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This study investigated the antioxidant and neuroprotective effects of nicotinamide combined with ischemic preconditioning against cerebral ischemia reperfusion (CIR) injury. Thirty-five Wistar albino male rats were randomly divided into five groups: sham, preconditioned ischemia/reperfusion (IP+IR), ischemia/reperfusion (IR), preconditioned ischemia/reperfusion + nicotinamide (IP+IR+N), and ischemia/reperfusion + nicotinamide (IR+N). CIR was achieved with bilateral common carotid artery occlusion. IP+IR and IP+IR+N groups 30 min before ischemia; Three cycles of 10 sec ischemia/30 sec reperfusion followed by 20 min IR were applied. The IP+IR+N and IR+N groups received 500 mg/kg nicotinamide intraperitoneally. After 24 h of reperfusion, a neurological evaluation was performed and vertıcal pole test. Biochemically, malondialdehyde (MDA), glutathione (GSH) levels and catalase (CAT) activity were measured in blood and brain tissue samples. Rates of red neurons, sateliosis and spongiosis were determined histopathologically in the prefrontal cortex areas. After CIR, MDA levels increased significantly in serum and brain tissue in the IR group compared to the sham group, while GSH and CAT activity decreased in the brain tissue (p < 0.05). MDA levels in the tissues were found significantly decreased in the IR+N group compared to the IR group (p < 0.05). Administration of nicotinamide together with IP significantly decreased MDA levels in brain tissue and increased GSH and CAT activity (p < 0.05). Compared to the IR group, the morphological and neurological damage in the prefrontal cortex areas decreased in the IP+IR, IP+IR+N, and IR+N groups (p < 0.05). In addition, red neuron, sateliosis and spongiosis rates increased significantly in the IR group compared to the Sham, IP+IR+N, IR+N groups (p < 0.001 for all). In neurological evaluation, while the neurological score increased and the time on the vertical pole decreased significantly in the IR group, preconditioning, and nicotinamide groups reversed (p < 0.05). The study's results show that nicotinamide administration with ischemic preconditioning alleviates cerebral ischemia/reperfusion injury., Competing Interests: Declaration of competing interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier B.V. All rights reserved.)

Cerebral ischemic preconditioning (CIP) has been shown to improve brain ischemic tolerance against subsequent lethal ischemia. Reactive astrocytes play important roles in cerebral ischemia-reperfusion. Recent studies have shown that reactive astrocytes can be polarized into neurotoxic A1 phenotype (C3d) and neuroprotective A2 phenotype (S100A10). However, their role in CIP remains unclear. Here, we focused on the role of N-myc downstream-regulated gene 2 (NDRG2) in regulating the transformation of A1/A2 astrocytes and promoting to brain ischemic tolerance induced by CIP. A Sprague Dawley rat model of middle cerebral artery occlusion/reperfusion (MCAO/R) was used. Rats were divided into the following six groups: (1) sham group; (2) CIP group: left middle cerebral artery was blocked for 10 min; (3) MCAO/R group: left middle cerebral artery was blocked for 90 min; (4) CIP + MCAO/R group: CIP was performed 72 h before MCAO/R; (5) AAV-NDRG2 + CIP + MCAO/R group: adeno-associated virus (AAV) carrying NDRG2 was administered 14 days before CIP + MCAO/R; (6) AAV-Ctrl + CIP + MCAO/R group: empty control group. The rats were subjected to neurological evaluation 24 h after the above treatments, and then were sacrificed for 2, 3, 5-triphenyltetraolium chloride staining, thionin staining, immunofluorescence and western blot analysis. In CIP + MCAO/R group, the neurological deficit scores decreased, infarct volume reduced, and neuronal density increased compared with MCAO/R group. Notably, CIP significantly increased S100A10 expression and the number of S100A10 + /GFAP + cells, and also increased NDRG2 expression. MCAO/R significantly decreased S100A10 expression and the number of S100A10 + /GFAP + cells yet increased C3d expression and the number of C3d + /GFAP + cells and NDRG2 expression, and these trends were reversed by CIP + MCAO/R. Furthermore, over-expression of NDRG2 before CIP + MCAO/R, the C3d expression and the number of C3d + /GFAP + cells increased, while S100A10 expression and the number of S100A10 + /GFAP + cells decreased. Meanwhile, over-expression of NDRG2 blocked the CIP-induced brain ischemic tolerance. Taken together, these results suggest that CIP exerts neuroprotective effects against ischemic injury by suppressing A1 astrocyte polarization and promoting A2 astrocyte polarization via inhibiting NDRG2 expression., (© 2024. The Author(s), under exclusive licence to Springer Science+Business Media, LLC, part of Springer Nature.)

Cerebral ischemic damage is prevalent and the second highest cause of death globally across patient populations; it is as a substantial reason of morbidity and mortality. Mesenchymal stromal cells (MSCs) have garnered significant interest as a potential treatment for cerebral ischemic damage, as shown in ischemic stroke, because of their potent intrinsic features, which include self-regeneration, immunomodulation, and multi-potency. Additionally, MSCs are easily obtained, isolated, and cultured. Despite this, there are a number of obstacles that hinder the effectiveness of MSC-based treatment, such as adverse microenvironmental conditions both in vivo and in vitro. To overcome these obstacles, the naïve MSC has undergone a number of modification processes to enhance its innate therapeutic qualities. Genetic modification and preconditioning modification (with medications, growth factors, and other substances) are the two main categories into which these modification techniques can be separated. This field has advanced significantly and is still attracting attention and innovation. We examine these cutting-edge methods for preserving and even improving the natural biological functions and therapeutic potential of MSCs in relation to adhesion, migration, homing to the target site, survival, and delayed premature senescence. We address the use of genetically altered MSC in stroke-induced damage. Future strategies for improving the therapeutic result and addressing the difficulties associated with MSC modification are also discussed., (© 2024 John Wiley & Sons Ltd.)

Several studies indicated that autophagy activation participates in brain ischemic tolerance (BIT) induced by cerebral ischemic preconditioning (CIP). However, the mechanism of autophagy activation during the process still remains unclear. The present study aimed to evaluate the role of p38 MAPK-peroxisome proliferator-activated receptor γ (PPARγ) signaling cascade in autophagy during the CIP-induced BIT. The results shown that, initially, autophagy activation was observed after CIP in the model of global cerebral ischemia in rats, as was indicated by the upregulation of Beclin 1 expression, an increase in LC3-II/LC3-I ratio, the enhanced LC3 immunofluorescence, and a rise in the number of autophagosomes in the neurons of the hippocampal CA1 area. Besides, the inhibitor of autophagy 3-methyladenine obliterated the neuroprotection induced by CIP. Furthermore, the upregulation of p-p38 MAPK and PPARγ expressions was earlier than autophagy activation after CIP. In addition, pretreatment with SB203580 (the inhibitor of p38 MAPK) reversed CIP-induced PPARγ upregulation, autophagy activation, and neuroprotection. Pretreatment with GW9662 (the inhibitor of PPARγ) reversed autophagy activation and neuroprotection, while it had no effect on p-p38 MAPK upregulation induced by CIP. These data suggested that the p38 MAPK-PPARγ signaling pathway participates in autophagy activation during the induction of BIT by CIP.

1 H Nuclear Magnetic Resonance (NMR) metabolomics is one of the fundamental tools in the fast-developing metabolomics field. It identifies and quantifies the most abundant metabolites, alterations of which can describe energy metabolism, activated immune response, protein synthesis and catabolism, neurotransmission, and many other factors. This paper summarizes our results of the 1 H NMR metabolomics approach to characterize the distribution of relevant metabolites and their alterations induced by cerebral ischemic injury or its combination with hyperhomocysteinemia in the affected tissue and blood plasma in rodents. A decrease in the neurotransmitter pool in the brain tissue likely follows the disordered feasibility of post-ischemic neurotransmission. This decline is balanced by the increased tissue glutamine level with the detected impact on neuronal health. The ischemic injury was also manifested in the metabolomic alterations in blood plasma with the decreased levels of glycolytic intermediates, as well as a post-ischemically induced ketosis-like state with increased plasma ketone bodies. As the 3-hydroxybutyrate can act as a likely neuroprotectant, its post-ischemic increase can suggest its supporting role in balancing ischemic metabolic dysregulation. Furthermore, the 1 H NMR approach revealed post-ischemically increased 3-hydroxybutyrate in the remote organs, such as the liver and heart, as well as decreased myocardial glutamate. Ischemic preconditioning, as a proposed protective strategy, was manifested in a lower extent of metabolomic changes and/or their faster recovery in a longitudinal study. The paper also summarizes the pre- and post-ischemic metabolomic changes in the rat hyperhomocysteinemic models. Animals are challenged with hyperglycemia and ketosis-like state. A decrease in several amino acids in plasma follows the onset and progression of hippocampal neuropathology when combined with ischemic injury. The 1 H NMR metabolomics approach also offers a high potential for metabolites in discriminatory analysis in the search for potential biomarkers of ischemic injury. Based on our results and the literature data, this paper presents valuable findings applicable in clinical studies and suggests the precaution of a high protein diet, especially foods which are high in Met content and low in B vitamins, in the possible risk of human cerebrovascular neuropathology.

We studied the influence of combined preconditioning (compound pQ-4 and moderate hypoxia) on morphometrical parameters of neuronal populations in hippocampal fields CA1 and CA3 in rats after bilateral ligation of the common carotid artery. Preconditioning produced a neuroprotective effect, improved survival of pyramidal neurons in the early and delayed periods of modeled ischemia, prevented the formation of necrotic and apoptotic neurons and hyperactivation of microglia, and protected endotheliocytes. The positive influence of preconditioning factors on the morphometric parameters of the brain under ischemic conditions agrees with the results of behavioral tests (open field and elevated plus maze) that demonstrated increased locomotor activity and exploratory behavior of animals., (© 2021. Springer Science+Business Media, LLC, part of Springer Nature.)

Ischemic preconditioning (IPC) significantly reduces ischemia-reperfusion injury in the brain by inducing ischemic tolerance. Although emerging evidence suggests that microRNAs (miRNAs) contribute to the pathogenesis of brain ischemia and IPC-induced neuroprotection, the role of miRNAs and their underlying mechanisms are still unclear. IPC was induced in male C57BL/6 mice by brief bilateral common carotid artery occlusion. After 24 h, mice underwent transient middle cerebral artery occlusion followed by 3 h of reperfusion. Expression levels of messenger RNAs (mRNAs) and proteins were examined in the ipsilateral cortex, and mimics and inhibitors of selective miRNAs were transfected into Neuro-2a cells before oxygen-glucose deprivation (OGD). Post-IPC miRNA expression profiling identified neuroprotection-associated changes in miRNA expression in the ipsilateral cortex after ischemic stroke. Among them, miR-33-5p and miR-135b-5p were significantly downregulated by IPC. Inhibition of miR-33-5p and miR-135b-5p expression protected Neuro-2a cells from OGD-induced apoptosis. Inhibition of these two miRNAs significantly increased mRNA and protein levels of ATP-binding cassette subfamily A member 1 (ABCA1), and a binding assay showed that these two miRNAs showed specificity for Abca1 mRNA. Overexpression of ABCA1 decreased the Bax/Bcl2 mRNA ratio and activation of caspase-9 and caspase-3, whereas knockdown of ABCA1 expression increased the Bax/Bcl2 mRNA ratio and the percentage of Neuro-2a cells with a loss of mitochondrial membrane potential after OGD-treatment. In conclusion, ABCA1 expression is regulated by miR-33-5p and miR-135b-5p. Increased ABCA1 expression following IPC exerts a protective influence against cerebral ischemia via suppression of a mitochondria-dependent apoptosis pathway.

We recently reported that isoflurane conditioning provided multifaceted protection against subarachnoid hemorrhage (SAH)-induced delayed cerebral ischemia (DCI), and this protection was through the upregulation of endothelial nitric oxide synthase (eNOS). SIRT1, an NAD-dependent deacetylase, was shown to be one of the critical regulators of eNOS. The aim of our current study is to examine the role of SIRT1 in isoflurane conditioning-induced neurovascular protection against SAH-induced DCI. Mice were divided into four groups: sham, SAH, or SAH with isoflurane conditioning (with and without EX-527). Experimental SAH via endovascular perforation was performed. Anesthetic conditioning was performed with isoflurane 2% for 1 h, 1 h after SAH. EX-527, a selective SIRT1 inhibitor, 10 mg/kg was injected intraperitoneally immediately after SAH in the EX-527 group. SIRT1 mRNA expression and activity levels were measured. Vasospasm, microvessel thrombosis, and neurological outcome were assessed. SIRT1 mRNA expression was downregulated, and no difference in SIRT1 activity was noted after isoflurane exposure. Isoflurane conditioning with and without EX-527 attenuated vasospasm, microvessel thrombosis and improved neurological outcomes. Our data validate our previous findings that isoflurane conditioning provides strong protection against both the macro and micro vascular deficits induced by SAH, but this protection is likely not mediated through the SIRT1 pathway.

Ischemic stroke is a refractory disease generally caused by cerebral ischemic injury. Remote ischemic preconditioning (RIPC) caused by transient ischemia and reperfusion of the femoral artery exerts a protective effect on ischemic stroke-induced brain injury. This study was designed to investigate the potential molecular mechanism of RIPC-mediated neuroprotection, namely, the biological effects of microRNA-144 on RIPC in mice with ischemic stroke and its effects on PTEN and Akt signaling pathways. Healthy adult C57BL6 mice were selected for the establishment of middle cerebral artery occlusion (MCAO). One hour before the start, remote ischemic preconditioning of limbs was performed in mice. Brain edema and infarct volume were measured. The expressions of microRNA-144, PTEN, and Akt were measured. The results showed that, compared with MCAO group, the RIPC group protected mice from cerebral ischemia-reperfusion injury, systemic accumulation of inflammatory cytokines, and accelerated apoptosis of parenchymal cells. In RIPC group, PTEN expression decreased, and mir-144 and Akt expression increased. The level of phosphorylated PTEN in the transfected microRNA-144 inhibitor group increased and the level of phosphorylated Akt reduced significantly. In conclusion, our results suggest that microRNA-144 may play a protective role in remote ischemic pretreatment by downregulating PTEN and upregulating Akt, suggesting that microRNA-144 via PTEN/Akt pathway may be of therapeutic significance in ischemic stroke.

Remote ischaemic conditioning (RIC) is achieved by repeated transient ischaemia of a distant organ/limb and is neuroprotective in experimental ischaemic stroke. However, the optimal time and methods of administration are unclear. Systematic review identified relevant preclinical studies; two authors independently extracted data on infarct volume, neurological deficit, RIC method (administration time, site, cycle number, length of limb occlusion (dose)), species and quality. Data were analysed using random effects models; results expressed as standardised mean difference (SMD). In 57 publications incorporating 99 experiments (1406 rats, 101 mice, 14 monkeys), RIC reduced lesion volume in transient (SMD -2.0; 95% CI -2.38, -1.61; p  < 0.00001) and permanent (SMD -1.54; 95% CI -2.38, -1.61; p  < 0.00001) focal models of ischaemia and improved neurological deficit (SMD -1.63; 95% CI -1.97, -1.29, p  < 0.00001). In meta-regression, cycle length and number, dose and limb number did not interact with infarct volume, although country and physiological monitoring during anaesthesia did. In all studies, RIC was ineffective if the dose was <10 or ≥50 min. Median study quality was 7 (range 4-9/10); Egger's test suggested publication bias ( p  < 0.001). RIC is most effective in experimental stroke using a dose between 10 and 45 min. Further studies using repeated dosing in animals with co-morbidities are warranted.

Ischemic stroke is a medical condition that arises because of poor blood supply to the brain. Reperfusion being salvage to the brain further causes, exacerbation of tissue injury, known as reperfusion injury. Ischemic preconditioning (IPC) has been known to provide benefits against ischemia reperfusion (I/R) injury. Dopamine D2/D3 receptor mediated several pathways are also reported as mediators in the IPC mediated neuroprotection. This study investigates the possible involvement of D2/D3 receptor activation in IPC mediated neuroprotection in the I/R brain. Global cerebral ischemia/reperfusion (GCI/R) injury in swiss albino mice was induced by occluding the common carotid arteries for 17 min, followed by 24 h reperfusion. IPC was provided by giving 3 episodes of I/R prior to Ischemia (17 min). Morris water maze (MWM) was used to assess the spatial learning, memory and Rota rod, lateral push test as well as inclined beam test were conducted to assess the motor functions in animals. Cerebral oxidative markers (thiobarbituric acid reactive species-TBARS, reduced glutathione-GSH, superoxide dismutase-SOD, and catalase-CAT), inflammatory markers (IL-6, IL-10, TNF-α, and myeloperoxidase-MPO), acetylcholinesterase activity-AChE, infarct size (% weight and % volume), and histopathological changes were also assessed. Haloperidol (0.05 mg/kg, i.p) was used to antagonize the effects of D2/D3 receptor activation. I/R animals showed reduction in memory, motor function, increase in cerebral oxidative stress, inflammation, AChE activity, infarct size and histopathological changes. Episodes of IPC prior to ischemia, attenuated the deleterious effects of I/R injury. Administration of haloperidol abolished the protective effects of IPC. Hence, it may be concluded that IPC mediated neuroprotection may involves dopamine D2/D3 receptor activation in mice., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Cardiac arrest is one of the major causes of death and disability. The aim of the study was to identify dynamic time-dependent metabolomic changes reflected in rat plasma induced by cerebral ischemia and reperfusion with the focus on the protective effect of ischemic preconditionig. Global cerebral ischemia in rats was induced by the four-vessel occlusion. Blood plasma was collected in three reperfusion times: an early post-acute 3 hr, then 24 hr, as an incipient time for delayed neuronal death induction and 72 hr as prolonged reperfusion period. The metabolomic measurements were conducted via untargeted nuclear magnetic resonance spectroscopy. Plasma of ischemized rats manifested dynamic metabolomic changes over the reperfusion time, such as increased levels of ketone bodies, decreased levels of pyruvate, alanine, and citrate. All three branched chain amino acids showed common pattern during reperfusion time: a decrease in 3 hr compared to sham, then a highest level in 24 hr and decrease in 72 hr reperfusion time, similar to their corresponding ketoacids. The protective effect of ischemic preconditioning was demonstrated by a faster tendency of plasma metabolites to normalize. Results also proved the remarkable metabolomic differences between the control (naïve) and sham-operated anesthetized animals, what warrants for critical evaluation of surgery/anaesthesy in the algorithm of metabolomic animal studies., (© 2020 International Union of Biochemistry and Molecular Biology.)

Remote ischemic perconditioning (RIPerC) results in collateral enhancement and a reduction in middle cerebral artery occlusion (MCAO) induced ischemia. RIPerC likely activates multiple metabolic protective mechanisms, including effects on matrix metalloproteinases (MMPs) and protein kinases. Here we explore if RIPerC improves neuroprotection and collateral flow by modifying the activities of MMP-9 and AMPK/e-NOS. Age matched adult male Sprague Dawley rats were subjected to MCAO followed one hour later by RIPerC (3 cycles of 15 min ischemia). Animals were euthanized 24 h post-MCAO. Haematoxylin and Eosin (H&E) staining 24 h post-MCAO revealed a significant (p < 0.02) reduction in the infarction volume in RIPerC treated animals (24.9 ± 5.4%) relative to MCAO controls (42.5 ± 4.2, %). TUNEL staining showed a 42.6% reduction in the apoptotic cells with RIPerC treatment (p < 0.01). Immunoblotting in congruence with RT-PCR and Zymography showed that RIPerC significantly reduced MMP-9 expression and activity in RIPerC + MCAO group compared to MCAO group (218.3 ± 19.1% vs. 148.9 ± 12.05% (p < 0.01). Immunoblotting revealed that RIPerC was associated with a significant 2.5-fold increase in activation of p-AMPK compared to the MCAO group (p < 0.01) which was also associated with a significant increase in the e-NOS activity (p < 0.01). RIPerC resulted in reduction of infarction volume, decreased apoptotic cell death and attenuated MMP-9 activity. This together with the increased activity of p-AMPK and increase in p-eNOS may, in part explain the neuroprotection and sustained increase in blood flow observed with RIPerC following acute stroke., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2020 Elsevier B.V. All rights reserved.)

Background: Remote ischemic conditioning during cerebral ischemia (remote ischemic perconditioning, RIPerC) refers to the application of several cycles of brief ischemia and reperfusion (I/R) commonly to a limb, and it represents a new paradigm in neuroprotection with multiple mechanisms of action in ischemic stroke (IS) patients during acute phase. Some clinical trials just finished, and a few others are still ongoing; gather the current knowledge and pull it down to influence the present and future studies was the goal of this paper., Methods: A systematic review of published research papers and/or registered clinical trials since 2000 was performed., Results: Nineteen studies were identified and only four studies were completed. All of them have demonstrated that RIPerC is safe, feasible and well tolerated in IS patients. However, a high heterogeneity of clinical trial characteristics was observed: five (26.3%) randomized clinical trials (RCTs) included only thrombolytic-treated patients, three (15.8%) RCTs only thrombectomy-treated patients, and five (26.3%) RCTs required radiological confirmation of IS. Temporal inclusion criteria vary from 4 h to 48 h. Most of the clinical trials used 4 cycles of RIPerC in the upper non-affected limb. Interestingly, only three (16.7%) RCTs applied RIPerC during the transportation in the ambulance. Neuroimaging outputs were the main endpoints when endovascular therapy was applied; functional outcome is also the main endpoint in large-medium size studies., Conclusions: This review summarizes the completed and ongoing clinical trials on RIPerC in IS patients, where RIPerC has been used alone or in combination with recanalization therapies. Ongoing clinical trials will provide new information on the best RIPerC intervention strategy and potentially improve the functional outcome of IS patients; definition of new RIPerC strategies would ideally aim at enhancing tissue preservation, promoting neurological recovery, and stratify patients to improve treatment feasibility.

The indirect use of the protective potential of stem cells in the form of cell secretomes has become an attractive strategy in regenerative medicine. In the present work, we studied the paracrine activity of blood cells that could be modulated towards a neuroprotective nature using in vivo remote conditioning (i.e. tolerant blood cells). The increased neuronal survival mediated by the tolerant secretome was clearly confirmed in vitro in a model of glutamate toxicity in a primary culture of rat cortical neurons and in vivo in a pre- and post-treatment of rats that were subjected to transient occlusion of the middle cerebral artery. Bioinformatic-based analysis of the protein profile revealed higher amounts of proteins released by the tolerant blood cells; 29 proteins were recognised as secreted. More than half of these secreted proteins were involved in the biological processes of the response to the stimulus (GO:0050896) and the response to chemicals (GO:0042221). The protective phenotype was most likely mediated by the synergistic effect of multiple identified proteins, including unique to the tolerant secretome (ceruloplasmin, D-3-phosphoglycerate dehydrogenase) and was promoted by the co-participation of several reaction pathways. The most probably of these pathways were post-translation protein modification, MAP2K and MAPK activation and platelet activation. Taken together, our results demonstrate that properly stimulated blood cells could serve as a source for cell-free-based therapies of regenerative medicine., (© 2019 International Society for Neurochemistry.)

Glutamate represents the main excitatory neurotransmitter in the mammalian brain; however, its excessive elevation in the extracellular space is cytotoxic and can result in neuronal death. The ischemia initiated brain damage reflects changes in glutamate concentration in peripheral blood. This paper investigated the role of the brain in blood efflux of the glutamate in an improved tolerance of the brain tissue to ischemic conditions. In the rat model of focal brain ischemia, the neuroprotection was initiated by rapid remote ischemic preconditioning (rRIPC). Our results confirmed a strong neuroprotective effect of rRIPC. We observed reduced infarction by about 78% related to improved neuronal survival by about 70% in the ischemic core. The level of tissue glutamate in core and penumbra dropped significantly and decreased to control value also in the core region of the contralateral hemisphere. Despite significant improvement of blood-brain barrier integrity (by about 76%), the additional gain of glutamate content in the peripheral blood was caused by rRIPC. Based on our results, we can assume that neuroprotection mediated by rapid remote ischemic preconditioning could lie in the regulated, whole-brain release of glutamate from nerve tissue to the blood, which preserves neurons from the exposure to glutamate toxicity and results in reduced infarction.

Remote ischemic preconditioning (RIPreC) and postconditioning (RIPostC) have been demonstrated to attenuate brain injury after ischemic stroke in healthy animals. This study investigated whether RIPreC and RIPostC exerted neuroprotection against cerebral ischemic injury in type 2 diabetic mice. RIPreC (24 h before ischemia) and RIPostC (immediately after reperfusion) were performed in an ischemia/reperfusion induced stroke model with type 2 diabetes. Ischemic outcomes, flow cytometry, multiplex cytokine assay, and western blotting were analyzed after 45 min of ischemia followed by 48 h of reperfusion. Our data indicated that RIPreC and RIPostC attenuated cerebral injuries and neurological deficits. RIPreC significantly reduced CD4 T cell and CD8 T cell infiltration and increased B cell infiltration into the ischemic brain. It also upregulated CD4 and CD8 T cell levels in the peripheral blood. However, RIPostC significantly decreased CD8 T cells infiltration and increased B cell infiltration into the ischemic brain. RIPreC inhibited IL-6 level in both the brain and blood, while RIPostC treatment attenuated IL-6 level upregulation in the peripheral blood. In addition, both RIPreC and RIPostC significantly increased p-ERK expression in the ipsilateral hemisphere in diabetic mice. This study indicated that RIPreC and RIPostC neuroprotection is present in type 2 diabetic mice via the modulation of brain ERK activity and inflammatory responses in both the peripheral blood and ischemic brain. However, the benefit was lower in RIPostC., Competing Interests: Declaration of competing interest The authors declare that they have no conflict of interest., (Copyright © 2020 Elsevier Ltd. All rights reserved.)

Objectives: This study was designed to investigate whether immunomodulation and Microglia polarization is involved in the anti-inflammatory and neuroprotective effect induced by hypoxic preconditioning (HPC) in the middle cerebral artery occlusion (MCAO) brain injury model., Methods: Longa method, (neurological disability status scale) NDSS method and TTC staining were used to evaluate the degree of cerebral infarction injury under different treatments (Sham, HPC, MCAO and co-treatment with HPC and MCAO). Western blot was used to detect expression profiles of apoptosis and related factors of neurological function. Flow cytometry was performed to analyze changes in the ratio of helper T cells, toxic T cells and NK cells in peripheral immune cells. And immunohistochemistry was used to examine the changes in microglial morphology. ELISA was used to evaluate the levels of nerve growth factors and neurogenesis conditions. Finally, RT-PCR was determined to analyze the transformation of microglia phenotype after HPC and MCAO treatment., Results: MCAO dramatically induced local formation of cerebral infarction. HPC relieved MCAO-induced cerebral infarction and increased rat cognition. HPC affected activation of microglia without significantly affecting in peripheral immune cell populations. After HPC co-treatment with MCAO, the M1 phenotype of microglia was changed and there was a transformation to M2., Conclusion: The treatment of HPC remarkably affected the polarization of microglia cells in MCAO rats, and reduced the cerebral nerve injury and played a protective role in MCAO model., (Copyright © 2019 The Authors. Published by Elsevier B.V. All rights reserved.)

Imbalances between cellular K + efflux and influx are considered to be involved in cerebral ischemia-reperfusion (I/R) injury. High-potassium pretreatment alleviates this injury, but the underlying molecular mechanism is unclear. In this study, we sought to investigate whether high-potassium preconditioning enhances cerebral tolerance to I/R injury through an anti-apoptotic mechanism. Adult male Sprague-Dawley rats were randomly divided into four groups (n = 40/group): a sham-operated group, normal saline group (3.2 ml/kg saline, intravenous (IV)), and low-dose and high-dose potassium chloride (KCl) groups (40 and 80 mg/kg KCl solution, IV, respectively). Subsequently, the rats underwent 90 min of middle cerebral artery occlusion (MCAO) followed by 24 hr of reperfusion (MCAO/R). Neurological deficit scores, 2,3,5-triphenyltetrazolium chloride (TTC) staining, hematoxylin and eosin staining, and TUNEL assay were used to assess neural injury. The expression of apoptotic proteins, brain potassium levels, mitochondrial function and oxidative stress were detected to explore the potential mechanism. After 24 hr of reperfusion, in both KCl treatment groups, neurological deficits and the cerebral infarct volume were reduced, and the apoptosis index of neurons was decreased. Furthermore, high-potassium preconditioning increased brain K + , adenosine triphosphate (ATP), cytochrome c oxidase (COX) levels, reduced malondialdehyde level, improved Na + /K + -ATPase, succinic dehydrogenase and superoxide dismutase activities, upregulated anti-apoptotic protein expression, and downregulated pro-apoptotic protein expression. This study suggests that high-potassium preconditioning enhanced cerebral tolerance to I/R injury in a rat MCAO/R model. The protective mechanism may involve apoptosis inhibition via preservation of intracellular K + and improvement of mitochondrial function., (© 2019 Wiley Periodicals, Inc.)

It is well-known that in ischemia-induced hypoxia, hypoxia-inducible factor -1α (HIF-1α) is critical in triggering expression of its downstream target genes to produce several products, such as erythropoietin (EPO), vascular endothelial growth factor (VEGF), nitric oxide synthesis (NOS), glucose transportor-1 (GLUT-1), insulin-like growth factor (IGF), which further promote erythropoiesis, angiogenesis, vasodilation and capitalization of glucose to overcome hypoxia. Meanwhile, as the factors with opposite effects on blood vessels, endothelin-1 (ET-1) and brain natriuretic peptide (BNP) also stand out strikingly in ischemic pathophysiology. To this day, several preconditioning manners have been used to induce tolerance to ischemia. During our research, exercise preconditioning was applied and it was demonstrated that HIF-1α triggered expression of ET-1 and BNP, which confirmed their downstream target genes for HIF-1α. And ET-1 may influcence expression of BNP to some degree but not the only factor which regulates BNP expression. Therefore, our findings suggest exercise preconditioning may provide protection to the ischemic brain tissue via HIF-1α which in turn increases expression of BNP to cause vasodilation in cooperation with some other factors, such as VEGF and EPO, to increase the blood flow in the ischemic area and then relieve the injuries induced by ischemia.

Ischemic preconditioning (IPC) exerts protective effects against ischemic cerebral injury. In the present study, an in vitro model of cerebral ischemia (oxygen and glucose deprivation, OGD) was established to investigate the neuroprotective mechanism of IPC. We found that conditioned medium (C.M.) from astrocytes rather than neurons nor microglia cell line BV2 exerted neuroprotection. Moreover, exosomes derived from OGD preconditioned astrocytes can be taken up by neurons and attenuated OGD-induced neuron death and apoptosis. High-throughput microRNA (miRNA) sequencing revealed that miR-92b-3p levels in exosomes released from preconditioned astrocytes were increased. Overexpression of miR-92b-3p in neurons with miR-92b-3p mimic achieved the same protective effects as C.M. from astrocytes. Thus, we propose that the mechanism of IPC may associate with astrocytes, and that exosome-mediated miR-92b-3p shuttle from preconditioned astrocytes to neurons participate in these process., (Copyright © 2019. Published by Elsevier B.V.)

Each year, thousands of patients are at risk of cerebral ischemic injury, due to iatrogenic responses to surgical procedures. Prophylactic treatment of these patients as standard care could minimize potential neurological complications. We have shown that protection of brain tissue, in a non-human primate model of cerebral ischemic injury, is possible through pharmacological preconditioning using the immune activator D192935. We postulate that preconditioning with D192935 results in neuroprotective reprogramming that is evident in the brain following experimentally induced cerebral ischemia. We performed quantitative proteomic analysis of cerebral spinal fluid (CSF) collected post-stroke from our previously published efficacy study to determine whether CSF protein profiles correlated with induced protection. Four groups of animals were examined: naïve animals (no treatment or stroke); animals treated with vehicle prior to stroke; D192935 treated and stroked animals, further delineated into two groups, ones that were protected (small infarcts) and those that were not protected (large infarcts). We found that distinct protein clusters defined the protected and non-protected animal groups, with a 16-member cluster of proteins induced exclusively in D192935 protected animals. Seventy percent of the proteins induced in the protected animals have functions that would enhance neuroprotection and tissue repair, including several members associated with M2 macrophages, a macrophage phenotype shown to contribute to neuroprotection and repair during ischemic injury. These studies highlight the translational importance of CSF biomarkers in defining mechanism and monitoring responses to treatment in development of stroke therapeutics.

Ischemic preconditioning (IPC) is a phenomenon in which a short-term sublethal ischemic exposure induces tolerance to a subsequent lethal ischemic insult; however, the detailed mechanism underlying IPC-induced neuroprotection remains obscure. Here, we applied middle cerebral artery occlusion, a preconditioning ischemic insult mouse model, to investigate the molecular mechanism underlying cerebral IPC. RNA sequencing and whole-genome bisulfite sequencing were performed to explore the gene expression profile and DNA methylation changes after cerebral IPC treatment. In this study, we identified 636 differentially expressed genes enriched for several pathways that were partially overlapping or interconnected in terms of similar gene function. The involvement of several genes in IPC-induced neuroprotection was first reported. Genes induced by IPC, including Arid5a, Nptx2 and Stc2, demonstrated a neuroprotective effect against oxygen-glucose deprivation induced neurotoxicity in vitro. Thus, our findings provide new insights into IPC signaling pathways and offer a novel therapeutic strategy towards stroke., (Copyright © 2019 The Author(s). Published by Elsevier Ltd.. All rights reserved.)

Background: Neurological complications after coronary artery bypass grafting (CABG) reduce quality of life, increase mortality, and inflate resource utilization. The risk of postoperative neurological complications parallels the increasing risk burden of the contemporary patient population. We evaluated the efficacy of remote ischemic preconditioning (RIPC) on inducing neuroprotection., Methods: Seventy patients undergoing first-time CABG were randomly assigned to RIPC or a sham procedure. Structural brain magnetic resonance imaging (MRI) was complemented with functional connectivity MRI to gain a whole-brain global connectivity analysis. Paired neurocognitive and MRI data were acquired pre- and postoperatively. The primary end point was a composite of new ischemic brain lesions and neurocognitive impairment. Secondary end points included brain connectivity profiles, pooled ischemic volumes, and individual components of the primary outcome. The Shapiro-Wilk test was used to determine whether a data set followed a normal distribution. The Fisher exact test was used to calculate the measures of association for categorical variables, whereas continuous data were tested with either the Mann-Whitney U test or the Student t test., Results: There was no between-group difference in the incidence of the primary end point (9 [27%] in the RIPC group vs 8 [24%] in the control group, odds ratio, 1.17 [95% confidence interval, 0.34-4.06]; P = 1.0). Although RIPC did not reduce the incidence of brain ischemia (8/33 [24%] vs 7/33 [21%]; P = 1.0), the pooled ischemic volume was lower in the RIPC group (157 [interquartile range, 125-231] vs 777 [interquartile range, 564-965] mm 3 ; P = .004). Postoperative neurocognition was marginally superior in the RIPC group as evidenced by a lower absolute number of abnormal neurocognitive tests in the RIPC group (7/99 [7%] vs 16/99 [16%]; odds ratio, 0.40 [95% confidence interval, 0.14-1.09]; P = .074). Robust reductions of functional connectivity profiles for the associative thalamus were documented in both groups, irrespective of RIPC (RIPC group, t = 3.31; P < .01; and the control group, t = 3.52; P < .01)., Conclusions: Silent brain ischemia occurs frequently after CABG. RIPC did not reduce the incidence of the primary outcome. However, RIPC significantly reduced the pooled volume of ischemic brain lesions. Surgery adversely affected global brain connectivity, with RIPC conferring no demonstrable protection. The association of RIPC with superior neurocognitive test scores failed to cross the threshold for significance., (Copyright © 2018. Published by Elsevier Inc.)

Cerebral preconditioning (PC) confers endogenous brain protection after stroke. Ischemic stroke patients with a prior transient ischemic attack (TIA) may potentially be in a preconditioned state. Although PC has been associated with the activation of pro-survival signals, the mechanism by which preconditioning confers neuroprotection is not yet fully clarified. Recently, we have described that PC-mediated neuroprotection against ischemic insult is promoted by p53 destabilization, which is mediated by its main regulator MDM2. Moreover, we have previously described that the human Tp53 Arg72Pro single nucleotide polymorphism (SNP) controls susceptibility to ischemia-induced neuronal apoptosis and governs the functional outcome of patients after stroke. Here, we studied the contribution of the human Tp53 Arg72Pro SNP on PC-induced neuroprotection after ischemia. Our results showed that cortical neurons expressing the Pro72-p53 variant exhibited higher PC-mediated neuroprotection as compared with Arg72-p53 neurons. PC prevented ischemia-induced nuclear and cytosolic p53 stabilization in Pro72-p53 neurons. However, PC failed to prevent mitochondrial p53 stabilization, which occurs in Arg72-p53 neurons after ischemia. Furthermore, PC promoted neuroprotection against ischemia by controlling the p53/active caspase-3 pathway in Pro72-p53, but not in Arg72-p53 neurons. Finally, we found that good prognosis associated to TIA within 1 month prior to ischemic stroke was restricted to patients harboring the Pro72 allele. Our findings demonstrate that the Tp53 Arg72Pro SNP controls PC-promoted neuroprotection against a subsequent ischemic insult by modulating mitochondrial p53 stabilization and then modulates TIA-induced ischemic tolerance.

Our previous studies have demonstrated that limb ischemic preconditioning (LIP) induced brain ischemic tolerance and up-regulated the expression of p38 MAPK and ERK in the hippocampal CA1 region in rats. The present study was undertaken to investigate the role of adenosine in brain protection and up-regulation of p38 MAPK and ERK induced by LIP. It was found that adenosine A1 receptor antagonist DPCPX dose-dependently inhibited the protective effect of LIP. The up-regulation of p38 MAPK and ERK induced by LIP could be blocked by DPCPX. Furthermore, we observed the effect of adenosine on the brain ischemia. The results showed that pre-administration of adenosine could partly mimic the neuroprotective effect on the brain, up-regulate the expression of p38 MAPK and ERK. Based on the above results, it can be concluded that adenosine participated in brain protection and up-regulation of the expression of p38 MAPK and ERK during the induction of brain ischemic tolerance after LIP., (Copyright © 2018. Published by Elsevier B.V.)

14-3-3γ is an important early ischemia-inducible protective factor against ischemic cell death in cerebral cortical neurons. We investigated the anti-apoptosis mechanism of enhanced 14-3-3γ mediated by preconditioning exercise-induced brain ischemic tolerance after stroke. Rats were assigned to four groups: exercise and ischemia (Ex group), ischemia and no exercise (No-Ex group), exercise and no ischemia (Ex-only group), and no exercise and ischemia (control group). Rats were trained on a treadmill for 5 days a week for 3 weeks (running speed, 25 m/min; running duration, 30 min/day). After the exercise program, stroke was induced by left middle cerebral artery occlusion. The infarct volume, neurological deficits, and motor function, as well as expression levels of hypoxia-induced factor-1α (HIF-1α), 14-3-3γ, P2X7 receptors, p-β-catenin Ser37, Bax, and caspase 3 were evaluated by immunohistochemistry and western blotting. The expression of HIF-1α and 14-3-3γ significantly increased in neurons and astrocytes in the Ex-only group. HIF-1α was co-expressed with P2X7 receptor- and GFAP-positive astrocytes. After stroke, the Ex group had significantly reduced brain infarction. HIF-1α and 14-3-3γ significantly increased in the Ex group compared to the No-Ex group. In addition, p-β-catenin Ser37 significantly increased following elevated 14-3-3γ; in contrast, Bax and caspase 3 were significantly reduced in the Ex group. Our findings suggest that preconditioning exercise prior to ischemia induces neuron- and astrocyte-mediated brain ischemic tolerance through increased expression of HIF-1α and 14-3-3γ, which are intrinsic protective factors; the upregulated 14-3-3γ induced by preconditioning exercise reduces ischemic neuronal cell death through the 14-3-3γ/p-β-catenin Ser37/Bax/caspase 3 anti-apoptotic pathway.

Background: Transient Ischemia/Reperfusion (I/R) is the main reason for brain injury and results in disruption of the Blood-Brain Barrier (BBB). It had been reported that BBB injury is one of the main risk factors for early death in patients with cerebral ischemia. Numerous investigations focus on the study of BBB injury which have been carried out., Objective: The objective of this study was to investigate the treatment function of the activation of the Hippo/Yes-Associated Protein (YAP) signaling pathway by combined Ischemic Preconditioning (IPC) and resveratrol (RES) before brain Ischemia/Reperfusion (BI/R) improves Blood-Brain Barrier (BBB) disruption in rats., Methods: Sprague-Dawley (SD) rats were pretreated with 20 mg/kg RES and IPC and then subjected to 2 h of ischemia and 22 h of reperfusion. The cerebral tissues were collected; the cerebral infarct volume was determined; the Evans Blue (EB) level, the brain Water Content (BWC), and apoptosis were assessed; and the expressions of YAP and TAZ were investigated in cerebral tissues., Results: Both IPC and RES preconditioning reduced the cerebral infarct size, improved BBB permeability, lessened apoptosis, and upregulated expressions of YAP and transcriptional co-activator with PDZ-binding motif (TAZ) compared to the Ischemia/Reperfusion (I/R) group, while combined IPC and RES significantly enhanced this action., Conclusion: combined ischemic preconditioning and resveratrol improved blood-brain barrier breakdown via Hippo/YAP/TAZ signaling pathway., (Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.net.)

Preconditioning with lipopolysaccharide (LPS) or opioid antagonists has a neuroprotective effect in ischemic insults. However, the co‑preconditioning effect of toll‑like receptor ligands and opioid antagonists has not been investigated. In this study we examined the neuroprotective effect of LPS and naltrexone (NTX) preconditioning and co‑preconditioning in unilateral selective hippocampal ischemia in rats to assess for possible synergistic protective effects. LPS and NTX were injected unilaterally into the left cerebral ventricle of male rats. Forty‑eight hours after LPS and twenty‑four hours after NTX injection, ipsilateral selective hippocampal ischemia was induced using a modified version of the photothrombotic method. Protective effects for LPS and NTX were assessed by evaluating infarct volume (using 2,3,5‑triphenyltetrazolium chloride staining), and cognitive function (using radial arm water maze and passive avoidance tests). Animals in the ischemic group had an infarct lesion and considerable cognitive impairment, compared with the sham group. LPS or NTX preconditioning significantly reduced the infarct size and improved cognitive function. Moreover, co‑preconditioning with LPS and NTX increased the protective effect compared with preconditioning with LPS or NTX alone. Our data showed that LPS and NTX preconditioning resulted in a neuroprotective effect in hippocampal ischemia. Furthermore, co‑preconditioning with LPS and NTX resulted in a synergistic protective effect.

Elevated homocysteine (Hcy) level is a well known risk factor for cardiovascular and neuropsychiatric diseases. In this study, we investigated metabolic changes in blood plasma in Hcy-treated rats. In combination with Hcy injections to induce hyperhomocysteinemia-like state, we used an animal model of global cerebral ischemia to investigate metabolic changes after 24 h reperfusion in rats. We also focused on the endogenous phenomenon known as ischemic tolerance induced by the preischemic treatment. The experiments were carried out on blood plasma samples as they are easily available and metabolically reflect the overall changes in injured organism. We observed significant changes in plasma metabolite levels of: pyruvate, citrate, acetate implicating alterations in energy metabolism, and increase in triacylglycerols, arginine and lysine, in Hcy-treated rats compared with naive animals. Ischemic insult with 24 reperfusion in Hcy-treated rats led to increase in plasma lactate, and decrease in plasma glucose, pyruvate, citrate and acetate. Complementary, an increase in ketone body 3-hydroxybutyrate was observed. The plasma metabolites: alanine, lactate, valine, glucose, leucine, isoleucine, acetate, citrate and 3-hydroxybutyrate were considered to reflect the response induced by ischemic preconditioning in Hcy rats, where the extent of postischemic damage was not as high as in the non-preconditioned rats. Our results provide evidence that nuclear magnetic resonance (NMR) spectra analysis can identify a specific group of metabolites present in plasma with the capability of discriminating between individual groups of animals. Regarding the effect of elevated Hcy level on plasma metabolome, we showed, that acetate, pyruvate and glucose had the excellent discriminatory power between Hcy-treated and naive rats plasma. Concerning ischemic insult in Hcy-treated animals, we also document the ideal discrimination of ischemic from non-ischemic rats by various groups of metabolites, that can be considered as a potential plasma biomarkers.

Background: Sevoflurane preconditioning induces brain ischemic tolerance, but the mechanism remains poorly elucidated. Nitration is an important form of post-translational modification in pathological signaling. This study was to investigate the role of thioredoxin-1 (Trx-1) nitration in neuroprotection effect induced by sevoflurane preconditioning in a transient stroke model in rats., Methods: Adult male Sprague-Dawley rats were preconditioned with 2% sevoflurane or vehicle oxygen exposure, 1 h per day, for 5 consecutive days. At 24 h after the last exposure, rats were subjected to focal brain ischemia induced by middle cerebral artery occlusion (MCAO) for 90 min, followed by 72-h reperfusion. Trx-1 expression and activity, as well as the content of nitrotyrosine at penumbra were detected at 24 h after preconditioning and 2, 8, 24, 72 h after MCAO. Nitrated Trx-1 was examined by immunoprecipitation at 8 h after MCAO. The role of Trx-1 nitration in ischemic tolerance was assessed by administration of nitrated human-Trx-1 prior to MCAO. Neurological scores, brain infarct volumes and TUNEL staining were evaluated at 24 h after reperfusion., Results: Ischemic stroke decreased Trx-1 activity but not the expression in penumbra tissue. The content of nitrotyrosine was elevated after MCAO. Preconditioning with sevoflurane increased Trx-1 activity and reduced its nitration at 8 h after MCAO in comparison with vehicle preconditioning. The decrement of Trx-1 activity was correlated with its nitration level. Exogenous administration of nitrated human-Trx-1 reversed the brain ischemic tolerance of sevoflurane preconditioning, exacerbating brain infarct volume, neurobehavioral defects and apoptosis, while administration of human-Trx-1 had no effect on the sevoflurane preconditioning-induced neuroprotection., Conclusion: Ischemic stroke reduces Trx-1 activity via post-translational nitrative modulation in rats. Sevoflurane preconditioning induces brain ischemic tolerance and anti-apoptosis by partially preserving Trx-1 activity via inhibiting nitration.

Despite great improvement during the past several decades, the management of stroke is still far from satisfactory, which warrants alternative or adjunctive strategies. Remote ischemic conditioning (RIC), an easy-to-use and noninvasive therapy, can be performed in various clinical scenarios (e.g., prehospital transportation, intrahospital, and at home), and it has been widely investigated for stroke management. RIC has been demonstrated to be well tolerated in patients with acute ischemic stroke and aneurysm subarachnoid hemorrhage, and it may benefit these patients by improving clinical outcomes; in patients with intracranial atherosclerosis, long-term repeated RIC could be safely performed and benefit patients by reducing recurrent ischemic stroke and transient ischemic attack, as well as improving cerebral perfusion status; long-term repeated RIC may also benefit patients with cerebral small vessel disease by slowing cognitive decline and reducing volume of white matter hyperintensities on brain MRI; in patients with severe carotid atherosclerotic stenosis undergoing stenting, preprocedural RIC could reduce the odds of new brain lesions on postprocedural MRI. Previous clinical studies suggest broad future prospects of RIC in the field of cerebrovascular diseases. However, the optimal RIC protocol and the mechanisms that RIC protects the brain is not fully clear, and there is lack of sensitive and specific biomarkers of RIC, all these dilemmas prevent RIC from entering clinical practice. This review focuses on recent advances in clinical studies of RIC in stroke management, its challenges, and the potential directions of future studies.

The brain consists of neurons and much higher number of glial cells. They communicate each other, by which they control brain functions. The brain is highly vulnerable to several insults such as ischemia, but has a self-protective and self-repairing mechanisms against these. Ischemic tolerance or preconditioning is an endogenous neuroprotective phenomenon, where a mild non-lethal ischemic episode can induce resistance to a subsequent severe ischemic injury in the brain. Because of its neuroprotective effects against cerebral ischemia or stroke, ischemic tolerance has been widely studied. However, almost all studies have been performed from the viewpoint of neurons. Glial cells are structurally in close association with synapses. Recent studies have uncovered the active roles of astrocytes in modulating synaptic connectivity, such as synapse formation, elimination and maturation, during development or pathology. However, glia-mediated ischemic tolerance and/or neuronal repairing have received only limited attention. We and others have demonstrated that glial cells, especially astrocytes, play a pivotal role in regulation of induction of ischemic tolerance as well as repairing/remodeling of neuronal networks by phagocytosis. Here, we review our current understanding of (1) glial-mediated ischemic tolerance and (2) glia-mediated repairing/remodeling of the penumbra neuronal networks, and highlight their mechanisms as well as their potential benefits, problems, and therapeutic application., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Cardiac arrest is one of the leading causes of death among adults in older age. Understanding mechanisms how organism responds to ischemia at global level is essential for the prevention and ischemic patient's treatment. In this study, we used a global cerebral ischemia induced by four-vessel occlusion as an established animal model for ischemic stroke to investigate metabolic changes after 24 h reperfusion, when transitions occur due to the onset of delayed neuronal death. We also focused on the endogenous phenomenon known as ischemic tolerance by the pre-ischemic treatment. The experiments were carried out on blood plasma samples as easily available and metabolically reflecting the overall changes in injured organism. Our results imply that disturbed glycolysis pathway, as a consequence of ischemic injury, leads to the increased level of ketone bodies (acetone, acetoacetate and β-hydroxybutyrate) along with increased utilization of triacylglycerols in plasma of ischemic and ischemically preconditioned rats. Complementary to, a decreased level of glycolytic intermediates (lactate, pyruvate, acetate) with increased level of glucose was found in ischemic and preconditioned animals. The protective effect of ischemic preconditioning on metabolome recovery was demonstrated by significantly increased level of creatine compared to ischemic, non-preconditioned rats. We also document that acetoacetate, pyruvate, lactate, and leucine have the best discriminatory power between ischemic and control plasma. Conclusively, our results provide evidence that NMR spectra analysis can identify specific group of metabolites present in plasma with the capability for discrimination between individual groups of animals. In addition, an excellent feasibility for the statistical discrimination among ischemic, preconditioned, and control rats can be applied regardless of native or deproteinated plasma and also regardless of noesy or cpmg NMR acquisition.

Brain ischemic preconditioning (IPC) with mild ischemic episodes is well known to protect the brain against subsequent ischemic challenges. However, the underlying mechanisms are poorly understood. Here we demonstrate the critical role of the master redox transcription factor, nuclear factor (erythroid-derived 2)-like 2 (Nrf2), in IPC-mediated neuroprotection and blood-brain barrier (BBB) preservation. We report that IPC causes generation of endogenous lipid electrophiles, including 4-hydroxy-2-nonenal (4-HNE), which release Nrf2 from inhibition by Keap1 (via Keap1-C288) and inhibition by glycogen synthase kinase 3β (via GSK3β-C199). Nrf2 then induces expression of its target genes, including a new target, cadherin 5, a key component of adherens junctions of the BBB. These effects culminate in mitigation of BBB leakage and of neurological deficits after stroke. Collectively, these studies are the first to demonstrate that IPC protects the BBB against ischemic injury by generation of endogenous electrophiles and activation of the Nrf2 pathway through inhibition of Keap1- and GSK3β-dependent Nrf2 degradation., (Copyright © 2018 The Authors. Published by Elsevier B.V. All rights reserved.)

Background: Remote ischemic preconditioning (RIPC) is a mechanism to protect tissues from injury during ischemia and reperfusion. We investigated the neuroprotective effects of RIPC in neonates undergoing cardiac surgery., Methods: The outcome was white matter injury (WMI), assessed by the change in volume of WMI from preoperative to postoperative brain magnetic resonance imaging (MRI). Patients were randomized to RIPC or SHAM. RIPC was induced prior to cardiopulmonary bypass by four 5-minute cycles of blood pressure cuff inflation to produce ischemia of the lower extremity. For patients randomized to SHAM, the cuff was positioned, but not inflated., Results: The study included 67 patients, with 33 randomized to RIPC and 34 randomized to SHAM. Preoperative and postoperative MRIs were available in 50 patients, including 26 of the 33 RIPC patients and 24 of the 34 SHAM patients. There were no differences in baseline and operative characteristics for either the overall study group or the group with evaluable MRIs. WMI was identified in 28% of the patients preoperatively and in 62% postoperatively. There was no difference in the prevalence of WMI by treatment group (p > 0.5). RIPC patients had an average change in WMI of 600 mL 3 , and SHAM subjects had an average WMI change of 107 mL 3 . There was no significant difference in the mean value of WMI change between patients who received RIPC and those who received SHAM treatments (p = 0.178)., Conclusions: In this randomized, blinded clinical trial, there was no evidence that use of RIPC provides neuroprotection in neonates undergoing repair of congenital heart defects with cardiopulmonary bypass., (Copyright © 2018 The Society of Thoracic Surgeons. Published by Elsevier Inc. All rights reserved.)

After ischaemic stroke, brain damage can be curtailed by rescuing the 'ischaemic penumbra' - that is, the severely hypoperfused, at-risk but not yet infarcted tissue. Current evidence-based treatments involve restoration of blood flow so as to salvage the penumbra before it evolves into irreversibly damaged tissue, termed the 'core'. Intravenous thrombolysis (IVT) can salvage the penumbra if given within 4.5 h after stroke onset; however, the early recanalization rate is only ~30%. Direct removal of the occluding clot by mechanical thrombectomy considerably improves outcomes over IVT alone, but despite early recanalization in > 80% of cases, ~50% of patients who receive this treatment do not enjoy functional independence, usually because the core is already too large at the time of recanalization. Novel therapies aiming to 'freeze' the penumbra - that is, prevent core growth until recanalization is complete - hold potential as adjuncts to mechanical thrombectomy. This Review focuses on nonpharmacological approaches that aim to restore the physiological balance between oxygen delivery to and oxygen demand of the penumbra. Particular emphasis is placed on normobaric oxygen therapy, hypothermia and sensory stimulation. Preclinical evidence and early pilot clinical trials are critically reviewed, and future directions, including clinical translation and trial design issues, are discussed.

Background OBJECTIVE: The cerebral ischemia/reperfusion greatly influences brain metabolism. Remote ischemia preconditioning (RIPC) is reported to confer neuroprotective effects against cerebral ischemia in animal models and human. This study aims to investigate the metabolomic profiles of cerebrospinal fluid (CSF) in patients treated with repetitive lower limb RIPC and provides an insight into possible mechanism underlying RIPC-induced neuroprotection., Method: Fifty healthy patients undergoing minor surgery under spinal anesthesia were randomly allocated to 2 groups: control group (Group C, n = 25) and RIPC treatment group (Group T,n = 25). Repetitive limb RIPC were performed 3 sessions, consisting of three 5-min cycles per session from the day before surgery to the morning on the surgery day. The CSF samples were collected from 48 patients before intrathecal injection of local anesthetic. A proton nuclear magnetic resonance ( 1 H NMR)-based metabonomics approach was used to obtain the CSF metabolic profiles of the samples (n = 24 each). The acquired data were processed with MestReNova and followed by statistical analysis with SIMCA-P., Results: The model obtained with the orthogonal partial least-squares discriminant analysis (OPLS-DA) identified difference of metabolite profiles between two groups. The validation of the discriminant analysis showed that the accuracy of the OPLS-DA model was 81.3%. Sixteen metabolites including glucose, amino-acids and organic acids et al. were identified as the most influential CSF biomarkers for the discrimination between two groups, which are involved in pathways of energy metabolism and amino-acids metabolism., Conclusion: 1 H NMR spectra combined with pattern recognition analysis offers a new and promising platform to investigate metabolic signatures in patients treated with RIPC. Our results suggest repetitive RIPC mainly changes energy metabolism and amino-acid metabolism in brain, which provides a potential mechanistic understanding of RIPC-induced tolerance to cerebral ischemia., (Copyright © 2018 Elsevier Ltd. All rights reserved.)

Stroke induces complex and dynamic, local and systemic changes including inflammatory reactions, immune responses, and repair and recovery processes. Mesenchymal stem cells (MSCs) have been shown to enhance neurological recovery after stroke. We hypothesized that serum factors play a critical role in the activation of bone marrow (BM) MSCs after stroke such as by increasing proliferation, paracrine effects, and rejuvenation. Human MSCs (hMSCs) were grown in fetal bovine serum (FBS), normal healthy control serum (NS), or stroke patient serum (SS). MSCs cultured in growth medium with 10% SS or NS exhibited higher proliferation indices than those cultured with FBS ( P < 0.01). FBS-, NS-, and SS-hMSCs showed differences in the expression of trophic factors; vascular endothelial growth factor, glial cell-derived neurotrophic factor, and fibroblast growth factor were densely expressed in samples cultured with SS ( P < 0.01). In addition, SS-MSCs revealed different cell cycle- or aging-associated messenger RNA expression in a later passage, and β-galactosidase staining showed the senescence of MSCs observed during culture expansion was lower in MSCs cultured with SS than those cultured with NS or FBS ( P < 0.01). Several proteins related to the activity of receptors, growth factors, and cytokines were more prevalent in the serum of stroke patients than in that of normal subjects. Neurogenesis and angiogenesis were markedly increased in rats that had received SS-MSCs ( P < 0.05), and these rats showed significant behavioral improvements ( P < 0.01). Our results indicate that stroke induces a process of recovery via the activation of MSCs. Culture methods for MSCs using SS obtained during the acute phase of a stroke could constitute a novel MSC activation method that is feasible and efficient for the neurorestoration of stroke.

Ischemic tolerance is the establishment of brain resistance to severe ischemic damage by a mild preconditioning stimulus, insufficient to irreversible tissue damage, but capable of initiating a defense response. We developed the model of focal-focal ischemic tolerance, in which the first local photothrombotic infarct (PTI) in the rat brain cortex reduced the infarct caused by second PTI applied to the contralateral cortex of the same rat 7 days later. Using antibody microarrays, we compared protein profiles in the penumbra surrounding the PTI core after single and double PTI. We observed up- or downregulation of several dozens of proteins that are aimed at neurodegeneration or neuroprotection. Both single and double PTI induced damaging processes in the rat cerebral cortex that included over-expression of various pro-apoptotic and signaling proteins and downregulation of other signaling proteins and regulators of proliferation, some components of actin, intermediate fiber and microtubular cytoskeletons, and proteins involved in vesicle transport and synaptic transmission. The simultaneous protective processes included the upregulation of different signaling and anti-apoptotic proteins, stimulators of proliferation, and proteins involved in remodeling of actin cytoskeleton. The elevated expression of some signaling proteins, such as calcium-dependent PLCγ1, PKVα1, CaMKIIα, calnexin, and calreticulin was preserved after double PTI. Less pro-survival proteins were downregulated in the penumbra after double than single impact.

A mild non-lethal ischemic episode can induce resistance to a subsequent severe ischemic injury in the brain. This phenomenon is termed ischemic tolerance or ischemic preconditioning, and is an endogenous mechanism that can provide robust neuroprotection. Because of its neuroprotective effects against cerebral ischemia or stroke, ischemic tolerance has been widely studied. However, almost all studies have been performed from the viewpoint of neurons. Accumulating evidence suggests that glial cells have various roles in regulation of brain function, including modulation of synaptic transmission, neuronal excitation, and neuronal structure. In addition, astrocytes are closely related to homeostasis, stability of brain function, and protection of neurons. However, glial cells have received only limited attention with regard to ischemic tolerance. Cross-ischemic preconditioning is a phenomenon whereby non-ischemic preconditioning such as mechanical, thermal, and chemical treatment can induce ischemic tolerance. Of these, chemical treatments that affect the immune system can strongly induce ischemic tolerance, suggesting that glial cells may have important roles in this process. Indeed, we and others have demonstrated that glial cells, especially astrocytes, play a pivotal role in the induction of ischemic tolerance. This glial-mediated ischemic tolerance provides a robust and long-lasting neuroprotection against ischemic injury. In this review, we discuss the mechanisms underlying glial-mediated ischemic tolerance, as well as its potential benefits, problems, and therapeutic application., (Copyright © 2017. Published by Elsevier B.V.)

Background: Isoflurane preconditioning (IPC) induces cerebral ischaemic tolerance, but the mechanism remains poorly understood. The aim of this study was to determine changes in mitochondrial function in the brain after IPC, and whether the cannabinoid receptor 1 (CB1R) could be involved in the mechanism of mitochondrial protection mediated by IPC., Methods: Adult male Sprague-Dawley rats were pretreated with isoflurane 2% for 1 h day -1 , for 5 days consecutively, and then subjected to 120 min right middle cerebral artery occlusion. Cannabinoid receptor 1 expression in the cellular and mitochondrial membrane was measured. The CB1R agonist HU-210 was administered alone, or the antagonists AM251 and SR141716A were given to the animals before each preconditioning. Neurological scores, infarct volume, apoptosis, and mitochondrial function were examined after middle cerebral artery occlusion., Results: Expression of CB1R on cellular and mitochondrial membranes was increased 6 h after preconditioning. Both IPC and HU-210 administration before middle cerebral artery occlusion improved neurological outcomes and reduced infarct volume. Isoflurane preconditioning increased the expression of the anti-apoptotic proteins Bcl-2 and Bcl-X L and reduced apoptosis in neurones. Isoflurane preconditioning and HU-210 also markedly preserved the activity of respiratory chain complexes, reduced mitochondrial radical generation, preserved mitochondrial membrane potential, and inhibited mitochondrial permeability transition pore opening. Cannabinoid receptor 1 antagonists abolished the improvement in mitochondrial function and the neuroprotective effects induced by IPC., Conclusions: Our results indicate that IPC elicits brain ischaemic tolerance and mitochondrial protection by activating the CB1R, which provides a new mechanism for IPC-induced neuroprotection against cerebral ischaemia., (© The Author 2017. Published by Oxford University Press on behalf of the British Journal of Anaesthesia. All rights reserved. For Permissions, please email: journals.permissions@oup.com)

Cardiac arrest-induced global cerebral ischemia is a main cause of neurological dysfunction in emergency medicine. Transplantation with bone marrow mesenchymal stem cells (MSCs) has been used in stroke models to repair the ischemic brain injury, but it is little studied in models with global cerebral ischemia. In the present study, a hypoxia precondition was used to improve the efficacy of MSC transplantation, given the low survival and migration rates and limited differentiation capacities of MSCs. We found that hypoxia can increase the expansion and migration of MSCs by activating the PI3K/AKT and hypoxia-inducible factor-1α/CXC chemokine receptor-4 pathways. By using a cardiac arrest-induced global cerebral ischemic model in rats, we found that transplantation of hypoxia-preconditioned MSCs promoted the migration and integration of MSCs and decreased neuronal death and inflammation in the ischemic cortex. © 2017 Wiley Periodicals, Inc., (© 2017 Wiley Periodicals, Inc.)

Conventional protein kinase C (cPKC)γ participated in cerebral hypoxic preconditioning-induced neuroprotection and affected the neurological outcome of ischemic stroked mice. As an independent predictor of ischemic stroke, the internal carotid artery occlusion (ICAO)-caused brain-regional ischemic injury may worsen the neurological outcome of patients. However, the brain-regional ischemic vulnerability and its underlying mechanism remain unclear. In this study, the bilateral ICAO (BICAO) model was applied in cPKCγ wild type (WT) and knockout (KO) mice to determine the cPKCγ impact on brain-regional ischemic vulnerability. The arterial spin labeling (ASL) imaging results showed that 7 days BICAO-induced global ischemia could cause significant blood perfusion loss in prefrontal cortex (69.13%), striatum (61.69%), hypothalamus (67.36%), hippocampus (69.82%) and midbrain (40.53%) of WT mice, along with neurological deficits. Nissl staining and Western blot results indicated that hypothalamus and midbrain had more severe neural cell loss than prefrontal cortex, striatum and hippocampus, which negatively coincided with endogenous cPKCγ protein levels but not blood perfusion loss and cPKCγ membrane translocation levels. Furthermore, we found that cPKCγ KO significantly aggravated the neuron loss in prefrontal cortex, striatum and hippocampus and abolish the regional ischemic vulnerability by using immunofluorescent staining with neuron-specific marker NeuN. Similarly, cPKCγ KO also significantly increased Caspase-3, -8 and -9 cleavage levels in prefrontal cortex, striatum, hippocampus, hypothalamus and midbrain of mice with 24 h BICAO. These results suggested that hypothalamus and midbrain are more vulnerable to ischemia, and endogenous cPKCγ affects the regional ischemic vulnerability through modulating Caspase-8 and -9 dependent cell apoptosis.

Collateral circulation is a key variable determining prognosis and response to recanalization therapy during acute ischemic stroke. Remote ischemic perconditioning (RIPerC) involves inducing peripheral ischemia (typically in the limbs) during stroke and may reduce perfusion deficits and brain damage due to cerebral ischemia. In this study, we directly investigated pial collateral flow augmentation due to RIPerC during distal middle cerebral artery occlusion (MCAo) in rats. Blood flow through pial collaterals between the anterior cerebral artery (ACA) and the MCA was assessed in male Sprague Dawley rats using in vivo laser speckle contrast imaging (LSCI) and two photon laser scanning microscopy (TPLSM) during distal MCAo. LSCI and TPLSM revealed that RIPerC augmented collateral flow into distal MCA segments. Notably, while control rats exhibited an initial dilation followed by a progressive narrowing of pial arterioles 60 to 150-min post-MCAo (constricting to 80-90% of post-MCAo peak diameter), this constriction was prevented or reversed by RIPerC (such that vessel diameters increased to 105-110% of post-MCAo, pre-RIPerC diameter). RIPerC significantly reduced early ischemic damage measured 6 h after stroke onset. Thus, prevention of collateral collapse via RIPerC is neuroprotective and may facilitate other protective or recanalization therapies by improving blood flow in penumbral tissue.

Our previous findings suggest that sphingosine kinase 2 (SPK2) mediates ischemic tolerance and autophagy in cerebral preconditioning. The aim of this study was to determine by which mechanism SPK2 activates autophagy in neural cells. In both primary murine cortical neurons and HT22 hippocampal neuronal cells, overexpression of SPK2 increased LC3II and enhanced the autophagy flux. SPK2 overexpression protected cortical neurons against oxygen glucose deprivation (OGD) injury, as evidenced by improvement of neuronal morphology, increased cell viability and reduced lactate dehydrogenase release. The inhibition of autophagy effectively suppressed the neuroprotective effect of SPK2. SPK2 overexpression reduced the co-immunoprecipitation of Beclin-1 and Bcl-2, while Beclin-1 knockdown inhibited SPK2-induced autophagy. Both co-immunoprecipitation and GST pull-down analysis suggest that SPK2 directly interacts with Bcl-2. SPK2 might interact to Bcl-2 in the cytoplasm. Notably, an SPK2 mutant with L219A substitution in its putative BH3 domain was not able to activate autophagy. A Tat peptide fused to an 18-amino acid peptide encompassing the native, but not the L219A mutated BH3 domain of SPK2 activated autophagy in neural cells. The Tat-SPK2 peptide also protected neurons against OGD injury through autophagy activation. These results suggest that SPK2 interacts with Bcl-2 via its BH3 domain, thereby dissociating it from Beclin-1 and activating autophagy. The observation that Tat-SPK2 peptide designed from the BH3 domain of SPK2 activates autophagy and protects neural cells against OGD injury suggest that this structure may provide the basis for a novel class of therapeutic agents against ischemic stroke.

Background and Purpose: Repeated episodes of limb ischemia and reperfusion (remote ischemic conditioning [RIC]) may improve outcome after acute stroke., Methods: We performed a pilot blinded placebo-controlled trial in patients with acute ischemic stroke, randomized 1:1 to receive 4 cycles of RIC within 24 hours of ictus. The primary outcome was tolerability and feasibility. Secondary outcomes included safety, clinical efficacy (day 90), putative biomarkers (pre- and post-intervention, day 4), and exploratory hemodynamic measures., Results: Twenty-six patients (13 RIC and 13 sham) were recruited 15.8 hours (SD 6.2) post-onset, age 76.2 years (SD 10.5), blood pressure 159/83 mm Hg (SD 25/11), and National Institutes of Health Stroke Scale (NIHSS) score 5 (interquartile range, 3.75-9.25). RIC was well tolerated with 49 out of 52 cycles completed in full. Three patients experienced vascular events in the sham group: 2 ischemic strokes and 2 myocardial infarcts versus none in the RIC group ( P =0.076, log-rank test). Compared with sham, there was a significant decrease in day 90 NIHSS score in the RIC group, median NIHSS score 1 (interquartile range, 0.5-5) versus 3 (interquartile range, 2-9.5; P =0.04); RIC augmented plasma HSP27 (heat shock protein 27; P <0.05, repeated 2-way ANOVA) and phosphorylated HSP27 ( P <0.001) but not plasma S100-β, matrix metalloproteinase-9, endocannabinoids, or arterial compliance., Conclusions: RIC after acute stroke is well tolerated and appears safe and feasible. RIC may improve neurological outcome, and protective mechanisms may be mediated through HSP27. A larger trial is warranted., Clinical Trial Registration: URL: http://www.isrctn.com. Unique identifier: ISRCTN86672015., (© 2017 American Heart Association, Inc.)

Preconditioning by brief ischemic episode induces tolerance to a subsequent lethal ischemic insult, and it has been suggested that reactive oxygen species are involved in this phenomenon. Thioredoxin 2 (Trx2), a small protein with redox-regulating function, shows cytoprotective roles against oxidative stress. Here, we had focused on the role of Trx2 in ischemic preconditioning (IPC)-mediated neuroprotection against oxidative stress followed by a subsequent lethal transient cerebral ischemia. Animals used in this study were randomly assigned to six groups; sham-operated group, ischemia-operated group, IPC plus (+) sham-operated group, IPC + ischemia-operated group, IPC + auranofin (a TrxR2 inhibitor) + sham-operated group and IPC + auranofin + ischemia-operated group. IPC was subjected to a 2 minutes of sublethal transient ischemia 1 day prior to a 5 minutes of lethal transient ischemia. A significant loss of neurons was found in the stratum pyramidale (SP) of the hippocampal CA1 region (CA1) in the ischemia-operated-group 5 days after ischemia-reperfusion; in the IPC + ischemia-operated-group, pyramidal neurons in the SP were well protected. In the IPC + ischemia-operated-group, Trx2 and TrxR2 immunoreactivities in the SP and its protein level in the CA1 were not significantly changed compared with those in the sham-operated-group after ischemia-reperfusion. In addition, superoxide dismutase 2 (SOD2) expression, superoxide anion radical ( O2-) production, denatured cytochrome c expression and TUNEL-positive cells in the IPC + ischemia-operated-group were similar to those in the sham-operated-group. Conversely, the treatment of auranofin to the IPC + ischemia-operated-group significantly increased cell damage/death and abolished the IPC-induced effect on Trx2 and TrxR2 expressions. Furthermore, the inhibition of Trx2R nearly cancelled the beneficial effects of IPC on SOD2 expression, O2- production, denatured cytochrome c expression and TUNEL-positive cells. In brief, this study shows that IPC conferred neuroprotection against ischemic injury by maintaining Trx2 and suggests that the maintenance or enhancement of Trx2 expression by IPC may be a legitimate strategy for therapeutic intervention of cerebral ischemia., (© 2016 International Society of Neuropathology.)

We explored the role of mTOR/autophagy pathway in the aggravation of cerebral ischemia-reperfusion nerve injury caused by intermittent hypoxia. Eighty male wistar rats were divided into four groups by the random number method: sham operation group (SO group, n=20), cerebral ischemia-reperfusion group (I/R group, n=20), intermittent hypoxia and cerebral ischemia-reperfusion group (IH+I/R group, n=20), intermittent hypoxia and cerebral ischemia-reperfusion group plus mTOR inhibitor group (inhibitor group, n=20).The results showed that compared with the SO group, HE staining showed structural damage of neurons at each time point, the immunohistochemical assay showed an increasing number of mTOR and beclin1 immune-positive cells (P<0.05) and RT-PCR showed enhanced expression of mTOR and beclin1 protein in the I/R group (P<0.05). Compared with the I/R group, HE staining showed exacerbating structural damage of neurons at each time point, the immunohistochemical assay showed an increasing number of mTOR and beclin1 immune-positive cells (P<0.05) and RT-PCR showed enhanced expression of mTOR and beclin1 protein in the IH+I/R group (P<0.05). Compared with the IH+I/R group, HE staining showed remissive structural damage of neurons at each time point, the immunohistochemical assay showed a decreasing number of mTOR immune-positive cells and a rising number of beclin1immune-positive cells (P<0.05) and RT-PCR showed weakened expression of mTOR protein and enhanced expression of beclin1 protein in the inhibitor group (P<0.05). Thence, the present study indicated that intermittent hypoxia preconditioning can aggravate the nerve injury of the global cerebral ischemia-reperfusion model, and the mechanism is associated with the activation of mTOR/autophagy pathway.

As remote limb preconditioning (RPC) ameliorates brain damage after ischemic cerebral stroke (ICS), the purpose of the present study was to explore the molecular mechanisms in the course of RPC. Results of TUNEL staining and cleaved caspase-3 expression showed that ischemia-induced neuronal apoptosis was inhibited by RPC. The expression changes in cleaved caspase-8, cFLIP, Bid itself, and its truncated form represented that RPC suppressed the activation of extrinsic apoptotic pathway during ICS. Then, the level of cytoplasmic cytochrome c was also decreased by RPC. In addition, RPC might partially suppress TNF-related apoptosis-inducing ligand (TRAIL)-induced extrinsic apoptosis through downregulation of TRAIL death receptors and upregulation of TRAIL decoy receptors. As a counterproof, immunoneutralization of TRAIL in dMCAO rats resulted in significant restraint of tissue damage and in a marked functional recovery. Our data complemented the knowledge of RPC neuroprotective mechanism and provided new evidence for its clinical application.