Research on attenuating the structural and functional deficits observed following ischemia-reperfusion has become increasingly focused on the therapeutic potential of ischemic postconditioning. In recent years, various methods and animal models of ischemic postconditioning have been utilized. The results of these numerous studies have indicated that the mechanisms underlying the neuroprotective effects of ischemic postconditioning may involve reductions in the generation of free radicals and inhibition of calcium overload, as well as the release of endogenous active substances, alterations in membrane channel function, and activation of protein kinases. Here we review the novel discovery, mechanism, key factors, and clinical application of ischemic postconditioning and discuss its implications for future research and problem of clinical practice.
Ischemic preconditioning has been widely adopted as a clinical strategy aimed at protecting the brain from subsequent, more serious ischemia-reperfusion insults. Ischemic preconditioning involves the application of a brief, subthreshold episode of ischemia prior to the occurrence of irreversible ischemic injury [
Ischemic postconditioning was initially defined in the field of myocardial ischemia as a series of brief mechanical occlusions and reperfusions [
Depending on the processing timeframe, ischemic postconditioning can be classified as either rapid ischemic postconditioning (RIPO) or delayed ischemic postconditioning (DIPO). RIPO is conducted within a few seconds to minutes following ischemia-reperfusion [
Ischemic postconditioning can also be defined according to the site of mechanical blockage as distal, proximal, or remote. Proximal ischemic postconditioning usually involves occlusion of the carotid artery, while distal ischemic postconditioning usually involves occlusion of the upper brachial artery. Remote ischemic postconditioning, on the other hand, involves occlusion of an artery in the lower limb. The bilateral arm ischemic preconditioning (BAIPC) device patented by Xunming Ji from Xuanwu Hospital can perform regular occlusion/reperfusion automatically and record the timing of every BAIPC process in real time. This device may be used in the RIPO clinical research (Figure
The BAIPC device patented by Xunming Ji from Xuanwu Hospital.
The BAIPC device and how it is used are as follows. The device can perform 5 minutes of ischemia followed by 5 minutes of reperfusion automatically and record the timing of every BAIPC process in real time. It can also record heart rate and blood pressure. The time intervals for the BAIPC can be adjusted based on the requirements of the study.
As in myocardial ischemia, the protective strength of cerebral ischemic postconditioning depends on temporal factors associated with the mechanical interruptions, such as the number of ischemia-reperfusion cycles and the durations of occlusion and reperfusion.
The therapeutic window for the beneficial effects of ischemic postconditioning also depends on the type of technique used. The therapeutic window for RIPO is usually a few minutes to several hours after ischemia/reperfusion, while that for DIPO is usually from several hours to days after ischemia/reperfusion. Both RIPO and DIPO can effectively reduce cerebral infarction volume and improve recovery of neural function when initiated within the therapeutic window.
However, there is no uniform standard regarding the therapeutic window of either form of ischemic postconditioning. To date, research has been conducted using various models of ischemia in a number of different experimental animals and under various experimental conditions. Thismakes it difficult to adopt an appropriate standard from the available literature.
Variations in the results of a number of previous studies highlight the need to establish such a standard. For example, Jang et al. developed a rabbit model of spinal cord ischemia-reperfusion via occlusion of the infrarenal aorta and observed that ischemic postconditioning initiated either 1 minute or 5 minutes following ischemia-reperfusion was effective in improving neurological function and preserving motor neurons. However, no such neuroprotective effects were observed when ischemic postconditioning was initiated 10 minutes after reperfusion [
Divergent results regarding the therapeutic window have been reported for DIPO as well. In a study by Burda et al., postconditioning performed by occlusion of the internal carotid artery 48 hours after ischemia-reperfusion in a rat model of cerebral ischemia resulted in significant improvements in the structure and function of nerve cells [
Though there is no uniform standard to define the therapeutic window for ischemic postconditioning, the results of the aforementioned studies indicate that the therapeutic window may be modified by changing the site of occlusion, number of cycles, or additional factors, thus enhancing the clinical potential of such treatment.
Ischemic postconditioning refers to the process of inducing a series of brief periods of ischemia and reperfusion following lethal ischemic injury to a specific organ in order to reduce the overall extent of ischemic injury. The event consisting of one ischemic period and one subsequent reperfusion period is defined as one cycle. In most animal models, three to ten cycles of postconditioning are performed, though some clinical research studies have utilized between three and five cycles.
In a 2008 study by Gao et al., focal ischemia was generated by permanent occlusion of the left distal middle cerebral artery (dMCA) combined with 30 minutes of occlusion of both common carotid arteries (CCAs) in rats [
Rezazadeh et al. developed a rat model of embolic stroke by embolizing a preformed clot into the MCA. This was followed by ischemic postconditioning involving blockage and release of the bilateral CCAs [
The proper time for the observation of the effects is another important factor when assessing the protective effects of ischemic postconditioning when determining whether the hypothetical neuroprotective effects have been achieved. The effects of postconditioning on some clinical indicators such as cerebral blood flow and infarct volume can usually be observed 2-3 days after ischemic stroke.
While research regarding RIPO is abundant, the study of DIPO has been relatively limited to date. Most previous studies have focused only on whether neuroprotective effects have occurred and have therefore utilized observation times of 2-3 days following stroke. Based on the onset time and the conditions of clinical emergencies, there is no doubt that DIPO has much higher potential for clinical application than RIPO. Most importantly, future research should stress the use of an appropriate therapy window to guide clinical practice. We could not make the conclusion that DIPO had no side effects when compared to RIPO. The mechanism of neuroprotection of DIPO, especially in the central nervous system, is another focus of study, as this effect is transferred from peripheral tissue to the central nervous system.
In summary, although it is possible to alter a number of factors involved in the control of ischemic postconditioning in the brain, it remains difficult to determine the appropriate solution for each individual. Due to the complexity of the clinical environment, it may be possible to adjust some factors, such as the postconditioning method (proximal or distal) or the number of cycles, in order to determine the ideal therapeutic schedule. Therefore, further studies regarding the mechanisms underlying the neuroprotective effects of and factors influencing ischemic postconditioning remain particularly important.
Preconditioning refers to the process of inducing brief periods of subthreshold ischemia in order to prevent or attenuate severe ischemic injury due to subsequent, prolonged periods of ischemia [
Despite the fact that ischemic preconditioning and postconditioning are applied along distinctly different time courses, both share several common protective mechanisms involving modification of key mitochondrial targets or activation of reperfusion injury salvage kinase (RISK) pathways. These may involve the Akt, extracellular signal-regulated kinase 1/2 (ERK1/2), and mitogen-activated protein kinase (MAPK) pathways [
The acute protective effects of IPC likely result from immediate posttranslational protein modifications (e.g., phosphorylation) within cell energetic or survival systems. In contrast, the protective effects of IPC likely result from protein synthesis of previously dormant genes involved in angiogenesis, energy metabolism, vasomotor control, inflammation, and cell survival (e.g., growth factors). Therefore, elucidation of the cell signaling pathways underlying the protective effects of ischemic preconditioning may provide insight into those underlying the effects of ischemic postconditioning.
Gao et al. investigated the effects of combining preconditioning with postconditioning treatment on ischemic damage [
Minimizing damage in the ischemic penumbra, which requires the attenuation/prevention of neural cell apoptosis, is the current primary therapeutic target in the treatment of acute stroke. Research indicates that remote ischemic postconditioning may protect against ischemic damage in the brain via the p38 MAPK signaling pathway, improve neuronal morphological changes in the area of the ischemic penumbra, and reduce neuronal cell apoptosis in rat models of focal cerebral ischemia/reperfusion (I/R) [
Recent studies have reported markedly increased autophagy following the upregulation of LC3/Beclin 1 and downregulation of p62 in the penumbra at various time intervals following ischemia. Furthermore, ischemic postconditioning performed at the onset of reperfusion reduces infarct size, mitigates brain edema, inhibits the induction of LC3/Beclin 1, and reverses decreases in p62 [
Other researchers have reported that the protective effects of remote limb ischemic postconditioning against cerebral I/R injury may be related to the attenuation of neuronal apoptosis and inflammation via activation of signal transducer and activator of transcription 3 (STAT3), as well as attenuation of tumor necrosis factor-
In a study by Joo et al., ischemic postconditioning consisted of a series of brief occlusions of the MCA after reperfusion in a mouse model of focal ischemia. As a result, spared infarct areas were observed in the border zones between the cortical territories of the ACA and MCA, as well as in the ventromedial and dorsolateral striatum. These regions have been confirmed to be affected by ischemia sequentially over longer periods following onset of ischemia in the dorsolateral striatum. Ischemia then progresses into the ventromedial striatum and the cerebral cortex in the MCA territory (Figure
RIPO acts at the cellular level to directly protect the vascular endothelium via KATP channel-dependent mechanisms [
Disturbances in cerebral blood flow (CBF) occur throughout the period of reperfusion following ischemic stroke. In fact, it has been reported that the clinical neuroprotective effects of remote ischemic conditioning (RIC) are partially related to improvements in CBF. Following reperfusion, there is a short period of hyperperfusion, followed by a longer period of hypoperfusion. A recent study indicates that combined ischemic postconditioning may stabilize CBF disturbances during the early hyperperfusion and later hypoperfusion periods [
Disruptions in the energy and material supply of brain tissue during cerebral ischemia, accompanied by the failure of ion pumps in the cell membrane, lead to cerebral edema. Numerous studies have reported that limb remote ischemic postconditioning (LRIP) significantly reduces cerebral infarct volume and relieves brain edema. Possible mechanisms underlying the protective effects of LRIP may include amelioration of endothelial dysfunction, maintenance of the integrity of the blood brain barrier, modulation of protein synthesis and nerve activity [
In remote postconditioning of cerebral ischemia in rats, downregulation of aquaporin 4 (AQP4), which is involved in water homeostasis in astrocytes, may attenuate cerebral damage after transient MCAO [
Chronic cerebral ischemia leads to cognitive dysfunction, although similar neuronal damage and dysfunction are also observed in vascular dementia, Alzheimer’s disease, and Binswanger’s disease [
The highest density of N-methyl-D-aspartate (NMDA) receptors is found in the hippocampal CA1 and CA3 areas and the dentate gyrus, which are areas closely associated with cognitive function. Ischemic postconditioning activates NMDA2A receptors, promotes the internal flow of calcium ions, influences the ERK pathway and the synthesis of NO, and restores hippocampal blood flow [
To date, hundreds of studies have reported the involvement of different signaling molecules and potential mechanisms underlying the effects of postconditioning under a wide range of experimental conditions. Studies have demonstrated the effects of ischemic postconditioning on activation of adenosine, bradykinin (BK), and endogenous protective molecules such as NO and G-protein-mediated kinases, which further act on the mitochondria, endoplasmic reticulum, or nucleus and produce neuroprotective effects in targeted tissues [
In animal models of brain ischemia, rapid ischemic postconditioning can be triggered by promoting the synthesis of eNOS and activating the PI3K/Akt signal transduction pathway, which act to protect vascular endothelial cells and promote vascular remodeling [
As is the case in myocardial ischemic reperfusion injury, while free radicals may be generated to a small extent during ischemia, far greater production of reactive oxygen intermediates occurs after reintroduction of oxygen during cerebral ischemic reperfusion. Most of the protective mechanisms of ischemic postconditioning are the same in the heart and the brain, although the effects of postconditioning on ROS are controversial.
In myocardial ischemic reperfusion injury, ROS signaling is an essential trigger of ischemic and pharmacological postconditioning. Chemically blocking the production of ROS abolishes the protective effect of ischemic postconditioning in the heart [
Another research also indicates that ischemic postconditioning exerts its neuroprotective effects via ROS suppression. A study involving rat models of local cerebral ischemia indicates that rapid initiation of ischemic postconditioning within 30 minutes of reperfusion reduces the levels of peroxides and lipid peroxides, in turn reducing free radical damage [
Xing et al. demonstrated that a decrease in the content of glutathione (GSH) together with an increase in myeloperoxidase (MPO) and proinflammatory markers may be observed in rats subjected to global cerebral ischemia/reperfusion [
Decreases in transient focal ischemia-induced infarct volume and rates of apoptosis have also been observed when ischemic postconditioning is induced within 24 hours of reperfusion following 2 hours of focal cerebral ischemia (Figure
It is known that T cells infiltrate areas of focal ischemia following stroke and that ischemic postconditioning effectively reduces the infiltration of T cells and total infarct volume (Figure
Rapid ischemic postconditioning significantly reduces the number of terminal deoxynucleotidyl transferase dUTP nick end labeled cells in the ischemic area when observed two days after stroke [
MAPK signaling pathways, including the ERK1/2, p38 lightning, and c-Jun amino terminal kinase (JNK) pathways, are closely related to the extent of ischemic injury and neuronal survival [
Endoplasmic reticulum (ER) stress in ischemia-reperfusion injury is one of the most important factors that lead to cell apoptosis. Following ischemic postconditioning, the ER stress response results in elevated levels of C/EBP homologous protein (CHOP). This affects the release of Bim and Bcl-2, which interfere with the cell apoptosis pathway. Ischemic postconditioning can also cause rapid increases in GRP78 expression, dephosphorylation of EIF2
Both ischemic preconditioning and postconditioning promote Akt phosphorylation and have neuroprotective effects. Research indicates that both rapid and delayed postconditioning influence important targets for neuroprotection (Figure
Phosphorylated Akt can also raise levels of mammalian target of rapamycin (mTOR) in order to promote neuroprotection. Ischemic postconditioning may result in time-dependent regulation of adenosine monophosphate-activated protein kinase (AMPK) activation and autophagy, and AMPK may strengthen the autophagy effect by inhibiting mTOR [
Akt may indirectly participate in the inhibition of the mitochondrial apoptosis pathway in order to ensure the survival of cells following ischemic injury by influencing the activity of Bim and the phosphorylation of PKC, thereby affecting mitochondrial ATP-dependent potassium channels. Therefore, Akt signaling pathways may play a vital role in mediating the protective effects of ischemic postconditioning.
Postconditioning leads to increased Hsp70 expression and decreased NF-
RIPO significantly upregulates the expression of nuclear factor erythroid 2-related factor 2, heme oxygenase-1, and quinone oxidoreductase-1 and the activity of superoxide dismutase, while downregulating the formation of malondialdehyde.
The standards for robust data on neuroprotective signaling have risen, and experiments utilizing single-dose antagonists are no longer satisfactory for the identification of steps within a signaling pathway. Unequivocal identification of a signaling step requires not only an appropriate conditioning protocol with infarct size (IS) as an endpoint, but also biochemical or immunoblotting data for signal activation. In fact, IS reduction following genetic ablation or pharmacological inhibition of the signal molecule is now routinely required.
To improve translation of experimental findings into clinically applicable standards, further insight into the mechanisms underlying postconditioning phenomena is required, although equal emphasis should be placed on the identification of novel signaling elements in potentially reductionist experimental models and on the translation of such novel, yet reductionist, findings into more complex and integrative models. In addition, future studies should focus on the identification of signaling elements involved in neuroprotection in the human CNS. Moreover, they should retrospectively evaluate experimental models that may have predicted these elements, develop standards for the identification of robust signaling elements that may serve as potential drug targets, and organize interactions between basic and clinical scientists in order to develop proof-of-concept clinical trials and to eventually carry out larger prospective multicenter trials.
The authors declare that they have no conflicts of interest.