百日咳毒素减轻实验性自身免疫性脑脊髓炎小鼠模型炎症反应的机制研究

目的探讨百日咳毒素(PTx)对实验性自身免疫性脑脊髓炎(EAE)小鼠模型的作用及机制。方法将C57BL/6小鼠随机分为正常对照组、EAE组和PTx治疗组,每组12只。EAE组和PTx治疗组用MOG 35-55诱导EAE模型。PTx组于建模后第7天给予腹腔注射1000 ng的PTx。随后观察两组临床症状,组织学染色评价炎症反应和脱髓鞘改变,并观察VEGF和血管新生现象,并用Western blot检测脊髓VEGF和Collagen IV的整体水平。在体外,用PTx刺激原代培养的神经元,评价PTx对在体外对神经元VEGF表达的影响。结果 PTx能减轻EAE模型的炎症反应和脱髓鞘改变,在大脑PTx将炎症评分从(3.4±0.55)分降至(1.2+0.45)分,P0.01;脱髓鞘评分从(3.6+0.55)分降至(1.4+0.55)分,P0.01。在脊髓PTx将炎症评分从(3.6+0.55)分降至(1.0+0.71)分,P0.01;脱髓鞘评分从(4.2+0.84)分降至(1.4+0.55)分,P0.01。Western blot检测显示,与正常对照组比较,EAE组VEGF下降49.0%(P0.01),Collagen IV下降36.0%(P0.01);PTx治疗后,与EAE组比较,VEGF上升59.5%(P0.01),Collagen IV上升45.0%(P0.01)。体外实验显示,PTx治疗24 h后VEGF在神经元上的表达增加,与正常对照相比,100 ng/ml组上调34.6%(P0.01),400 ng/ml组上调76.9%(P0.01)。结论 PTx可上调神经元内源VEGF的表达和血管新生现象,继而在EAE模型中起保护作用。     

钙内流与癫痫

钙内流与阵发性去极化漂移、神经元同步放电和抑制性突融后电位形成有关,因而在癫痫发生中起重要的作用。过度的钙离子内流主要通过钙通道(包括电压依赖性钙通道和受体门控性钙通道)的改变得以实现,某些钙通道拮抗剂对癫痫的治疗有一定的临床价值。     

姜黄素对AMPA/KA受体介导大鼠海马神经元钙内流的影响

目的 探讨姜黄素对α-氨基-3-羧基-5-甲基异恶唑-4-丙酸(AMPA)/海人酸(KA)受体介导大鼠海马神经元钙内流的影响.方法 选用胚胎17dSD鼠分离海马,离体培养海马神经元,借助活体钙荧光染色和激光共聚焦钙成像技术观察100μmol/LKA刺激海马神经元内钙的变化,不同浓度(5、10、15、30、50 μmol/L)姜黄素预孵育海马神经元30min对100μmol/L KA刺激下细胞内钙变化的影响,15 μmol/L姜黄素对不同浓度(10、30、50、100、200、300 μmol/L)KA刺激海马神经元内钙变化的影响.应用钴染色技术观察(30、100 μmol/L KA)刺激后海马神经元钴阳性染色细胞变化.姜黄素预孵育30min对KA刺激导致钴阳性染色细胞变化的影响.结果 不同浓度姜黄素预孵育30 min均可以明显缓解100 μmol/L或30 μmol/L KA导致的细胞内钙升高程度.差异均有统计学意义(P<0.05),其中15 μmol/L姜黄素作用最为明显.30μmol/L或100 μmol/LKA刺激均可以引起海马神经元钴染色阳性细胞增加,15 μmol/L姜黄素预处理30 min后明显减少钴染色阳性细胞,差异有统计学意义(P<0.05),而其他浓度(5 μmol/L或30 μmol/L)姜黄素未见明显影响.结论 一定浓度的姜黄素可以影响AMPA/KA受体介导大鼠海马神经元钙内流.这可能是姜黄素抗癫痫作用的一个机制.     

抑制容量性钙内流对U87侵袭及迁移力的影响

目的探讨抑制容量性钙内流(CCE)对胶质瘤U87细胞侵袭及迁移力的影响。方法采用免疫印迹(Western blot)检测基质相互作用分子1(STIM1)RNA干涉(RNAi)后U87细胞STIM1表达变化;活细胞钙成像检测CCE抑制剂(SKF-96365)处理及STIM1 RNAi后U87细胞CCE的改变;细胞划痕和Transwell侵袭实验检测CCE对U87细胞侵袭及迁移力的影响。结果 SKF-96365处理及STIM1RNAi均能明显抑制U87细胞的CCE(P0.05);与对照组比较,抑制CCE后U87细胞迁移及侵袭能力明显降低(P0.05)。结论抑制CCE可有效抑制U87细胞迁移及侵袭力,其机制可能与调控细胞内钙稳态相关。     

爪蟾顶盖神经元的大微抑制性突触后电流（mIPSCs）依赖从钙储池释放的钙？

目的　用影响突触前细胞外钙内流及或细胞内钙储池释放钙的措施,研究大、小微抑制性突触后电流(mIPSCs)的一些特性。方法　采用盲法电压钳全细胞记录技术。结果　①无钙液中加或不加EGTA(200mmol/L)灌流时,均可逆地使大mIPSCs较正常含钙液灌流时明显增多,小mIPSCs的平均频率明显下降;增加细胞外液钙(4mmol/L)浓度,小mIPSCs的频率增加,大mIPSCs变化不明显。Cd2+ (100μmol/L),仅轻度降低小mIPSCs平均频率至对照组的(87. 3±30. 0)% (n=13)。②carbachol(100μmol/L)使小mIPSCs增加至对照组的315. 63%。然而,无钙液中加carbachol,小mIPSCs不增加。含钙液和无钙液中加carbachol均可使大mIPSCs的比例增加。③Thapsigargin(8μmol/L)可使小mIPSCs的平均频率增加至对照组的(132. 1±27. 4)%。④U73122 (40μmol/L)可使小mIPSCs的平均频率降低至对照组的(74. 7±29. 6)%。⑤Procaine(2mmol/L)及咖啡因(10mmol/L)均可降低小mIPSC的平均频率。较高浓度的兰尼定(30 50μmol/L)也降低小mIPSCs的频率。结论　改变灌流液的钙浓度,小mIPSCs的频率受到明显影响。大mIPSCs的出现或增加主要与钙储池释放Ca2+的机制有关。     

意向运动疗法与脑缺血后损伤可塑性关系的研究

缺血性脑卒中后通常伴有肢体功能的障碍,康复锻炼能够影响相关蛋白的表达,从而提高大脑的可塑性,进而促进肢体功能的恢复;miRNA在蛋白质的表达过程中起着重要的调节作用.意向运动是一种主动运动方式,它能最大限度的调动患者的主观能动性,以获得最佳的康复效果.本文主要就脑可塑性机制、意向运动过程中蛋白质变化、缺血性卒中后miRNA表达变化以及意向运动促进脑可塑性的可能分子生物学机制进行综述.     

急性缺血性脑卒中患者血清钙调蛋白水平检测及意义

目的 研究急性缺血性脑卒中患者血清钙调蛋白(CaM)水平及临床意义.方法 采用病例对照研究,选取92例急性缺血性脑卒中患者,入院后按照美国国立卫生研究院卒中量表评分(NIHSS评分)分为NIHSS≤5分组和NIHSS ＞5分组.选择45例健康体检者作为对照组.运用酶联免疫吸附法(ELISA法)检测所有入组者血清CaM水平,并分析CaM水平与NIHSS评分的相关性.结果 (1)急性缺血性脑卒中患者血清CaM水平明显高于健康对照组,有统计学意义(t =0.296,P＜0.01);(2) NIHSS＞5分组患者血清CaM水平显著高于NIHSS≤5分组有统计学意义(=2.417,P＜0.05);(3)急性缺血性脑卒中患者血清CaM水平与NIHSS评分呈正相关(r=0.318,P=0.002).结论 急性缺血性脑卒中患者血清CaM水平明显增高,可作为判定病情严重程度的重要指标.     

细胞毒素相关蛋白A阳性的幽门螺杆菌感染与缺血性脑卒中及其亚型的关系的Meta分析

目的探讨细胞毒素相关蛋白A(CagA)阳性的幽门螺杆菌感染与缺血性脑卒中(IS)及其亚型的关系。方法检索MEDLINE、EMbase、Cochrance Collaboration database、中国期刊全文数据库及维普中文科技期刊全文数据库,纳入相关的病例-对照研究的文献。采用RevMan5.0软件进行Meta分析;同时进行文献的异质性检验、敏感性分析以及发表偏倚分析。结果共纳入8篇相关文献,其中与大动脉硬化性脑卒中(LAA)相关文献5篇。Meta分析结果显示,CagA阳性菌株感染有增加IS发病风险的趋势(OR=2.31,95%CI:1.89～2.82,P<0.01),CagA阴性菌株感染亦与IS相关(OR=0.57,95%CI:0.47～0.70,P<0.01)。进一步分析显示,CagA阳性及阴性菌株均与LAA亚型相关(OR=2.87,95%CI:2.19～3.77,P<0.01;OR=0.51,95%CI:0.39～0.67,P<0.01)。对纳入的各文献中混杂因素进一步校正后再次合并OR总效应值显示,CagA阳性菌株感染与IS及LAA仍存在相关性(OR=2.36,95%CI:1.84～3.02,P<0.01;OR=3.10,95%CI:2.29～4.19,P<0.01)。异质性检验示CagA阳性菌株与IS及其亚型的研究不存在异质性(均I2=0%)。敏感性分析示结果较稳定。漏斗图分析示存在发表偏倚。结论 CagA阳性的幽门螺杆菌感染是IS的危险因素之一,尤其是LAA亚型。     

肉毒杆菌毒素A在脑卒中后肢体痉挛中的应用

Ａ型肉毒毒素是近年来治疗脑卒中后肢体痉挛的一个新进展,国外已有多项试验证实了它的有效性及安全性,国内的研究也正在展开,本文综述了肉毒毒素的特点及其在脑卒中后肢体痉挛治疗的临床应用状况。     

肌苷促进脑梗死后神经功能重塑的实验研究

目的探讨腹腔注射肌苷对脑梗死后运动功能缺损的治疗作用.方法成年雄性SD大鼠72只,随机分为肌苷治疗组、对照组和假手术组,采用电凝法建立MCAO(大脑中动脉闭塞,Middle Cerebral Artery Occlusion)的动物模型,腹腔注射肌苷后使用HPLC(高效液相色谱,High-performance Liquid Chroma-tography)技术测定梗死灶周围肌苷的浓度,并用免疫组织化学染色技术观察SYP(突触素,Synaptophysin)、MAP-2(微管相关蛋白-2,Microtubule-associated protein-2)和CAP-23(细胞骨架相关蛋白-23,Cytoskeleton-associated protein-23)的表达情况,同时评估大鼠运动功能的恢复情况.结果腹腔注射肌苷能使梗死灶周围的肌苷浓度升高近3倍,并能增强该部位SYP、MAP-2和CAP-23的表达,促进大鼠运动功能的恢复.结论腹腔注射肌苷能促进大鼠脑梗死后的神经重塑,一定程度地改善脑梗死后的运动功能缺损.     

Nimodipine in acute ischemic stroke: a double-blind controlled study

Nimodipine (BAY e 9736), a new dihydropyridine derivative, has been shown to reduce neurological deficits and mortality induced by acute cerebral ischemia in experimental studies. We investigated the effects of this calcium antagonist in patients with acute ischemic stroke through a randomized, double-blind, parallel-designed trial in which nimodipine was compared with placebo. Forty-one of 54 screened cases were found to fulfil the inclusion criteria (sudden occurrence of a focal neurological deficit secondary to an acute ischemic event in the carotid area diagnosed after a complete neurological work-up) and entered the study. Nineteen of them were treated with nimodipine (40 mg t.i.d. administered for 28 days) and the remaining 22 with placebo, given in identical tablets. In all patients the treatment started within 12 h after the onset of the symptoms. Course and intensity of the neurological deficit were evaluated by the Mathew Scale (slightly modified). Forty patients concluded the trial. Nimodipine was withdrawn in one case following the occurrence of a skin rash whose causative relation with the test drug could not be clarified. Altogether, however, nimodipine was well tolerated and no severe cardiovascular adverse reactions were observed. In terms of efficacy, the scores obtained by the Mathew Scale showed a higher rate of improvement on nimodipine than on placebo, thus indicating that patients receiving the latter drug did not fare as well as those receiving the test medication. Our data suggest that nimodipine may be beneficial in the treatment of acute stroke.     

Acetylated α-tubulin alleviates injury to the dendritic spines after ischemic stroke in mice

Background and Aim

Functional recovery is associated with the preservation of dendritic spines in the penumbra area after stroke. Previous studies found that polymerized microtubules (MTs) serve a crucial role in regulating dendritic spine formation and plasticity. However, the mechanisms that are involved are poorly understood. This study is designed to understand whether the upregulation of acetylated α-tubulin (α-Ac-Tub, a marker for stable, and polymerized MTs) could alleviate injury to the dendritic spines in the penumbra area and motor dysfunction after ischemic stroke.

Methods

Ischemic stroke was mimicked both in an in vivo and in vitro setup using middle cerebral artery occlusion and oxygen–glucose deprivation models. Thy1-YFP mice were utilized to observe the morphology of the dendritic spines in the penumbra area. MEC17 is the specific acetyltransferase of α-tubulin. Thy1 Cre^ERT2-eYFP and MEC17^fl/fl mice were mated to produce mice with decreased expression of α-Ac-Tub in dendritic spines of pyramidal neurons in the cerebral cortex. Moreover, AAV-PHP.B-DIO-MEC17 virus and tubastatin A (TBA) were injected into Thy1 Cre^ERT2-eYFP and Thy1-YFP mice to increase α-Ac-Tub expression. Single-pellet retrieval, irregular ladder walking, rotarod, and cylinder tests were performed to test the motor function after the ischemic stroke.

Results

α-Ac-Tub was colocalized with postsynaptic density 95. Although knockout of MEC17 in the pyramidal neurons did not affect the density of the dendritic spines, it significantly aggravated the injury to them in the penumbra area and motor dysfunction after stroke. However, MEC17 upregulation in the pyramidal neurons and TBA treatment could maintain mature dendritic spine density and alleviate motor dysfunction after stroke.

Conclusion

Our study demonstrated that α-Ac-Tub plays a crucial role in the maintenance of the structure and functions of mature dendritic spines. Moreover, α-Ac-Tub protected the dendritic spines in the penumbra area and alleviated motor dysfunction after stroke.     

糖尿病与进展性缺血性脑卒中相关性研究

目的 探讨糖尿病与进展性缺血性脑卒中的相关性.方法 我院神经内科2006-05～2009-05收治的缺血性脑卒中患者130例为研究对象,其中伴糖尿病患者64例为糖尿病组,非糖尿病患者66例为非糖尿病组,比较2组发展为进展性缺血性脑卒中的差异.结果 糖尿病组64例中诊断为进展性卒中41例,占64.1%;非糖尿病组66例诊...     

Elevated bilirubin after acute ischemic stroke linked to the stroke severity

Background

Our previous study demonstrated that the level of serum bilirubin after acute ischemic stroke (AIS) was correlated to the severity of stroke, also there has the evidence of hyperbilirubinemia prevalent in AIS. We aimed to identify the exact change of bilirubin in the early phase of AIS, and study if this kind of change linked to the severity of stroke.

Methods

Bilirubin and other biochemical indexes were measured in 608 AIS patients and 188 transient ischemic attack (TIA) patients which set as the control group. National Institutes of Health Stroke Scale (NIHSS) scores were assessed simultaneously with blood collection. First, the level of bilirubin and its distribution were compared between the AIS and control group. According to a cut-off point, we next analyzed the impacted factors of elevated bilirubin including the direct bilirubin (Dbil) and total bilirubin (Tbil), especially the correlation between elevated bilirubin and the severity of stroke. Finally, we compared the difference of concentration and percent of elevated bilirubin among the Oxford Community Stroke Project (OCSP) subtypes.

Results

The level of serum Dbil and Tbil was significantly higher in the AIS group than that in the TIA group. Different distribution was observed between the two groups, which manifested as the percent of low bilirubin level group was lower while high level group was higher in AIS than that in TIA, the p value were 0.043 and 0.078 in Dbil and Tbil, respectively. When the cut-off point of elevated bilirubin was selected as Dbil ≥ 6.84 μmol/L and Tbil ≥ 22.2 μmol/L, we found that both NIHSS score and relative severity of AIS were significantly associated with elevated bilirubin whenever in Dbil or Tbil, so did the OCSP subtypes. This trend was still maintained by multivariable logistic regression analysis adjust for relative influence factors. In regard of OCSP subtypes, the highest level of bilirubin was found in TACI, so did the highest rate of elevated bilirubin.

Conclusion

The serum levels of Dbil and Tbil were increased after AIS, which linked to the severity of stroke.     

Predictors of outcome after acute ischemic stroke

The aim of this study was to determine which variables should be the predictors for clinical outcome at discharge and sixth month after acute ischemic stroke. METHODS: Two hundred and sixty-six consecutive patients, each with an acute ischemic cerebrovascular disease, were evaluated within 24 h of symptom onset. We divided our patients into two groups; 1 - Independent (Rankin scale RS < or = 2) and, 2 - Dependent (RS>3) and death. Baseline characteristics, clinical variables, risk factors, infarct subtypes and radiologic parameters were analyzed. RESULTS: Canadian Neurological Scale (CNS) on admission <6.5 [odds ratio (OR) 22] and posterior circulation infarction (OR 4.2) were associated with a poor outcome at discharge from hospital whereas only a CNS score <6.5 (OR 14) was associated with a poor outcome at 6 months. CONCLUSIONS: Severity of neurologic deficit is the most important indicator for clinical outcome in acute ischemic stroke both at short-term and at sixth month, whereas posterior circulation infarction also predicts a poor outcome at discharge.     

Inhibiting leukocyte-endothelial cell interactions by Chinese medicine Tongxinluo capsule alleviates no-reflow after arterial recanalization in ischemic stroke

Aims

Despite successful vascular recanalization in stroke, one-fourth of patients have an unfavorable outcome due to no-reflow. The pathogenesis of no-reflow is fully unclear, and therapeutic strategies are lacking. Upon traditional Chinese medicine, Tongxinluo capsule (TXL) is a potential therapeutic agent for no-reflow. Thus, this study is aimed to investigate the pathogenesis of no-reflow in stroke, and whether TXL could alleviate no-reflow as well as its potential mechanisms of action.

Methods

Mice were orally administered with TXL (3.0 g/kg/d) after transient middle cerebral artery occlusion. We examined the following parameters: neurological function, no-reflow, leukocyte-endothelial cell interactions, HE staining, leukocyte subtypes, adhesion molecules, and chemokines.

Results

Our results showed stroke caused neurological deficits, neuron death, and no-reflow. Adherent and aggregated leukocytes obstructed microvessels as well as leukocyte infiltration in ischemic brain. Leukocyte subtypes changed after stroke mainly including neutrophils, lymphocytes, regulatory T cells, suppressor T cells, helper T type 1 (Th1) cells, Th2 cells, B cells, macrophages, natural killer cells, and dendritic cells. Stroke resulted in upregulated expression of adhesion molecules (P-selectin, E-selectin, and ICAM-1) and chemokines (CC-chemokine ligand (CCL)-2, CCL-3, CCL-4, CCL-5, and chemokine C-X-C ligand 1 (CXCL-1)). Notably, TXL improved neurological deficits, protected neurons, alleviated no-reflow and leukocyte-endothelial cell interactions, regulated multiple leukocyte subtypes, and inhibited the expression of various inflammatory mediators.

Conclusion

Leukocyte-endothelial cell interactions mediated by multiple inflammatory factors are an important cause of no-reflow in stroke. Accordingly, TXL could alleviate no-reflow via suppressing the interactions through modulating various leukocyte subtypes and inhibiting the expression of multiple inflammatory mediators.     

Young ischemic stroke in Tunisia: a multicentric study

Purpose: There is wanting data regarding young ischemic stroke in developing countries, especially in Tunisia. The purpose of this study was to investigate risk factors and etiologies of young ischemic stroke in Tunisian and make a comparison with previous reports. Materials and methods: A total of 102 young ischemic stroke patients (15–45 years old) were admitted, between January 1996 and August 2007, to 11 departments of internal medicine in different Tunisian hospitals. The risk factors for stroke were documented and assessed. Diagnosis workup consisted of anamnesis, complete physical examination and extensive laboratory, radiologic, immunologic, neurologic and cardiologic examination. Stroke etiologies were classified according the Trial of ORG 10172 in acute stroke treatment. Results: There were 42 men (41.2%) and 60 women (58.89%) with a mean age at onset of 35.7 years. As regards stroke subtype, large-artery atherosclerosis was diagnosed in 6.9% of cases, cardioembolism in 11.8%, small-vessel occlusion in 8.8%, other determined etiology in 37.3% and undetermined etiology in 35.3%. Concerning the traditional risk factors, smoking (31.4%), hypertension and diabetes mellitus (12.7% for each one) and a family history of stroke (10.8%) were the most common. The mean follow-up period was 30.5 months. Conclusions: In our study, traditional risk factors were not-so-uncommon in young adults with ischemic stroke suggesting that prevention can go through controlling these factors. Stroke of other determined etiology was the most common among our patients, so that a broad and detailed diagnostic workup is crucial to puzzle out the etiology for more and better stroke prevention.     

缺血性脑卒中与血脂水平关系的初步分析

目的研究血脂异常与缺血性脑卒中及其各亚型的关系。方法收集516例缺血性脑卒中患者及131例非缺血性卒中对照组个体的血脂情况,包括血总胆固醇（TC）、甘油三酯（TG）、低密度脂蛋白（LDL）、高密度脂蛋白（HDL）等指标,对其进行统计分析比较。并将其中131例缺血性脑卒中患者根据TOAST分型标准进行分型,将各亚组的血脂水平与对照组再进行比较。结果缺血性脑卒中组的HDL水平明显低于对照组,TC／HDL比值明显高于较对照组。其余血脂成分,包括TC、TG、LDL则未发现存在显著差异。亚组比较中,大动脉粥样硬化性卒中（LAA）组、心源性脑栓塞（CE）组、小动脉闭塞性卒中或腔隙性卒中（SAO）组的HDL水平明显低干对照组,而他们的TC／HDL比值则明显高于对照。其余血脂成分比较未见统计学差异。其他原因卒中（SOE）及不明原因卒中（SUE）均显示与血脂各成分无相关性。结论HDL在缺血性脑卒中患者明显降低,其保护因素降低可能是卒中发生的原因之一。TC／HDL比值可作为衡量血脂异常与缺血性卒中关系的指标之一。需要进一步进行基于卒中分型基础上的更大样本量的研究。     

Percutaneous mastoid electrical stimulator alleviates autonomic dysfunction in patients with acute ischemic stroke

ABSTRACT

Background and Purpose: Poststroke prognosis is associated with autonomic status. The purpose of our study was to determine whether percutaneous mastoid electrical stimulator (PMES) can alleviate abnormal heart rate variability (HRV) and improve clinical outcome.

Methods: This prospective, randomized, double-blinded, placebo-controlled study enrolled a total of 140 patients with autonomic dysfunction within 3d after acute ischemic stroke. The patients were treated with PMES or sham stimulation once daily over a period of 2 weeks. HRV was primarily assessed by the fractal dimension (FD) at admission and 2 weeks. All patients were followed up for 3 months. The clinical outcome was death and major disability (modified Rankin Scale score≥ 3) at 3 months after acute ischemic stroke.

Results: FD of the 2-week treatment period increased in PMES groups. PMES can significantly alleviate abnormal HRV. The difference in FD of the 2-week treatment period between the PMES and sham groups was significant (1.14 ± 0.27 vs. 1.00 ± 0.23; P = 0.001). In fully adjusted models, PMES was associated with reduced 3-month mortality (adjusted odds ratio, 0.32; 95% confidence interval, 0.11–0.93; P = 0.036). No significant group differences were seen in three major disability and composite outcome (P > 0.05).

Conclusions: PMES was a safe, effective, and low-cost therapy to alleviate HRV and could significantly reduce mortality in the early recovery phase after acute ischemic stroke.