2% to 76.8% ± 0.8% in Groups IP (p < 0.05) and returned to preischmic levels with reperfusion. However, following 60 minutes of ischemia, pulsatile reperfusion prevented the significant increase in MWC that accrued after linear reperfusion (80.7% ± 1.5% to 84.0% ± 0.7%, p < 0.05). These data indicate that pulsatile reperfusion initiated after an ischemic injury that results in a 50% or greater depletion of myocardial ATP stores does not restore myocardial nucleotide levels or enhance coronary blood flow, although the pathological increase in MWC may be avoided.

Despite meticulous adherence to presently known principles of myocardial preservation, reperfusion after aortic cross-clamping results in a unique injury manifested by decreasing high-energy phosphate levels and increased coronary resistance. We hypothesize that platelet deposition into the coronary microvasculature is a major factor in reperfusion injury. To differentiate platelet deposition due to subendocardial hemorrhage from deposition due to vascular entrapment, we infused 111In-labeled platelets together with 51Cr-labeled erythrocytes into 15 dogs that were on normothermic bypass and subjected to 60 minutes of global ischemia followed by 30 minutes of reperfusion. Platelet deposition is indicated only when the proportion of platelets to erythrocytes in tissue exceeds that measured by peripheral blood. Myocardial biopsy specimens were obtained after 10 minutes of bypass, 120 minutes of continuous bypass (Group I), and at the end of reperfusion after global ischemia (Group II). In five dogs (Group III), dipyridamole (1 mg/kg), an antiplatelet activation agent, was administered in the preischemic period. Platelet deposition was expressed as the number of radioactive-labeled platelets deposited per gram of tissue. Bypass for 120 minutes resulted in only a minimal increase in platelet deposition. However, normothermic ischemia followed by reperfusion resulted in over a twofold increase in platelet deposition compared to controls. Pretreatment with dipyridamole appeared to avoid platelet deposition. These data indicate that platelet deposition in the coronary microcirculation following surgically induced myocardial ischemia may be associated with reperfusion injury and that antiplatelet drugs alter this sequence.

The effect of varying the mode of cardioplegic delivery and the presence of regional wall motion abnormalities on myocardial protection by crystalloid cardioplegic solutions was assessed in 68 patients undergoing coronary artery bypass grafting. Serial transmural biopsy specimens from the left ventricular apex were assayed for adenosine triphosphate. All patients had more than 75% stenosis of the left anterior descending coronary artery. They were prospectively randomized into Groups I and II to receive (I) all cardioplegic solution infused via the aortic root or (II) reinfusions of cardioplegic solution given both centrally and through the completed distal left anterior descending anastomosis. Patients were also stratified as to the presence of normal (N) or impaired (Ab) apicoanterior regional wall motion. Inadequate delivery of cardioplegia during ischemia in Group I was manifested by a 41% (p < 0.01) depletion of adenosine triphosphate stores in abnormally contracting myocardium distal to the left anterior descending stenosis that was not repleted after restoration of coronary flow and a 27% (p < 0.05) decline in ATP stores during reperfusion in myocardium with normal preoperative wall motion. In contrast, nucleotide stores were preserved at preischemic levels throughout ischemia and reperfusion in Group II regardless of preoperative wall motion. Preservation of ATP did not correlate with duration of ischemia, highest recorded septal temperature, or volume of cardioplegic solution infused. Two patients in each group had a new perioperative infarction. However, 38% of patients in Group IAb required transient inotropic support versus 5% in Group IIAb (p < 0.05). These data emphasize that reinfusion of cardioplegic solutions distal to coronary obstructions is mandatory for optimal myocardial protection during coronary revascularization.

We have observed that high-resolution ultrasonic imaging can be used to detect and measure the size and sedimentation of large human platelet aggregates formed in vitro. At progressively lower shear rates, individual platelet aggregates grew in cross-sectional area from the limit of ultrasonic detectability of less than 1 mm2 at moderate shear rates to an approximate mean area of 25 mm2 at zero shear. Platelet aggregates tended to sediment at mean shear rates higher than zero shear. At a shear rate of 5.5 s-1, 70% of platelet aggregates had settled, and at 1.6 s-1, 90% had settled to the dependent half of horizontal tubes in less than 15 s. Ultrasonic aggregometry of platelets may be a valuable technique for investigation of platelet dynamics under controlled flow conditions in vitro.

Developmental differences in ischemic and potassium cardioplegic arrest were evaluated in newborn (birth to 7 day old) and adult (6 to 12 month old) New Zealand white rabbit hearts isolated and perfused by Langendorff’s method. An extracellular space washout technique was used to measure intracellular sodium and calcium in the two age groups after ischemia alone, after normothermic and hypothermic cardioplegia, and after cardioplegia with reperfusion. Although the intracellular ionic contents of nonreperfused adult hearts after 30 and 40 minutes of ischemia were identical, there was a twofold elevation in intracellular sodium level after 40 minutes of ischemia in the newborn hearts. Adult hearts arrested by normothermic potassium cardioplegia demonstrated no alteration in the intracellular ionic content, whereas in the newborn hearts, potassium cardioplegia produced excess intracellular calcium loading before reperfusion, which was greater than that occurring with ischemia alone. When hypothermia (12° C) was combined with cardioplegic arrest, a prereperfusion influx of sodium and calcium was not observed in the newborn hearts, and ionic reperfusion injury was blunted in both newborn and adult hearts. These studies demonstrate that the newborn heart is more susceptible than the adult to both ischemia and cardioplegia. This may be due to age-dependent differences in transmembrane passive diffusion, sodium/calcium exchange, or calcium slow channel properties and suggests alternative myocardial protective strategies for the newborn infant.

Using high-resolution real-time ultrasound to monitor platelet aggregation and plasma clotting, the effect of protamine on platelets was evaluated in a dynamic system of plasma in vitro. Protamine induced platelet aggregation preceding plasma clotting at both low (1.6 s-1) and moderate (22.6 s-1) shear rates. The onset of aggregation and clotting was accelerated at the higher shear rate. Low-shear clots were heterogeneous on ultrasonic imaging, whereas moderate-shear clots were more densely homogeneous. Protamine-induced platelet aggregation was reversed by additional heparin. Pretreatment of plasma with pros-tacyclin prevented protamine-induced aggregation, but clot formation occurred. However, such clots were less echogenic. EGTA and hirudin inhibited both aggregation and clotting. Our ultrasonic aggregometry showed that platelet aggregation was induced by protamine before clots formed, and that not only shear rate but platelet activation might affect the rate and composition of plasma clots.

Previous studies assessing the efficacy of myoprotective regimens have compared preischemic and postischemic myocardial oxygen consumption within a limited range of cardiac performance. However, recent data suggest that ischemia-induced perturbations in myocardial energetics may occur only when the left ventricle develops physiologic pressures. Therefore, in canine hearts supported by cardiopulmonary bypass, myocardial oxygen consumption (ml oxygen × 10 2/beat/100 gm left ventricular weight) was determined during incremental isovolumic pressure-volume loading before and 30 minutes after 2 hours of cardioplegic arrest The ischemic insult was graded by maintaining myocardial temperature at 12° C (Group I, n = 6), 20° C (Group II, n = 7), or 28° C (Group III, n = 6). Postischemic Starling curves were unchanged in Groups I and II but depressed 53% in Group III hearts (p < 0.005). In Group I, postischemic myocardial oxygen consumption at specific peak developed pressures was similar to preischemic oxygen consumption. In contrast, postischemic Group II and III hearts consumed 39% more oxygen than preischemically when peak developed pressure exceeded 75 mm Hg (p < 0.01). Postischemic hearts demonstrated reciprocal changes in arteriovenous oxygen content difference (24%, 30%, and 34% lower than preischemic values for Groups I, II, and III, respectively) and coronary blood flow (156%, 195%, and 192% higher than preischemic values for Groups I, II, and III, respectively). Only in Group II and III hearts did the increased coronary blood flow offset the defect in oxygen extraction such that myocardial oxygen consumption was increased. These data suggest that inefficient utilization of oxygen when the heart is developing physiologic pressures is a sensitive marker for myocardial injury after crystalloid cardioplegic arrest.

Neutrophils are known to undergo a burst of metabolic activity in response to particulate and soluble stimuli. Most of these stimuli depend upon calcium in the extracellular medium to exert their maximal effects. We have shown that 24 mMsodium fluoride (NaF) stimulates the metabolic burst in canine neutrophils as measured by chemiluminescence and CO2formation from the carbon-1 position of glucose. Because these phenomena are calcium dependent, we determined the uptake of calcium by neutrophils activated with NaF. Neutrophils suspended in buffer were incubated with 45CaCl2and exposed to NaF. This caused an increase in cell-associated calcium from 1.89 ± 0.43 × 10−10mol/107cells to 1.06 ± 0.15 × 10−8mol/107cells in 2 min. In comparison, the calcium ionophore A23187 (10−6M) caused only a modest increase in cell calcium, from 1.89 ± 0.43 × 10−10mol/107cells to 4.89 ± 0.85 × 10−10mol/107cells. When treated with EGTA (3 mM), NaF-stimulated neutrophils rapidly lost 90% of their cell-associated calcium. Verapamil (3 × 10−4M), a calcium channel blocker, inhibited NaF stimulation of neutrophil metabolism but did not decrease the magnitude or the rate of the calcium association. These studies indicate that the mechanism of NaF stimulation of neutrophils involves massive calcium binding to the external membrane and a subsequent movement of some of this membrane calcium into the cell.