Use of microscopic interferometry for measuring changes in water content of small samples of tissue

Oedema following periods of ischaemic arrest and subsequent reperfusion has been shown experimentally and clinically to affect the functional state of the heart. Tissue water content has been measured in myocardial sections by microscopic interferometry and densitometry, and the results correlated with those obtained by wet and dry weight analysis (r = 0.87; p less than 0.001). Microscopic interferometry also revealed the distribution of the water in the tissue. Experimentally induced ischaemic arrest in isolated rat hearts resulted in predominantly intra-fibrillar oedema, whilst subsequent reperfusion resulted in interfibrillar oedema. Microscopic interferometry facilitates accurate measurement of water content in tissue samples as small as 2 mg wet weight and shows (as conventional wet/dry weight analysis cannot) the distribution of the water in the tissue.     

亚低温联合镁剂对脑缺血再灌注大鼠保护作用的研究

目的:rt-PA溶栓为缺血性卒中最有效的治疗方法,脑血流再通后挽救濒临死亡的神经细胞同时,也可能发生更为严重而持久的脑缺血再灌注损伤。本研究探讨联合应用局部亚低温(32-35℃)及硫酸镁对局灶性脑缺血再灌注大鼠的保护作用及其可能机制。方法:通过线栓法建立大鼠大脑中动脉阻塞(MCAO)及再通模型,将50只雄性Wistar大鼠随机分为假手术组、常温组、亚低温组、硫酸镁组、亚低温+硫酸镁组,每组10例,采用Longa神经功能评分、TTC染色、干湿重法、TUNEL技术,检测和比较各组脑缺血再灌注后大鼠的神经功能、脑梗死体积、脑组织含水量及凋亡细胞数。结果:与常温组相比,亚低温组与亚低温+硫酸镁组的梗死体积、神经功能评分、脑组织含水量、凋亡细胞数均明显降低,差异有显著意义(P0.05);而与亚低温组相比,亚低温+硫酸镁组局灶脑缺血大鼠的脑梗死体积、神经功能评分、脑组织含水量、凋亡细胞数均显著减少,差异有显著意义(P0.05)。结论:与单独应用亚低温相比,局部亚低温与硫酸镁联合应用,对局灶性脑缺血再灌注大鼠可发挥更有效的脑保护作用。其机制可能与抑制脑缺血再灌注后凋亡及减轻脑水肿有关。二者联用可能为缺血性卒中患者提供一种减轻溶栓后再灌注损伤的有效脑保护方法。     

Edaravone ameliorates oxidative stress associated cholinergic dysfunction and limits apoptotic response following focal cerebral ischemia in rat

Stroke is a life-threatening disease with major cause of mortality and morbidity worldwide. The neuronal damage following cerebral ischemia is a serious risk to stroke patients. Oxidative stress and apoptotic damage play an important role in cerebral ischemic pathogenesis and may represent a target for treatment. The objective of this study was to test the hypothesis that administration of edaravone (Edv) maintains antioxidant status in brain, improves the cholinergic dysfunction and suppresses the progression of apoptosis response in rat. To test this hypothesis, male Wistar rats were subjected to middle cerebral artery occlusion (MCAO) of 2 h followed by reperfusion for 22 h. Edv was administered (10 mg/kg bwt) intraperitoneally 30 min before the onset of ischemia and 1 h after reperfusion. After reperfusion, rats were tested for neurobehavioral activities and were sacrificed for the infarct volume, estimation of oxidative damage markers. Edv treatment significantly reduced ischemic lesion volume, improved neurological deficits, contended oxidative loads, and suppressed apoptotic damage. In conclusion, treatment with Edv ameliorated the neurological and histological outcomes with elevated endogenous anti-oxidants status as well as reduced induction of apoptotic responses in MCA occluded rat. We theorized that Edv is among the pharmacological agents that reduce free radicals and its associated cholinergic dysfunction and apoptotic damage and have been found to limit the extent of brain damage following stroke.     

Elevated production of 20-HETE in the cerebral vasculature contributes to severity of ischemic stroke and oxidative stress in spontaneously hypertensive rats

Hypertension is a major risk factor for stroke, but the factors that contribute to the increased incidence and severity of ischemic stroke in hypertension remain to be determined. 20-hydroxyeicosatetraenoic acid (20-HETE) has been reported to be a potent constrictor of cerebral arteries, and inhibitors of 20-HETE formation reduce infarct size following cerebral ischemia. The present study examined whether elevated production of 20-HETE in the cerebral vasculature could contribute to the larger infarct size previously reported after transient middle cerebral artery occlusion (MCAO) in hypertensive strains of rat [spontaneously hypertensive rat (SHR) and spontaneously hypertensive stroke-prone rat (SHRSP)]. The synthesis of 20-HETE in the cerebral vasculature of SHRSP measured by liquid chromatography-tandem mass spectrometry was about twice that seen in Wistar-Kyoto (WKY) rats. This was associated with the elevated expression of cytochrome P-450 (CYP)4A protein and CYP4A1 and CYP4A8 mRNA. Infarct volume after transient MCAO was greater in SHRSP (36+/-4% of hemisphere volume) than in SHR (19+/-5%) or WKY rats (5+/-2%). This was associated with a significantly greater reduction in regional cerebral blood flow (rCBF) in SHR and SHRSP than in WKY rats during the ischemic period (78% vs. 62%). In WKY rats, rCBF returned to 75% of control following reperfusion. In contrast, SHR and SHRSP exhibited a large (166+/-18% of baseline) and sustained (1 h) postischemic hyperperfusion. Acute blockade of the synthesis of 20-HETE with N-hydroxy-N'-(4-butyl-2-methylphenyl)-formamidine (HET0016; 1 mg/kg) reduced infarct size by 59% in SHR and 87% in SHRSP. HET0016 had no effect on the fall in rCBF during MCAO but eliminated the hyperemic response. HET0016 also attenuated vascular O2*- formation and restored endothelium-dependent dilation in cerebral arteries of SHRSP. These results indicate the production of 20-HETE is elevated in the cerebral vasculature of SHRSP and contributes to oxidative stress, endothelial dysfunction, and the enhanced sensitivity to ischemic stroke in this hypertensive model.     

Intra-Arterial Transplantation of Allogeneic Mesenchymal Stem Cells Mounts Neuroprotective Effects in a Transient Ischemic Stroke Model in Rats: Analyses of Therapeutic Time Window and Its Mechanisms

Objective

Intra-arterial stem cell transplantation exerts neuroprotective effects for ischemic stroke. However, the optimal therapeutic time window and mechanisms have not been completely understood. In this study, we investigated the relationship between the timing of intra-arterial transplantation of allogeneic mesenchymal stem cells (MSCs) in ischemic stroke model in rats and its efficacy in acute phase.

Methods

Adult male Wistar rats weighing 200 to 250g received right middle cerebral artery occlusion (MCAO) for 90 minutes. MSCs (1×10⁶cells/ 1ml PBS) were intra-arterially injected at either 1, 6, 24, or 48 hours (1, 6, 24, 48h group) after MCAO. PBS (1ml) was intra-arterially injected to control rats at 1 hour after MCAO. Behavioral test was performed immediately after reperfusion, and at 3, 7 days after MCAO using the Modified Neurological Severity Score (mNSS). Rats were euthanized at 7 days after MCAO for evaluation of infarct volumes and the migration of MSCs. In order to explore potential mechanisms of action, the upregulation of neurotrophic factor and chemotactic cytokine (bFGF, SDF-1α) induced by cell transplantation was examined in another cohort of rats that received intra-arterial transplantation at 24 hours after recanalization then euthanized at 7 days after MCAO for protein assays.

Results

Behavioral test at 3 and 7 days after transplantation revealed that stroke rats in 24h group displayed the most robust significant improvements in mNSS compared to stroke rats in all other groups (p’s<0.05). Similarly, the infarct volumes of stroke rats in 24h group were much significantly decreased compared to those in all other groups (p’s<0.05). These observed behavioral and histological effects were accompanied by MSC survival and migration, with the highest number of integrated MSCs detected in the 24h group. Moreover, bFGF and SDF-1α levels of the infarcted cortex were highly elevated in the 24h group compared to control group (p’s<0.05).

Conclusions

These results suggest that intra-arterial allogeneic transplantation of MSCs provides post-stroke functional recovery and reduction of infarct volumes in ischemic stroke model of rats. The upregulation of bFGF and SDF-1α likely played a key mechanistic role in enabling MSC to afford functional effects in stroke. MSC transplantation at 24 hours after recanalization appears to be the optimal timing for ischemic stroke model, which should guide the design of clinical trials of cell transplantation for stroke patients.     

Human amniotic fluid stem cells can improve cerebral vascular remodelling and neurological function after focal cerebral ischaemia in diabetic rats

Diabetes causes vascular injury and carries a high risk of ischaemic stroke. Human amniotic fluid stem cells ( hAFSCs) can enhance cerebral vascular remodelling and have the potential to improve neurological function after stroke in diabetic rats. Five groups of female rats were examined: (1) normal control, (2) type 1 diabetic (T1DM) rats induced by streptozotocin injection (DM), (3) non-DM rats receiving 60-minute middle cerebral artery occlusion (MCAO), (4) T1DM rats receiving 60-minute MCAO (DM + MCAO) and (5) T1DM rats receiving 60-minute MCAO and injection with 5 × 10⁶ hAFSCs at 3 h after MCAO (DM + MCAO + hAFSCs). Neurological function was examined before, and at 1, 7, 14, 21 and 28 days, and cerebral infarction volume and haemorrhage, cerebral vascular density, angiogenesis and inflammatory were examined at 7 and 28 days after MCAO. hAFSCs treatment caused a significant improvement of neurological dysfunction, infarction volume, blood-brain barrier leakage, cerebral arterial density, vascular density and angiogenesis and a reduction of brain haemorrhage and inflammation compared with non-treatment. Our results showed that the effect of hAFSCs treatment against focal cerebral ischaemia may act through the recovery of vascular remodelling and angiogenesis and the reduction of inflammation in ischaemic brain.     

Synergistic Effect of Selenium and Melatonin on Neuroprotection in Cerebral Ischemia in Rats

The synergistic scavenger effects of selenium and melatonin collectively we called Se-Mel was studied on the prevention of neuronal injury induced by ischemia/reperfusion. Male Wistar rats were treated with sodium selenite (0.1 mg/kg, i.p.) and melatonin (10 mg/kg, i.p.) 30 min before the middle carotid artery occlusion (MCAO) and immediately after MCAO to male Wistar rats and was continued for 3 days once daily at the interval of 24 h. Behavioral activity (spontaneous motor activity and motor deficit) was improved in Se-Mel-treated rats as compared to MCAO group rats. The level of glutathione and the activity of antioxidant enzymes was depleted significantly while the content of thiobarbituric acid reactive substances, protein carbonyl, and nitric oxide radical (NO^·) was increased significantly in MCAO group. Systemic administration of Se-Mel ameliorated oxidative stress and improves ischemia/reperfusion-induced focal cerebral ischemia. Se-Mel also inhibited inducible nitric oxide synthase expression in Se-Mel+MCAO group as compared to MCAO group rats. Thus, Se-Mel has shown an excellent neuroprotective effect against ischemia/reperfusion injury through an anti-ischemic pathway. In conclusion, we demonstrated that the pretreatment with Se-Mel at the onset of reperfusion, reduced post-ischemic damage, and improved neurological outcome following transient focal cerebral ischemia in male Wistar rat.     

SD大鼠与Wistar大鼠脑胶质瘤动物模型的建立及比较

目的比较利用SD大鼠、Wistar大鼠建立脑胶质瘤动物模型的不同,为研究脑胶质瘤的发病机制及治疗方法提供操作平台。方法利用立体定向仪建立SD大鼠、Wistar大鼠大脑皮层接种C6细胞（2.5×105个细胞/只）,建立脑胶质瘤动物模型,利用组织病理学、免疫组织化学以及核磁共振成像等技术,比较两种动物模型在成瘤率、肿瘤生长状况、死亡率以及动物一般情况等方面的异同。结果SD大鼠组、Wistar大鼠组的成瘤率均为100%,两组均未见转移;但SD大鼠组肿瘤成瘤时间较长,且部分肿瘤有自愈倾向,而Wistar大鼠组则未出现类似情况。结论Wistar大鼠大脑皮层脑胶质瘤动物模型的肿瘤性状更接近于人的脑胶质瘤,因此更适合探索和研究脑胶质瘤的发病机制和治疗方法;而SD大鼠的肿瘤由于性状类似转移瘤,且有自愈倾向,不适合作为上述相关研究的动物模型。     

缺血后处理对大鼠局灶性脑缺血再灌注损伤时TLR4 信号通路表达的影响

目的:观察缺血后处理对大鼠局灶性脑缺血再灌注损伤后TLR4通路表达的影响。方法:成年健康雄性SD大鼠110只,随机分为假手术组(sham组)(n=10)、缺血再灌注组(I/R组)和后处理组(IP组),后两组又依据缺血再灌注6h、12h、24h、48h、72h不同的时间点再分五个亚组。对各组行神经行为学评分,脑组织梗死体积测量,TUNEL技术检测神经细胞凋亡的情况,免疫组织化学技术观察各组大鼠脑组织TLR4、NF-κB和TNF-α蛋白的表达,原位杂交方法检测各组大鼠脑组织TLR4mRNA、NF-κBmRNA的表达。结果:缺血后处理可下调TLR4、NF-κB、TNF-α细胞炎性因子的表达,抑制细胞凋亡、减少脑梗死体积,改善神经行为。结论:后处理可通过抑制TLR4信号通路表达,减少脑梗死体积,改善神经功能。     

Calcitonin gene-related peptide prevents blood-brain barrier injury and brain edema induced by focal cerebral ischemia reperfusion

Cerebral ischemia is one of the diseases that most compromise the human species. Therapeutic recovery of blood-brain barrier (BBB) disruption represents a novel promising approach to reduce brain injury after stroke. To determine the effects of calcitonin gene-related peptide (CGRP) on the BBB participate in stroke progression, rat cerebral ischemia reperfusion injury was induced by a 2-hour left transient middle cerebral artery occlusion (MCAO) using an intraluminal filament, followed by 46h of reperfusion. CGRP (1μg/ml) at the dose of 3μg/kg (i.p.) was administered at the beginning of reperfusion. Subsequently, 48h after MCAO, arterial blood pressure, infarct volume, water content, BBB permeability, BBB ultrastructure, levels of aquaporin-4 (AQP4) and its mRNA were evaluated. CGRP could reduce arterial blood pressure (P<0.001), infarct volume (P<0.05), cerebral edema (P<0.01), BBB permeability (P<0.05), AQP4 mRNA expression (P<0.05) and AQP4 protein expression (P<0.01). Furthermore, CGRP treatment improved ultrastructural damage of capillary endothelium cells and decreased the loss of the tight junction observed by transmission electronic microscopy (TEM) after 46h of reperfusion. Our findings show that CGRP significantly reduced postischemic increase of brain edema with a 2-hour therapeutic window in the transient model of focal cerebral ischemia. Moreover, it seems that at least part of the anti-edematous effects of CGRP is due to decrease of BBB disruption by improving ultrastructural damage of capillary endothelium cells, enhancing basal membrane, and inhibiting AQP4 and its mRNA over-expression. The data of the present study provide a new possible approach for acute stroke therapy by administration of CGRP.     

Continuous oral administration of atorvastatin ameliorates brain damage after transient focal ischemia in rats

Aims

Pre-treatment with statins is known to ameliorate ischemic brain damage after experimental stroke, and is independent of cholesterol levels. We undertook pre- vs post-ischemic treatment with atorvastatin after focal cerebral ischemia in rats.

Main methods

Male Sprague–Dawley rats underwent transient 90-min middle cerebral artery occlusion (MCAO). Atorvastatin (20 mg/kg/day) or vehicle was administered orally. Rats were divided into vehicle-treated, atorvastatin pre-treatment, atorvastatin post-treatment, and atorvastatin continuous-treatment groups. In the pre-treatment, rats were given atorvastatin or vehicle for 7 days before MCAO. In the post-treatment, rats received atorvastatin or vehicle for 7 days after MCAO. Measurement of infarct volume, as well as neurological and immunohistochemical assessments, were done 24 h and 7 days after reperfusion.

Key findings

Each atorvastatin-treated group demonstrated significant reductions in infarct and edema volumes compared with the vehicle-treated group 24 h after reperfusion. Seven days after reperfusion, infarct volumes in the post-treatment group and continuous-treatment group (but not the pre-treatment group) were significantly smaller than in the vehicle-treated group. Only the continuous-treatment group had significantly improved neurological scores 7 days after reperfusion compared with the vehicle group. Post-treatment and continuous-treatment groups had significantly decreased lipid peroxidation, oxidative DNA damage, microglial activation, expression of tumor necrosis factor-alpha, and neuronal damage in the cortical ischemic boundary area after 7 days of reperfusion.

Significance

These results suggest that continuous oral administration (avoiding withdrawal) with statins after stroke may reduce the extent of post-ischemic brain damage and improve neurological outcome by inhibiting oxidative stress and inflammatory responses.     

Effect of combined therapy with hyperbaric oxygen and antioxidant on infarct volume after permanent focal cerebral ischemia

The aim of the present study was to evaluate the efficiency of combination of hyperbaric oxygen (HBO) and an antioxidant on permanent focal cerebral ischemia. Male Wistar rats underwent permanent middle cerebral artery occlusion (MCAO). Then, animals were randomly assigned to one of four groups: the control group (n=9) received no treatment, HBO group (n=9) was treated for 90 min at 2.5 absolute atmosphere for 3 days, the U-74389G group (n=8) received single U-74389G injection (3 mg/kg), the HBO + U-74389G group (n=8) received both HBO and U-74389G treatments. Treatments were initiated within the first 10 min after MCAO. After 3 days, the infarct volumes in rat brains were measured. The infarct ratios were 25.6+/-6.5 % for the control group, 21.9+/-6.4 % for the HBO group, 15.7+/-5.7 % for U-74389G group and 12.5+/-3.8 % for HBO + U74389G group. The infarct volumes were significantly reduced in rats treated with U-74389G (p<0.05) and combination therapy (p<0.05). HBO failed to reduce infarct volume significantly. We concluded that 1) U-74389G is more beneficial than HBO on permanent MCAO in rats, and 2) a combined therapy failed to significantly improve infarct volume more than either single treatment.     

局部亚低温联合硫酸镁对局灶性脑缺血大鼠的保护作用

目的:探讨联合应用局部亚低温(32-35℃)及硫酸镁对局灶性脑缺血大鼠的保护作用及其可能机制。方法:通过线栓法建立大鼠大脑中动脉阻塞(MCAO)模型,将40只雄性Wistar大鼠随机分为假手术组、常温组、亚低温组、硫酸镁组、亚低温+硫酸镁组,每组8例,采用Longa神经功能评分、TTC染色、TUNEL技术,检测和比较各组脑缺血后大鼠的神经功能、脑梗死体积、凋亡细胞数。结果:与常温组相比,亚低温组与亚低温+硫酸镁组的梗死体积、神经功能评分、凋亡细胞数均明显降低,差异有显著意义(P0.05);而与亚低温组相比,亚低温+硫酸镁组局灶脑缺血大鼠的脑梗死体积、神经功能评分、凋亡细胞数均显著减少,差异有显著意义(P0.05)。结论:与单独应用亚低温相比,局部亚低温与硫酸镁联合应用,对局灶性脑缺血大鼠可发挥更有效的脑保护作用,其机制可能与抑制脑缺血后凋亡有关。     

Effects of Acupuncture at GV20 and ST36 on the Expression of Matrix Metalloproteinase 2, Aquaporin 4, and Aquaporin 9 in Rats Subjected to Cerebral Ischemia/Reperfusion Injury

Background/Purpose

Ischemic stroke is characterized by high morbidity and mortality worldwide. Matrix metalloproteinase 2 (MMP2), aquaporin (AQP) 4, and AQP9 are linked to permeabilization of the blood-brain barrier (BBB) in cerebral ischemia/reperfusion injury (CIRI). BBB disruption, tissue inflammation, and MMP/AQP upregulation jointly provoke brain edema/swelling after CIRI, while acupuncture and electroacupuncture can alleviate CIRI symptoms. This study evaluated the hypothesis that acupuncture and electroacupuncture can similarly exert neuroprotective actions in a rat model of middle cerebral artery occlusion (MCAO) by modulating MMP2/AQP4/APQ9 expression and inflammatory cell infiltration.

Methods

Eighty 8-week-old Sprague-Dawley rats were randomly divided into sham group S, MCAO model group M, acupuncture group A, electroacupuncture group EA, and edaravone group ED. The MCAO model was established by placement of a suture to block the middle carotid artery, and reperfusion was triggered by suture removal in all groups except group S. Acupuncture and electroacupuncture were administered at acupoints GV20 (governing vessel-20) and ST36 (stomach-36). Rats in groups A, EA, and ED received acupuncture, electroacupuncture, or edaravone, respectively, immediately after MCAO. Neurological function (assessed using the Modified Neurological Severity Score), infarct volume, MMP2/AQP4/AQP9 mRNA and protein expression, and inflammatory cell infiltration were all evaluated at 24 h post-reperfusion.

Results

Acupuncture and electroacupuncture significantly decreased infarct size and improved neurological function. Furthermore, target mRNA and protein levels and inflammatory cell infiltration were significantly reduced in groups A, EA, and ED vs. group M. However, MMP2/AQP levels and inflammatory cell infiltration were generally higher in groups A and EA than in group ED except MMP2 mRNA levels.

Conclusions

Acupuncture and electroacupuncture at GV20 and ST36 both exercised neuroprotective actions in a rat model of MCAO, with no clear differences between groups A and EA. Therefore, acupuncture and electroacupuncture might find utility as adjunctive and complementary treatments to supplement conventional therapy for ischemic stroke.     

Testosterone increases neurotoxicity of glutamate in vitro and ischemia-reperfusion injury in an animal model.

Increasing evidence has demonstrated striking sex differences in the outcome of neurological injury. Whereas estrogens contribute to these differences by attenuating neurotoxicity and ischemia-reperfusion injury, the effects of testosterone are unclear. The present study was undertaken to determine the effects of testosterone on neuronal injury in both a cell-culture model and a rodent ischemia-reperfusion model. Glutamate-induced HT-22 cell-death model was used to evaluate the effects of testosterone on cell survival. Testosterone was shown to significantly increase the toxicity of glutamate at a 10 microM concentration, whereas 17beta-estradiol significantly attenuated the toxicity at the same concentration. In a rodent stroke model, ischemia-reperfusion injury was induced by temporal middle cerebral artery occlusion (MCAO) for 1 h and reperfusion for 24 h. To avoid the stress-related testosterone reduction, male rats were castrated and testosterone was replaced by testosterone pellet implantation. Testosterone pellets were removed at 1, 2, 4, or 6 h before MCAO to determine the duration of acute testosterone depletion effects on infarct volume. Ischemic lesion volume was significantly decreased from 239.6 +/- 25.9 mm(3) in control to 122.5 +/- 28.6 mm(3) when testosterone pellets were removed at 6 h before MCAO. Reduction of lesion volume was associated with amelioration of the hyperemia during reperfusion. Our in vitro and in vivo studies suggest that sex differences in response to brain injury are partly due to the consequence of damaging effects of testosterone.     

Propofol Protects Against Focal Cerebral Ischemia via Inhibition of Microglia-Mediated Proinflammatory Cytokines in a Rat Model of Experimental Stroke

Ischemic stroke induces microglial activation and release of proinflammatory cytokines, contributing to the expansion of brain injury and poor clinical outcome. Propofol has been shown to ameliorate neuronal injury in a number of experimental studies, but the precise mechanisms involved in its neuroprotective effects remain unclear. We tested the hypothesis that propofol confers neuroprotection against focal ischemia by inhibiting microglia-mediated inflammatory response in a rat model of ischemic stroke. Sprague-Dawley rats were subjected to middle cerebral artery occlusion (MCAO) for 2 h followed by 24 h of reperfusion. Propofol (50 mg/kg/h) or vehicle was infused intravenously at the onset of reperfusion for 30 minutes. In vehicle-treated rats, MCAO resulted in significant cerebral infarction, higher neurological deficit scores and decreased time on the rotarod compared with sham-operated rats. Propofol treatment reduced infarct volume and improved the neurological functions. In addition, molecular studies demonstrated that mRNA expression of microglial marker Cd68 and Emr1 was significantly increased, and mRNA and protein expressions of proinflammatory cytokines tumor necrosis factor-α, interleukin-1β and interleukin-6 were augmented in the peri-infarct cortical regions of vehicle-treated rats 24 h after MCAO. Immunohistochemical study revealed that number of total microglia and proportion of activated microglia in the peri-infarct cortical regions were markedly elevated. All of these findings were ameliorated in propofol-treated rats. Furthermore, vehicle-treated rats had higher plasma levels of interleukin-6 and C-reactive protein 24 h after MCAO, which were decreased after treatment with propofol. These results suggest that propofol protects against focal cerebral ischemia via inhibition of microglia-mediated proinflammatory cytokines. Propofol may be a promising therapeutic agent for the treatment of ischemic stroke and other neurodegenerative diseases associated with microglial activation.     

Does nitric oxide contribute to iron-dependent brain injury after experimental cerebral ischaemia?

Experimental and clinical data suggest that iron has a key role in cerebral ischaemia. We measure infarct volume and analyse the nitric oxide responses to brain injury in rat stroke model after increased oral iron intake. Permanent middle cerebral artery occlusion (MCAO) was performed in a group of 20 male Wistar rats, 10 of which were fed with a control diet and 10 of which were fed with iron-enriched diet containing 2.5% carbonyl iron for 9 weeks. L-arginine and nitric oxide metabolites were determined in blood samples before and at 2, 6, 8 and 48 h after MCAO. Infarct volume, thiobarbituric acid reaction substances (TBARS) and tissue iron were measured at 48 h. Infarct volume was 66% greater in the iron-fed rats than in the control group. Iron-fed animals showed significantly higher levels of TBARS. Liver iron stores (3500 +/- 199 vs 352 +/- 28 microg Fe/g, p<0.0001) but not brain iron stores (131 +/- 11 vs 139 +/- 8 microg Fe/g, p=0.617), were significantly higher in the iron-fed group. L-arginine levels were slightly lower in iron-fed rats and decreased significantly in both groups at 6 and 8 hours after MCAO. The levels of the stable end products of NOS (NOx = nitrite + nitrate) were significantly higher in iron-fed rats before MCAO (16.2 +/- 2.2 vs. 9.6 +/- 0.8 micromol x L(-1), p<0.05), with a further increase during the six first hours after MCAO in both groups. These results suggest that the iron overload that increases both superoxide and nitric oxide production leads to peroxynitrite formation, thus enhancing brain damage.     

毛蕊异黄酮通过抑制calpain-1的表达发挥抗脑缺血再灌注损伤的研究

目的:探讨毛蕊异黄酮抗脑缺血再灌注损伤的作用是否与抑制calpain-1的表达有关。方法:将SD大鼠随机分为假手术组、模型组以及药物组,采用线栓法建立大鼠大脑中动脉阻断(MCAO)模型,于缺血再灌注前30 min腹腔注射给予20 mg/kg毛蕊异黄酮或等体积的溶剂。再灌注24 h后,行神经功能学评分、脑梗死面积以及神经元凋亡检测;再灌注12 h、24 h时,采用免疫组化和蛋白印迹技术检测大鼠脑皮层calpain-1的表达。结果:与假手术组大鼠比较,MCAO模型组大鼠再灌注24 h后神经功能学评分、梗死面积、神经元凋亡率及calpain-1的表达均明显升高(P0.05),而毛蕊异黄酮能够降低模型组大鼠再灌注24 h后神经功能学评分、梗死面积、神经元凋亡率以及calpain-1的表达(P0.05)。结论:毛蕊异黄酮可能通过抑制calpain-1的表达发挥抗脑缺血再灌注损伤作用。     

Lipocalin‐2 released in response to cerebral ischaemia mediates reperfusion injury in mice

Thrombolysis remains the only effective therapy to reverse acute ischaemic stroke. However, delayed treatment may cause serious complications including hemorrhagic transformation and reperfusion injury. The level of lipocalin‐2 (LCN2) is elevated in the plasma of ischaemic stroke patients, but its role in stroke is unknown. Here, we show that LCN2 was acutely induced in mice after ischaemic stroke and is an important mediator of reperfusion injury. Increased levels of LCN2 were observed in mouse serum as early as 1 hr after transient middle cerebral artery occlusion (tMCAO), reaching peak levels at 23 hrs. LCN2 was also detected in neutrophils infiltrating into the ipsilateral hemisphere, as well as a subset of astrocytes after tMCAO, but not in neurons and microglia. Stroke injury, neurological deficits and infiltration of immune cells were markedly diminished in LCN2 null mice after tMCAO, but not after permanent MCAO (pMCAO). In vitro, recombinant LCN2 protein induced apoptosis in primary cultured neurons in a dose‐dependent manner. Our results demonstrate that LCN2 is a neurotoxic factor secreted rapidly in response to cerebral ischaemia, suggesting its potential usage as an early stroke biomarker and a novel therapeutic target to reduce stroke‐reperfusion injury.