Résumé
Aim To investigate the dynamic time-course changes in neuronal cytoskeleton after acute ischemia and reperfusion in rats. Methods Reperfusion was performedin rats by blocking the middle cerebralarteryfor 90 min, then therats wereobserved and collected at different time points. The brain damage wasobserved by Nissl staining,and neurobehavioural function was evaluated with neurological deficit score and forelimb placement test. The cellular changes in the alternations of cytoskeletal elements including microtubule associated protein 2 (MAP2) and neurofilament heavy chain (NF-H) were observed by immunohistochemistry staining and Western blot. Impaired axons, dendrites and cytoskeletal alternations were detected by electron microscope. Results Brain damage and neurobehavioural function were gradually aggravated with the prolongation of reperfusion. Brain damage appeared earlier and more severe in striatum than in cortex. Moreover, decreased MAP2-related and increased NF-H-related immunoreactive intensities were found in the ischemic areas. Impaired cytoskeletal arrangement and reduced dense were indicated. Damaged cytoskeletal components such as microtubules and neurofilament arrangement, decreased axonal filament density, and swelled dendrites were observed after cerebral ischemia reperfusion by ultrastructural observations. Conclusions Different brain regions have diverse tolerance to ischemia-reperfusion injury. Major elements of neuronal cytoskeleton show dynamic responses to ischemia and reperfusion, which may further contribute to brain damage and neurological impairment following MCAO and reperfusion.
Résumé
Aim To observe cellular damage and astrocyte activation at different time points of cerebral ischemia and reperfusion. Methods The middle cerebral artery of male SpragueDawley rats was occluded for 90 min followed by different time points of reperfusion. Eighty-five SPF male SD rats were randomly divided into control group (Sham), IR3, 6, 12, 24 and IR48h (MCAO followed by 48 h of reperfusion) group. Cerebral ischemia and reperfusion injury was observed by HE staining, and the structure of astrocytes was estimated with transmission electron microscopy (TEM). GFAP expression was detected by immunofluorescence staining and Western blot. Results Cerebral ischemia following by different time points of reperfusion led to different degrees of cellular damage, which was the most serious at 24 h of reperfusion. TEM showed destruction of astrocytes structure, swollen organelles and broken mitochondrial ridge. After cerebral ischemia-reperfusion, the expression levels of GFAP were significant up-regulated in the ischemic penumbra cortex and the highest was at 48 h of reperfusion, indicating astrocytes were activated. In addition, the results showed the gradual decrease in GFAP expression in the infarct core. Conclusions After cerebral ischemia-reperfusion, cellular damage is aggravated, and astrocytes are gradually activated in the ischemic penumbra. With the extension of reperfusion time, the boundaries of infarct area and ischemic area are gradually clear, and scarring may occur.
Résumé
Objective:To explore the effect of hyperbaric oxygen (HBO) on the blood-brain barrier via the silent information regulator 1 (SIRT1)/Forkhead box O1(FoxO1) signaling pathway after cerebral ischemia and reperfusion using a rat model.Methods:Forty Wistar rats were randomly assigned into sham, cerebral ischemia-reperfusion (CIR), CIR+ HBO and CIR+ HBO+ EX527 groups, each of 10. The cerebral ischemia-reperfusion model was established in all groups except the sham group by right middle cerebral artery occlusion using the modified thread-occlusion method. The sham group was not ligated. Both the CIR+ HBO and CIR+ HBO+ EX527 groups were given HBO 1, 9, 21, 45 and 69 hours after the reperfusion. The CIR+ HBO+ EX527 group was additionally injected with 5mg/kg of EX527(a SIRT1inhibitor) peritoneally 4, 12, 24, 48 and 72 hours after the reperfusion. Then 2% Evens blue (EB) was injected into the tail vein an hour before the rats were sacrificed. The content of EB and the expression of SIRT1, FoxO1, ZO-1, Occludin, Claudin-5 mRNA and their proteins were determined using spectrophotometry, reverse transcription-polymerase chain reactions and Western blotting.Results:The average EB content of the hippocampal brain tissue from the CIR, CIR+ HBO and CIR+ HBO+ EX527 rats was significantly greater than the Sham group′s average 72h after reperfusion. The average expression of SIRT1, FoxO1, ZO-1, Occludin and Claudin-5 mRNA and their proteins was significantly lower, with the CIR + HBO + EX 527 group′s average significantly lower than that of the CIR+ HBO group.Conclusions:HBO can increase the expression of tight junction protein via the SIRT1/FoxO1 pathway. It helps to protect the blood-brain barrier in CIR injury situations.
Résumé
Objective@#To observe the effect of electroacupuncture on the volume of cerebral infarction, apoptosis of cerebral cells and the expression of protein kinase A (PKA) in the cerebral cortex of rats after ischemia and reperfusion so as to explore how electroacupuncture stimulates brain protection.@*Methods@#One hundred and twenty healthy, adult, male Sprague-Dawley rats were randomly divided into a sham operation group, a model group, an electroacupuncture group and an electroacupuncture with pre-stimulation group. All except the rats in the sham operation group received occulusion of the left middle cerebral artery using the intraluminal thread method for 2h and then reperfusion. Before the operation, the rats in the electroacupuncture with pre-stimulation group were given 30 minutes of electroacupuncture at the baihui, dazhui and right neiguan points every day for 5 days. After the operation both the electroacupuncture group and the pre-stimulated group were given that same electroacupuncture regimen. The other two groups received no special treatment. Garcia scoring was used to evaluate the neurological deficits of all of the rats 5 and 10 days after the intervention. Meanwhile, the ischemic volume, apoptosis of cortical cells and PKA-positive cells were determined using flow cytometry and immunohistochemistry after triphenyltetrazolium chloride (TTC) staining.@*Results@#The neurological function of the injured rats was severely impaired, while no neurological deficit was found in the sham operation group. The average Garcia score, cerebral infarction volume, cerebral apoptosis rate and PKA-positive cell expression rate of the electroacupuncture and electroacupuncture with pre-stimulation groups were all significantly better than those of the model group at the same time points. The averages of the electroacupuncture with pre-stimulation group were all significantly superior to those of the electroacupuncture group at the same time points.@*Conclusions@#Pre-stimulation using electroacupuncture can promote the recovery of injured nerves after cerebral ischemia and reperfusion, at least in rats. Electroacupuncture′s protective mechanism may be related to its reducing the infarcted volume, inhibiting apoptosis of brain cells and promoting PKA expression.
Résumé
Objective: To investigate the protective effects of the combination of Xuesaitong (XST) and aspirin on cerebral ischemia and reperfusion injury (CIRI) in rats, and further explore the underlying mechanisms. Methods: A total of 150 male Sprague-Dawley (SD) rats were randomly divided into five groups with 30 rats in each group: sham group, middle cerebral artery occlusion/reperfusion (MCAO/R) model group, XST group, aspirin group, and XST + aspirin group. Rats were pretreated with XST, aspirin, or XST + aspirin for 7 d. One hour after the last administration, a model of CIRI was induced by MCAO/R. Neurological deficits were assessed using Longa's five-point scale. Cerebral edema was detected by the measurement of brain water content. The volume of cerebral infarction was determined by 2,3,5-triphenyltetrazolium chloride (TTC) staining. The activities of superoxide dismutase (SOD), catalase (CAT), and glutathione peroxidase (GSH-Px), as well as levels of malonaldehyde (MDA) were detected by commercial kits. Enzyme-linked immunosorbent assay (ELISA) was used to determine the levels of interleukin-1 (IL-1β), interleukin-4 (IL-4), interleukin-6 (IL-6), interleukin-10 (IL-10), tumor necrosis factor-alpha (TNF-α), monocyte chemotactic protein 1 (MCP-1), and kynurenine in serum, cerebral cortex, and hippocampus of MCAO/R rats. The protein expression of nuclear factor erythroid 2-related factor (Nrf2), heme oxygenase-1 (HO-1), I-kappa B alpha (IκBα), and nuclear factor kappa B (NF-κB)/p65 in the cortex were analyzed by western blotting. Results: Treatment of XST, aspirin, and XST + aspirin significantly alleviated the neurological deficits, cerebral edema, and cerebral infarct volume induced by MCAO/R. Treatment of XST, aspirin, and XST + aspirin also reduced MDA, IL-1β IL-6,TNF-α MCP-1, and kynurenine levels, and increased SOD, CAT, GSH-Px, IL-4, and IL-10 levels in serum, cerebral cortex, and hippocampus of MCAO/R rats. Furthermore, treatment of XST, aspirin, and XST + aspirin decreased the expression of nuclear NF-κB/p65 and increased the expression of IκBα nuclear Nrf2, and HO-1. Importantly, the combination of XST and aspirin enhanced the protective effects of XST or aspirin treatment alone on CIRI in rats. Conclusion: The combination of XST and aspirin significantly inhibited oxidative stress and inflammation in serum, cerebral cortex, and hippocampus of MCAO/R rats. The combination of XST and aspirin exerted more protective effects than XST or aspirin treatment alone. The combination of XST and aspirin might provide the synergistic therapeutic effects on CIRI, and deserve further clinical investigation.
Résumé
Objective:To investigate the effects of high thoracic epidural anesthesia (HTEA) on the cerebral blood flow (CBF) and hippocampal apoptosis-related proteins Bcl-2 and Bax during global cerebral ischemia and reperfusion (GCI) in rats.Methods:Fifteen-minute global ischemia was established by 4-vessel occlusion and epidural catheterization was performed through T4-5 intervertebral spaces in adult male Wistar rats.According to the different drugs infused into the epidural space,the rats were randomly divided into four groups:Sham group (0.9 ％ NaC1),Sham-HTEA group (0.25 ％ bupivacaine),GCI group (global cerebral ischemia,0.9 ％ NaC1) and HTEA group (global cerebral ischemia,0.25 ％ bupivacaine).And 0.25 ％bupivacaine or 0.9 ％ saline (20 μL·h-1) was infused continuously to the thoracic epidural space from 15 minutes before ischemia to 24 hours after reperfusion.Mean arterial pressure (MAP),heart rate (HR) and cerebral blood flow (CBF) were determined until 2 hours after reperfusion,and the hippocampal Bcl-2 and Bax proteins at 24 hours after reperfusion were examined by Western-blot.Results:Compared with the GCI group,HTEA group has no significant difference on MAP and HR during ischemia and 2 hours after reperfusion,andcompared with the Sham group,MAP in GCI group increased in ischemia 0 min and decreased in reperfusion 0 min.The CBF in HTEA group was significantly lower than that in GCI group (123.1％± 35.2％ vs 177.5％± 32.4％,P＜0.01) in reperfusion 10 min,and higher than that in GCI group during the hypoperfusion of 60 to 120 minutes after reperfusion (P＜0.05),and the ratio of Bax/Bcl-2 in hippocampus was significantly decreased in HTEA group 24 hours after reperfusion (P＜0.01).Conclusions:Continuous HTEA infusion of 0.25 ％ bupivacaine 20 μL ·h-1 could maintain the hemodynamic stability,and improve the CBF of hypoperfusion period in rats,as well as reduce the ratio of Bax/Bcl-2 at 24 hours after reperfusion.
Résumé
Objective To observe the effect of scalp needling at different time points on focal cerebral microcirculation in rats with focal ischemia and reperfusion ( I-R). Methods One hundred SD rats were randomized into normal group, sham operation group, model group and scalp needling group. The model group and scalp needling group were divided I-R 12, 24, 48 and 72 h subgroups according to the time of ischemia and reperfusion, 10 rats in each subgroup. The normal group had no oper ation, sham operation only had the operation for vessel isolation, and model group and scalp needling group received middle cerebral artery occlusion with thread. Scalp needling group also had electro-acupuncture on scalp points along vertex and temple anterior linea oblique and posterior linea oblique. And then we observed the neurological severity scores (NSS) and detected the cerebral blood flow ( CBF) in the focal ischemic brain with laser Doppler blood stream meter before I-R and 12, 24, 48 and 72 h after I-R. Immunofluorescence method was used for counting the focal ischemic cerebral microvascular endothelial cells, and then the correlation of cerebral microvascular endothelial cells count with the cerebral blood flow volume was evaluated with Pearson correlation analysis. Results NSS in scalp needling group was lower than that in the model group (P0.05). Except for reperfusion for 24 hours, CBF in scalp needling group was higher than that in the model group at different time points ( P<0.05 or P<0.01). Microvascular endothelial cells count in scalp needling group was higher than that in the model group after reperfusion for 24, 48 and 72 hours ( P<0.05 or P<0.01). In the model group and scalp needling group, cerebral blood flow volume was positively correlated with microvascular endothelial cells count. Conclusion Scalp needling shows obvious effect on improving the microcirculation, increasing CBF and cerebral microvascular endothelial cells count, and promoting the recovery of ischemic tissues of rats after I-R.
Résumé
ObjectiveTo observe the protection of Baweichenxiang powder on cerebral ischemia-reperfusion injury in rats.MethodsSD rats were randomly divided into six groups,namely,sham operation group,model group,nimodipine group,Baweichenxiang powder high,medium and low-dose group.Cerebral ischemia-reperfusion rat model was established by gavage.Observe the effects of different doses of Baweichenxiang powder on behavior in rats,brain water content,infarct size,SOD,MDA and morphological changes.ResultsCompared with the control group,Baweichenxiang powder significantly reduced behavioral score (P＜0.05),reduced brain edema,decreased infarct size,improved brainmorphological changes,increased SOD activity(P＜0.05,P＜0.01)and reduced the MDA content(P＜0.05).ConclusionBaweichenxiang powder of high dose and middle dose have significant protective effect for cerebral ischemia,the mechanism may be related to its improving levels of free radicals and lipid peroxidation.
Résumé
Objective To assess the influence of transcranial electric stimulation (TES) on the recovery of motor function after cerebral focal ischemia and reperfusion and to explore the mechanisms in terms of neural plasticity.Methods An acute focal ischemia-reperfusion model was established by transient occlusion of the right middle cerebral artery (MCAO).Seventy-two male Sprague-Dawley rats were randomly divided into a TES group,a model group,a sham-operation group and a normal group.The TES group was given TES 24 h after MCAO;the model group received the operation without any treatment.Forelimb placing (FPT) and beam walking (BWT) were mea-sured at the 3rd,7th,14th and 28th day after reperfusion.Microtubule-associated protein-2 (MAP-2) and growth-associated protein-43 (GAP-43) and grey levels of reaction products in the peri-infarct region were examined by immunohistochemical techniques.Results The TES group rats had markedly better FPT and BWT performance at the 7th,14th and 28th day after MCAO,compared with the model group.Expression of MAP-2 had increased significantly more at the 14th and 28th day in the peri-infarct region in the TES group compared with the model group.Expression of GAP-43 was significantly elevated in the peri-infarct region in the TES group compared with the model group at all time points.Conclusions TES can improve motor function and neural plasticity following cerebral ischemia and reperfusion damage.The functional enhancement may be partly due to up-regulation of the expression of GAP-43 and MAP-2 in the peri-infarct region.
Résumé
Objective To observe the expression of glial fibrillary acidic protein ( GFAP), and the pathological and ultrastructnral changes of astrocytes in the CA1 subfield of the hippocampus following global cerebral ischemia and reperfusion, and to explore the neuroprotective mechanism of mild hypothermia. Methods Global cerebral ischemia was established in rats by a modified version of Pulsinelli's method. Ninety-six rats were divided into three groups including a sham-operated group, a normothermic ischemic reperfusion (IR) group and a hypothermic ischemic reperfusion (HIR) group. Each group had four subgroups which were sacrificed for 6, 12 or 24 hours, or 4 days after reperfusion (for each subgroup n = 8 ). Hematoxylin-eosin (HE) staining was used to observe morphological changes in neurons in the CA1 subfield of the hippocampus. TUNEL methods were used to detect apoptosis among those neurons. Immunohistochemical staining was used to detect the expression of GFAP in the CA1 subfield and the mechanism of astrocyte pathology. GFAP TUNEL double-labeled immunohistochemistry was used with both the shamoperated and experimental groups. Electron microscopy was also used to evaluate morphological changes in astrocytes 24 hours and 4 days after ischemia and reperfusion. Results Compared with the sham-operated group, expression of GFAP immunoreactive positive cells increased gradually in the CA1 subfield of the IR group rats. Compared with the IR group, expression of GFAP immunoreactive positive cells was significantly lower in the HIR group at all time points. Microscopic observation at the 4th day showed that some astrocytes in the CA1 subfield had died through oncosis. Conclusions Mild hypothermia can significantly decrease the expression of GFAP immunoreactive positive cells and the number of apoptotic neurons in the CA1 subfield of the hippocampus, minimize cell oedema and provide protection for neurons. Oncosis kills astrocytes following global cerebral ischemia and reperfusion.
Résumé
Objective To observe the influence of δ-opioid reperfusion in rats with change in serum protein S-100B level and to explore the neuroprotective effect of DADLE during cerebral resuscitation. Method The model of global cerebral ischemia and reperfusion was induced by bilateral common carol id artery occlusion combined with hypotension. Fifty SD rats were randomly divided into five groups: sham operation group, model group, DADLE pretreated group, DADI.E treated postischemia group, DADLE treatment during reperfusion group ( n = 10 for each group) .In sham operation group,the rats were operated without ischemia and treatment; in model group, rats had global cerebral ischemia and reperfusion model up without any treament; in DADLE pretreated group, rats received DADlE before ischemia; in DADLE treatment postischemia group,rats had DADLE immediately after ischemia; in DADLE treatment during reperfusion group,rats got DADLE during early reperfusion. After the establishment of model, serum protein S-JOOB was measured by using ELlSA.One-way analysis of variance and SNK test were used for comparison between groups. Results The serum protein S-100B level was (475.56±1.93) pg/ml in sham operational group and that was much lower than that in model group and DADLE treatment groups. While the levels of serum protein S-100B in all DADLE treatment groups were reduced significantly in comparison with model group. There were no differences in the levels of serum protein KS-100B between DADLE treatment groups. Conclusions The δ-opioid receptor DADLE exerts neuroprotective effects on global cerebral ischemia and reperfusion in rats.
Résumé
Objective:To investigate the pathological changes following the focal cerebral ischemia and reperfusion in rats and the effect of acupuncture on them. Methods: A model of ischemia and reperfusion was established by thread occlusion, and the brain-activating acupuncture was applied, and then the morphological changes of the cerebral cortex on the ischemic side were observed under electron and light microscopes. Results:Cerebral ischemia and reperfusion could lead to the structural injury of neurons, gliacytes and capillary vessels in rat's brain, and acupuncture could improve the ultra-structural injury in the area around the cerebral ischemia. It was also found that acupuncture intervention at 3 hours following reperfusion was more ideal than that at any other time points. Conclusion: Acupuncture has a protective effect on the ultra-structural injury of neurons associated with focal cerebral ischemia and reperfusion in rats, and acupuncture given within 3 hours following reperfusion can produce a satisfactory effect.
Résumé
OBJECTIVE:To study the effects of 5-hydroxymethylfurfural on learning-memory and cerebral free radical metabolism in cerebral ischemia and reperfusion model mice. METHODS: The mice were divided into sham operation group, model group, 5-HMF low dose group, 5-HMF high dose group, and dihydroergotamine mesilate positive control group. After administration of the corresponding drugs, all groups except the sham operation group were subjected to construction of cerebral ischemia and reperfusion model. The learning-memory of mice was examined by Morris water maze and step-down tests, and cerebral superoxide dismutase (SOD) activity and malondialdehyde (MDA) content were determined as well. RESULTS: In model group compared with sham operation group the escape latency and swimming distance were significantly prolonged in Morris water maze test (P
Résumé
Objective To study the effect of sodium ?-aescin on the matrix metalloproteinase-9 (MMP-9) and laminin (LN) expression in rats after focal cerebral ischemia and reperfusion, and provide experimental evidence for the relationship of MMP-9 and LN expression change with the neuroprotection of sodium ?-aescin. Methods The Spraque-Dawley (SD) rat model of focal cerebral ischemia and reperfusion of cerebral middle artery was prepared. Fifty-six rats were randomly divided into the normal, sham, ischemia and reperfusion, as well as ischemia and reperfusion-treated with sodium ?-aescin groups. The latter two were further divided into six subgroups according to reperfusion time interval of 3h, 6h, 12h, 24h, 72h and 7d after brain ischemia, with 4 rats in each subgroup. Immunohistochemical staining and image analysis were used to measure the expression of MMP-9 and LN in the ischemic region. Results The expression of MMP-9 started to increase at 6h, reached the peak at 12h, decreased in 24h, and reached the lowest on 7d in cerebral ischemia and reperfusion group. The expression of MMP-9 in the same time groups of the cerebral ischemia and reperfusion-treated with sodium ?-aescin group all obviously decreased (P
Résumé
Objective To observe the effects of mild hypothermia on heat shock protein 70 (HSP70) and glial fibrillary acidic protein (GFAP) expression in cerebral tissue after cerebral ischemia and reperfusion. Methods 30 male Wistar rats were randomly divided into sham-operated group, normal temperature group and mild hypothermia group. The model of focal cerebral ischemia was made by middle cerebral artery occlusion (MCAO) method. After 2 h of MCAO following 48 h of reperfusion, HSP70 and GFAP positive neurons were detected by immunohistochemistry technique in ischemic regions of rats. Results Many necrotic neurons were appeared in normal temperature group and no necrotic neurons in sham-operated group. Fewer necrotic neurons were fond in mild hypothermia group than in normal temperature group. The positive cells of HSP70 and GFAP in sham-operated group and mild hypothermia group were less than normal temperature group. Image analysis revealed that the average optical density of HSP70 and GFAP in sham-operated group and mild hypothermia group was significantly lower than in normal temperature group (all P