RESUMEN
Sevoflurane postconditioning has been shown to provide neuroprotection against cerebral hypoxia-ischemia injury, but the mechanisms remain elusive. Microtubule-associated protein 2 (MAP2) is implicated in early neuronal hypoxia-ischemia injury. This study aimed to investigate whether the neuroprotective effects of sevoflurane postconditioning are related to the Akt/GSK-3ß pathway and its downstream target MAP2 in zebrafish hypoxia/reoxygenation (H/R) model. Sevoflurane postconditioning or GSK-3ß inhibitor TDZD-8 were used to treat H/R zebrafish. The cerebral infarction, neuronal apoptosis, and mitochondrial changes were evaluated using TTC staining, TUNEL staining, and transmission electron microscopy, respectively. The distribution of MAP2 in the brain was determined by immunofluorescence imaging. The levels of Akt, p-Akt, GSK-3ß, p-GSK-3ß, and MAP2 proteins were evaluated by Western blotting. The neurobehavioral recovery of zebrafish was assessed based on optokinetic response behavior. Our results indicated that sevoflurane postconditioning and TDZD-8 significantly reduced the cerebral infarction area, suppressed cell apoptosis, and improved mitochondrial integrity in zebrafish subjected to H/R. Furthermore, sevoflurane postconditioning and TDZD-8 elevated the ratios of p-Akt/Akt and p-GSK-3ß/GSK-3ß. However, the neuroprotective effect of sevoflurane postconditioning was effectively abolished upon suppression of MAP2 expression. In conclusion, sevoflurane postconditioning ameliorated cerebral H/R injury and facilitated the restoration of neurobehavioral function through the activation of Akt/GSK-3ß pathway and promotion of MAP2 expression.
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Glucógeno Sintasa Quinasa 3 beta , Proteínas Asociadas a Microtúbulos , Fármacos Neuroprotectores , Proteínas Proto-Oncogénicas c-akt , Sevoflurano , Transducción de Señal , Pez Cebra , Animales , Sevoflurano/farmacología , Glucógeno Sintasa Quinasa 3 beta/metabolismo , Proteínas Proto-Oncogénicas c-akt/metabolismo , Fármacos Neuroprotectores/farmacología , Transducción de Señal/efectos de los fármacos , Proteínas Asociadas a Microtúbulos/metabolismo , Apoptosis/efectos de los fármacos , Daño por Reperfusión/tratamiento farmacológico , Daño por Reperfusión/metabolismo , Daño por Reperfusión/patología , Poscondicionamiento Isquémico/métodos , Hipoxia-Isquemia Encefálica/metabolismo , Hipoxia-Isquemia Encefálica/tratamiento farmacológico , Hipoxia-Isquemia Encefálica/patología , Proteínas de Pez Cebra/metabolismo , Modelos Animales de Enfermedad , Mitocondrias/efectos de los fármacos , Mitocondrias/metabolismo , MasculinoRESUMEN
Hypoxic-ischemic brain damage (HIBD) frequently induces cognitive impairments. Investigating the role of sevoflurane postconditioning (SPC) in HIBD, we conducted experiments involving HIBD modeling, SPC treatment, and interventions with the PERK inhibitor GSK2656157 or the PERK activator CCT020312, administered 30 min before modeling, followed by SPC treatment. Behavioral testing using the Morris water maze test and Neurological Deficiency Scale (NDS) was conducted. Additionally, Nissl staining assessed hippocampal CA1 area neuronal density, TUNEL staining evaluated hippocampal CA1 area neuronal apoptosis, and Western blot determined hippocampal CA1 area protein levels, including Bax, Bcl-2, p-PERK/PERK, p-eIF2/eIF2, ATF4, CHOP, GRP78, Bax, and Bcl-2 protein levels. Following SPC treatment, HIBD rats exhibited improved spatial learning and memory abilities, reduced neuronal apoptosis, increased neuronal density in the hippocampal CA1 area, elevated Bcl-2 protein level, decreased Bax protein levels, and decreased levels of endoplasmic reticulum stress pathway related proteins (p-PERK/PERK, p-eIF2/eIF2, ATF4, CHOP and GRP78). Pre-modeling treatment with the PERK inhibitor treatment improved outcomes in HIBD rats. However, pre-modeling treatment with the PERK activator CCT020312 counteracted the protective effects of SPC against HIBD in rats. In conclusion, SPC alleviates neuronal apoptosis in the hippocampus CA1 area of HIBD rats by inhibiting the endoplasmic reticulum stress pathway PERK/ATF4/CHOP, thereby mitigating HIBD in rats.
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Chaperón BiP del Retículo Endoplásmico , Estrés del Retículo Endoplásmico , Hipoxia-Isquemia Encefálica , Sevoflurano , Animales , Ratas , Apoptosis , Proteína X Asociada a bcl-2/metabolismo , Estrés del Retículo Endoplásmico/efectos de los fármacos , Factor 2 Eucariótico de Iniciación/metabolismo , Factor 2 Eucariótico de Iniciación/farmacología , Hipocampo/metabolismo , Hipoxia-Isquemia Encefálica/metabolismo , Proteínas Proto-Oncogénicas c-bcl-2/metabolismo , Ratas Sprague-Dawley , Sevoflurano/farmacologíaRESUMEN
To determine whether sevoflurane postconditioning protects against cerebral ischemia reperfusion (I/R) injury and its potential mechanism, we employed bioinformatic analysis, neurological assessments, and western blot analysis, as well as triphenyl tetrazolium chloride, hematoxylin and eosin, Nissl, and immunofluorescence staining. We identified 103 differentially expressed genes induced by cerebral I/R, including 75 upregulated genes and 28 downregulated genes enriched for certain biological processes (involving regulation of inflammatory responses, cellular responses to interleukin 1, and chemokine activity) and signaling pathways (such as transcriptional misregulation in cancer, interleukin-17 signaling, rheumatoid arthritis, MAPK signaling, and Toll-like receptor signaling). As a typical path in Toll-like receptor signaling pathway, in the current study, we investigated the protective effect of sevoflurane postconditioning in cerebral I/R rats and further explore the role of TLR4/MyD88/TRAF6 signaling pathway in it. The results showed cerebral I/R-induced neurological deficits were comparatively less severe following sevoflurane postconditioning. In addition, TLR4/MyD88/TRAF6 signaling pathway-related proteins and neuropathic damage were ameliorated in aged rats following sevoflurane postconditioning, while the TLR4 agonist lipopolysaccharide aggravated these changes. Together, these findings suggest that sevoflurane postconditioning ameliorates cerebral I/R injury by a mechanism involving inhibition of the TLR4/MyD88/TRAF6 signaling pathway to suppress neuroinflammatory responses.
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Isquemia Encefálica , Daño por Reperfusión , Ratas , Animales , Sevoflurano/farmacología , Factor 88 de Diferenciación Mieloide/metabolismo , Receptor Toll-Like 4/metabolismo , Factor 6 Asociado a Receptor de TNF/metabolismo , Ratas Sprague-Dawley , Transducción de Señal , Daño por Reperfusión/metabolismo , Isquemia Encefálica/tratamiento farmacológico , Isquemia Encefálica/metabolismoRESUMEN
BACKGROUND: Although extensive reports have demonstrated the neuroprotection of sevoflurane postconditioning in cases of focal and global cerebral ischemia/reperfusion, the underlying mechanisms are not completely elucidated. This study investigated whether this effect is related to endothelial nitric oxide synthase (eNOS) and mediated by the phosphoinositide-3-kinase pathway in a rat model of hemorrhagic shock and resuscitation. METHODS: Adult male Sprague Dawley rats were subjected to hemorrhagic shock for 60 minutes and then resuscitation for 30 minutes in experimental groups. Sevoflurane postconditioning was performed at the beginning of resuscitation to completion. At 24 hours after resuscitation, the brain infarct volume was evaluated by 2,3,5-triphenyltetrazolium chloride staining. The neuronal morphological changes and apoptosis were determined by hematoxylin and eosin staining and immunohistochemistry analysis, respectively. The activity of phosphorylated Akt and eNOS was evaluated by Western blot analysis. RESULTS: Brain injuries such as the cerebral infarct volume and pathological neuronal changes as well as cell apoptosis were observed in the hippocampus after hemorrhagic shock and resuscitation. Postconditioning with 2.4% sevoflurane significantly attenuated brain injuries. Wortmannin prevented the improvements of neuronal characteristics elicited by sevoflurane postconditioning as well as the hyperactivity of eNOS and phosphorylated Akt. CONCLUSIONS: Sevoflurane postconditioning could attenuate brain injury induced by hemorrhagic shock and resuscitation, and this neuroprotective effect may be partly by upregulation of eNOS through the phosphoinositide-3-kinase/Akt signaling pathway.
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Fármacos Neuroprotectores/administración & dosificación , Óxido Nítrico Sintasa de Tipo III/biosíntesis , Fosfatidilinositol 3-Quinasa/biosíntesis , Proteínas Proto-Oncogénicas c-akt/biosíntesis , Sevoflurano/administración & dosificación , Choque Hemorrágico/metabolismo , Anestésicos por Inhalación/administración & dosificación , Animales , Modelos Animales de Enfermedad , Poscondicionamiento Isquémico/tendencias , Masculino , Ratas , Ratas Sprague-Dawley , Resucitación/tendencias , Choque Hemorrágico/prevención & control , Choque Hemorrágico/terapia , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiologíaRESUMEN
Post-conditioning with sevoflurane, a volatile anesthetic, has been proved to be neuroprotective against hypoxic-ischemic brain injury (HIBI). Our previous research showed that autophagy is over-activated in a neonatal HIBI rat model, and inhibition of autophagy confers neuroprotection. There is increasing recognition that autophagy can be stimulated by activating endoplasmic reticulum (ER) stress. Herein, we purposed to explore: i) the association of ER stress with autophagy in the setting of neonatal HIBI; and ii) the possible roles of ER stress-triggered autophagy, as well as IRE1 signaling in the neuroprotection of sevoflurane post-conditioning against neonatal HIBI. Seven-day-old rats underwent ligation of the left common artery, and a subsequent 2 h hypoxia (8% O2/92% N2). The association of ER stress with autophagy was examined by ER stress inducer (tunicamycin), 4-PBA (ER stress inhibitor), or 3-MA (autophagy inhibitor). Rats in the sevoflurane post-conditioning groups were treated with 2.4% sevoflurane for 30 min after HIBI stimulation. The roles of ER stress-mediated autophagy, as well as the IRE1-JNK-beclin1 signaling cascade in the neuroprotection afforded by sevoflurane were explored by ER stress inducer (tunicamycin) and the IRE1 inhibitor (STF-083010). HIBI over-activated ER stress and autophagy in neonatal rats. HIBI-induced autophagy was significantly aggravated by tunicamycin but blocked by 4-PBA; however, HIBI-induced ER stress was not affected by 3-MA. Sevoflurane post-conditioning significantly alleviated ER stress, autophagy, cell apoptosis, and cognitive impairments, which were remarkably abolished by tunicamycin. Also, tunicamycin blocked sevoflurane-induced downregulation of IRE1-JNK-beclin1 signaling pathway. Whereas, IRE1 inhibitor could reverse the effects of tunicamycin. ER stress contributes to autophagy induced by HIBI. Furthermore, sevoflurane post-conditioning significantly protects against HIBI in neonatal rats by inhibiting ER stress-mediated autophagy via IRE1-JNK-beclin1 signaling cascade.
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Autofagia/fisiología , Estrés del Retículo Endoplásmico/fisiología , Hipoxia-Isquemia Encefálica/tratamiento farmacológico , Poscondicionamiento Isquémico/métodos , Proteínas de la Membrana/antagonistas & inhibidores , Proteínas Serina-Treonina Quinasas/antagonistas & inhibidores , Sevoflurano/administración & dosificación , Animales , Animales Recién Nacidos , Autofagia/efectos de los fármacos , Estrés del Retículo Endoplásmico/efectos de los fármacos , Hipoxia-Isquemia Encefálica/metabolismo , Aprendizaje por Laberinto/efectos de los fármacos , Aprendizaje por Laberinto/fisiología , Proteínas de la Membrana/metabolismo , Inhibidores de Agregación Plaquetaria/administración & dosificación , Proteínas Serina-Treonina Quinasas/metabolismo , Ratas , Ratas Sprague-Dawley , Transducción de Señal/efectos de los fármacos , Transducción de Señal/fisiologíaRESUMEN
OBJECTIVES: Myocardial ischemia-reperfusion injury is a phenomenon that promotes myocardial damage when the blood supply returns to the tissue after a period of ischemia. Anesthetic postconditioning involves myocardial protection against myocardial I/R injury. The effects of atorvastatin (ATV) on sevoflurane postconditioning against myocardial ischemia-reperfusion injury have not been thoroughly studied. The present study aimed to investigate if ATV interacts synergistically with sevoflurane postconditioning against myocardial infarction in rabbit hearts in vivo. METHODS: Twenty-eight male rabbits underwent 30 min of left anterior descending coronary artery occlusion that was followed by reperfusion for 180 min under ketamine/xylazine (K/X) anesthesia. Rabbits were randomly assigned to four groups that included Group K/X (under K/X anesthesia only), Group POST (sevoflurane exposure at initial reperfusion), Group ATV (ATV 5 mg/kg/day administered before ischemia), and Group ATV + POST (POST intervention with atorvastatin administered once daily for 3 days). At the end of reperfusion, the myocardial infarct size and the area at risk were both measured. RESULTS: The mean infarct sizes in the POST, ATV, and ATV + POST groups were significantly smaller compared to those in the K/X group. Furthermore, the mean infarct size in Group ATV + POST was significantly smaller than was that in Group POST and significantly smaller compared to that in Group ATV. CONCLUSION: The combination of sevoflurane postconditioning and pre-administration of ATV further reduced the myocardial infarction size compared to that observed with sevoflurane postconditioning alone or ATV alone. Our data suggest that sevoflurane postconditioning and ATV may function additively to enhance cardioprotection.
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Anestésicos por Inhalación , Éteres Metílicos , Animales , Atorvastatina/farmacología , Masculino , Conejos , Ratas , Ratas Sprague-Dawley , SevofluranoRESUMEN
Sevoflurane postconditioning (SPC) has been widely reported to attenuate brain injury after hypoxia-ischemia encephalopathy (HIE) by inhibiting neural necrosis and autophagy. Moreover, recent reports revealed that sevoflurane facilitated hippocampal reconstruction via regulating migration. Yet, it remains unclear whether the promotion of neural migration by SPC repairs the hippocampal injury after HIE. Here, we hypothesize that SPC exerts a neuroprotective effect by ameliorating neuronal migration disorder after HIE and regulating Reelin expression. Furthermore, the downstream Reelin/Dab1 pathway may be involved. The classical Rice-Vannucci model of hypoxia-ischemia was performed on postnatal day 7 rat pups, which was followed by SPC at 1 minimum alveolar concentration (MAC 2.5%) for 30 min. Piceatannol, causing Reelin aggregation in vivo, was used to detect whether Reelin/Dab1 was involved in the neuroprotection effect of SPC. Hippocampal-dependent learning ability tests were conducted to assess the long-term effects on locomotor activity and spatial learning ability. Our findings suggest that hypoxia-ischemia injury inhibited neurons migrated outward from the basal zone of dentate gyrus, disrupted cytoarchitecture of the dentate gyrus (DG), and led to long-term cognition deficits. However, SPC could relieve the restricted hippocampal neurons and repair the hippocampal-dependent memory function damaged after HIE by attenuating the overactivation of the Reelin/Dab1 pathway. These results demonstrate that SPC plays a pivotal role in ameliorating neuronal migration disorder and maintaining normal cytoarchitecture of the DG via inhibiting overactivated Reelin expression. This process may involve overactivated Reelin/Dab1 signaling pathway and spatial learning ability by regulating the Reelin expression which may associate with its neuroprotection.
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Proteínas Adaptadoras Transductoras de Señales/antagonistas & inhibidores , Cognición/efectos de los fármacos , Hipoxia-Isquemia Encefálica/tratamiento farmacológico , Malformaciones del Desarrollo Cortical del Grupo II/tratamiento farmacológico , Proteínas del Tejido Nervioso/antagonistas & inhibidores , Proteína Reelina/antagonistas & inhibidores , Sevoflurano/administración & dosificación , Proteínas Adaptadoras Transductoras de Señales/biosíntesis , Animales , Animales Recién Nacidos , Movimiento Celular/efectos de los fármacos , Movimiento Celular/fisiología , Cognición/fisiología , Hipoxia-Isquemia Encefálica/metabolismo , Hipoxia-Isquemia Encefálica/patología , Poscondicionamiento Isquémico/métodos , Masculino , Malformaciones del Desarrollo Cortical del Grupo II/metabolismo , Malformaciones del Desarrollo Cortical del Grupo II/patología , Proteínas del Tejido Nervioso/biosíntesis , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Neuronas/patología , Fármacos Neuroprotectores/administración & dosificación , Inhibidores de Agregación Plaquetaria/administración & dosificación , Ratas , Ratas Sprague-Dawley , Proteína Reelina/biosíntesis , Factores de TiempoRESUMEN
BACKGROUND: Sevoflurane postconditioning (SevP) effectively relieves myocardial ischemia/reperfusion (I/R) injury but performs poorly in the diabetic myocardium. Previous studies have revealed the important role of increased oxidative stress in diabetic tissues. Notably, mitochondrial fission mediated by dynamin-related protein 1 (Drp1) is an upstream pathway of reactive oxygen production. Whether the ineffectiveness of SevP in the diabetic myocardium is related to Drp1-dependent mitochondrial fission remains unknown. This study aimed to explore the important role of Drp1 in the diabetic myocardium and investigate whether Drp1 inhibition could restore the cardioprotective effect of SevP. METHODS: In the first part of the study, adult male Sprague-Dawley rats were divided into 6 groups. Rats in the diabetic groups were fed with high-fat and high-sugar diets for 8 weeks and injected intraperitoneally with streptozotocin (35 mg/kg). Myocardial I/R was induced by 30 min of occlusion of the left anterior descending branch of the coronary artery followed by 120 min of reperfusion. SevP was applied by continuous inhalation of 2.5 % sevoflurane 1 min before reperfusion, which lasted for 10 min. In the second part of the study, we applied mdivi-1 to investigate whether Drp1 inhibition could restore the cardioprotective effect of SevP in the diabetic myocardium. The myocardial infarct size, mitochondrial ultrastructure, apoptosis index, SOD activity, MDA content, and Drp1 expression were detected. RESULTS: TTC staining and TUNEL results showed that the myocardial infarct size and apoptosis index were increased in the diabetic myocardium. However, SevP significantly alleviated myocardial I/R injury in the normal myocardium but not in the diabetic myocardium. Additionally, we found an elevation in Drp1 expression, accompanied by more severe fission-induced structural damage and oxidative stress in the diabetic myocardium. Interestingly, we discovered that the beneficial effect of SevP was restored by mdivi-1, which significantly suppressed mitochondrial fission and oxidative stress. CONCLUSIONS: Our study demonstrates the crucial role of mitochondrial fission dependent on Drp1 in the diabetic myocardium subjected to I/R, and strongly indicates that Drp1 inhibition may restore the cardioprotective effect of SevP in diabetic rats.
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Anestésicos por Inhalación/farmacología , Diabetes Mellitus Experimental/metabolismo , Dinaminas/metabolismo , Infarto del Miocardio/prevención & control , Daño por Reperfusión Miocárdica/prevención & control , Miocitos Cardíacos/efectos de los fármacos , Sevoflurano/farmacología , Animales , Apoptosis/efectos de los fármacos , Diabetes Mellitus Experimental/complicaciones , Diabetes Mellitus Experimental/patología , Masculino , Mitocondrias Cardíacas/efectos de los fármacos , Mitocondrias Cardíacas/metabolismo , Mitocondrias Cardíacas/patología , Dinámicas Mitocondriales/efectos de los fármacos , Infarto del Miocardio/complicaciones , Infarto del Miocardio/metabolismo , Infarto del Miocardio/patología , Daño por Reperfusión Miocárdica/complicaciones , Daño por Reperfusión Miocárdica/metabolismo , Daño por Reperfusión Miocárdica/patología , Miocitos Cardíacos/metabolismo , Miocitos Cardíacos/patología , Estrés Oxidativo/efectos de los fármacos , Ratas Sprague-DawleyRESUMEN
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.
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Enfermedades Desmielinizantes/terapia , Hipoxia-Isquemia Encefálica/terapia , Poscondicionamiento Isquémico/métodos , Macrófagos/efectos de los fármacos , Microglía/efectos de los fármacos , Sevoflurano/administración & dosificación , Animales , Animales Recién Nacidos , Axones/efectos de los fármacos , Axones/metabolismo , Encéfalo/irrigación sanguínea , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Enfermedades Desmielinizantes/metabolismo , Femenino , Hipoxia-Isquemia Encefálica/metabolismo , Macrófagos/metabolismo , Masculino , Aprendizaje por Laberinto/efectos de los fármacos , Aprendizaje por Laberinto/fisiología , Microglía/metabolismo , Neuronas/efectos de los fármacos , Neuronas/metabolismo , Inhibidores de Agregación Plaquetaria/administración & dosificación , Ratas , Ratas Sprague-DawleyRESUMEN
Inhalation anesthetics have been demonstrated to have protective effects against myocardial ischemia reperfusion injury (MIRI). O-linked GlcNAcylation (O-GlcNAc) modifications have been shown to protect against MIRI. This study aimed to investigate whether O-GlcNAcylation and necroptosis signaling were important for sevoflurane postconditioning (SPC) induced cardioprotective effects. Apart from rats in the SHAM and sevoflurane (SEVO) group, rats underwent 30 min ischemia followed by 2 h reperfusion. Cardiac hemodynamics and function were determined. In addition, myocardial infarction size, cardiac function parameters, myocardial lactic dehydrogenase (LDH) content, myocardium histopathological changes, necrotic myocardium, O-GlcNAcylation, and protein expression levels of necroptosis biomarkers were measured, together with co-immunoprecipitation experiments using proteins associated with the necroptosis pathway and O-GlcNAcylation. SPC reduced myocardial infarction size, ameliorated cardiac function, restored hemodynamic performance, improved histopathological changes, and reduced receptor-interacting protein kinase 1 (RIPK1)/receptor-interacting protein kinase 3 (RIPK3)/mixed lineage kinase domain-like (MLKL) mediated necroptosis. In addition, SPC up-regulated O-GlcNAc transferase (OGT) mediated O-GlcNAcylation, increased O-GlcNAcylated RIPK3, and inhibited the association of RIPK3 and MLKL. However, OSMI-1, an OGT inhibitor, abolished SPC mediated cardioprotective effects and inhibited OGT mediated up-regulation of O-GlcNAcylation and down-regulation of RIPK3 and MLKL proteins induced by SPC. Our study demonstrated that SPC restrained MIRI induced necroptosis via regulating OGT mediated O-GlcNAcylation of RIPK3 and lessening the formulation of RIPK3/MLKL complex.
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Anestésicos por Inhalación/farmacología , Daño por Reperfusión Miocárdica/patología , Proteína Serina-Treonina Quinasas de Interacción con Receptores/metabolismo , Sevoflurano/farmacología , Transducción de Señal/efectos de los fármacos , Acetilación , Animales , Masculino , Infarto del Miocardio/patología , Daño por Reperfusión Miocárdica/metabolismo , N-Acetilglucosaminiltransferasas/metabolismo , Necroptosis/efectos de los fármacos , Ratas , Ratas Sprague-Dawley , Transducción de Señal/fisiología , Regulación hacia ArribaRESUMEN
Mitochondrial autophagy is involved in myocardial protection of sevoflurane postconditioning (SPostC) and in diabetic state this protective effect is weakened due to impaired HIF-1 signaling pathway. Previous studies have proved that deferoxamine (DFO) could activate impaired HIF-1α in diabetic state to restore the cardioprotective of sevoflurane, while the specific mechanism is unclear. This study aims to investigate whether HIF-1/BNIP3-mediated mitochondrial autophagy is involved in the restoration of sevoflurane postconditioning cardioprotection in diabetic state. Ischemia/reperfusion (I/R) model was established by ligating the anterior descending coronary artery and sevoflurane was administered at the first 15 min of reperfusion. Myocardial infarct size, mitochondrial ultrastructure and autophagosome, ATP content, mitochondrial membrane potential, ROS production, HIF-1α, BNIP3, LC3B-II, Beclin-1, P62, LAMP2 protein expression were detected 2 h after reperfusion, and cardiac function was evaluated by ultrasound at 24 h after reperfusion. Our results showed that with DFO treatment, SPostC up-regulated the expression of HIF-1α and BNIP3, thus reduced the expression of key autophagy proteins LC3B-II, Beclin-1, p62, and increased the expression of LAMP2. Furthermore, it reduced the accumulation of autophagosomes and ROS production, increased the content of ATP, and stabilized the membrane potential. Finally, the myocardial infarction size was reduced and cardiac function was improved. Taken together, DFO treatment combined with SPostC could alleviate myocardial ischemia reperfusion injury in diabetic rats by restoring and promoting HIF-1/BNIP3-mediated mitochondrial autophagy.
RESUMEN
BACKGROUND: Hemorrhagic shock induces the cognitive deficiency. Sevoflurane postconditioning has been documented to provide neuroprotection against ischemic-reperfusion injury by suppressing apoptosis. Mitochondrial permeability transition pore (mPTP) plays an important role in apoptosis, but it is unknown if the protective effect of sevoflurane postconditioning on hemorrhagic shock and resuscitation is associated with the change of mPTP opening. Hence, the aim of the study was to find out the precise mechanism of the sevoflurane postconditioning. METHODS: Sprague Dawley rats were subjected to hemorrhage shock for 60 min and then exposed to 2.4% sevoflurane for 30 min at the instant of reperfusion. Additionally, an opener (atractyloside) or an inhibitor (cyclosporine A) of mPTP was used in the animal model before sevoflurane postconditioning. Rats were randomly assigned into groups of Sham, Shock, Shock+Sevoflurane, Shock+Atractyloside, Shock+Sevoflurane+Atractyloside, Shock+Cyclosporin A, and Shock+Sevoflurane+Cyclosporin A treatment. Rat behavior was assessed for 5 days by Morris water maze 72 hr after surgery, and then hippocampus CA1 region was immunohistochemically stained. Brains were harvested 24 hr after surgery to detect the protein expression levels of Bcl-2, Bax, and cytochrome C by Western blot, the changes of mPTP opening, and mitochondrial membrane potential (MMP). RESULTS: We found that sevoflurane postconditioning significantly improved rats' spatial learning and memory ability, down-regulated the expression of Bax, cytochrome C, and caspase-3, up-regulated the expression of Bcl-2, decreased the mPTP opening, and increased the MMP. The neuroprotective effect of sevoflurane postconditioning was abolished by atractyloside, but cyclosporin A played the similar protective role as sevoflurane postconditioning. CONCLUSION: These findings proved that sevoflurane postconditioning improved spatial learning and memory ability in hemorrhage shock and resuscitation rats, the mechanism of which may be related to block mPTP opening, increase MMP, and reduce neuron apoptosis in the hippocampus.
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Poscondicionamiento Isquémico/métodos , Memoria/efectos de los fármacos , Poro de Transición de la Permeabilidad Mitocondrial/metabolismo , Sevoflurano/farmacología , Aprendizaje Espacial/efectos de los fármacos , Animales , Apoptosis , Modelos Animales de Enfermedad , Masculino , Fármacos Neuroprotectores , Ratas , Ratas Sprague-Dawley , Choque Hemorrágico/complicaciones , Choque Hemorrágico/metabolismoRESUMEN
OBJECTIVE: Various experimental studies have reported neuroprotective effects of sevoflurane postconditioning against cerebral ischemia-reperfusion injury. We therefore investigated its neuroprotective effects on hyperperfusion-related transient neurologic deterioration, called symptomatic cerebral hyperperfusion (SCH), and also identified predictive factors for SCH in patients with moyamoya disease after revascularization surgery. METHODS: A total of 152 adult patients with moyamoya disease undergoing anastomosis of the superficial temporal artery to middle cerebral artery were randomly allocated into 2 groups. The postconditioning group (group S, n = 76) inhaled sevoflurane of 1 minimum alveolar concentration for 15 minutes immediately after reperfusion and then washed it out slowly for 15 minutes. The control group (group C, n = 76) received no intervention. The incidence of SCH was compared between the 2 groups. RESULTS: The incidence of SCH was not significantly different between groups S and C (53.3% vs. 43.4%, respectively; P = 0.291). The incidence of postoperative complications and the Glasgow Outcome Scale score at hospital discharge also did not differ significantly. Predictive factors for SCH included a decreased vascular reserve in preoperative single-photon emission computed tomography scan (odds ratio [OR], 7.18; 95% confidence interval [CI], 1.78-29.02; P = 0.006), an operation performed on the dominant hemisphere (OR, 3.32; 95% CI, 1.57-6.98; P = 0.002), temporal occlusion time (OR, 1.06; 95% CI, 1.01-1.11; P = 0.017), and intraoperative minimum partial pressure of carbon dioxide (PaCO2) (OR, 0.86; 95% CI, 0.78-0.94; P = 0.001). CONCLUSIONS: Sevoflurane postconditioning did not reduce the incidence of SCH after revascularization surgery in patients with moyamoya disease. Rather, a decreased vascular reserve, operation on the dominant hemisphere, increased temporal occlusion time, and decreased intraoperative minimum PaCO2 were associated with SCH in these patients.
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Anestésicos por Inhalación/uso terapéutico , Isquemia Encefálica/prevención & control , Revascularización Cerebral/métodos , Poscondicionamiento Isquémico/métodos , Arteria Cerebral Media/cirugía , Enfermedad de Moyamoya/cirugía , Complicaciones Posoperatorias/prevención & control , Sevoflurano/uso terapéutico , Arterias Temporales/cirugía , Adulto , Isquemia Encefálica/epidemiología , Isquemia Encefálica/fisiopatología , Dióxido de Carbono , Femenino , Humanos , Incidencia , Masculino , Persona de Mediana Edad , Enfermedad de Moyamoya/diagnóstico por imagen , Oportunidad Relativa , Presión Parcial , Complicaciones Posoperatorias/epidemiología , Complicaciones Posoperatorias/fisiopatología , Factores de Tiempo , Tomografía Computarizada de Emisión de Fotón ÚnicoRESUMEN
This study aimed to investigate the effect of miR-206 on apoptosis and autophagy of cardiomyocytes in ischaemia/reperfusion (I/R) injured rats treated with sevoflurane post-conditioning (SP) through the AMPK/Nampt signalling pathway. Rat models of myocardial I/R injury were established. The combination of SP, miR-206 inhibitor, AMPK activator AICAR and inhibitor Compound C was induced in rats, and their effects on I/R injury were determined with detection of malondialdehyde (MDA), hydroxyproline (HYP) and superoxide dismutase (SOD) levels in cardiomyocytes, autophagy, and apoptosis. We also conducted experiments to determine p62, Beclin1, Bax and Bcl-2 expression and the mRNA and expression pattern of AMPK/Nampt signalling pathway-related genes. Myocardial I/R injured rats revealed decreased SOD activity and elevated MDA content, autophagy and apoptosis. With the combined performance of SP, miR-206 inhibitor and AMPK activator AICAR, the rats presented higher SOD level and lower MDA and HYP levels, suppressed autophagy and apoptosis. Meanwhile, miR-206 inhibition also contributed to elevated expression of Nampt and the extent of AMPK phosphorylation, increased Bcl-2/Bax ratio, but degraded expression of p62 and Beclin1. Collectively, inhibition of miR-206 could activate the AMPK/Nampt signalling pathway, thus protecting against myocardial I/R injury on the basis of SP.
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Proteínas Quinasas Activadas por AMP/metabolismo , MicroARNs/genética , Daño por Reperfusión Miocárdica/genética , Daño por Reperfusión Miocárdica/patología , Nicotinamida Fosforribosiltransferasa/metabolismo , Animales , Apoptosis/efectos de los fármacos , Autofagia/efectos de los fármacos , Beclina-1/biosíntesis , Genes bcl-2/fisiología , Miocitos Cardíacos/efectos de los fármacos , Estrés Oxidativo/efectos de los fármacos , Péptidos/metabolismo , ARN Mensajero/biosíntesis , Ratas , Ratas Sprague-Dawley , Sevoflurano/farmacología , Proteína X Asociada a bcl-2/biosíntesisRESUMEN
Subject: Cardiovascular disease, as a very common and serious coexisting disease in diabetic patients, and is one of the risk factors that seriously affect the prognosis and complications of surgical patients. Previous studies have shown that sevoflurane post-conditioning (SPostC) exerts a protective effect against myocardial ischemia/reperfusion injury by HIF-1α, but the protective effect is weakened or even disappeared under hyperglycemia. This study aims to explore whether regulating the HIF-1α/MIF/AMPK signaling pathway can restore the protective effect and reveal the mechanism of SPostC on cardiomyocyte hypoxia/reoxygenation injury under high glucose conditions. Methods: H9c2 cardiomyocytes were cultured in normal and high-concentration glucose medium to establish a hypoxia/reoxygenation (H/R) injury model of cardiomyocytes. SPostC was performed with 2.4% sevoflurane for 15 min before reoxygenation. Cell damage was determined by measuring cell viability, lactate dehydrogenase activity, and apoptosis; Testing cell energy metabolism by detecting reactive oxygen species (ROS) generation, ATP content and mitochondrial membrane potential; Analysis of the change of HIF-1α, MIF and AMPKα mRNA expression by RT-PCR. Western blotting was used to examine the expression of HIF-1α, MIF, AMPKα and p-AMPKα proteins. HIF-1α and MIF inhibitors and agonists were administered 40 min before hypoxia. Results: 1) SPostC exerts a protective effect by increasing cell viability, reducing LDH levels and cell apoptosis under low glucose (5 µM) after undergoing H/R injury; 2) High glucose concentration (35 µM) eliminated the cardioprotective effect of SPostC, which is manifested by a significantly decrease in the protein and mRNA expression level of the HIF-1α/MIF/AMPK signaling pathway, accompanied by decreased cell viability, increased LDH levels and apoptosis, increased ROS production, decreased ATP synthesis, and decreased mitochondrial membrane potential; 3. Under high glucose (35 µM), the expression levels of HIF-1α and MIF were up-regulated by using agonists, which can significantly increase the level of p-AMPKα protein, and the cardioprotective effect of SPostC was restored. Conclusion: The signal pathway of HIF-1α/MIF/AMPK of H9c2 cardiomyocytes may be the key point of SPostC against H/R injure. The cardioprotective of SPostC could be restored by upregulating the protein expression of HIF-1α and MIF under hyperglycemia.
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BACKGROUND: Sevoflurane postconditioning (SpostC) can alleviate hypoxia-reoxygenation injury of cardiomyocytes; however, the specific mechanism remains unclear. This study aimed to investigate whether SpostC promotes mitochondrial autophagy through the hypoxia-inducible factor-1 (HIF-1)/BCL2/adenovirus E1B 19-kDa-interacting protein 3 (BNIP3) signaling pathway to attenuate hypoxia-reoxygenation injury in cardiomyocytes. METHODS: The H9C2 cardiomyocyte hypoxia/reoxygenation model was established and treated with 2.4% sevoflurane at the beginning of reoxygenation. Cell damage was determined by measuring cell viability, lactate dehydrogenase activity, and apoptosis. Mitochondrial ultrastructural and autophagosomes were observed by transmission electron microscope. Western blotting was used to examine the expression of HIF-1, BNIP3, and Beclin-1 proteins. The effects of BNIP3 on promoting autophagy were determined using interfering RNA technology to silence BNIP3. RESULTS: Hypoxia-reoxygenation injury led to accumulation of autophagosomes in cardiomyocytes, and cell viability was significantly reduced, which seriously damaged cells. Sevoflurane postconditioning could upregulate HIF-1α and BNIP3 protein expression, promote autophagosome clearance, and reduce cell damage. However, these protective effects were inhibited by 2-methoxyestradiol or sinBNIP3. CONCLUSION: Sevoflurane postconditioning can alleviate hypoxia-reoxygenation injury in cardiomyocytes, and this effect may be achieved by promoting mitochondrial autophagy through the HIF-1/BNIP3 signaling pathway.
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Background: When neonatal rats suffer hypoxic-ischemic brain injury (HIBI), autophagy is over-activated in the hippocampus, and inhibition of autophagy provides neuroprotection. The aim of this study was to investigate the possible roles of autophagy and Ezh2-regulated Pten/Akt/mTOR pathway in sevoflurane post-conditioning (SPC)-mediated neuroprotection against HIBI in neonatal rats. Methods: Seven-day-old Sprague-Dawley rats underwent left common artery ligation followed by 2 h hypoxia as described in the Rice-Vannucci model. The roles of autophagy and the Ezh2-regulated Pten/Akt/mTOR signaling pathway in the neuroprotection conferred by SPC were examined by left-side intracerebroventricular injection with the autophagy activator rapamycin and the Ezh2 inhibitor GSK126. Results: SPC was neuroprotective against HIBI through the inhibition of over-activated autophagy in the hippocampus as characterized by the rapamycin-induced reversal of neuronal density, neuronal morphology, cerebral morphology, and the expression of the autophagy markers, LC3B-II and Beclin1. SPC significantly increased the expression of Ezh2, H3K27me3, pAkt, and mTOR and decreased the expression of Pten induced by HI. The Ezh2 inhibitor, GSK126, significantly reversed the SPC-induced changes in expression of H3K27me3, Pten, pAkt, mTOR, LC3B-II, and Beclin1. Ezh2 inhibition also reversed SPC-mediated attenuation of neuronal loss and behavioral improvement in the Morris water maze. Conclusion: These results indicate that SPC inhibits excessive autophagy via the regulation of Pten/Akt/mTOR signaling by Ezh2 to confer neuroprotection against HIBI in neonatal rats.
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Autofagia/efectos de los fármacos , Proteína Potenciadora del Homólogo Zeste 2/metabolismo , Hipoxia-Isquemia Encefálica/tratamiento farmacológico , Fármacos Neuroprotectores/farmacología , Sevoflurano/farmacología , Animales , Animales Recién Nacidos , Proteína Potenciadora del Homólogo Zeste 2/antagonistas & inhibidores , Femenino , Hipoxia-Isquemia Encefálica/metabolismo , Hipoxia-Isquemia Encefálica/patología , Indoles/farmacología , Masculino , Piridonas/farmacología , Ratas , Ratas Sprague-DawleyRESUMEN
Hypoxic-ischemic brain injury (HIBI) in neonates is one of the major contributors of newborn death and cognitive impairment. Numerous animal studies have demonstrated that autophagy is substantially increased in HIBI and that sevoflurane postconditioning (SPC) can attenuate HIBI. However, if SPC-induced neuroprotection inhibits autophagy in HIBI remains unknown. To investigate if cerebral protection induced by SPC is related to decreased autophagy in the setting of HIBI. Postnatal rats at day 7 (P7) were randomly assigned to 7 different groups: Sham, HIBI, SPC-HIBI, HIBI + rapamycin, SPC-HIBI + rapamycin, HIBI + p-extracellular signal-regulated kinase (p-ERK) inhibitor, and SPC-HIBI + p-ERK inhibitor. To induce HIBI, neonatal rats underwent left common carotid artery ligation, followed by 2 h of hypoxia (8% O2). Rats in the SPC groups were treated with 1 minimum alveolar concentration ([MAC], 2.4%) SPC for 30 min after HIBI induction. Markers of autophagy and expression of ERK cascade components were measured in the rat brains after 24 h. Spatial learning and memory function were examined 29-34 days after administration of an autophagy agonist or a p-ERK inhibitor. The expression of microtubule-associated proteins 1A/1B, light chain 3B II (LC3-II) and tuberous sclerosis complex 2 (TSC2) were decreased in the SPC-HIBI group compared to the HIBI group. Expression of the p62 sequestosome 1 (P62/SQSTM1) protein, p-ERK/ERK, phospho-mammalian target of rapamycin (p-mTOR) and phospho-p70S6 were increased in SPC-HIBI group. Rats within the SPC-HIBI groups that also received the p-ERK inhibitor or autophagy inhibitor demonstrated reduced cross platform times and increased escape latency. Approximately 30 min of 2.4% SPC treatment in the P7 rat HIBI model attenuated excessive autophagy in the brain by elevating the ERK cascade. This finding provides additional insight into HIBI and identifies new targets for therapeutic approaches to treat HIBI.
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Autofagia/efectos de los fármacos , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Hipoxia-Isquemia Encefálica/tratamiento farmacológico , Poscondicionamiento Isquémico , Sevoflurano/farmacología , Animales , Encéfalo/efectos de los fármacos , Encéfalo/metabolismo , Lesiones Encefálicas/tratamiento farmacológico , Quinasas MAP Reguladas por Señal Extracelular/efectos de los fármacos , Hipoxia-Isquemia Encefálica/metabolismo , Memoria/efectos de los fármacos , Proteínas Proto-Oncogénicas c-akt/metabolismo , Ratas , Transducción de Señal/efectos de los fármacos , Serina-Treonina Quinasas TOR/metabolismoRESUMEN
Severe hemorrhagic shock induces cognitive dysfunction by promoting cell death mediated by activating endoplasmic reticulum (ER) stress. Sevoflurane postconditioning prevents neuronal apoptosis against cerebral ischemia/reperfusion injury. It is unknown if this protective effect on hemorrhagic shock and resuscitation rats (HSR) is associated with ER stress attenuation. Male adult Sprague-Dawley rats were subjected HSR by removing 40% blood volume within 30 min, and 60 min later the animals were resuscitated with infusion of the removing blood in 30 min. Sevoflurane postconditioning was performed by inhaling sevoflurane at three different concentrations (0.5, 1.0, 1.5 MAC) at the onset of resuscitation for 30 min. Severe hypotension (mean arterial pressure 40-45 mmHg) occurred in the shock session for 60 min accompanying with significantly elevated lactate, decreased BE and pH values in arterial blood gas analysis. There were impaired spatial learning and memory following HSR indicated by persistently longer escape latency and lower correct rate, as well as less duration and crossing in the target quadrant by using Morris water maze and Y-maze tests. In the hippocampal CA1 region, there was significantly higher activity of caspase-3 induced by HSR. HSR also elevated the expression of inositol-requiring enzyme 1α (IRE1α) and caspase-12 in the hippocampus by western blot analysis. Sevoflurane postconditioning at 1.0 and 1.5 MAC significantly reversed these changes. These findings suggested that sevoflurane postconditioning could improve spatial learning and memory deficits induced by severe hemorrhagic shock and subsequent resuscitation. The suppression of endoplasmic reticulum stress provided critical contribution in neural apoptosis mediated by IRE1α-caspase-12 pathway.