Myocardial ischemic conditioning: Pathogenesis

INTRODUCTION — The myocardium possesses adaptive mechanisms that render it resistant to potentially lethal ischemia and reperfusion injury. (See "Reperfusion injury of the heart", section on 'Definition'.)

Myocardial ischemic conditioning refers to interventions that protect the heart from injury due to exposure to prolonged ischemia. The conditioning may be delivered before, during or after the prolonged ischemic insult. Other protective mechanisms include the long-term development of coronary collateral vessels, and myocardial hibernation and stunning. (See "Coronary collateral circulation" and "Pathophysiology of stunned or hibernating myocardium".)

The definition and pathogenesis of ischemic preconditioning and postconditioning will be reviewed here. The clinical implications of these phenomena in the human myocardium and possible future therapeutic applications are discussed separately. (See "Myocardial ischemic conditioning: Clinical implications".)

ISCHEMIC PRECONDITIONING — Myocardial ischemic preconditioning refers to the protection conferred to ischemic myocardium by brief periods of sublethal ischemia separated by periods of short bursts of reperfusion delivered before the ischemic insult [1]. It provides the myocardium with a powerful means of protecting against acute myocardial ischemia.

Ischemic preconditioning has been reproducibly demonstrated in all animal species studied as well as humans [1-5]. It may occur as part of some naturally occurring ischemic syndromes [6]. Furthermore, in addition to enhanced tolerance to lethal cell injury, preconditioning is protective against other end points of ischemia-reperfusion injury, including postischemic contractile dysfunction [7] and ischemia- and reperfusion-induced ventricular arrhythmias [8,9]. Ischemic preconditioning also decreases apoptosis in animals, another mechanism that can contribute to cell death after myocardial ischemia and reperfusion [10]. (See "Reperfusion injury of the heart", section on 'Apoptosis'.)

The protection afforded by ischemic preconditioning is apparent immediately after the preconditioning ischemia, but is short-lived and wanes after one to two hours [11,12]. After 12 to 24 hours following the preconditioning ischemia, a delayed phase of myocardial protection is induced that, although not as powerful as the early phase, is more prolonged and lasts up to 72 hours [13-15]. This delayed phase of resistance to ischemic injury has been termed the "second window of protection" [16], distinguishing it from early or "classic" preconditioning.

Most laboratory studies of ischemic preconditioning have been performed in animals with "normal" hearts; these observations do not necessarily permit conclusions about the influence of coexisting pathology [17]. Preclinical (animal and human) studies have shown that the threshold for protection in co-morbid settings such as diabetes and age is raised and although it is still possible to protect the myocardium, it requires a stronger conditioning stimulus [18-20]. More experiments have shown that hypertrophied myocardium is amenable to preconditioning [21,22]. Some reports have found that the preconditioning response is conserved in senescent myocardium [23], while others have not [24,25]. However, the demonstration of cardioprotection by preconditioning in the clinical setting suggests that patients with ischemic heart disease are amenable to preconditioning. (See "Myocardial ischemic conditioning: Clinical implications".)

Duration — The reduction in infarct size resulting from ischemic preconditioning is a graded phenomenon. A threshold must be reached during preconditioning ischemia to achieve protection during prolonged ischemia. In one animal study, ischemia lasting less than two minutes did not reduce infarct size, even when there were multiple cycles of ischemia; in contrast, infarct size was significantly and progressively reduced by a single ischemic episode lasting 3 or 10 minutes (infarct size 17 percent without ischemic preconditioning versus 9 and 2 percent with 3 and 10 minutes of ischemic preconditioning, respectively) [26]. In other studies, extending the duration of preconditioning ischemia beyond 10 minutes abolished the protection or even has deleterious consequences during the prolonged ischemia [27,28].

Mediators — The mechanisms underlying ischemic preconditioning have not been completely elucidated (algorithm 1) [29]. A number of early theories for the preconditioning effect have been thoroughly investigated and ruled out, including recruitment of collateral vessels [1,2,5,30,31], myocardial stunning [32], or depletion of glycogen [31,33,34]

Considerable progress has been made toward the identification of potential cell triggers, intracellular signaling cascades, and end-effectors involved in ischemic preconditioning [35,36]. There are several lines of evidence suggesting a role for endogenous paracrine mediators, released during the brief period of ischemia and acting on local receptors, as triggers of ischemic preconditioning. Various models of preconditioning in different species have implicated the involvement of substances such as adenosine [2], acetylcholine [37], catecholamines [38], angiotensin II [39], bradykinin [40], endothelin [41], opioids [42], and reactive oxygen species [43].

The relative importance of the mediators may vary with the duration of preconditioning ischemia and timing. In one study, for example, bradykinin was essential when preconditioning ischemia lasted for three minutes while adenosine is more important during preconditioning ischemia lasting 10 minutes [26].

There appears to be an additive interaction of individual preconditioning triggers to achieve the protective threshold. This was illustrated in a report using human atrial muscle in which two individual triggers (a subthreshold hypoxic insult and elevated bradykinin levels by angiotensin converting enzyme inhibition) were insufficient to induce preconditioning when administered separately but, in combination, produced a full protective effect [44].

With respect to the time at which mediators of ischemic preconditioning act, mediators such as adenosine and protein kinase C are released or activated in response to the preconditioning stimulus and therefore act prior to or during the index ischemic period. In contrast, reactive oxygen species and prosurvival kinases act both prior to the index ischemic period and at the time of reperfusion after the index ischemic period, while inhibition of opening of the mitochondrial permeability transition pore occurs only during reperfusion.

Mediators of ischemic preconditioning likely lead to intracellular changes that protect individual myocytes from ischemic reperfusion injury. A cascade of intracellular kinases and subsequent modifications of mitochondrial function, though opening of adenosine triphosphate-sensitive potassium channels and closure of mitochondrial permeability transition pores, have been observed [45,46].

Adenosine — The relative importance of the triggers implicated in preconditioning seems to be dependent upon the species and the end point of ischemia-reperfusion injury studied. Adenosine, an endogenous compound formed from the breakdown of adenosine triphosphate (ATP) in myocytes during periods of ischemia has been consistently shown to be involved in triggering preconditioning in most animal species studied, including humans [47]. An inability to produce adenosine endogenously is associated with refractoriness of the myocardium to ischemic preconditioning while stimulation of myocardial adenosine A1 receptors by an exogenously administered agonist re-induces cardioprotection of preconditioning [48]. Furthermore, the myocardium can be maintained in a preconditioned state against ischemia-reperfusion by repeated activation of the adenosine A1 receptor over a 10-day period, without the development of tachyphylaxis [49].

Protein kinase C — A number of these triggers, such as adenosine, norepinephrine, acetylcholine, and bradykinin, are coupled to pertussis toxin sensitive inhibitory G-proteins (Gi). It has been proposed that activation of these Gi-protein coupled receptors by various ligands results in activation of a number of kinase cascades at the center of which lies protein kinase C (PKC) [50].

One animal study reported that a small increase in intracellular calcium concentration that results from myocardial ischemia and reperfusion can also activate PKC [51]. In another report, a brief intracoronary infusion of calcium chloride mimicked infarct size reduction seen with preconditioning; this effect could be blocked by an inhibitor of calcium influx [52]. Activation of PKC can occur by alpha adrenoreceptor stimulation during ischemic preconditioning and this also requires calcium [53]. This group of enzymes is therefore thought to be an important intermediate in the signal transduction pathway of ischemic preconditioning, resulting in phosphorylation and activation of possible end-effector(s).

A number of subsequent studies supported this hypothesis. Ischemic preconditioning is blocked by pretreatment with specific PKC inhibitors [53,54] and the substitution of preconditioning ischemia with PKC activators mimics the infarct-limiting effects of ischemic preconditioning [54]. The time course of classic preconditioning also parallels the transient translocation of PKC to the sarcolemmal membrane following activation that lasts for about 60 minutes. Which PKC isoenzyme(s) are activated during preconditioning ischemia is not known.

It is also likely that other protein kinases, in parallel with or following activation by PKC, may play a role in classic ischemic preconditioning. Activation of tyrosine protein kinases may be a crucial step in the signaling cascade since genistein, a selective tyrosine kinase inhibitor, abolishes the protective effects of preconditioning [55]. Protein kinase A has also been implicated [56]. The relative positions of PKC, tyrosine kinase, and other kinases in the signal transduction pathway of ischemic preconditioning are currently under investigation. Tyrosine kinase may be downstream of PKC and tyrosine phosphorylation may activate a member of the mitogen activated protein kinase (MAPK) family [55,57].

ATP-dependent potassium channels — ATP-dependent potassium channels (KATP) appear to be involved in the metabolic adaptation to ischemia seen during preconditioning [58]. There is evidence that mitochondrial KATP channels rather than surface sarcolemmal KATP channels are the likely effectors [59,60]. Opening of these channels results in a repolarizing current that reduces the duration of the cardiac action potential, leading to reduced calcium entry into the myocyte [61]. This results in a reduction in cardiac workload, an enhancement of myocardial viability, and a reduction in cardiac sympathetic nerve injury [62].

The potential importance of these channels can be illustrated by the following observations:

●In an experimental model, the transfer of cardiac KATP channel subunits to KATP channel-deficient cells resulted in protection from hypoxia-reoxygenation [63].

●Dysfunction of the mitochondrial KATP channel may be responsible for the failure to precondition the diabetic heart [64].

The KATP channels may interact with the other mediators described above. PKC activates these channels in human and rabbit ventricular myocytes [65], and there may be synergistic action of adenosine and PKC on KATP channels and shortening of action potential duration [66]. (See "Cardiac excitability, mechanisms of arrhythmia, and action of antiarrhythmic drugs".) Further support for the interaction between these factors comes from one study that found that adenosine receptor activation primes the opening of mitochondrial KATP channels in a PKC-dependent manner [67].

Opioid receptors — The activation of delta opioid receptors within cardiac myocytes triggers ischemic preconditioning in animal models and in human atrial tissue [68]; their stimulation mimics ischemic preconditioning while their blockade abolishes it [69-71]. The opioid receptors are coupled to G(i/o) proteins and can activate PKC and mitochondrial KATP channels [72].

Reactive oxygen species — In response to a preconditioning stimulus, the mitochondrial production of reactive oxygen species (ROS) [73], prior to the index ischemic period, acts as a mediator of the preconditioning signal through the activation of various protein kinases such as PKC [43], extracellular-signal regulated kinases (Erk1/2) [74], and p38 MAPK [55,75].

The following observations illustrated the importance of ROS in preconditioning:

●The preconditioning stimulus can induce mitochondrial release of ROS [73].

●Exogenous ROS can mimic the protective effect of ischemic preconditioning [76].

●The presence of an antioxidant during the preconditioning phase can diminish the degree of protection [43,77].

On the other hand, reducing oxidative stress normally generated during reperfusion may be protective [78].

Prosurvival kinases — Signaling through the prosurvival, phosphatidyl inositol 3-OH kinase (PI3K)-Akt cascade [79-81] and ERK1/2 [82,83] during the preconditioning phase appears to contribute to the protective effect of ischemic preconditioning with further suggestions that specific isoforms of P13K, ie, PI3Ka, play a direct cardioprotective role [84]. Signaling through this pathway then results in the activation of endothelial nitric oxide synthase, opening of the mitochondrial KATP channel [85], and cardioprotection due to the mitochondrial generation of ROS [85].

Prosurvival kinases such as Akt and ERK1/2 also appear to contribute to ischemic preconditioning protection at the time of reperfusion, termed the Reperfusion Injury Salvage Kinase (RISK) pathway [82], acting in part by inhibition of mitochondrial permeability transition pore (MPTP) opening [86,87].

An alternative cardioprotective signaling pathway recruited at the time of reperfusion that has been shown to contribute to IPC has been described. Termed the Survivor Activating Factor Enhancement (SAFE) pathway, this cascade involves TNF-α and the JAK-STAT3 pathway, and also appears to terminate on the mitochondrial permeability transition pore (MPTP) [88].

Inhibition of MPTP opening — It has been proposed that a common pathway for ischemic preconditioning, PKC activation, and mitochondrial KATP channel opening to protect the myocardium occurs via inhibition of MPTP opening during reperfusion, thereby preventing uncoupling of the mitochondria [86,87]. The MPTP is a nonspecific channel of the inner mitochondrial membrane that opens during the first few minutes of postischemic reperfusion [89], and is a critical determinant of both apoptotic and necrotic cell death in the setting of ischemia-reperfusion injury [90]. Inhibition of MPTP opening with ischemic preconditioning is believed to be mediated by activation of the prosurvival kinases [86,87].

Second window of protection — The triggers and signaling pathways mentioned above have also been implicated in the delayed phase (second window) of ischemic preconditioning [16]. In animals, preconditioning induces late protective effects against endothelial injury after ischemia and reperfusion [91].

As with classic preconditioning, the specific triggers and mediators involved depend upon the species and the end points studied. As an example, adenosine is an important trigger for the delayed protection against myocardial infarction in the rabbit and rat [92,93]. In a study in rats, adenosine A1 receptor activation induced delayed myocardial protection, mediated by the mitochondrial antioxidant manganese superoxide dismutase [93].

In contrast, delayed preconditioning against stunning, originally described in the pig heart, involves free radicals and increased formation of nitric oxide via the inducible form of nitric oxide synthase [94,95], but not adenosine [96,97]. Nitric oxide directly activates the mitochondrial, but not sarcolemmal, KATP channels; this provides a link between nitric oxide-induced cardioprotection and the KATP channels [98]. The delayed antiarrhythmic effect following preconditioning in the canine myocardium is abolished by pretreatment by dexamethasone, an effect that may be due to inhibition of nitric oxide or products of the cyclooxygenase pathway [99].

Further evidence for a role of the cyclooxygenase pathway in the second window of protection comes from studies in rabbits showing that delayed ischemic preconditioning induces upregulation of cyclooxygenase-2 (COX-2), leading to increased prostaglandin synthesis in the myocardium [100,101]; COX-2 upregulation may be mediated by nitric oxide [102]. Administration of selective COX-2 inhibitors in this rabbit model abolishes both the increase in tissue prostaglandins and the cardioprotective effects of ischemic preconditioning [100,101]; this observation has implications for the use of COX-2 inhibitor therapy in patients with ischemic heart disease. (See "Myocardial ischemic conditioning: Clinical implications", section on 'Therapeutic applications'.)

ISCHEMIC POSTCONDITIONING — Ischemic postconditioning refers to the ability of a series of brief occlusions of either the coronary or peripheral arterial circulation after a severe ischemic insult to protect against ischemic reperfusion injury of the myocardium.

In animal models, ischemic postconditioning (using the coronary circulation) is almost as effective as ischemic preconditioning and involves similar pathogenetic mechanisms [103,104]. Ischemic postconditioning appears to affect cardioprotection by activating survival protein kinases of the reperfusion-injury salvage kinase pathway [105,106]. Postconditioning reduces the number of necrotic, apoptotic, and autophagic cells [107]. (See 'Ischemic preconditioning' above.)

Initial insights into the potential timing of ischemic postconditioning in humans were provided by a forearm model of ischemic postconditioning [108]. After 20 minutes of sustained forearm ischemia, and at the onset of 20 minutes of reperfusion, an ischemic postconditioning protocol was applied comprising three 10- or 30-second cycles of alternate ischemia and reperfusion. Ischemic postconditioning improved endothelial function. No protection was observed if application of the postconditioning protocol was delayed for one minute, a finding that supports preclinical studies in which the postconditioning protocol needs to be instituted within one minute of full reflow [109].

The potential clinical use of ischemic postconditioning is discussed separately. (See "Myocardial ischemic conditioning: Clinical implications", section on 'Therapeutic applications'.)

REMOTE ISCHEMIC CONDITIONING — The major limitation of myocardial ischemic conditioning is that it requires a protective stimulus to be applied to the heart directly. However, it has been discovered that the heart can be protected against acute myocardial infarction by applying brief cycles of (reversible) ischemia and reperfusion to a vascular bed, organ, or tissue away from the heart, termed remote ischemic conditioning [110-112].

The mechanism through which the cardioprotective stimulus is relayed from the remote organ or tissue to the heart is unclear, and neurogenic, hormonal, and systemic factors have been proposed. Furthermore, it has been shown that there may be a co-dependence of both neural and humoral pathways in the overall mechanism of this form of conditioning [113]. It has been suggested that a bloodborne transferable protective factor is released into the circulation in response to the remote ischemic preconditioning stimulus to the organ or tissue [45,46,84,110]. Potential mediators include:

●Neurogenic pathway involving the somato-sensory system, the spinal cord, and the autonomous nervous system: adenosine, bradykinin, and calcitonin.

●Humoral pathway (a circulating cardioprotective factor): opioids, endocannabinoids, and angiotensin-2 receptors.

●Systemic pathway: suppression of proinflammatory genes, expression in leucocytes, and reduced neutrophilic adhesion.

Remote ischemic conditioning has been applied before (remote ischemic preconditioning), during (remote ischemic preconditioning), and after (remote ischemic postconditioning) the onset of sustained myocardial ischemia [84]. Crucially, the protective stimulus can be applied in a noninvasive manner using a cuff placed on the hind limb to induce brief cycles of ischemia and reperfusion to the limb, facilitating its application in the clinical setting.

These findings have been shown to be applicable to humans by simply inflating and deflating a blood pressure cuff placed on the upper arm or thigh to induce brief ischemia and reperfusion. In one report, three five-minute cycles of ischemia to one upper extremity prevented endothelial dysfunction induced by subsequent ischemia to the contralateral upper extremity [114]. The potential use of remote ischemic preconditioning has now been evaluated in the clinical setting. (See "Myocardial ischemic conditioning: Clinical implications", section on 'Therapeutic applications'.)

SUMMARY

●Definition – Ischemic conditioning describes an endogenous phenomenon, in which one or more brief episodes of nonlethal ischemia and reperfusion confer protection against a sustained lethal episode of ischemia and reperfusion.

●Conditioning ischemic stimulus

•Timing – The conditioning stimulus can be applied before (ischemic preconditioning), during (ischemic preconditioning) or after the onset of (ischemic postconditioning) ischemia, or at the transition from sustained ischemia to reperfusion. (See 'Ischemic preconditioning' above and 'Ischemic postconditioning' above.)

•Anatomic location – The conditioning stimulus can be applied either directly to the heart or to a distant organ or tissue (remote ischemic conditioning). (See 'Remote ischemic conditioning' above.)

●Pharmacologic ischemic conditioning – Elucidation of the mechanistic pathways underlying ischemic conditioning has identified potential pharmacological cardioprotective strategies (pharmacological conditioning). (See 'Mediators' above.)