Ischemic Heart Disease and its Consequences

Myocardial Stunning

Myocardial stunning is defined as ‘the mechanical dysfunction that persists after reperfusion despite the absence of irreversible damage’. It was first described by Heyndrickx and colleagues in 1975, when the authors reported that coronary occlusions in the dog lasting for periods not long enough to induce cell death produced prolonged periods of contractile dysfunction. Subsequent studies demonstrated that reversible contractile dysfunction occurs in experimental models of demand ischemia in the hypertrophic heart, in isolated hearts subjected to global ischemia, and after cardiac surgery. Although the traditional definition of cardiac stunning indicated that the ischemic insult was non-lethal (i.e. not of sufficient duration to induce cell death), cardiac stunning also occurs after prolonged coronary artery occlusion resulting from myocardial infarction, as evidenced by myocardium salvaged by reperfusion, which can remain stunned with delayed contractile recovery. Importantly, experimental work has demonstrated the occurrence of stunning in all animal species investigated, suggesting that, despite the differences in contractile physiology, this is a universal phenomenon.

Reperfusion Arrhythmias

Reperfusion arrhythmias result from complex cellular and humoral reactions that accompany the opening of a blocked coronary artery. Early insights into the conditions that produce reperfusion arrhythmias came from experiments that subjected the hearts of large animals to varying periods of coronary artery occlusion followed by reperfusion. In these experiments, it was noted that the incidence of reperfusion-induced ventricular fibrillation increased when the ischemic period extended from five minutes to 20 or 30 minutes. In contrast, the incidence of reperfusion-induced ventricular fibrillation declined when reperfusion was delayed beyond 30 to 60 minutes. These studies also found that reperfusion-induced ventricular fibrillation was more frequent when severe arrhythmias developed during the ischemic period. In humans, the most common reperfusion arrhythmia is an accelerated idioventricular rhythm. However, ventricular tachycardia and ventricular fibrillation remain the most important causes of sudden death following spontaneous restoration of antegrade flow.

Endothelial Dysfunction

Endothelial dysfunction is a systemic pathological state of the endothelium characterized by an impairment in endothelium-dependent vasodilation and an exacerbation in the response to endothelium-dependent vasoconstrictors. Endothelial dysfunction occurs during the early reperfusion period and persists. Experimental studies have identified that vascular injury begins with an endothelial triggering phase followed by a neutrophil amplification phase. For instance, within five minutes of reperfusion the endothelium becomes dysfunctional as the production of nitric oxide (NO) decreases. By 20 minutes of reperfusion, leukocytes start to adhere to the endothelium and neutrophils begin to migrate across the endothelium into the damaged tissue. Once in the tissue, activated neutrophils release cytotoxic and chemotactic substances, such as cytokines, proteases, leukotrienes, and oxygen-derived free radicals. In the experimental setting, endothelial dysfunction after ischemia–reperfusion injury can be identified after 4–12 weeks. A combination of endothelial dysfunction, microvascular obstruction (downstream microembolism of platelets, de novo thrombosis, and/or neutrophil capillary plugging), edema, and oxidative stress is responsible for the pathogenesis of microvascular dysfunction following ischemia–reperfusion injury. Sometimes, severe microvascular dysfunction limits adequate perfusion after reperfusion. This is known as the ‘no-reflow’ phenomenon and is characterized by the absence of tissue perfusion despite both epicardial coronary artery patency and flow. Although the underlying mechanisms of this phenomenon have not been fully elucidated, the result is microvascular damage produced by microvascular vasoconstriction and obstruction associated with reperfusion injury. No-reflow after coronary revascularization therapy is associated with incomplete ST-segment recovery and increases the incidence of acute myocardial infarction, myocardial rupture, and death. Therefore, the recognition of no-reflow affords an opportunity for therapeutic intervention designed to augment tissue perfusion and maintain the viability of myocardium at risk for infarction.

Irreversible Cell Death

While arrhythmias, contractile impairment, and impaired coronary blood flow can be either reversible or irreversible, myocardial cell death or infarction is irreversible. As such, myocardial infarction is the most robust endpoint of all studies designed to investigate myocardial ischemia–reperfusion injury. Studies demonstrate that the myocardium undergoes rapid ultrastructural changes after the onset of ischemia. These alterations tend to be reversible if reperfusion is established promptly. However, if blood flow is not rapidly restored (20–30 minutes of ischemia), these reversible alterations undergo a transition to a state of irreversible tissue injury characterized by cell death. Originally, cardiac myocyte cell death following ischemia–reperfusion injury was hypothesized to occur through the process of necrosis. However, studies over the last 20 years have challenged this dogma and established that apoptotic cell death also contributes in a meaningful way to the development of myocardial infarction after ischemia–reperfusion injury.

Apoptosis is characterized by cytoplasmic shrinkage, plasma membrane blebbing, nuclear condensation, and fragmentation of both the cytoplasm and nucleus into membrane-enclosed apoptotic bodies. These bodies are subjected to phagocytosis by macrophages or even neighboring cells, thereby avoiding an inflammatory response. Apoptosis is an actively regulated form of cell death that is mediated by two pathways: extrinsic and intrinsic pathways ( Figures 5.3 and 5.4 ). The extrinsic pathway utilizes cell surface receptors to signal through death effector domains to cleave and activate downstream procaspases. The activated caspases then amplify apoptotic signaling by cutting and stimulating a variety of additional pro-apoptotic mediators. In contrast, the intrinsic pathway involves the mitochondria and endoplasmic reticulum. A key feature of the intrinsic pathway involves the permeabilization of the outer mitochondrial membrane, which permits the release of several apoptogens (e.g. bcl-1, bax) into the cytosol. In the cytosol, these apoptogens trigger downstream targets to facilitate the activation of caspases. Therefore, both pathways lead to the activation of caspases. Interestingly, most of the proteins that are released from the mitochondria in response to apoptotic signaling have important physiological functions in healthy cells, but exhibit pathological properties when discharged into the cytosol. Both the extrinsic and intrinsic apoptosis pathways have been shown to play an important role in the development of myocardial infarction following ischemia–reperfusion injury. For example, mice deficient in the death receptor, Fas, display distinct reductions in infarct size after myocardial ischemia–reperfusion when compared to wild-type controls. Additionally, mice with a cardiac-specific overexpression of the anti-apoptogene, Bcl-2, also display distinct reductions in infarct size, as well as cardiac myocyte apoptosis and left ventricular dysfunction after myocardial ischemia–reperfusion when compared to wild-type mice. Similarly, mice deficient in the pro-apoptogene, Bax, display smaller infarcts following myocardial ischemia–reperfusion and treatment with various caspase inhibitors leads to a reduction in infarct size.

In contrast to apoptosis, necrosis has been the conventional example of a type of unregulated cell death. Although a significant proportion of necrotic deaths are passive, evidence has emerged to indicate that much like apoptosis, necrosis can also be regulated. The exact parts of the necrotic cell death pathway that are unregulated as opposed to those parts that are regulated have not been established. However, regulated necrosis has clearly been shown to be an important component of myocardial cell death after ischemia–reperfusion injury. Currently, efforts are underway to determine the molecular components of the necrosis-signaling cascade and to determine if it can be targeted with pharmacological intervention. A key feature of necrosis is the opening of the mitochondrial permeability transition pore (MPTP), a non-selective channel found in the inner mitochondrial membrane. While the components of the MPTP still remain unknown, cyclophilin D clearly plays a regulatory role in its opening. The MPTP opens in response to elevated matrix calcium (Ca ²⁺ ) concentration, oxidative stress, elevated phosphate concentration, and adenine nucleotide depletion. As a result, there is a loss of the electrical potential difference (Δψ _m ) across the inner mitochondrial membrane, which ultimately leads to ATP depletion. Additionally, opening of the MPTP causes mitochondrial swelling due to an influx of water down its osmotic gradient into the mitochondrial matrix. Furthermore, swelling of the inner mitochondrial membrane can cause the rupture of the outer mitochondrial membrane resulting in the release of mitochondrial apoptogens into the cytosol. Whether or not the release of these factors adds to necrotic cell death is currently not known. However, the rupturing of the outer mitochondrial membrane under these conditions is different from the permeabilization that occurs during apoptosis. Opening of the MPTP is the most apparent link between myocardial ischemia with or without reperfusion and necrosis. Studies have demonstrated that events during ischemia and events that occur after reperfusion contribute to the opening of the MPTP.

Cyclophilin D regulates the opening of the MPTP without being a component of it. Studies have shown that hearts from cyclophilin-D-deficient mice display smaller infarct sizes after myocardial ischemia–reperfusion when compared to wild-type control mice. Necrotic cell death can also be activated through serine/threonine protein kinases in the receptor interacting protein (RIP) family. Inhibition of RIP1 with the small molecule inhibitor, necrostatin-1, has been reported to decrease infarct size after myocardial ischemia–reperfusion injury. Interestingly, necrostatin-1 did not reduce infarct size further in mice deficient in cyclophilin D, suggesting that there is a link between the RIP kinase-dependent necrotic pathway and the opening of the MPTP. Although the molecular nature of such a connection remains to be determined, one possibility is the generation of oxidative stress from the activation of metabolic pathways by RIP during necrosis.