We have previously shown that a brief episode of ischemia slows the rate of ATP depletion during subsequent ischemic episodes. Additionally, intermittent reperfusion may be beneficial to the myocardium by washing out catabolites that have accumulated during ischemia. ' Thus, we proposed that multiple brief ischemic episodes might actually protect the heart from a subsequent sustained ischemic insult. To test this hypothesis, two sets of experiments were performed. In the first set, one group of dogs (n = 7) was preconditioned with four 5 min circumflex occlusions, each separated by 5 min of reperfusion, followed by a sustained 40 min occlusion. The control group (n = 5) received a single 40 min occlusion. In the second study, an identical preconditioning protocol was followed, and animals (n = 9) then received a sustained 3 hr occlusion. Control animals (n = 7) received a single 3 hr occlusion. Animals were allowed 4 days of reperfusion thereafter. Histologic infarct size then was measured and was related to the major baseline predictors of infarct size, including the anatomic area at risk and collateral blood flow. In the 40 min study, preconditioning with ischemia paradoxically limited infarct size to 25% of that seen in the control group (p < .001). Collateral blood flows were not significantly different in the two groups. In the 3 hr study, there was no difference between infarct size in the preconditioned and control groups. The protective effect of preconditioning in the 40 min study may have been due to reduced ATP depletion and/or to reduced catabolite accumulation during the sustained occlusion. These results suggest that the multiple anginal episodes that often precede myocardial infarction in man may delay cell death after coronary occlusion, and thereby allow for greater salvage of myocardium through reperfusion therapy. Circulation 74, No. 5, 1124-1136, 1986 OCCLUSION of a major coronary artery in the dog induces injury that is reversible for the first 15 to 20 min, in that complete tissue recovery ensues after reperfusion. However, if the occlusion is maintained beyond this time, the injury becomes irreversible; some cells die despite reperfusion. Ischemic cell death begins first in the subendocardial region and then progresses with time toward the subepicardium.2 A previous study from our laboratory' has shown that repeated brief episodes of ischemia do not have a cumulative deleterious effect. Four 10 min occlusions produced no more ATP depletion than a single occlusion, and did not cause necrosis, although 40 min of sustained ischemia has been associated with severe

A B S T R A C T The role of microvascular damage in the genesis of the "no-reflow" phenomenon was investigated in the left ventricular myocardium of dogs subjected to temporary occlusions of a major coronary artery for 40 and 90 min. Intravenous carbon black or thioflavin S (a fluorescent vital stain for endothelium) were used to demonstrate the distribution of coronary arterial flow in control and damaged myocardium. These tracers were injected simultaneously with release of the coronary occlusion or after 5 or 20 min of reflow of coronary arterial blood. After 40 min of ischemia plus arterial reperfusion, usually the tracers were evenly distributed throughout the damaged tissue at each time of reperfusion. On the other hand, when reflow was allowed after 90 min of ischemia, portions of the inner half of damaged myocardium were not penetrated by the tracers. Electron microscopic study of this poorly perfused tissue revealed severe capillary damage; endothelial cells with large intraluminal protrusions and decreased pinocytic vesicles were common. Also, occasional intraluminal fibrin thrombi were noted, as well as extravascular fibrin deposits and erythrocytes. Myocardial cells were swollen in both poorly perfused and well-perfused irreversibly injured tissue. Contraction bands and mitochondrial Ca"+ accumulation were prominent features of irreversible injury with reflow at 40 min but were not noted after 90 min of ischemia in areas with poor perfusion. These results suggest that 40 min of ischemia were tolerated by the capillary bed of the dog heart without serious capillary damage or perfusion defects, but that 90 min of ischemic injury was associated with the "no-reflow" phenomenon, i.e., failure to achieve uniform reperfusion. This failure of reflow was associated with extensive capillary damage and myocardial cell swelling. Death of severely ischemic myoMr. Kloner was a predoctoral fellow during the time of this study (National Institutes of Health grant GM 00131).

Irreversible ischemic myocardial cell injury developes in an increasing number of cells as the duration of coronary occlusion is prolonged. The present study quantitates myocardial necrosis produced by 40 minutes, 3 hours, or 6 hours of temporary circumflex coronary occlusion (CO) followed by 2 to 4 days of reperfusion, or by 24 or 96 hours of permanent circumflex ligation in pentobarbital anesthetized open chest dogs. After 40 minutes of ischemia, myocyte necrosis was subendocardial but with increasing duration of coronary occlusion, irreversible injury progressed as a wavefront toward the subepicardium. Transmural necrosis was 38 +/- 4% after 40 min, 57 +/- 7% after 3 hours, 71 +/- 7% after 6 hours and 85 +/- 2% after 24 hours of ischemic injury. These results document the presence of a subepicardial zone of ischemic but viable myocardium which is available for pharmacologic or surgical salvage for at least three and perhaps six hours following circumflex occlusion in the dog.

We have shown previously that preconditioning myocardium with four 5-minute episodes of ischemia and reperfusion dramatically limited the size of infarcts caused by a subsequent 40-minute episode of sustained ischemia. The current study was undertaken to assess whether the same preconditioning protocol slowed the loss of high energy phosphates, limited catabolite accumulation, and/or delayed ultrastructural damage during a sustained ischemic episode. Myocardial metabolites and ultrastructure in the severely ischemic subendocardial regions were compared between control and preconditioned canine hearts. Hearts (four to 10 per group) were excised after 0, 5, 10, 20, or In an attempt to separate the effects of catabolite accumulation from those of ATP depletion, we have examined the effects of repeated coronary occlusions. We hypothesized that repeated coronary occlusions might cause partial depletion of ATP, whereas intermittent reperfusion would wash out ischemic catabolites. In a recent study,' we proposed that multiple brief ischemic episodes actually might protect the heart from a subsequent sustained ischemic insult. To test this hypothesis, dogs were "preconditioned" with four 5-minute episodes of ischemia, each separated by 5 minutes of reperfusion, and then subjected immediately to a sustained occlusion of either 40 minutes or 3 hours. Control animals received either a single 40-minute or 3-hour occlusion. In the 40-minute study, infarcts in preconditioned hearts were smaller than controls at any level of collateral flow, averaging 25% of control infarct size. In the 3-hour study, preconditioning had no effect on infarct size.by guest on May 12, 2018