小鼠脑微血管内皮细胞在氧糖剥夺条件下的差异蛋白质组学研究

目的对参与血-脑屏障组成的脑微血管内皮细胞在模拟缺血性脑损伤条件下的蛋白质组学变化特征进行研究．为深入理解缺血性脑损伤过程中血-脑屏障应答的分子学机制提供新的数据。方法采用比较蛋白质组学方法．对正常培养条件和氧糖剥夺培养条件下的小鼠脑微血管内皮细胞系（bEnd．3细胞）进行研究．分析二者之间的差异蛋白质表达谱。结果于氧糖剥夺培养条件下，小鼠脑微血管内皮细胞系中共发现52个蛋白质点的表达发生改变，通过串联飞行质谱成功鉴定45个蛋白质点，其中表达水平上调的41个．下调4个。根据基因本体论功能注释结果，这些蛋白质的功能分为糖代谢与能量产生、抗氧化损伤、细胞骨架重排、信号转导、可变剪接、蛋白折叠与降解等。其中约1／4的蛋白质与以往文献报道相似．即与缺血、缺氧损伤密切相关。结论这些差异蛋白质的发现可促进对血-脑屏障病理生理机制的了解．为深入理解缺血性脑损伤的分子学机制提供了新的数据，并将为后续研究提供参考。     

脑肿瘤细胞的B2受体表达水平决定缓激肽诱发之血脑屏障开放的程度

目的：探讨缓激肽能选择性地诱发肿瘤血脑屏障开放的机理及影响此种开放的因素。方法：定性、定量评价不同类型大鼠脑肿瘤模型中B2受体表达的部位及水平，阐明它与缓激肽诱发的肿瘤血脑屏障开放程度的相关关系。结果：脑和肿瘤组织内的毛细血管内皮细胞不表达B2受体，而肿瘤细胞则表达大量的B2受体；B2受体表达水平与缓激肽诱发的肿瘤血脑屏障开放的程度呈显著的正相关关系。结论：脑肿瘤细胞的B2受体表达水平决定缓激肽诱发的血脑屏障开放的程度。     

大鼠C6脑胶质瘤血-脑屏障的硝酸镧示踪电镜研究

目的 研究C6脑胶质瘤不同区域血-脑屏障（BBB）的超微结构特征及血-脑肿瘤屏障（BBTB）通透性增高的途径。方法 运用成瘤3周SD大鼠C6脑胶质瘤动物模型。取肿瘤中心、肿瘤边缘、距肿瘤边界2mm以内及2mm以外的同侧大脑半球和对侧大脑半球脑皮质等五个部位的脑组织，采用外源性硝酸镧透射电镜示踪法，观察BBB／BBTB超微结构特征。结果 La^3＋在肿瘤中心与边缘部充填并通过血管内皮细胞之紧密连接至血管腔外一脑间质裂隙内。距肿瘤边界2mm以内的同侧大脑半球脑组织也有La^3＋渗出至局部基底膜，而其它观察脑区La^3＋则完全封闭于血管腔内；各部位脑血管内皮细胞胞浆内均无La^3＋；BM呈线状连续，管腔内无瘤细胞。结论 C6胶质瘤紧密连接开放导致BBTB通透性增高；C6胶质瘤诱导的屏障功能破坏随其至肿瘤距离的增加而减弱。     

Stathmin在脑胶质瘤血管内皮细胞中的表达及意义

目的 观察Stathmin及CD105在脑胶质瘤血管内皮细胞中的表达,探讨其在脑胶质瘤血管形成中的作用. 方法 用SP免疫组化法检测10例正常脑组织和78例脑胶质瘤血管内皮细胞中Stathmin和CD105蛋白表达,并通过检测肿瘤微血管密度(MVD)分析肿瘤血管形成.结果 正常脑组织血管内皮细胞中Stathmin和CD105均无表达,脑胶质瘤血管内皮细胞中二者均呈高表达;随着胶质瘤病理级别的增高,Stathmin和CD105表达上调,MVD值增高,Stathmin-MVD和CD105-MVD与脑胶质瘤病理分级均成正相关(r=0.912,P<0.05;P<0.936,P<0.05);且Stathmin-MVD和CD105-MVD之间也存在正相关(r=0.996,P<0.05). 结论 Stathmin与CD105在脑胶质瘤血管内皮细胞中均呈高表达,在肿瘤微血管形成过程起重要作用.     

MMP-2和MMP-9对人脑胶质瘤侵袭微生态系统中血瘤屏障的影响

目的 探讨基质金属蛋白酶MMP-2和MMP-9对人脑胶质瘤侵袭微生态系统的影响。方法 应用透射电镜观察脑胶质瘤侵袭微生态系统中血瘤屏障的超微结构改变；用免疫组织化学S-P法检测MMP-2和MMP-9在胶质瘤中的表达情况，并与透射电镜观察结果进行对照比较。结果 （1）电镜下可见瘤组织中多数微血管基膜的胶质膜侧出现大小不等，多少不一的虫蚀样空洞或血管周围间隙扩大；有的血管内皮细胞呈不同程度的增生，以出芽方式生成新的肿瘤性血管伴血浆渗出。（2）在胶质瘤瘤体内的基膜内皮细胞及瘤细胞可见MMP-2和MMP-9的表达，而在正常脑组织中的血管基膜内皮细胞只见MMP-2表达，瘤体组织中MMP-2染色阳性强者其相应的电镜下观察见基膜上虫蚀样空洞明显，而MMP-9染色阳性强者血管外周围间隙扩大明显。结论 MMP-2，MMP-9与人脑胶质瘤侵袭微生态系统中血瘤屏障的改造重建有关。     

TrkB-shiRNA质粒转染心脏微血管内皮细胞

背景：脑源性神经营养因子与其受体TrkB在心脏与骨骼肌内皮细胞存在表达,在心脏血管系统的发育中及骨骼肌缺血时能有效促进血管新生,但其促血管新生的细胞与分子机制尚不清楚。 目的：应用针对脑源性神经营养因子受体TrkB的shiRNA质粒对心脏微血管内皮细胞进行转染,观察TrkB 3种亚型的表达及心脏微血管内皮细胞的生长状况和形态,初步探讨脑源性神经营养因子-TrkB通路在心脏微血管内皮细胞中的调控作用。 方法：使用TrkB-shiRNA质粒转染大鼠心脏微血管内皮细胞,应用荧光实时定量PCR检测TrkB的3个亚型,TrkB-FL、TrkB-T1及TrkB-T2 mRNA的表达,并观察经转染的心脏微血管内皮细胞的增殖情况。 结果与结论：应用TrkB-shiRNA质粒转染心脏微血管内皮细胞后,TrkB 3种亚型在转染后的4 d内mRNA表达均下降(P < 0.05或P < 0.01),且经转染的心脏微血管内皮细胞的生长变慢。说明TrkB-shiRNA质粒可沉默心脏微血管内皮细胞的TrkB-FL、TrkB-T1及TrkB-T2的表达,且TrkB表达被抑制可能会影响心脏微血管内皮细胞的增殖。     

冰片促血脑屏障开放与一氧化氮含量改变的关系初探

目的 :探讨内皮细胞型一氧化氮合成酶 (e NOS)与冰片促血脑屏障 (BBB)开放作用的关系。方法 :采用链菌素亲生物素 -过氧化氢酶法 (S- P法 )染色和抗 e NOS抗体的免疫组化方法 ,观察正常与脑外伤情况下服用冰片后大鼠脑微血管内皮细胞 (EC)中 e NOS表达量的变化。结果 :脑外伤时 ,因 EC受损 ,e NOS表达减弱 ,冰片可增加生理、病理状态下大鼠脑微血管内皮细胞中 e NOS的表达量。结论 :NO与冰片促 BBB开放作用有一定联系     

胶质细胞对共培养的脑微血管内皮细胞增殖及功能的影响

目的 研究体外培养条件下，胶质细胞对脑微血管内皮细胞(BMEC)增殖及功能的影响。方法 模拟血-脑脊液屏障结构及内皮细胞、胶质细胞间相互影响的途径，建立内皮细胞与胶质细胞共培养模型，采用细胞计数、细胞活性检测、酶含量与细胞吞饮量测定对内皮细胞增殖和功能进行研究。结果 共培养和条件培养时，内皮细胞增殖能力减弱，细胞活性以及酶含量增高，细胞吞饮量则无明显变化。结论 胶质细胞可通过两种途径影响内皮细胞的生长。胶质细胞可诱导和维持微血管内皮细胞的脑表型，但并不能促进内皮细胞生长。     

脑胶质瘤动物模型中血脑屏障紧密连接的变化

目的 探讨脑胶质瘤对血脑屏障内皮细胞间紧密连接的影响及其分子机制. 方法 60只Wistar大鼠采用随机数字表法分为正常组(n=30)和脑肿瘤组(n=30),脑肿瘤组按常规方法 制作C6脑胶质瘤模型,造模后第21天行MRI检查后,取正常脑组织、肿瘤中心、肿瘤边缘及肿瘤远隔部位(边缘外2mm以上)组织,采用透射电镜观察两组大鼠血脑屏障紧密连接的超微结构特征,应用免疫组织化学染色和RT-PCR分别检测脑组织中紧密连接蛋白Claudia-5和Claudin-5 mRNA表达的变化. 结果 C6肿瘤细胞接种21 d后,MRI可见大鼠右脑形成肿瘤.在透射电镜下,正常脑组织微血管相邻内皮细胞间可见连续条带状的紧密连接,细胞间未见裂隙;在肿瘤中心组织,仅有22.23%微血管相邻内皮细胞间可见条带状紧密连接.其余内皮细胞间可见明显裂隙.部分内皮细胞间连接为缝隙连接;在肿瘤边缘组织中,有57.15%微血管内皮细胞间存在紧密连接,其余内皮细胞间局部可见裂隙.部分内皮细胞间可见缝隙连接.免疫组化染色显示正常脑组织微血管内皮细胞Claudin-5呈强阳性表达;肿瘤中心组织微血管内皮细胞Claudia-5表达呈阴性;肿瘤边缘微血管内皮细胞Claudin-5呈弱阳性表达.RT-PCR结果 示肿瘤中心Claudia-5 mRNA表达较肿瘤边缘、肿瘤远隔部位、正常组脑组织降低,差异均有统计学意义(P均<0.05). 结论 在胶质瘤的发生、发展过程中,胶质瘤细胞可以导致血脑屏障内皮细胞紧密连接蛋白Claudia-5的表达下降及血脑屏障紧密连接结构的破坏.     

高糖对心肌微血管内皮细胞表达神经调节蛋白1的影响

背景：糖尿病心肌病的发病机制尚不清楚,神经调节蛋白1具有促进血管再生等作用,糖尿病心肌病的发病是否和心肌微血管内皮细胞表达神经调节蛋白1下降有关值得进一步研究。 目的：观察高糖对心肌微血管内皮细胞表达神经调节蛋白1的影响。 方法：取生长良好的第2代心肌微血管内皮细胞,高糖组加入10 mmol/L的葡萄糖,高糖+胰岛素组加入10 mmol/L的葡萄糖及10-5 U/L的胰岛素,对照组正常培养。培养24 h后收集细胞,RT-PCR及Western blot方法检测神经调节蛋白1 mRNA和蛋白的表达。 结果与结论：与对照组比较,高糖组心肌微血管内皮细胞神经调节蛋白1 mRNA和蛋白的表达明显下降(P < 0.05);与高糖组比较,高糖+胰岛素组心肌微血管内皮细胞神经调节蛋白1 mRNA和蛋白的表达明显升高(P < 0.05),与对照组间差异无显著性意义(P > 0.05)。说明高糖可抑制心肌微血管内皮细胞表达神经调节蛋白1 mRNA和蛋白,而胰岛素可拮抗此作用。     

Epilepsy and anomalies of neuronal migration: MRI and clinical aspects

Neuronal migration disorders are the result of disturbed brain development. In such disorders, neurons are abnormally located. In diagnosing these conditions, magnetic resonance imaging is superior to any other imaging technique. This enables us to improve our knowledge of the clinical correlates of neuronal migration. With reference to migrational disorder, a retrospective study of all 303 patients with epileptic seizures referred for magnetic resonance imaging during a 3-year period was performed, 13 patients (aged 12-41, mean age 27) were identified. They represent 4.3% of the entire study group. Of the patients with known epilepsy, 6.7% and of the mentally retarded, 13.7% had migrational disorders. Four patients had schizencephaly as the dominant finding, one was classified as hemimegalencephaly, 2 had isolated heterotopias, and 6 had localized pachy- and/or poly-microgyria. The clinical pictures are complex. Ectopias of grey matter are recognised foci of epilepsy, but from an epileptological and a clinical viewpoint little attention has been given to these disorders. The present study shows that malmigration is not rare in epilepsy patients, especially not in the mentally retarded.     

Hepatic Considerations in the Use of Antiepileptic Drugs

Summary: Virtually all of the major antiepileptic drugs (AEDs) can cause hepatotoxicity, although fatal hepatic reactions are rare. The mechanisms, incidences, and risk profiles for such reactions differ from drug to drug. With carbamazepine and phenytoin, hepatotoxicity may be due to drug hypersensitivity. Although the profiles of patients at risk have not been well-defined for these two antiepileptic drugs, it would appear from reports in the literature that older adolescents and adults are at higher risk than children of developing serious or fatal hepatotoxicity. Once hepatotoxicity develops, mortality rates are 10–38% with phenytoin and 25% for carbamazepine. The risk profile for valproate fatal hepatotoxicity has been more clearly defined. Those at primary risk of fatal hepatic dysfunction are children under the age of 2 years who are receiving multiple anticonvulsants and also have significant medical problems in addition to severe epilepsy. The risk is considerably lower for patients over the age of 2 years on valproate monotherapy. In contrast to the risk profile with other AEDs, adults receiving valproate as monotherapy have the lowest risk of hepatotoxicity. Fatal hepatic dysfunction coincident with valproate may be the result of aberrant drug metabolism. Concomitant use of AEDs that induce microsomal P450 enzymes (e.g., phenytoin and phenobarbital) may enhance the production of a toxic metabolite, and hence the greater risk of hepatotoxicity with polypharmacy.     

Vascular Malformations and Epilepsy: Clinical Considerations and Basic Mechanisms

Summary: Vascular malformations (VMs) are associated with epilepsy. The natural history of the various VMs, clinical presentation, and tendency to provoke epilepsy determine treatment strategies. Investigations have probed the mechanisms of epileptogenesis associated with these lesions. Electrophysiologic changes are associated with epileptogenic cortex adjacent to VMs. Putative pathophysiologic mechanisms of epileptogenesis include neuronal cell loss, glial proliferation and abnormal glial physiology, altered neurotransmitter levels, free radical formation, and aberrant second messenger physiology.     

Classification of methods in transcranial Electrical Stimulation (tES) and evolving strategy from historical approaches to contemporary innovations

Transcranial Electrical Stimulation (tES) encompasses all methods of non-invasive current application to the brain used in research and clinical practice. We present the first comprehensive and technical review, explaining the evolution of tES in both terminology and dosage over the past 100 years of research to present day. Current transcranial Pulsed Current Stimulation (tPCS) approaches such as Cranial Electrotherapy Stimulation (CES) descended from Electrosleep (ES) through Cranial Electro-stimulation Therapy (CET), Transcerebral Electrotherapy (TCET), and NeuroElectric Therapy (NET) while others like Transcutaneous Cranial Electrical Stimulation (TCES) descended from Electroanesthesia (EA) through Limoge, and Interferential Stimulation. Prior to a contemporary resurgence in interest, variations of transcranial Direct Current Stimulation were explored intermittently, including Polarizing current, Galvanic Vestibular Stimulation (GVS), and Transcranial Micropolarization. The development of these approaches alongside Electroconvulsive Therapy (ECT) and pharmacological developments are considered. Both the roots and unique features of contemporary approaches such as transcranial Alternating Current Stimulation (tACS) and transcranial Random Noise Stimulation (tRNS) are discussed. Trends and incremental developments in electrode montage and waveform spanning decades are presented leading to the present day. Commercial devices, seminal conferences, and regulatory decisions are noted. We conclude with six rules on how increasing medical and technological sophistication may now be leveraged for broader success and adoption of tES.     

Carbamazepine Efficacy and Utilization in Children

Summary: Carbamazepine is effective for preventing partial and generalized tonic-clonic seizures in children. Although absence epilepsies are more common in children than adults, an estimated 80% of children with epilepsy have seizure types or epilepsies that are potentially responsive to carbamazepine. The differential diagnosis of ictal staring is an especially important issue in children because absence and atypical absence seizures are more prevalent in children than adults. Age-related pharmacokinetic differences and drug interactions are major considerations in children. On average, children have higher clearance rates of carbamazepine, shorter half-lives, and higher ratios of carbamazepine-10, 11-epoxide to carbamazepine than adults. In addition, children with severe epilepsy are more likely to require multiple-drug therapy, which can lead to complex drug interactions. When carbamazepine is administered along with valproate, drug protein binding interactions can cause intermittent side effects.     

Neonatal Seizures: Problems in Diagnosis and Classification

Summary: The clinical identification of neonatal seizures is critical for the recognition of brain dysfunction; however, diagnosis is often difficult because of the poorly organized and varied nature of these behaviors. Current classification systems are limited in their ability to communicate motor, autonomic, and electroencephalo-graphic features of seizures precisely and to provide a basis for uniform effective diagnosis, therapy, and determination of prognosis. Recent investigations of neonates, utilizing bedside electroencephalographic/polygraphic/ video monitoring techniques, have provided the basis for improved diagnosis and classification of seizures in the newborn. These studies have demonstrated that not all clinical phenomena currently considered to be seizures require electrocortical epileptiform activity for their initiation or elaboration. In addition, the specific clinical character of the phenomena considered to be seizures, the clinical state of the infant, and the character of the EEG indicate the probable pathophysiological mechanisms involved and suggest probable etiologies, prognosis, and therapy. Similarities between animal models that demonstrate reflex physiology and neonates with motor automatisms and tonic posturing suggest that these clinical behaviors may not be epileptic in origin but, rather, primitive movements of progression and posture mediated by brainstem mechanisms. Although not all clinical behaviors currently considered to be neonatal seizures may have similar pathophysiological mechanisms, they are clinically significant because they all indicate brain dysfunction.     

Valproate Monotherapy in the Management of Generalized and Partial Seizures

Summary: For decades, therapeutic tradition has promoted the concept of polypharmacy in the management of epilepsy. In recent years, however, studies have shown that, for most patients, monotherapy can provide comparable or better seizure control than administration of multiple anticonvulsants, while diminishing the potential for adverse reactions, drug interactions, and poor compliance. Valproate is an important monotherapeutic agent that is highly effective in the control of idiopathic primary and secondarily generalized epilepsies, and partial seizures that do not generalize. Comparative studies have found that valproate is at least as effective as phenytoin and carbamazepine in the treatment of generalized and partial seizures. Given the similar efficacy, other factors such as pharmacokinetics and side effects may therefore determine anticonvulsant selection for monotherapy.     

Un nouveau cadre conceptuel de travail pour la psychiatrie

In an attempt to place psychiatric thinking and the training of future psychiatrists more centrally into the context of modern biology, the author outlines the beginnings of a new intellectual framework for psychiatry that derives from current biological thinking about the relationship of mind to brain. The purpose of this framework is twofold. First, it is designed to emphasize that the professional requirements for future psychiatrists will demand a greater knowledge of the structure and functioning of the brain than is currently available in most training programs. Second, it is designed to illustrate that the unique domain which psychiatry occupies within academic medicine, the analysis of the interaction between social and biological determinants of behavior, can best be studied by also having a full understanding of the biological components of behavior.