Alpha-lipoic acid enhances ischemic postconditioning-mediated improvement of myocardial infarction and apoptosis in diabetic rats with ischemia/reperfusion injury.

This work evaluated the combined effects of alpha-lipoic acid (ALA) and ischemic postconditioning (Post) on myocardial infarction and cell death in rats with chronic type-II diabetes following ischemia/reperfusion injury. The rats received a high-fat diet and were given one intraperitoneal injection of 35 mg/kg streptozotocin to induce chronic diabetes. They were then pretreated with ALA (100 mg/kg/day, orally) for 5 weeks before undergoing ischemia/reperfusion (I/R) insult. The hearts experienced 35 min regional ischemia through ligating the left anterior descending coronary artery, followed by 60 min reperfusion. The Post protocol involved 6 cycles of a 10/10 s algorithm, applied during the early stage of reperfusion. The use of Post alone did not significantly alter lactate dehydrogenase and infarct size levels, while ALA showed positive effects. Similar findings were observed for apoptotic changes with single treatments. However, the concurrent administration of ALA and Post significantly reduced the protein expressions of Bax, Bax/Bcl2, and cleaved caspase-3 while increasing Bcl2 expression. Additionally, the histopathological findings of the combined therapy were superior to those of single treatments. The concomitant use of ALA and Post effectively inhibited apoptosis, leading to cardiac recovery after I/R injury in diabetic conditions. This strategy could improve outcomes for preserving diabetic hearts following I/R insults.

Effect of remote limb ischemic post-conditioning on the expression of miR-21-5p/PirB in the brain of rats with focal cerebral ischemia.

Cerebral ischemia causes great damage to ischemic brain regions. Remote limb post-conditioning (RLIPostC) has neuroprotective effects on cerebral ischemia. Paired immunoglobulin receptor B (PirB) has been confirmed to affect the cerebral ischemia injury. Therefore, this study investigated the mechanism of RLIPostC on cerebral ischemia injury. The middle cerebral artery occlusion (MCAO) model was established in rats, followed by RLIPostC (by blocking the blood flow in the distal limb for 10 min after MCAO, and restored for 10 min, three cycles), or in vivo lentivirus infection of miR-21-5p agomir, sh-PirB and oe-PirB. The locomotor abilities were assessed using the foot fault test and balance beam walking test at 48 h, 7 and 21 days after operation, and neurological function was measured on day 21. The cerebral infarct area, Nissl bodies and the pathology of brain tissues were examined using histological staining, followed by the assessment of miR-21-5p and PirB levels and the binding relationship between miR-21-5p and PirB. miR-21-5p was poorly expressed in MCAO rats. Both RLIPostC and overexpression of miR-21-5p reduced motor dysfunction, neurological function score, cerebral infarction area and brain edema; increased Nissl bodies in MCAO rats and thus alleviated the brain damage caused by cerebral ischemia. RLIPostC promoted miR-21-5p expression. PirB was highly expressed in cerebral ischemia, and miR-21-5p targeted PirB. Overall, RLIPostC promotes miR-21-5p expression and then inhibits PirB to alleviate cerebral ischemia injury, which provides theoretical basis for basic research on motor dysfunction and neuron injury after cerebral ischemia and the development of neuroscience.

Network-Assisted Systems Biology Analysis of the Mitochondrial Proteome in a Pre-Clinical Model of Ischemia, Revascularization and Post-Conditioning.

Infarct size is the major risk predictor for developing heart failure after an acute myocardial infarction (AMI). The discovery of the conditioning phenomena (i.e., repetitive brief cycles of ischemia applied either before or after a prolonged ischemic insult) has highlighted the existence of endogenous protective mechanisms of the heart potentially limiting infarct size after revascularization. However, most cardioprotective strategies, aiming at infarct size reduction, have failed in clinical studies. Thus, cardioprotection is an unmet clinical need. In the present study, we took a network-assisted systems biology approach to explore the mitochondrial proteomic signature of the myocardium after ischemia, ischemia with direct revascularization, and ischemia with re-establishment of blood flow by post-conditioning in a swine model of AMI. Furthermore, network extension with the ENCODE project human regulatory data allowed the prediction of potential transcription factors at play in the response to post-conditioning of the myocardium. Collectively, our results identify cardiac metabolism as a driver of cardioprotection, highlighting a dual role for post-conditioning promoting metabolic reprogramming of the myocardium, and a protective response mediated by VDAC2 and DJ-1 in the mitochondria.

Cardioprotection by post-conditioning with exogenous triiodothyronine in isolated perfused rat hearts and isolated adult rat cardiomyocytes.

Ischemic post-conditioning (iPoCo) by coronary re-occlusion/reperfusion during immediate reperfusion after prolonged myocardial ischemia reduces infarct size. Mechanical manipulation of culprit lesions, however, carries the risk of coronary microembolization which may obscure iPoCo's cardioprotection. Pharmacological post-conditioning with exogenous triiodothyronine (T3) could serve as an alternative conditioning strategy. Similar to iPoCo, T3 may activate cardioprotective prosurvival pathways. We aimed to study T3's impact on infarct size and its underlying signal transduction. Hearts were isolated from male Lewis rats (200-380 g), buffer-perfused and subjected to 30 min/120 min global zero-flow ischemia/reperfusion (I/R). In additional hearts, either iPoCo (2 × 30 s/30 s I/R) was performed or T3 (100-500 µg/L) infused at reperfusion. Infarct size was demarcated with triphenyl tetrazolium chloride staining and calculated as percent of ventricular mass. Infarct size was reduced with iPoCo to 16 ± 7% vs. 36 ± 4% with I/R only. The maximum infarct size reduction was observed with 300 µg/L T3 (14 ± 2%). T3 increased the phosphorylation of protein kinase B and mitogen extracellular-regulated-kinase 1/2, both key enzymes of the reperfusion injury salvage kinase (RISK) pathway. Pharmacological RISK blockade (RISK-BL) during reperfusion abrogated T3's cardioprotection (35 ± 10%). Adult ventricular cardiomyocytes were isolated from buffer-perfused rat hearts and exposed to 30 min/5 min hypoxia/reoxygenation (H/R); reoxygenation was initiated without or with T3, respectively, and without or with RISK-BL, respectively. Maximal preservation of viability was observed with 500 µg/L T3 after H/R (27 ± 4% of all cells vs. 5 ± 3% in time-matched controls). Again, RISK-BL abrogated protection (11 ± 3%). Mitochondria were isolated at early reperfusion from buffer-perfused rat hearts without or with iPoCo or 300 µg/L T3, respectively, at reperfusion. T3 improved mitochondrial function (i.e.: increased respiration, adenosine triphosphate production, calcium retention capacity, and decreased reactive oxygen species formation) to a similar extent as iPoCo. T3 at reperfusion reduces infarct size by activation of the RISK pathway. T3's protection is a cardiomyocyte phenomenon and targets mitochondria.

Sevoflurane Post-Conditioning Ameliorates Neuronal Deficits and Axon Demyelination After Neonatal Hypoxic Ischemic Brain Injury: Role of Microglia/Macrophage.

Microglia/macrophages have been identified to be highly polarized after ischemia. Interestingly, the polarization of these microglia/macrophages varies immensely under differing disease conditions. Post-conditioning using sevoflurane, a volatile anesthetic, could provide long-term neuroprotection to neonatal rats after hypoxic-ischemic brain injury (HIBI). Thus, the current study aimed at investigating the effects of sevoflurane post-conditioning (SPC) on microglia/macrophage polarization after HIBI induction in neonatal rats. Additionally, we aimed at identifying the underpinning mechanisms specifically related to autophagy and lysosomal protease enzyme, cathepsin B. To develop a HIBI model, 7-day-old Sprague-Dawley rats underwent left common carotid artery ligation followed by 2 h of hypoxia. The role of microglia/macrophages in the neuroprotection conferred by SPC was examined by left-side intra-cerebroventricular injection with adenovirus vector carrying catB-GFP or rapamycin. The number of interleukin (IL)-1ß+ cells, cathepsin B+ cells, light chain 3B positive (LC3B+) cells among ionized calcium binding adaptor molecule 1(Iba1+)cells to investigate microglia polarization, neuronal apoptosis to assess neuronal death in the acute phase were tested at 24 h after HIBI. Behavioral tests including suspension test, Morris water maze tests were performed to investigate the long-term effects of SPC, at 21 to 34 days post HIBI. Nissl staining and myelin basic protein (MBP) immunostaining to assess the long-term neuronal and myelin damage were performed at 34 days after HIBI. Based on the obtained results post HIBI, we observed the cells that were positive for IL-1ß, cathepsin B, and LC3B among Iba1 positive cell population in the hippocampus were significantly decreased after SPC treatment. SPC significantly attenuated the HIBI-induced increase in neuronal apoptosis, improved long-term cognitive function, and attenuated HI-induced decrease of Nissl-positive cells and MBP expression. However, these trends were reversed by injection of adenovirus vector carrying catB-GFP and rapamycin. SPC attenuated microglia polarization towards neurotoxic phenotypes, alleviates neuronal death and axon demyelination after HIBI in neonatal rats by regulating microglia autophagy and cathepsin B expression, and therefore provided long-term cognitive, learning and memory protection.

Dexmedetomidine post-conditioning attenuates cerebral ischemia following asphyxia cardiac arrest through down-regulation of apoptosis and neuroinflammation in rats.

BACKGROUND: Neuroprotection strategies after cardiac arrest (CA)/cardiopulmonary resuscitation (CPR) remain key areas of basic and clinical research. This study was designed to investigate the neuroprotective effects of dexmedetomidine following resuscitation and potential mechanisms. METHODS: Anesthetized rats underwent 6-min asphyxia-based cardiac arrest and resuscitation, after which the experimental group received a single intravenous dose of dexmedetomidine (25 µg/kg). Neurological outcomes and ataxia were assessed after the return of spontaneous circulation. The serum levels and brain expression of inflammation markers was examined, and apoptotic cells were quantified by TUNEL staining. RESULTS: Neuroprotection was enhanced by dexmedetomidine post-conditioning after the return of spontaneous circulation. This enhancement was characterized by the promotion of neurological function scores and coordination. In addition, dexmedetomidine post-conditioning attenuated the serum levels of the pro-inflammatory cytokine tumor necrosis factor (TNF)-α at 2 h, as well as interleukin IL-1ß at 2, 24, and 48 h. TUNEL staining showed that the number of apoptotic cells in the dexmedetomidine post-conditioning group was significantly reduced compared with the control group. Further western blot analysis indicated that dexmedetomidine markedly reduced the levels of caspase-3 and nuclear factor-kappa B (NF-κB) in the brain. CONCLUSIONS: Dexmedetomidine post-conditioning had a neuroprotective effect against cerebral injury following asphyxia-induced cardiac arrest. The mechanism was associated with the downregulation of apoptosis and neuroinflammation.

Sphingosine 1-phosphate signaling in ischemia and reperfusion injury.

Ischemia and reperfusion injury is a complex hemodynamic pathological phenomenon that engages the metabolic to inflammatory machinery in development of disease conditions like heart failure, stroke and acute kidney failure. Target specific therapeutic approaches for ischemia reperfusion injury remains critical despite the extensive studies contributing to the understanding of its pathogenesis. Ischemic or pharmacological conditionings have been long established manipulations to harness the endogenous protective mechanisms against ischemia reperfusion injury that fostered the development of potential therapeutic targets such as sphingolipids signaling. Sphingosine 1-phosphate has been emerged as a crucial metabolite of sphingolipids to regulate the cell survival, vascular integrity and inflammatory cascades in ischemia reperfusion injury. Sphingosine 1-phosphate signaling process has been implicated to downgrade the mitochondrial dysfunction, apoptotic assembly along with upregulation of RISK and SAFE pro-survival pathways. It also regulates the endothelial dysfunction and immune cells behavior to control the vascular permeability and immune cells infiltration at ischemia reperfusion injury site. Targeting the signaling of this single moiety holds the vast potential to extensively influence the detrimental signaling of ischemia reperfusion injury. This review highlights the role and significance of S1P signaling that can be therapeutically exploit to treat ischemia reperfusion injury mediated pathological conditions in different organs.

rAAV9-Mediated MEK1 Gene Expression Restores Post-conditioning Protection Against Ischemia Injury in Hypertrophic Myocardium.

PURPOSE: We investigated whether increased expression of activated mitogen-activated protein kinase (MAPK) kinases 1 (MEK1) restores ischemic post-conditioning (IPostC) protection in hypertrophic myocardium following ischemia/reperfusion (I/R) injury. METHODS: C57Bl/6 mice received recombinant adeno-associated virus type 9 (rAAV9)-mediated activated MEK1 gene delivery systemically, then following the induction of cardiac hypertrophy via transverse aortic constriction for 4 weeks. In a Langendorff model, hypertrophic hearts were subjected to 40 min/60 min I/R or with IPostC intervention consisting of 6 cycles of 10 s reperfusion and 10 s no-flow before a 60-min reperfusion. Hemodynamics, infarct size (IS), myocyte apoptosis and changes in expression of reperfusion injury salvage kinase (RISK) pathway were examined. RESULTS: rAAV9-MEK1 gene delivery led to a 4.3-fold and 2.7-fold increase in MEK1 mRNA and protein expression in the heart versus their control values. I/R resulted in a larger IS in hypertrophic than in non-hypertrophic hearts (52.3 ± 4.7% vs. 40.0 ± 2.5%, P < 0.05). IPostC mediated IS reduction in non-hypertrophic hearts (27.6 ± 2.6%, P < 0.05), while it had no significant effect in hypertrophic hearts (46.5 ± 3.1%, P=NS) compared with the IS in non-hypertrophic or hypertrophic hearts subjected to I/R injury only, respectively. Hemodynamic decline induced by I/R was preserved by IPostC in non-hypertrophic hearts but not in hypertrophic hearts. rAAV9-MEK1 gene delivery restored IPostC protection in hypertrophic hearts evidenced by reduced IS (32.0 ± 2.8% vs. 46.5 ± 3.1%) and cardiac cell apoptosis and largely preserved hemodynamic parameters. These protective effects were associated with significantly increased phosphorylation of ERK1/2 and ribosomal protein S6 kinases (p70S6K), but it had no influence on Akt and glycogen synthase kinase-3ß. CONCLUSION: These results demonstrated that rAAV9-mediated activated MEK1 expression restores IPostC protection in the hypertrophic heart against I/R injury through the activation of ERK pathway.

Definition of hidden drug cardiotoxicity: paradigm change in cardiac safety testing and its clinical implications.

Unexpected cardiac adverse effects are the leading causes of discontinuation of clinical trials and withdrawal of drugs from the market. Since the original observations in the mid-90s, it has been well established that cardiovascular risk factors and comorbidities (such as ageing, hyperlipidaemia, and diabetes) and their medications (e.g. nitrate tolerance, adenosine triphosphate-dependent potassium inhibitor antidiabetic drugs, statins, etc.) may interfere with cardiac ischaemic tolerance and endogenous cardioprotective signalling pathways. Indeed drugs may exert unwanted effects on the diseased and treated heart that is hidden in the healthy myocardium. Hidden cardiotoxic effects may be due to (i) drug-induced enhancement of deleterious signalling due to ischaemia/reperfusion injury and/or the presence of risk factors and/or (ii) inhibition of cardioprotective survival signalling pathways, both of which may lead to ischaemia-related cell death and/or pro-arrhythmic effects. This led to a novel concept of 'hidden cardiotoxicity', defined as cardiotoxity of a drug that manifests only in the diseased heart with e.g. ischaemia/reperfusion injury and/or in the presence of its major comorbidities. Little is known on the mechanism of hidden cardiotoxocity, moreover, hidden cardiotoxicity cannot be revealed by the routinely used non-clinical cardiac safety testing methods on healthy animals or tissues. Therefore, here, we emphasize the need for development of novel cardiac safety testing platform involving combined experimental models of cardiac diseases (especially myocardial ischaemia/reperfusion and ischaemic conditioning) in the presence and absence of major cardiovascular comorbidities and/or cotreatments.

Remote Ischemic Post-Conditioning may Improve Post-Stroke Cognitive Impairment: A Pilot Single Center Randomized Controlled Trial.

BACKGROUND AND PURPOSE: We aimed to demonstrate the tolerability and feasibility and the effect of remote ischemic post-conditioning on cognitive functioning in patients with post-stroke cognitive impairment. METHODS: This was a single-center, randomized, outcome-blinded, placebo-controlled trial, randomized 1:1 to receive 4 cycles of remote ischemic post-conditioning or a sham procedure for 7 days. The primary outcome measure was tolerability and feasibility of remote ischemic post-conditioning. Secondary outcomes to measure the neurological function with national institute of health stroke scale and the cognitive impairment with Montreal Cognitive Assessment scale and Alzheimer's disease assessment scale-cognitive (at baseline, 90 days, 180 days). RESULTS: 48 patients (24 RIPC and 24 Control) were recruited. remote ischemic post-conditioning was well tolerated with 90 out of 96 cycles completed in full. 4 patients experienced vascular events in the control group: 3 cerebrovascular and 1 cardiovascular event versus only 2 cerebrovascular events in the RIPC group. We showed the similar result in the neurological function with national institute of health stroke scale score with no statistically significant differences between RIPC and control group at baseline (P = 0.796) and 90 days (P = 0.401) and 180 days (P = 0.695). But compare with baseline, it was significantly difference in the control and RIPC group at 90 days (P < 0.05) and 180 days (P < 0.05). The comparison of Montreal Cognitive Assessment scale between two groups both showed that P > 0.05 at baseline which was no statistical difference, but P < 0.05 at 90 days and 180 days which were significant statistical difference. The comparison of Alzheimer's disease assessment scale-cognitive between two groups showed that P > 0.05 at baseline (P = 0.955) and 90 days (P = 0.138) was no statistical difference, but P = 0.005<0.05 at 180 days was significant statistical difference. CONCLUSIONS: The remote ischemic post-conditioning for post-stroke cognitive impairment was well tolerated, safe and feasible. The remote ischemic post-conditioning may improve neurological and cognitive outcomes in patients with post-stroke cognitive impairment. A larger trial is warranted. (Clinical Trial Registration-URL: http://www.clinicaltrials.gov. Unique identifier: ChiCTR1800015231.).

Repeated cell transplantation and adjunct renal denervation in ischemic heart failure: exploring modalities for improving cell therapy efficacy.

Enthusiasm for cell therapy for myocardial injury has waned due to equivocal benefits in clinical trials. In an attempt to improve efficacy, we investigated repeated cell therapy and adjunct renal denervation (RDN) as strategies for augmenting cardioprotection with cardiosphere-derived cells (CDCs). We hypothesized that combining CDC post-conditioning with repeated CDC doses or delayed RDN therapy would result in superior function and remodeling. Wistar-Kyoto (WKY) rats or spontaneously hypertensive rats (SHR) were subjected to 45 min of coronary artery ligation followed by reperfusion for 12-14 weeks. In the first study arm, SHR were treated with CDCs (0.5 × 106 i.c.) or PBS 20 min following reperfusion, or additionally treated with CDCs (1.0 × 106 i.v.) at 2, 4, and 8 weeks. In the second arm, at 4 weeks following myocardial infarction (MI), SHR received CDCs (0.5 × 106 i.c.) or CDCs + RDN. In the third arm, WKY rats were treated with i.c. CDCs administered 20 min following reperfusion and RDN or a sham at 4 weeks. Early i.c. + multiple i.v. dosing, but not single i.c. dosing, of CDCs improved long-term left ventricular (LV) function, but not remodeling. Delayed CDC + RDN therapy was not superior to single-dose delayed CDC therapy. Early CDC + delayed RDN therapy improved LV ejection fraction and remodeling compared to both CDCs alone and RDN alone. Given that both RDN and CDCs are currently in the clinic, our findings motivate further translation targeting a heart failure indication with combined approaches.

The role of PI3K-mediated AMPA receptor changes in post-conditioning of propofol in brain protection.

BACKGROUND: We aimed to study the role of amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor (AMPAR) glutamate receptor 2 (GluR2) subunit trafficking, and activity changes in short-term neuroprotection provided by propofol post-conditioning. We also aimed to determine the role of phosphoinositide-3-kinase (PI3K) in the regulation of these processes. METHODS: Rats underwent 1 h of focal cerebral ischemia followed by 23 h of reperfusion were randomly divided into 6 groups (n = 36 per group): sham- operation (S), ischemia-reperfusion (IR), propofol (P group, propofol 20 mg/kg/h at the onset of reperfusion for 2 h after 60 min of occlusion), and LY294002 (PI3K non-selective antagonist) + sham (L + S, LY294002 of 1.5 mg/kg was infused 30 min before sham operation), LY294002+ ischemia-reperfusion (L + IR, LY294002 of 1.5 mg/kg was infused 30 min before middle cerebral artery occlusion), LY294002 + IR + propofol (L + P, LY294002 of 1.5 mg/kg was infused 30 min before middle cerebral artery occlusion and propofol 20 mg/kg/h at the onset of reperfusion for 2 h after 60 min of occlusion). RESULTS: Compared with group IR, rats in group P had significant lower neurologic defect scores and infarct volume. Additionally, consistent with enhanced expression of PI3K-AMPAR GluR2 subunit complex substances in ipsilateral hippocampus, GluR2 subunits showed increased levels in both the plasma and postsynaptic membranes of neurons, while pGluR2 expression was reduced in group P. Furthermore, LY294002, the PI3K non-selective antagonist, blocked those effects. CONCLUSION: These observations demonstrated that propofol post-conditioning revealed acute neuroprotective role against transient MCAO in rats. The short-term neuroprotective effect was contributed by enhanced GluR2 subunits trafficking to membrane and postsynaptic membranes of neurons, as well as down-regulated the expression of pGluR2 in damaged hippocampus. Finally, the above-mentioned protective mechanism might be contributed by increased combination of PI3K to AMPAR GluR2 subunit, thus maintained the expression and activation of AMPAR GluR2 in the ipsilateral hippocampus.

Neuroprotective Mechanism of Hypoxic Post-conditioning Involves HIF1-Associated Regulation of the Pentose Phosphate Pathway in Rat Brain.

Post-conditioning is exposure of an injured organism to the same harmful factors but of milder intensity which mobilizes endogenous protective mechanisms. Recently, we have developed a novel noninvasive post-conditioning (PostC) protocol involving three sequential episodes of mild hypobaric hypoxia which exerts pronounced neuroprotective action. In particular, it prevents development of pathological cascades caused by severe hypobaric hypoxia (SH) such as cellular loss, lipid peroxidation, abnormal neuroendocrine responses and behavioural deficit in experimental animals. Development of these post-hypoxic pathological effects has been associated with the delayed reduction of hypoxia-inducible factor 1 (HIF1) regulatory α-subunit levels in rat hippocampus, whereas PostC up-regulated it. The present study has been aimed at experimental examination of the hypothesis that intrinsic mechanisms underlying the neuroprotective and antioxidant effects of PostC involves HIF1-dependent stimulation of the pentose phosphate pathway (PPP). We have observed that SH leads to a decrease of glucose-6-phosphate dehydrogenase (G6PD) activity in the hippocampus and neocortex of rats as well as to a reduction in NADPH and total glutathione levels. This depletion of the antioxidant defense system together with excessive lipid peroxidation during the reoxygenation phase resulted in increased oxidative stress and massive cellular death observed after SH. In contrast, PostC led to normalization of G6PD activity, stabilization of the NADPH and total glutathione levels and thereby resulted in recovery of the cellular redox state and prevention of neuronal death. Our data suggest that stabilization of the antioxidant system via HIF1-associated PPP regulation represents an important neuroprotective mechanism enabled by PostC.

Age-related difference in protective effect of early post-conditioning on ischemic brain injury: possible involvement of MAP-2/Synaptophysin role.

Brain Ischemia/Reperfusion (I/R) injury leads to the failure of the microtubules function and neuronal death. Ischemic post-conditioning is defined as a series of rapid alternating interruptions of blood flow in the first seconds of reperfusion. In the present study, the caspase-3, Microtubule-Associated Protein-2 (MAP-2), Protein Kinase C α (PKCα), c-fos, and synaptophysin were evaluated in the hippocampus of focal I/R post-conditioning model in a time -dependent study in aged and young rats. Adult and aged rats were subjected to right MCAO for 30 min and post-conditioned (10 s) for 3 cycles. Sensory-motor tests were performed, and locomotion and anxiety-like behavior were evaluated. Molecular tests were done by detection kit, RT-PCR, and Western blotting techniques. Ninety-six hours after I/R post-conditioning, neurological signs, locomotion, anxiety-like behavior, and ischemic area were improved in young rats compared to 6 h after I/R post-conditioning (P < 0.001). Caspase-3 activity declined in the hippocampus and cortex of I/R post-conditioned young rats in 96 h after I/R post-conditioning compared with 6 h after I/R post-conditioning (P < 0.001). Also, MAP-2 mRNA, MAP-2 protein level, PKCα, c-fos and synaptophysin protein levels were enhanced during post-conditioning in young rats in 96 h after I/R post-conditioning compared with 6 h after induction of I/R post-conditioning. The results of the present study suggested that, early post-conditioning might be considered as a candidate for therapeutic methods against I/R in the adult animals not aged rats. Moreover, inhibition of cell death in post-conditioned ischemic rats was found to be regulated by some neuroprotective molecules as well as MAP-2 and c-fos in young rats. Graphical abstract Graphical abstract representing the post-conditioning (PC) treatment timeline in adult and old rats.

Propofol post-conditioning after temporary clipping reverses oxidative stress in aneurysm surgery.

PURPOSE: Animal studies have demonstrated that propofol post-conditioning produces long-term neuroprotection in focal cerebral ischemia-reperfusion injury. However, whether propofol post-conditioning provides neuroprotection in human beings has never been explored. The aim of this study was to evaluate the role of propofol post-conditioning on oxidative stress and post-operative cognitive function following aneurysm clipping. MATERIALS AND METHODS: Sixty patients undergoing intracranial aneurysm clipping were randomized into a propofol post-conditioning group or a sevoflurane group. Sevoflurane (0.5-2%) was used for maintenance anesthesia in both groups. In the propofol post-conditioning group, the inhaled concentration of sevoflurane was reduced after temporary clip removal to keep the bispectral index (BIS) value between 40 and 60, and propofol (Cp 1.2 µg/mL) was subsequently started. Blood samples were drawn at six time points: before induction, immediately after clip removal, at the end of the operation, 24-h post-surgery, 3 days post-surgery, and 7 days post-surgery. Oxidative stress and cognitive function were measured. RESULTS: Between the conclusion of the operation to 7 days after surgery, propofol post-conditioning decreased the serum concentration of •OH and 8-isoprostane and increased γ-tocopherol and SOD. Reduced micronuclei and nucleoplasmic bridges were observed in the propofol post-conditioning group. Mini-Mental State Examination (MMSE) and Montreal Cognitive Assessment (MoCA) scores were improved by propofol post-conditioning compared to the group that received no propofol. CONCLUSIONS: Together, our data suggest that propofol post-conditioning (Cp 1.2 µg/mL) may protect the brain from oxidative stress injury up to 7 days post-surgery after temporary parent artery clipping. Furthermore, the neuroprotection induced by propofol post-conditioning may contribute to improvement in cognitive function.

Systemic redistribution of the intramyocardially injected mesenchymal stem cells by repeated remote ischaemic post-conditioning.

We investigated the effect of repeated remote ischaemic post-conditioning (RIPoC) on the organ distribution of the intramyocardially injected MSCs in rat myocardial ischemia model. Myocardial ischemia of adult female Sprague-Dawley rats was induced by 30-min. obstruction of the left anterior descending coronary artery. Repeated RIPoC was induced after ischemia with three cycles of 5-min. occlusion and reperfusion of the limb with the frequency of half a day, 1 or 2 days, respectively. Compared with that by single RIPoC, repeated RIPoC transiently reduced oxidative stress, lipid peroxidation and inflammation in ischaemic myocardium; the gene expression of stromal cell-derived factor-1 alpha (SDF-1α) was consistently induced by repeated RIPoC procedures. A total of 4 × 106 male bone marrow-derived MSCs were intramyocardially injected into ischaemic myocardium at 1 week after reperfusion. Three weeks later, immunohistological examination and quantitative reverse transcriptase polymerase chain reaction demonstrated that repeated RIPoC significantly increased MSCs retention in myocardium and decreased MSCs distribution over the lungs, spleen and liver; echocardiography assessment revealed that further cardiac function enhancement imposed by repeated RIPoC procedure. Furthermore, blockade with the anti-CXCR4 antibody before cell transplantation markedly attenuated the benefits of therapeutic efficacy and cardiac function. Repeated RIPoC enhanced MSCs engraftment in ischaemic myocardium and reduced the distribution of MSCs over peripheral organs in a frequency-effect way, but it reached a ceiling of maximal effect when applied with every 1 day. The SDF1α-CXCR4 interaction played a major role in MSCs systemic redistribution induced by repeated RIPoC.

Neuroprotection by Propofol Post-Conditioning: Focus on PKMζ/KCC2 Pathway Activity.

Critical and major operations are often accompanied by brain ischemic complications. Previous studies found that propofol post-conditioning provided neuroprotective functions through upregulating the expression of potassium chloride cotransporter 2 (KCC2) in gamma-aminobutyric acid (GABA) interneurons. Membrane expression and phosphorylation represents KCC2 activity, which were modulated by a protein kinase C (PKC)-dependent mechanism. However, the role of propofol in increasing KCC2 phosphorylation and the involvement of protein kinase Mζ (PKMζ), a major subtype of PKC, in the KCC2 pathway remained unclear. In this study, we established middle cerebral artery occlusion model in rats to evaluate the long-term recovery of brain functions using behavioral experiments. KCC2 and PKMζ were assessed via western blot. We used the selective inhibitor, zeta inhibitory peptide (ZIP), to investigate the relationship between KCC2 and PKMζ. Intracellular chloride concentration in the hippocampal CA1 area was measured to determine KCC2 activity. We found that propofol, infused at a speed of 20 mg kg-1 h-1 for 2 h at the onset of reperfusion, improved neurological deficits and cognitive dysfunction following ischemia/reperfusion injury. PKMζ expression was significantly upregulated, which improved KCC2 membrane expression and phosphorylation in the ischemic hippocampal CA1 area, and these effects could last up to 28 days. But ZIP inhibited this process. Ultimately, we showed that propofol increased KCC2 phosphorylation and PKMζ was the upstream of KCC2. Propofol led to long-term recovery of brain functions by upregulating the activity of the PKMζ/KCC2 pathway.

Remote ischaemic conditioning for prevention of acute kidney injury after valvular heart surgery: a randomised controlled trial.

BACKGROUND: Repeated remote ischaemic conditioning (RIC) during weaning from cardiopulmonary bypass and in the early postoperative period may confer protection against acute kidney injury (AKI). We evaluated the effect of repeated RIC on the incidence of AKI in patients undergoing valvular heart surgery. METHODS: Patients were randomised into either the RIC (n=120) or control (n=124) group. A pneumatic tourniquet was placed on each patient's thigh. Upon removal of the aortic cross-clamp, three cycles of inflation for 5 min at 250 mm Hg (with 5 min intervals) were applied in the RIC group. Additionally, three cycles of RIC were repeated at postoperative 12 and 24 h. AKI was diagnosed based on the Kidney Disease: Improving Global Outcomes guideline. The incidences of renal replacement therapy, permanent stroke, sternal wound infection, newly developed atrial fibrillation, mechanical ventilation >24 h, and reoperation for bleeding during hospitalisation were recorded. RESULTS: The incidences of AKI were not significantly different between the control (19.4%) and RIC (15.8%) groups (a difference of 3.5 percentage points; 95% confidence interval: -6.8%-13.9%; P=0.470). Perioperative serum creatinine concentrations were similar in the control and RIC groups (P=0.494). Fluid balance, urine output, blood loss, transfusion, and vasopressor/inotropic requirements were not significantly different between the groups (all P>0.05). The occurrences of a composite of morbidity and mortality endpoints were not significantly different between the control (46.0%) and RIC (39.2%) groups (a difference of 6.8 percentage points; 95% confidence interval: -6.4%-20.0%; P=0.283). CONCLUSIONS: The results of our study do not support repeated RIC to decrease the incidence of AKI after valvular heart surgery. CLINICAL TRIAL REGISTRATION: NCT02720549.

Gene expression in retinal ischemic post-conditioning.

PURPOSE: The pathophysiology of retinal ischemia involves mechanisms including inflammation and apoptosis. Ischemic post-conditioning (Post-C), a brief non-lethal ischemia, induces a long-term ischemic tolerance, but the mechanisms of ischemic post-conditioning in the retina have only been described on a limited basis. Accordingly, we conducted this study to determine the molecular events in retinal ischemic post-conditioning and to identify targets for therapeutic strategies for retinal ischemia. METHODS: To determine global molecular events in ischemic post-conditioning, a comprehensive study of the transcriptome of whole retina was performed. We utilized RNA sequencing (RNA-Seq), a recently developed, deep sequencing technique enabling quantitative gene expression, with low background noise, dynamic detection range, and discovery of novel genes. Rat retina was subjected to ischemia in vivo by elevation of intraocular pressure above systolic blood pressure. At 24 h after ischemia, Post-C or sham Post-C was performed by another, briefer period of ischemia, and 24 h later, retinas were collected and RNA processed. RESULTS: There were 71 significantly affected pathways in post-conditioned/ischemic vs. normals and 43 in sham post conditioned/ischemic vs. normals. Of these, 28 were unique to Post-C and ischemia. Seven biological pathways relevant to ischemic injury, in Post-C as opposed to sham Post-C, were examined in detail. Apoptosis, p53, cell cycle, JAK-STAT, HIF-1, MAPK and PI3K-Akt pathways significantly differed in the number as well as degree of fold change in genes between conditions. CONCLUSION: Post-C is a complex molecular signaling process with a multitude of altered molecular pathways. We identified potential gene candidates in Post-C. Studying the impact of altering expression of these factors may yield insight into new methods for treating or preventing damage from retinal ischemic disorders.

RLIPostC protects against cerebral ischemia through improved synaptogenesis in rats.

OBJECTIVE: Remote limb ischemic post-conditioning (RLIPostC) has been shown to be neuroprotective in cerebral ischemia, whereas the effect of RLIPostC on synaptogenesis remains elusive. In the present study, we investigated the effects of RLIPostC on synaptogenesis in an experimental stroke rat model. METHODS: Sprague-Dawley rats were subjected to left middle cerebral artery occlusion (MCAO) and were randomly divided into a control group, an RLIPostC group and a sham group. The RLIPostC group received three cycles of RLIPostC treatment immediately after reperfusion (ten minutes ischemia and ten minutes reperfusion in bilateral femoral artery). The neurological function was assessed by neurological deficit scores and the foot fault test at days 7 and 14 after MCAO. At day 14 after MCAO, the infarct volume and oedema were determined by cresyl violet (CV) staining and by measuring brain water content, respectively. Synaptogenesis was evaluated by western blotting and immunofluorescence staining. RESULTS: Our results showed that RLIPostC treatment significantly promoted the recovery of behavioural function, reduced infarct volume and brain oedema, and increased the expressions of SYN1, PSD95 and GAP43. CONCLUSIONS: These results confirmed that RLIPostC treatment for cerebral ischemia was safe and effective. A possible molecular mechanism of the beneficial effects of RLIPostC treatment may be the promotion of synaptogenesis.