丹参酮ⅡA通过抑制TLR4/NF-κB途径减轻糖氧剥夺对大鼠脑微血管内皮细胞的炎症损伤

目的 探讨丹参酮ⅡA对大鼠脑微血管内皮细胞糖氧剥夺炎症损伤的影响及其机制。方法 原代培养雄性SD大鼠脑微血管内皮细胞，分为对照组、糖氧剥夺组和丹参酮ⅡA组。对照组用DMEM培养，糖氧剥夺组细胞用无糖DMEM培养，丹参酮ⅡA组用在无糖DMEM基础上加用含8 μg/ml丹参酮ⅡA培养。利用CCK8检测细胞活力，酶联免疫吸附试验检测细胞炎症因子IL-6和TNF-α水平；实时PCR和免疫印迹法检测细胞TLR4和NF-κB mRNA和蛋白表达水平。结果 与对照组相比，糖氧剥夺组细胞活力明显降低（P<0.05），IL-6和TNF-α水平明显增加（P<0.05），TLR4和NF-κB mRNA和蛋白表达均明显增加（P<0.05）；丹参酮ⅡA明显抑制糖氧剥夺作用（P<0.05），但未恢复到对照组水平（P<0.05）。结论 丹参酮ⅡA可减轻糖氧剥夺对大鼠脑微血管内皮细胞的炎症损伤，其作用机制可能与抑制TLR4-NF-κB信号通路有关     

Netrin-1促进氧糖剥夺后脑微血管内皮细胞存活

目的通过建立体外氧糖剥夺模型模拟缺血缺氧环境,观察轴突导向因子netrin-1对脑微血管内皮细胞氧糖剥夺损伤后的作用。方法体外培养脑微血管内皮细胞,免疫荧光法检测其netrin-1受体结直肠癌缺失基因(deleted in colorectal cancer,DCC)及uncoordinated(UNC)5H2的表达;并使用不同浓度netrin-1作用于氧糖剥夺损伤的脑微血管内皮细胞,Cell Counting Kit-8(CCK-8)试剂盒及Annexin V-FITC/PI细胞凋亡双染试剂盒检测其细胞活性。结果脑微血管内皮细胞表达UNC5H2受体,无DCC受体表达;与对照组相比,浓度为10、50、100 ng/m L的netrin-1作用于氧糖剥夺损伤的脑微血管内皮细胞后细胞活性增强(10 ng/m L:1.25±0.20,存活率63.33%,P0.05;50 ng/m L:1.41±0.17,存活率79.40%,P0.05;100 ng/m L:1.33±0.27,存活率75.98%,P0.05),且在其浓度为50 ng/m L时细胞活性最强。结论 Netrin-1对氧糖剥夺损伤的脑微血管内皮细胞具有剂量相关的保护作用,这种保护作用可能由其受体UNC5H2介导。     

大鼠脑微血管内皮细胞氧糖剥夺模型的构建及黄体酮的保护作用

目的：体外研究氧糖剥夺对大鼠脑微血管内皮细胞的损伤机制及黄体酮的保护作用。方法体外培养大鼠脑微血管内皮细胞（brain microvascular endothelial cell ,BMEC）,并构建氧糖剥夺（oxygen and glucose deprivation ,OGD）模型。流式细胞技术检测BMEC在OGD后细胞间黏附分子-1（intercellular adhesion molecule-1,ICAM-1）、血管细胞黏附分子-1（vascular cell adhesion molecule-1,VCAM-1）的表达情况。利用Western blot方法检测大鼠BMEC氧糖剥夺后核转录因子-B （nuclear factor-kappa B ,NF-κB）p65亚基含量,染色质免疫沉淀分析（Chromatin immunoprecipitation assay kit ,ChIP）技术检测NF-κB的DNA结合活性。结果 OGD后6 h可诱导BMEC的ICAM-1、VCAM-1和NF-κB表达上调,黄体酮对此有明显抑制作用。氧糖剥夺能显著提高NF-κB的DNA结合活性。黄体酮对此有明显抑制作用。结论 OGD可通过活化NF-κB、上调ICAM-1和VCAM-1的表达、黄体酮能减少OGD对BMEC造成的损伤。     

脑微血管内皮细胞TNFR差异表达对rhTNF开放血肿瘤屏障的影响

目的观察脑微血管内皮细胞的肿瘤坏死因子受体(TNFR)差异表达对rhTNF开放血肿瘤屏障的影响。方法建立TNFR1/TNFR2高表达的脑微血管内皮细胞株,并与C6胶质瘤细胞一起通过Transwell小室构建体外血肿瘤屏障模型。免疫荧光技术检测脑微血管内皮细胞TNFR的表达水平。荧光分析法测定给予rhTNF后的血肿瘤屏障通透性变化。结果脑微血管内皮细胞的TNFR1高表达时,rhTNF开放血肿瘤屏障的程度最大。而TNFR2高表达时,血肿瘤屏障开放程度与对照组相比无明显差异。结论 rhTNF可能是通过与脑微血管内皮细胞的TNFR1结合而引起血肿瘤屏障开放的。     

西洛他唑对大鼠脑微血管内皮细胞缺血再灌注损伤的保护作用

目的研究西洛他唑对大鼠脑微血管内皮细胞缺血再灌注损伤的保护作用机制。方法以原代方法培养大鼠脑微血管内皮细胞并传代,将第3代传代细胞随机分为正常对照组、西洛他唑组、溶剂对照组和缺血再灌注模型组(再灌模型组)4组,对后3组大鼠建立脑微血管内皮细胞糖氧剥夺后复糖氧模型,模拟"缺血再灌注"过程,并对西洛他唑组和溶剂对照组在造模同时分别以终浓度1×10-5mmol/L西洛他唑〔二甲基亚砜(DMSO)为溶剂〕和0.05%(体积分数)DMSO进行干预。糖氧剥夺3 h复糖氧24 h后,测定各组细胞上清液中诱导型一氧化氮合酶(iNOS)和内皮型一氧化氮合酶(eNOS)水平及细胞内环磷腺苷酸(cAMP)水平,并以四唑盐(MTT)比色实验测定细胞活力。结果西洛他唑组与再灌模型组比较,eNOS水平明显提高,iNOS水平明显降低,cAMP水平及细胞存活率显著提高(均P<0.05)。结论西洛他唑对大鼠脑微血管内皮细胞的缺血再灌注损伤有保护作用,其作用机制可能与促进eNOS水平升高和iNOS水平降低有关。     

黄体酮对大鼠脑微血管内皮细胞氧糖剥夺后的保护作用

目的研究黄体酮(progesterone,PROG)对大鼠脑微血管内皮细胞氧糖剥夺后的保护作用机制。方法体外培养大鼠脑微血管内皮细胞(brain microvascular endothelial cell,BMEC),并构建氧糖剥夺(oxygen and glucose deprivation,OGD)模型。以3-(4,5-二甲基噻唑-2)-2,5-二苯基四氮唑溴盐[3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide,MTT]法检测BMEC存活率,比色法测定乳酸脱氢酶(lactate dehydrogenase,LDH)漏出率反映细胞损伤程度。为反映BMEC的氧化应激损伤水平,利用生物化学法测定细胞培养液中及细胞内丙二醛(malondialdelyde,MDA)水平及细胞内超氧化物歧化酶(superoxide dismutase,SOD)和谷胱甘肽过氧化物酶(glutathione peroxidase,GSH-Px)活性。结果 OGD 6h明显降低BMEC的存活率,并增加LDH释放率。OGD 6h可明显增加BMEC培养基中及细胞内MDA含量,减少BMEC内SOD、GSH-Px活性。与氧糖剥夺组相比,黄体酮可明显降低培养基中和细胞内的MDA含量,提高BMEC内SOD、GSH-Px活性。结论 OGD可通过诱导氧化应激等途径损伤BMEC,黄体酮能减少OGD对BMEC造成的损伤。     

LncRNA MEG3靶向调节miR-17-5p对缺氧缺血性脑损伤新生小鼠血-脑屏障的影响

目的 探讨LncRNA MEG3通过调节miR-17-5p对缺氧缺血性脑损伤(HIBD)新生小鼠血-脑屏障的影响。方法 90只C57BL小鼠按照随机数字表法分为对照组、模型组、si-NC组、si-LncRNA MEG3组、si-LncRNA MEG3+anti-miR-NC组、si-LncRNA MEG3+anti-miR-17-5p组,每组15只。除对照组外,其余组小鼠构建HIBD模型,并采用Longa评分评估小鼠神经损伤情况;电镜观察脑组织周围血管内皮细胞超微结构;EB法测定小鼠血-脑屏障通透性;qRT-PCR法测定各组小鼠脑组织LncRNA MEG3、miR-17-5p水平;Western blotting法测定小鼠脑组织ZO-1、Occludin蛋白水平。结果 与对照组相比,模型组小鼠血管内皮薄厚不均、多处呈现水肿状态;与模型组相比,si-LncRNA MEG3组小鼠血管内皮细胞逐渐连接紧密、薄厚较为均匀,水肿减少;与si-LncRNA MEG3组相比,si-LncRNA MEG3+anti-miR-17-5p组血管内皮细胞损伤加剧。与对照组相比,模型组小鼠神经功能缺损评分、脑...     

急性脑损伤脑微血管三维构型和超微结构实验研究

通过血管铸型方法观察急性脑损伤动物模型脑微血管三维结构和血管内皮细胞的超微结构改变。结果发现：脑损伤后，脑微血管内皮细胞发生显著变性，脑微血管痉挛，小动脉、毛细血管断裂以及出现多处无毛细血管区。本实验证实，脑损伤以后，发生明显的脑微血管系统出血性和缺血性改变，病灶区血液循环障碍。这些发现有力地支持继发性脑损害血管源性机制的假说。     

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。     

MALAT1 介导 β-catenin 通路参与 Salvinorin A减轻脑卒中后血脑屏障损伤的机制

目的 探讨MALAT1介导β-catenin通路参与Salvinorin A减轻脑卒中后血脑屏障(BBB)损伤的机制.方法 选取SD大鼠制作大脑中动脉栓塞模型(MCAO),进行Salvinorin A减轻脑卒中后脑损伤相关实验.选取人脑微血管内皮细胞(HBMECs)制作糖氧剥夺(OGD)模型,进行MALAT1介导β-c...     

Diagnostic Difficulties and Treatment Implications

Summary: Differentiation between types of epileptic seizures has been aided in recent years by the introduction of intensive neurodiagnostic techniques and the development of increasingly detailed classification systems. Paradoxically, these developments have not simplified the task of matching the appropriate antiepileptic drug to a particular seizure type. It is reasonable to assume that anticonvulsant drugs will have different effects on different types of seizures, but faulty, circular reasoning can enter the picture if one also assumes that responses of seizures to different drugs signify different seizure types. There are several examples of differential diagnoses that can fall prey to this problem, including the diagnosis between partial seizures with secondary generalization and generalized tonic-clonic seizures, and the diagnosis between complex partial seizures and absence seizures with automatisms, among others. Considerations of etiology in future classification systems can further complicate the problem: should one then choose an anticonvulsant drug on the basis of individual seizure type or on the basis of the type of epilepsy? Ramifications of this issue extend even to the drug approval process. Official sanction is not given for use of a drug for a seizure type not included in the original efficacy studies, even if later scientific evidence shows that seizure type to be related to a type that is included. New trials must be undertaken. These problems arise from how we choose to classify seizures.     

Cognitive Dysfunction Associated with Antiepileptic Drug Therapy

Summary: Epilepsy is frequently associated with cognitive dysfunction. However, the reasons for this correlation are unclear. Possible influential factors include patient age; duration, frequency, etiology, and type of seizures; hereditary factors; psychosocial issues; and antiepileptic drug (AED) therapy. Whereas many of these factors are beyond the physician's control, AED therapy is one element that can be addressed in treatment decisions by recognizing the potential cognitive effects of particular AEDs. For example, phenobarbital impairs memory and concentration; phenytoin affects attention, problem solving ability, and performance of visuomotor tasks. In contrast, carbamazepine may affect concentration, while valproate would appear to have minimal effects on cognition. Moreover, cognitive effects of AEDs are amplified with coadministration of multiple anticonvulsants (polytherapy). A review of studies on the cognitive effects of monotherapy with AEDs, as opposed to those of polytherapy, provides evidence that drug-related cognitive dysfunction can be reversed if patients are switched to a simpler therapeutic regimen. Future research should be directed toward developing reliable measures for assessing and monitoring cognition, and understanding the particular cognitive side effects of each AED. Physicians also need to revise their opinions about which side effects are "tolerable" for epileptic patients.     

Carbamazepine Efficacy in Adults With Partial and Generalized Tonic-Clonic Seizures

Summary: Carbamazepine and phenytoin are drugs of choice in initial monotherapy for adult partial and secondarily generalized tonic-clonic seizures. These designations reflect the results of the Veterans Administration Epilepsy Cooperative Study Group of 1985. An earlier comparative study of carbamazepine and phenytoin by Ramsay and associates found both drugs equally effective in controlling new-onset seizures. Among the advantages of carbamazepine is that it causes relatively few cognitive and dysmorphic side effects. Its disadvantages are its unavailability in parenteral formulation and its metabolic autoinduction. The latter must be compensated for by planned dosage increases to maintain therapeutic plasma steady-state levels during the first 2 or 3 months of treatment. Carbamazepine is judged a drug of choice in the treatment of these secondarily generalized tonic-clonic seizures, and the drug of choice in children, adolescents, and women susceptible to the dysmorphic side effects associated with other anticonvulsant agents.     

Etiologic and Preventive Aspects of Epilepsy in the Child–Bridging the Gap Between Laboratory and Clinic

Summary: Four broad categories of basic phenomena are pertinent to developing ways to prevent epilepsy. These include mechanisms of epileptogenesis, ictal initiation and temporary entrainment by the seizure discharge of normally functioning brain, seizure propagation, and control mechanisms that function both to restrain the cascade of epileptic events culminating in a seizure and to arrest the epileptic event and restore the interictal state. In newborns and children, hypoxia-ischemia is a major factor leading to epileptogenesis, and several schemes are proposed to classify, quantify, and prevent hypoxic-ischemic encephalopathy. Control mechanisms must be better understood in order to develop prophylactic recommendations for epilepsy, and an experimental model of "kindling antagonism" may increase our understanding of these. Programs of prevention of seizures in children will evolve only if basic researchers and clinicians work productively together to develop an adequate understanding of factors important in epileptogenesis and antiepileptogenic control mechanisms.     

Predisposing and Causative Factors in Childhood Epilepsy

Summary: We review information from large studies of defined populations, examining the role of known factors and especially of prenatal and perinatal factors in contributing to nonfebrile seizure disorders of early childhood. We depend especially, but not exclusively, on the recently completed analyses from the Collaborative Perinatal Project of the National Institute of Neurological and Communicative Disorders and Stroke, the NCPP. About 4% of children in the NCPP who had at least one non-febrile nonsymptomatic seizure by the age of 7 years had a previous seizure during acute neurologic illness, such as meningitis or during the acute illness after trauma. Many such seizures should potentially be preventable. Of children with seizures, 10% had had a neonatal seizure and 13% had had a febrile seizure. Among the hundreds of prenatal and perinatal factors explored as predictors of childhood seizure disorders, the principal predictors identified were congenital malformations of the fetus, cerebral and noncerebral; family history of certain neurologic disorders; and neonatal seizures. In agreement with the British National Child Development Study, labor and delivery factors in the NCPP appeared to contribute very little to childhood seizure disorders. Maldevelopment, rather than damage at birth to an initially intact nervous system, appeared to be the more common mechanism. Most seizure disorders of early childhood remained unexplained by the large set of prenatal and perinatal characteristics examined.     

Anticonvulsant Drugs and Cognitive Function: A Review of the Literature

Summary: Alterations of cognitive function are separate from disturbances of behavior seen in association with epilepsy. The nature of the cognitive disability may to a certain extent depend on the seizure type. Partial seizures, mainly derived from a temporal lobe focus, impair memory tasks, while generalized seizures seem to have more effect on attentional abilities. A number of studies, reviewed in this paper, suggest that anticonvulsant drugs further impair cognitive function. Maximal impairments are seen in patients receiving polytherapy: rationalization of polytherapy improves cognitive abilities. Studies in children and adults have allowed differentiation of the effects of various commonly used antiepileptic agents. Maximal cognitive deficits are seen with. phenytoin, while phenobarbital and sodium valproate induce moderate disturbances, and carbamazepine seems relatively free from such toxicity. Further research is needed on the interrelationship between types of seizure disorders, types of anticonvulsant medications, and cognitive function.     

Effects of NMD A- and AMPA-Receptor Antagonists on Different Forms of Epileptiform Activity in Rat Temporal Cortex Slices

Summary: Lowering extracellular magnesium induces different patterns of epileptiform activity in rat hippocampus and entorhinal cortex. Short recurrent epileptiform discharges in the hippocampus are stable over time, whereas seizurelike events (SLEs) in the entorhinal cortex, the subiculum, and the neighboring neocortex develop into late recurrent discharges which are not blocked by clinically employed antiepileptic drugs. We tested the sensitivity of the different epileptiform discharge patterns to. /V-methyl-D-aspartate (NMDA)- and non-NMDA-receptor antagonists. As NMDA-receptor antagonist we used dextrorphan, ket-amine, and 2-aminophosphonovalerate (2APV); as α-amino-3-hydroxy-5-methyl-4-isoxazole-propionic acid (AMPA)-receptor antagonist we employed the quinoxaline derivative glutamate 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX). The findings show that the different patterns of epileptiform activity, including the late recurrent discharges, are sensitive to all NMDA-receptor antagonists. However, when dextrorphan was employed to suppress seizure-like events, later recurrent discharges did not develop during the remaining time course of the experiment. CNQX reversibly suppressed recurrent discharges in the hippocampus and SLEs in the entorhinal cortex. However, late recurrent discharges become insensitive to CNQX, even at a high concentration of 60 μM m. This finding suggests a prominent role for NMDA receptors in the generation of late recurrent discharges.     

Treatment Considerations in Anticonvulsant Monotherapy

Summary: The long-standing practice of polypharmacy in treating epilepsy is giving way to use of monotherapy. Monotherapy can improve seizure control as well as reduce the risk of serious idiosyncratic reactions, dose-related side effects, and complex drug interactions. Monotherapy also offers improved compliance and cost-effectiveness. The basis of monotherapy is accurate diagnosis and assessment of the patient's seizure type(s), followed by selection of a single appropriate anticonvulsant drug. Many patients currently treated with multiple anticonvulsants can be successfully converted to monotherapy with a carefully monitored program in which troublesome and redundant drugs are gradually withdrawn from the therapeutic regimen.     

The influence of comorbid depression on seizure severity

PURPOSE: To determine the relation between depressive symptoms and seizure severity among people with epilepsy. METHODS: A postal questionnaire was used to survey a nationwide community sample about seizures and depression. The Seizure Severity Questionnaire (SSQ) assessed the severity and bothersomeness of seizure components. The Centers for Epidemiological Studies-Depression scale categorized levels of depression. RESULTS: Respondents categorized as having current severe (SEV, n = 166), mild-moderate (MOD, n = 74), or no depression (NO, n = 443) differed significantly in SSQ scores (all p < 0.0001). People with SEV or MOD reported significantly worse problems than did those with NO depression for overall seizure recovery (mean, 5.3, 4.9, 4.5, respectively); overall severity (5.0, 4.5, 4.2); and overall seizure bother (5.3, 4.8, 4.4) (all p < 0.005). Cognitive, emotional, and physical aspects of seizure recovery also were rated worse among people with SEV than with NO depression (all p < 0.05). Symptoms of depression were significantly correlated with higher levels of all components of generalized tonic-clonic seizure severity (r = 0.33-0.48; all p < 0.0001), and partial seizures (r = 0.31-0.38; all p < 0.01). CONCLUSIONS: Clinically depressed people with epilepsy reported higher levels of perceived severity and bother from seizures, as well as greater problems with overall seizure recovery than did nondepressed people experiencing similar types of seizures. The pervasive influence of depressive symptoms on reports of seizure activity suggests that people with epilepsy should be screened for depression. These data highlight the importance of detecting and treating depression among people with epilepsy.     

Dextromethorphan: Cellular Effects Reducing Neuronal Hyperactivity

Summary: Dextromethorphan is a dextrorotary morphinan without affinity for opioid receptors, commonly used as an antitussive medication. During the past 5 years, interest in the compound and its demethylated derivative, dextrorphan, has been revived because additional neuroprotective and an-tiepileptic properties were found in in vitro studies, animal experiments, and a few clinical cases. Both morphinans are able to inhibit N -methyl-D-aspartate (NMDA) receptor channels and voltage-operated calcium and sodium channels with different potencies. The inhibition of the NMDA receptor is believed to be the predominant mechanism of action responsible for the anticonvulsant and neuroprotective properties of the compounds.