Postconditioning: a new or old option after ischemic stroke?

The mechanisms of brain ischemia include glutamate excitoxicity, calcium toxicity, free radicals, nitric oxide and inflammatory reactions, as well as dysfunctions of endoplasmic reticulum and mitochondrion [1–4]. These injury cascades are interconnected in complex ways, thus it is hard to compare their pathogenic importance in ischemia models [5]. The research in cellular and molecular pathways has spurred the studies in potential neuroprotections mainly in pharmacological fields, such as anti-excitotoxic treatment, calcium-channel antagonism, approaches for inhibition of oxidation, inflammation and apoptosis [6,7]. Besides other protective interventions, including thrombolysis, arteriogenesis and regeneration therapy, ischemia preconditioning or postconditioning are also under investigation [6]. However, new therapeutic neuroprotective approaches are needed to substantially improve the treatment of stroke in the near future [8]. Postconditioning may be such an option.

Ischemic postconditioning is initially referred to when a stuttering reperfusion is performed immediately after reperfusion, for preventing ischemia/reperfusion injury in both myocardial and cerebral infarction. It has evolved into a concept that can be induced by a broad range of stimuli or triggers, and may even be performed as late as 6 h after focal ischemia and 2 days after transient global ischemia [9]. The concept is thought to be derived from ischemic preconditioning or partial/gradual reperfusion, but in fact the first experiment for postconditioning was carried out much earlier than that of preconditioning or partial/gradual reperfusion, in the research on myocardial ischemia.

Definition of postconditioning

Brief interruption (10–60 s, depending on the model) of the blood flow applied immediately after the onset of reperfusion following an ischemic event, is observed to provide protection [10]. In the initial study in an anesthetized canine model of coronary artery occlusion–reperfusion, three cycles of coronary artery reperfusion alternating with 30 s of reocclusion were associated with a significant reduction in both the infarct size and the endothelial dysfunction [11]. Similar findings are now also demonstrated in the rat’s brain [12]. This ‘postconditioning’, is so named because the stimulus is applied after the ischemia and is now also conferred by other species.

The protective effect of postconditioning can be achieved by occluding the ipsilateral common carotid artery (CCA), which is clinically relevant, for the ipsilateral CCA is accessible [5]. In addition, postconditioning can also be induced via volatile anesthetics [13,14]. Isoflurane, for example, reduces the infarct size by 50%, if administered early during reperfusion. Likely, this effect is mediated via the PI3K pathway [15]. Other pharmacological agents administered at the start of reperfusion have been shown to be cardioprotective. These include:

• • Adenosine

• • Nitric oxide

• • Cytokines

• • Complement inhibitors

However, whether this approach should be described as ‘pharmacological postconditioning’ or as a ‘post-reperfusion treatment’ has been debated. Since the mechanisms of postconditioning have not been fully elucidated, and the importance of the alternating cycles of ischemia–reperfusion has not been appreciated nor determined to be critical to protection, direct association to ‘postconditioning’ (whatever ‘conditioning’ means at reperfusion) should be debated.

Experimental studies support the neuroprotective potency of postconditioning to reduce the infarct size, endothelial dysfunction and neutrophil accumulation in the jeopardized area [12]. These experimental results suggest that within the first minutes of reperfusion, endogenous processes are initiated that help reduce reperfusion injury after a limited duration of ischemia. The multiplicity of cell types that are affected by postconditioning (i.e., vascular endothelial cells and inflammatory cells) reflects the complexity of reperfusion injury and suggests a wide network of effects within this complex interactive web of responses. In addition, delayed postconditioning, which is clinically more relevant, also improved glucose uptake, inhibited edema and mitigated blood–brain barrier leakage in the penumbra, and lastly, attenuated the exacerbating effect of tissue plasminogen activator (t-PA) [5].

Basis of the hypothesis of postconditioning

Since clinical trials of pharmacological neuroprotective strategies in stroke have been disappointing, attention has turned to the brain’s own endogenous strategies for neuroprotection. Two endogenous mechanisms have been characterized so far, namely ischemic preconditioning and ischemic postconditioning. The neuroprotective concept of preconditioning is based on the observation that a brief, noninjurious episode of ischemia is able to protect the brain from a subsequent longer ischemic insult. Recently, a hypothesis has been offered that modified reperfusion subsequent to a prolonged ischemic episode may also confer ischemic neuroprotection, a phenomenon termed postconditioning. Many pathways have been proposed as plausible mechanisms to explain the neuroprotection offered by preconditioning and postconditioning. Unfortunately, so far, none of them has clearly identified the mechanism involved in preconditioning and postconditioning.

The neuroprotective effects of postconditioning depend mainly on the degree of ischemia [12], the onset time of postconditioning [16,17], and the cycle number of occluding and releasing the blood vessels [5,16].

Mechanisms of postconditioning

Reperfusion injury is a complex response that involves many of the cells in the integrated structure of the brain. The effects of postconditioning on each of these cell types alone, and then their interaction, must still be described.

It is suggested that limitation of oxygen radicals or of calcium overload is involved in the protection. Postconditioning seems to reduce the oxidative stress, in particular by reduction of the superoxide load that is generated, and in mitochondrial peroxide production and the depletion of gluthathione [18]. Other mechanisms to induce postconditioning are guanylyl cyclase activation, opening of the mitochondrial KATP channels, inhibiting opening of mitochondrial permeability transition pores (mats) [13], and the activation of specific protein pathways (endothelial nitric oxide synthetase [eNOS] and nitric oxide; ERK ½; PI3K – act) and opioid receptors [19]. Regarding the modulation of these endogenous autacoids, ischemia has been shown to increase the generation of each of these substances. Reperfusion may wash out (adenosine) or reduce the generation (opioids) of these autacoids so that they are no longer available to exert their endogenous cardioprotection. By mechanisms that have not yet been elucidated, postconditioning delays the washout of adenosine and may thereby increase its exposure to the vascular space in which adenosine may exert potent protection [20,21]. For opidoids, postconditioning may promote generation of precursor molecules by unclear mechanisms [22].

Tsang et al. have proposed that there are ‘passive’ and ‘active’ phases to the mechanisms of postconditioning [23]. The passive portion is initiated via stepwise reperfusion that reduces oxygen radicals (presumably the delivery), mitochondrial calcium overload, and neutrophils. By the active portion, the ‘reperfusion injury salvage’ kinases (RISK) pathway is activated by endogenous stimulators, for example, adenosine, opioids or other as yet unidentified endogenous substances, and stressors, such as ischemia–reperfusion. Here, two interactive pathways might be responsible at least for protection: first, activation of PI3K, Akt and eNOS to inhibit opening of the 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridins, and second, activation of MEK ½ and ERK ½. In turn, Akt together with ERK ½ activate p70s6K, which finally initiates protein translation.

Neuroprotective potential of postconditioning

Postconditioning treatment reduces infarct volume by up to 50% in vivo and by approximately 30% in vitro[17]. A duration of 10 min of postconditioning ischemia after 10 min of reperfusion produces the most effective postconditioning condition both in vivo and in vitro[17]. The degree of neuroprotection after postconditioning is equivalent to that observed in models of ischemic preconditioning. However, subjecting the brain to both preconditioning as well as postconditioning has not caused greater protection than each treatment alone [17]. The prosurvival protein kinases ERK, MAPK and Akt show prolonged phosphorylation in the cortex of postconditioned rats.

Postconditioning of ischemic tissue, via mechanical or pharmacological manipulation, offers an exciting avenue towards amelioration of ischemia–reperfusion injury. Postconditioning has been shown to be successful in reducing ischemia–reperfusion injury in both animal models and clinical trials. Human studies are presently limited to cardiac studies, but there is scope for research into other organ systems with potential beneficial effects, particularly within the field of vascular surgery where ischemia–reperfusion occurs by nature of both the disease and the intervention.

Reperfusion damage is a complex process involving several cell types, soluble proinflammatory mediators, oxidants, ionic and metabolic dyshomeostasis, and cellular and molecular signals. Novel neuroprotective strategies are required to target this form of injury [2,12]. The neuroprotective potential of ischemic preconditioning has not been realized in clinical practice because it necessitates intervention applied before the onset of ischemic stroke, which is difficult to predict. A more amenable approach to neuroprotection is to intervene at the onset of reperfusion, the timing of which is under the control of the operator. In this regard, these new findings of postconditioning in the brain may open a window to improve stroke treatment or prevention [8]. In contrast to preconditioning, which requires a knowledge of the ischemic event, prior postconditioning can be applied at the onset of reperfusion at the point of clinical service. Interestingly, experimental studies suggest that ischemic preconditioning and postconditioning activate the same signaling pathway at the time of reperfusion, thereby offering a common target for neuroprotection [12]. The pharmacologic recruitment of this signaling pathway at the time of cerebral reperfusion might allow one to harness the neuroprotective potential of ischemic preconditioning and postconditioning and therefore substantially improve the outcome in extracranial–intracranial bypass and neurovascular surgery [3,4,24,25].

Remote postconditioning

A short time after the term ‘remote preconditioning’ was generally established during the last few years, ‘remote post-conditioning’ was recently experimentally demonstrated for the first time in the heart: occlusion of the renal artery immediately before the onset of reperfusion of a coronary artery clearly reduced the size of the myocardial infarct [26]. These investigations provide some results of interest that may be similar in the brain:

• • The benefit is likely initiated within the first minutes of the reperfusion of the cerebral artery;

• • The inter-organ remote postcondition is likely mediated via adenosine liberated from the ischemic–reperfused remote organ since neuroprotection is abrogated by an adenosine receptor blocker (8-p-sulfophenyl theophylline).

It is not known whether the adenosine receptors were localized in the brain or in a remote organ and whether neurological mediation of the adenosine stimulus is involved [26].

The concept of remote preconditioning is not merely a laboratory curiosity; it may have clinical application. One may ‘stimulate’ postconditioning via another organ, such as the leg. This ‘organ’ is much more accessible than the kidney as used in the original study introducing the concept [26]. A tourniquet could be applied to the leg during transport to the emergency room or to the cath-lab to set up a strong postconditioning signal. The tourniquet could then be removed just before completion of the angioplasty procedure and in advance of the onset of reperfusion, being certain to miss the window of neuroprotection afforded in the first minutes of reperfusion [26].

Expert commentary & five-year view

Postconditioning protects against focal ischemia in animal studies. It appears to provide long-term protection and improved neurological function, and partially reversed the detrimental effect of t-PA [5]. Such a novel neuroprotective model offers an alternative and promising avenue for studying therapeutic strategies against stroke. Further research is needed to find new pharmacological agents that would mimic postconditioning in order to treat all patients with ongoing acute ischemic stroke. Of course, the potential of postconditioning must be rigorously tested in clinical trials, first for its safety and feasibility and then subsequently for its efficacy and therapeutic potential. It will be mandatory in such clinical trials to carefully control for confounding variables, such as the size of the area at risk, the duration of the preceding ischemic insult and collateral status. Neglect of these confounding variables has probably contributed to the failure to translate experimentally validated principles of neuroprotection to the clinical arena (e.g., adenosine receptor activation) in the past.

Financial & competing interests disclosure

The authors have no relevant affiliations or financial involvement with any organization or entity with a financial interest in or financial conflict with the subject matter or materials discussed in the manuscript. This includes employment, consultancies, honoraria, stock ownership or options, expert testimony, grants or patents received or pending, or royalties.

No writing assistance was utilized in the production of this manuscript.