血管内皮生长因子-C在大脑皮质发育障碍中的作用及机制研究
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摘要
大脑皮质发育障碍(MCD)是药物难治性癫痫的重要病因之一。随着高清影像学技术的普及,MCD的临床检出率不断提高,现已证实约有40%的药物难治性癫痫是由MCD所致,而在小于3岁的儿童药物难治性癫痫病人中更高达80%,因此MCD已成为药物难治性癫痫研究领域内重点和热点的问题。MCD是一类病理学表现类似疾病的总称,主要包括结节性硬化(TSC)、局灶性皮质发育不良(FCD)、多小脑回、无脑回畸形及Sturge-Weber综合征等亚型,其中TSC和FCD是最常见的MCD亚型。尽管MCD根据不同的临床表现具有不同的分类,但都具有共同的组织病理学特点,表现为大脑皮层层状结构和柱状结构紊乱、脑回结构破坏,出现大量异形神经元(DNs)、巨形神经元(GNs)、气球样细胞(BCs)以及巨细胞(GCs)等,这些异常的细胞被称作异构神经元(MNs)。临床资料表明伴有异构神经元的MCD患者更易出现癫痫发作,早期出现耐药。因此,异构神经元在MCD的致痫性中发挥“核心作用”。目前,对异构神经元的病理发生机制尚无明确定论,较为公认的观点认为,异构神经元的产生可能是由胚胎发育期间神经元迁移,成熟和／或细胞死亡过程中出现异常所致。此外,围产期和出生后早期的脑损伤可能也参与了异构神经元的病理发生。总之,目前对MCD及异构神经元病理发生机制所知有限,对涉及其中的关键因子、信号途径仍待阐明。
VEGF-C是公认的淋巴系统发生调节因子,通过激活两个特异性受体VEGFR-2, VEGFR-3发挥生物学效应,其中VEGFR-3为VEGF-C的主力型受体。在正常情况下,VEGF-C及VEGFR-3局限性表达于淋巴内皮,而它的另一受体VEGFR-2则在血管内皮和淋巴内皮上均有表达,发挥其促脉管系统发生的作用。
由于脑内淋巴系统缺如,既往很少有研究关注VEGF-C在中枢神经系统的作用。直到2006年,有学者发现当小鼠敲除VEGF-C后,表达VEGFR-3受体的神经前体细胞增殖明显受阻,而且胚胎视神经中少突胶质细胞前体细胞选择性缺失,证明VEGF-C信号通路参与了中枢神经系统的生长发育、发挥神经营养功能。近两年来陆续有研究揭示了VEGF-C信号系统在神经系统中的新作用,如VEGF-C/VEGFR-3能够介导胶质前体细胞的增殖,迁移。新近报道脑缺血后,定位于海马齿状回颗粒细胞层最内侧带的细胞高度表达VEGFR-3,这些细胞同时表达与细胞迁移相关的神经前体细胞标记物PSA-NCAM,表明VEGF-C系统参与脑缺血后海马神经发生。此外,脑卒中后,患侧SVZ区内VEGFR-3的表达水平显著上调,表型鉴定实验证实绝大多数表达VEGFR-3的细胞主要为具有旺盛增殖活力的神经祖细胞,另有小部分VEGFR-3阳性细胞为具备迁徙能力的神经前体细胞。在本研究预实验中,我们发现手术切除的MCD标本中VEGF-C mRNA表达上调。那么,VEGF-C及其受体在MCD中的具体表达模式如何?是否表达于异构神经元?高表达的VEGF-C是否参与MCD所致癫痫发作?这些问题值得我们关注。
为了阐明VEGF-C在MCD中的作用及其可能的机制,首先,我们利用半定量RT-PCR、western blot、免疫组织化学技术观察了VEGF-C及其受体在MCD(TSC与FCD亚型)临床标本中的表达分布情况；其次,观测了VEGF-C受体信号通路下游相关信号分子的表达活化情况；第三,观察了与VEGF-C高度同源的VEGF-D在MCD中的表达情况；最后,为了直观的了解VEGF-C对MCD致痫效应的影响,我们建立MCD大鼠模型,利用膜片钳全细胞记录技术,观察VEGF-C对MCD大鼠异构神经元兴奋性谷氨酸受体(NMDA受体)的调节效应。通过这些研究,我们得到以下结果：
一、VEGF-C及其受体在MCD中的表达分布
1.通过RT-PCR, western blot分析,从基因和蛋白水平证明正常皮层中不同程度地表达VEGF-C、VEGFR-2及VEGFR-3;免疫组化结果表明VEGF-C及其受体主要分布于皮层锥体神经元；
2.在TSC皮层结节组织匀浆中,VEGF-C及其受体VEGFR-2、VEGFR-3的mRNA及蛋白表达水平较正常对照组明显升高。免疫组化结果显示,VEGF-C, VEGFR-2及VEGFR-3特征性的高表达于DNs、GCs及反应型胶质细胞；免疫荧光双标结果表明：①绝大多数VEGF-C免疫阳性的DNs, GCs共表达神经丝蛋白标记物NF-200；然而未见共表达与另一种神经元标记物NeuN以及星形胶质细胞特异性标志物GFAP共表达；TSC组织中可见CD68阳性的巨噬细胞浸润,但是这些细胞不表达VEGF-C;②VEGFR-2与VEGFR-3的细胞定位表达模式类似,多数VEGFR-2或VEGFR-3免疫阳性的DNs与NF-200共表达,而共表达VEGFR-2 (VEGFR-3)与NF-200的GCs略少,未见VEGFR-2 (VEGFR-3)免疫阳性的DNs, GCs共表达GFAP。
3.在另一常见的MCD亚型-FCD中,VEGF-C及受体VEGFR-2、VEGFR-3的mRNA及蛋白表达水平较正常对照组同样显著升高。VEGF-C及其受体高表达于DNs, GNs,BCs及胶质细胞。免疫双标结果显示：①VEGF-C免疫阳性的异构神经元不表达GFAP;与TSC中表达模式不同的是,FCD中的DNs, GNs, BCs能够同时表达VEGF-C与NeuN,提示其具有成熟神经元属性。②VEGFR-3免疫阳性的异构神经元既不表达NF-200,又不表达GFAP,表明其特殊的分子表型。
二、MCD病灶内VEGF-C下游相关信号通路的表达活化
1. RT-PCR结果表明,MCD中Akt的基因表达上调；利用western blot观察Akt蛋白亚型表达,发现三种Akt亚型的表达均上调,其中Akt-1的表达上调最为明显,表明Akt信号通路参与了MCD的病理生理机制。
2.正常皮层及MCD中均可见Bad的表达,且表达水平基本相同；然而,MCD病灶内的磷酸化Bad (p-Bad)表达显著升高,提示MCD中抗凋亡通路活化。
3.ERK表达于MCD病灶组织和正常皮层中,呈现特征性的44kDa和42 kDa双条带(ERK 1/2),并可见ERK1/2在MCD中的表达明显上调。
三、VEGF-D在MCD中的表达分布
1.正常皮层组织匀浆中,可探及VEGF-D mRNA及蛋白表达；免疫组织化学结果表明VEGF-D主要表达于锥体神经元,且免疫反应程度较轻。
2.MCD组织匀浆中,VEGF-D mRNA及蛋白表达水平上调。免疫组织化学实验发现：①TSC亚型中VEGF-D表达于多数DNs、GCs及反应性胶质细胞,但免疫阳性强度不一；免疫双标未发现VEGF-D阳性异构神经元与NeuN, NF-200, GFAP及CD68共表达；②FCD亚型中可见VEGF-D表达于DNs, GNs, BCs及反应性胶质细胞,免疫双标结果表明VEGF-D免疫阳性的异构神经元不表达成神经纤维蛋白NF-200,亦不表达成熟星型胶质细胞标记物GFAP。
四、VEGF-C对MCD大鼠异构神经元NMDA受体功能的调节效应
1.建立MCD动物模型,制备活体脑片后进行膜片钳全细胞记录,结果表明①MCD大鼠异构神经元可产生动作电位,但动作电位形态各异,峰率适应性降低或消失,提示其电生理功能发育不完善；②异构神经元中可记录到evoked-EPSC,但电流形态明显发生改变,通道电流成分混杂,电流面积明显增加,提示MCD异构神经元突触传递效能改变；③MCD异构神经元中NMDA/AMPA受体介导电流的比率显著上调,这种比率改变主要是由NMDA受体电流成分增多引起,表明异构神经元中NMDA受体功能上调。
2.MCD大鼠异构神经元中VEGF-C及其受体VEGFR-2的表达明显上调,VEGFR-3的表达亦有增加。
3.100μmol/L NMDA作用于异构神经元后,可诱发典型的快反应电流,经过100ng/mL VEGF-C孵育后,相同浓度NMDA诱发的电流显著增大,提示VEGF-C对NMDA受体产生协同作用；当给予VEGFR-2受体阻断剂Ki8751预处理后,这种协同效应被阻断,表明VEGF-C对NMDA受体的调节效应主要是由VEGFR-2受体介导的。
综上所述,本研究结果表明：第一、人MCD组织中VEGF-C及其受体VEGFR-2,VEGFR-3表达显著上调,且特征性地高表达于异构神经元；第二、MCD中VEGF-C下游通路中抗凋亡相关信号分子激活,有益地提示VEGF-C可能通过抗凋亡相关信号通路调节MCD内异构神经元的生物学行为；第三、VEGF-D在MCD中的表达上调；第四、VEGF-C通过VEGFR-2增强MCD大鼠异构神经元NMDA受体活性,易化异构神经元的兴奋性。总之,我们的结果提示VEGF-C信号通路参与了MCD的病理生理机制。
Malformations of cortical development (MCD) represents a well-recognized cause of intractable epilepsy, with a poor response to the currently available anti-epileptic drugs (AEDs). With the development of high-resolution imageology, the detection rate of MCD is continuously rising during the past decade in clinic.It has been demonstrated that approximately 40 percent of adult patients and more than 80 percent of pediatric with intractable epilepsy are associated with MCD.Thus, the pathogenesis of MCD has became a central concern in the field of intractable epilepsy. MCD encompasses a large spectrum of disorders related to abnormal cortical development, including tuberous sclerosis complex (TSC), focal cortical dysplasia (FCD), microgyria, congenital agyria and Sturge-Weber syndrome. Among these subsets, two of the most common components are TSC and FCD. Despite the fact that these subsets with varied genetic etiologies, anatomic abnormalities, and clinical manifestations, they often share similar histological features such as disorganization or lack of the normal six-layered cortical lamination structure, and the occurrence of malformed neurons (MNs) known as dysplastic neurons (DNs), giant neurons (GNs),balloon cells (BCs) and giant cells (GCs). Mounting clinical evidence has revealed that epilepsy readily occurs in patients with MNs, and appears drug resistance in the early stage, suggesting the dominant role of MNs in epileptogenesis of MCD. Until now, the pathogenesis of MNs remains unclear. A more popular view is that MNs is a malformation due to abnormal migration,mature and cell death during embryonic brain development. Moreover, recent pathological studies on surgical or autoptic human tissue suggest a role for perinatal and early postnatal brain injury in the formation of MNs. In spite of the fact that there are many reports on the subject of MCD, the exact mechanism of this disease has not yet been elucidated.
VEGF-C,a well-recognized regulator of lymphangiogenesis, binds to and activates the specific receptors VEGFR-2 and VEGFR-3, and the affinity of VEGFR-3 is much more than that of VEGFR-2. Previous studies show VEGF-C and VEGFR-3 are restrictly expresssed in lymphatic endothelial cells, and VEGFR-2 is expressed in both vascular endothelial cells and lymphatic endothelial cells.
Due to the deficiency of the lymphatic system in the brain, few studies have focused on the role of VEGF-C in the CNS. Until to 2006, an important study demonstrats that VEGF-C and VEGFR-3 are present in the neural progenitor cells of Xenopus laevis and mouse embryos. Furthermore, a lack of VEGF-C results in a severe defect in the proliferation of neuronal progenitor cells expressing VEGFR-3, suggesting that VEGF-C is a trophic factor in neuronal progenitor cells in the vertebrate brain. Recently, several studies revealed VEGF-C mediate multiple effects in CNS. For example,activation of the VEGF-C/VEGFR-3 signaling pathway was shown to mediate proliferation and chemotaxis in glial precursor cells. In addition, after a short period of brain ischemia, the innermost layer of granular cells in the hippocampal dental gyrus displayed increased levels of VEGFR-3 expression, these VEGFR-3 immunoreactive cells are also expressed PSA-NCAM, this might be related to neurogenesis in the hippocampus after cerebral ischemia. A recent study showed that the expression of VEGFR-3 increased in subventricular zone, and most of VEGFR-3 positive cells were neural progenitors, were highly proliferative. To our knowledge, there have been no reports about the roles of VEGF-C in in seizure disorders, especially those induced by MCD. In our preliminary experiment,we found that VEGF-C mRNA exhibited high levels in MCD. However, the detailed expression pattern of VEGF-C and it receptors in MCD, and the role of VEGF-C signal system in the epileptogenesis of MCD are unclear.
To address these concerns:First,we identified the distribution of VEGF-C system in MCD from patients with medically intractable epilepsy by means of RT-PCR, western blot and immunohistochemistry. Second,we tested the expression and activation of signaling molecule involved in VEGF-C pathway. Third, we investigated the expression and cellular distribution of VEGF-D,which is highly homologous with VEGF-C, in MCD. Fourth, to reveal the role of VEGF-C in the epileptogenesis of MCD, a MCD rat model was employed, and we investigated the effect of VEGF-C on NMDA receptor(NMDAR)of MNs by using whole-cell patch-clamp recordings technique on brain slice of MCD rat model. The results show as following:
Ⅰ. Expression and cellular distribution of VEGF-C and VEGF receptors 2 and 3 in MCD
1. RT-PCR and western blot analysis showed VEGF-C and its receptors mRNA and protein expression in total homogenates of normal control cortex (CTX); immunohistochemical experiments revealed immunostaining was mainly observed in pyramidal neurons.
2. There was a statistically significant increase of VEGF-C, VEGFR-2 and VEGFR-3 mRNA and protein levels in cortical tubers of TSC, in comparison with CTX. The immunostaining results demonstrated that the high-level expression of protein, including VEGF-C and its receptors, was mainly localized within GCs, DNs, and reactive astrocytes. The results of double-labeling immunofluorescence showed:①the co-localization of VEGF-C immunostaining with the neuronal marker NF-200 in majority of DNs and GCs. Intriguingly, we could not detect any co-immunostaining of VEGF-C with another neuronal marker, NeuN, in both DNs and GCs in all cases. Also, GFAP and CD68 immunostaining was not detected in any VEGF-C-positive DNs and GCs.②VEGFR-2 and VEGFR-3 shared a similar cellular distribution pattern in TSC, VEGFR-2 (VEGFR-3) immunoreactivity co-localized with NF-200 in the majority of DNs,the frequency of VEGFR-2 (VEGFR-3) and NF-200 co-expression in GCs was less than that in DNs. VEGFR-2 (VEGFR-3) and GFAP co-immunolabeling was not detected in any DNs and GCs, although prominent co-labeling of reactive astrocytes was noted in all of the TSC cases examined.
3. We found an obvious increase of VEGF-C and it receptors mRNA and protein expression in FCD specimens compared with CTX. The high-level expression of protein, including VEGF-C and its receptors, was mainly localized within DNs,GNs,GCs and reactive astrocytes. The results of double-labeling immunofluorescence showed:①GFAP and VEGF-C co-immunolabeling was not detected in any MNs. Interestingly,we detected co-immunostaining of VEGF-C with NeuN, which is usually recognized as a marker of mature neurons, in MNs.②VEGFR-3 and GFAP/NF-200 co-immunolabeling was not observed in any MNs,suggesting a complex phenotype of MNs.
Ⅱ. The expression and activation of signaling molecule involved in VEGF-C pathway
1. In comparison with CTX, RT-PCR analysis showed a increase of Akt mRNA levels in MCD specimens. Immunoblots revealed a upward trend in the three types of Akt subsets and the most obvious change was observed in the expression of Akt1.These findings suggested the Akt pathway involved in the pathogenesis of MCD.
2. A similar expression levels of Bad was detected in both CTX and MCD specimens, but we detected a significant increase of phosphorylated Bad (P-Bad) in MCD.
3. We observed representative immunoblot double-bands of ERK1/2 (42 kDa and 44 kDa) in homogenates from MCD and CTX, and the ERK1/2 protein level in MCD was much high than that in CTX.
Ⅲ. Expression and cellular distribution of VEGF-D in MCD
1. We detected the VEGF-D mRNA and protein expression in total homogenates of CTX; immunohistochemical results revealed a weak immunostaining was restricted to pyramidal neurons.
2. VEGF-D mRNA and protein levels were significantly increased in MCD compared with CTX. The immunostaining results demonstrated:①In TSC specimens, the high-level expression of VEGF-D protein was mainly localized within most of DNs,GCs, and reactive astrocytes; confocal images showed no co-localization of VEGF-D with NeuN, NF-200, GFAP and CD68 in MNs;②In FCD subtype, we found the VEGF-D was highly expressed in DNs,GNs,BCs and reactive astrocytes; the results of double-labeling showed no co-localization of VEGF-D with NF-200 and GFAP in these MNs;
V.The modulatory effects of VEGF-C on NMDAR function in MNs from MCD rats model.
1. We employed a well-established MCD rat model by in utero irradiation, and In vitro brain slices were obtained from these rats.The results of patch clamp recording showed:①With step-wise depolarization, MNs of MCD responded initially with action potentials (APs),but the shape of APs varied, the spike frequence adaption (SFA) was damaged,even disappeared,suggesting a immature characteristic in electrophysiology.②We successfully observed evoked-EPSC in MNs, and the amplitude and area of evoked-EPSC of MNs were clearly increased,suggesting a complex response comprising multiple inward currents;③The current ratio of NMDAR/AMPAR was increased in MNs compared with pyramidal neurons from normal control, mainly induced by the increase of NMDAR-mediated current, revealing a upregulation of the function of NMDAR.
2. The immunostaining results showed there was a significant increase of VEGF-C and VEGFR-2 expression in MNs from MCD rats; the expression level of VEGFR-3 was also statistically increased.
3. 100μmol/L NMDA could evoke the inward. After treatment with 100 ng/mL VEGF-C,100μmol/L NMDA caused a pronounced increase of transmembrane current in MNs, the data indicated a synergistic action between VEGF-C and NMDAR in MNs from MCD rats. We also found that Ki8751 (1 nmol/L), a VEGFR-2 inhibitor, abolished the upregulative effects of VEGF-C on NMDA receptor channel, this finding illuminated that the potentiation effects of VEGF-C on NMDAR is mediated by VEGFR-2.
In summary, in the present study, we found an obvious increase of VEGF-C and it receptors mRNA and protein expression in MCD specimens compared with the control sample. The high-level expression of protein, including VEGF-C and its receptors, was mainly localized within MNs(DNs,GNs,GCs and BCs). Subsequently, we observed the activation of signaling molecule Akt, Bad, and ERK in MCD, which suggested the VEGF-C miediated anti-apoptotic mechanism might involve in the pathogenesis of MCD. Moreover, our data showed that NMDA receptor-mediated current is markedly potentiated by VEGF-C in the MNs from MCD rats, which suggests that a synergistic action between VEGF-C and glutamate systems exists in MCD. Altogether, these findings illuminate the VEGF-C system involves in the pathogenesis of MCD.
VEGF-C是公认的淋巴系统发生调节因子,通过激活两个特异性受体VEGFR-2, VEGFR-3发挥生物学效应,其中VEGFR-3为VEGF-C的主力型受体。在正常情况下,VEGF-C及VEGFR-3局限性表达于淋巴内皮,而它的另一受体VEGFR-2则在血管内皮和淋巴内皮上均有表达,发挥其促脉管系统发生的作用。
由于脑内淋巴系统缺如,既往很少有研究关注VEGF-C在中枢神经系统的作用。直到2006年,有学者发现当小鼠敲除VEGF-C后,表达VEGFR-3受体的神经前体细胞增殖明显受阻,而且胚胎视神经中少突胶质细胞前体细胞选择性缺失,证明VEGF-C信号通路参与了中枢神经系统的生长发育、发挥神经营养功能。近两年来陆续有研究揭示了VEGF-C信号系统在神经系统中的新作用,如VEGF-C/VEGFR-3能够介导胶质前体细胞的增殖,迁移。新近报道脑缺血后,定位于海马齿状回颗粒细胞层最内侧带的细胞高度表达VEGFR-3,这些细胞同时表达与细胞迁移相关的神经前体细胞标记物PSA-NCAM,表明VEGF-C系统参与脑缺血后海马神经发生。此外,脑卒中后,患侧SVZ区内VEGFR-3的表达水平显著上调,表型鉴定实验证实绝大多数表达VEGFR-3的细胞主要为具有旺盛增殖活力的神经祖细胞,另有小部分VEGFR-3阳性细胞为具备迁徙能力的神经前体细胞。在本研究预实验中,我们发现手术切除的MCD标本中VEGF-C mRNA表达上调。那么,VEGF-C及其受体在MCD中的具体表达模式如何?是否表达于异构神经元?高表达的VEGF-C是否参与MCD所致癫痫发作?这些问题值得我们关注。
为了阐明VEGF-C在MCD中的作用及其可能的机制,首先,我们利用半定量RT-PCR、western blot、免疫组织化学技术观察了VEGF-C及其受体在MCD(TSC与FCD亚型)临床标本中的表达分布情况；其次,观测了VEGF-C受体信号通路下游相关信号分子的表达活化情况；第三,观察了与VEGF-C高度同源的VEGF-D在MCD中的表达情况；最后,为了直观的了解VEGF-C对MCD致痫效应的影响,我们建立MCD大鼠模型,利用膜片钳全细胞记录技术,观察VEGF-C对MCD大鼠异构神经元兴奋性谷氨酸受体(NMDA受体)的调节效应。通过这些研究,我们得到以下结果：
一、VEGF-C及其受体在MCD中的表达分布
1.通过RT-PCR, western blot分析,从基因和蛋白水平证明正常皮层中不同程度地表达VEGF-C、VEGFR-2及VEGFR-3;免疫组化结果表明VEGF-C及其受体主要分布于皮层锥体神经元；
2.在TSC皮层结节组织匀浆中,VEGF-C及其受体VEGFR-2、VEGFR-3的mRNA及蛋白表达水平较正常对照组明显升高。免疫组化结果显示,VEGF-C, VEGFR-2及VEGFR-3特征性的高表达于DNs、GCs及反应型胶质细胞；免疫荧光双标结果表明：①绝大多数VEGF-C免疫阳性的DNs, GCs共表达神经丝蛋白标记物NF-200；然而未见共表达与另一种神经元标记物NeuN以及星形胶质细胞特异性标志物GFAP共表达；TSC组织中可见CD68阳性的巨噬细胞浸润,但是这些细胞不表达VEGF-C;②VEGFR-2与VEGFR-3的细胞定位表达模式类似,多数VEGFR-2或VEGFR-3免疫阳性的DNs与NF-200共表达,而共表达VEGFR-2 (VEGFR-3)与NF-200的GCs略少,未见VEGFR-2 (VEGFR-3)免疫阳性的DNs, GCs共表达GFAP。
3.在另一常见的MCD亚型-FCD中,VEGF-C及受体VEGFR-2、VEGFR-3的mRNA及蛋白表达水平较正常对照组同样显著升高。VEGF-C及其受体高表达于DNs, GNs,BCs及胶质细胞。免疫双标结果显示：①VEGF-C免疫阳性的异构神经元不表达GFAP;与TSC中表达模式不同的是,FCD中的DNs, GNs, BCs能够同时表达VEGF-C与NeuN,提示其具有成熟神经元属性。②VEGFR-3免疫阳性的异构神经元既不表达NF-200,又不表达GFAP,表明其特殊的分子表型。
二、MCD病灶内VEGF-C下游相关信号通路的表达活化
1. RT-PCR结果表明,MCD中Akt的基因表达上调；利用western blot观察Akt蛋白亚型表达,发现三种Akt亚型的表达均上调,其中Akt-1的表达上调最为明显,表明Akt信号通路参与了MCD的病理生理机制。
2.正常皮层及MCD中均可见Bad的表达,且表达水平基本相同；然而,MCD病灶内的磷酸化Bad (p-Bad)表达显著升高,提示MCD中抗凋亡通路活化。
3.ERK表达于MCD病灶组织和正常皮层中,呈现特征性的44kDa和42 kDa双条带(ERK 1/2),并可见ERK1/2在MCD中的表达明显上调。
三、VEGF-D在MCD中的表达分布
1.正常皮层组织匀浆中,可探及VEGF-D mRNA及蛋白表达；免疫组织化学结果表明VEGF-D主要表达于锥体神经元,且免疫反应程度较轻。
2.MCD组织匀浆中,VEGF-D mRNA及蛋白表达水平上调。免疫组织化学实验发现：①TSC亚型中VEGF-D表达于多数DNs、GCs及反应性胶质细胞,但免疫阳性强度不一；免疫双标未发现VEGF-D阳性异构神经元与NeuN, NF-200, GFAP及CD68共表达；②FCD亚型中可见VEGF-D表达于DNs, GNs, BCs及反应性胶质细胞,免疫双标结果表明VEGF-D免疫阳性的异构神经元不表达成神经纤维蛋白NF-200,亦不表达成熟星型胶质细胞标记物GFAP。
四、VEGF-C对MCD大鼠异构神经元NMDA受体功能的调节效应
1.建立MCD动物模型,制备活体脑片后进行膜片钳全细胞记录,结果表明①MCD大鼠异构神经元可产生动作电位,但动作电位形态各异,峰率适应性降低或消失,提示其电生理功能发育不完善；②异构神经元中可记录到evoked-EPSC,但电流形态明显发生改变,通道电流成分混杂,电流面积明显增加,提示MCD异构神经元突触传递效能改变；③MCD异构神经元中NMDA/AMPA受体介导电流的比率显著上调,这种比率改变主要是由NMDA受体电流成分增多引起,表明异构神经元中NMDA受体功能上调。
2.MCD大鼠异构神经元中VEGF-C及其受体VEGFR-2的表达明显上调,VEGFR-3的表达亦有增加。
3.100μmol/L NMDA作用于异构神经元后,可诱发典型的快反应电流,经过100ng/mL VEGF-C孵育后,相同浓度NMDA诱发的电流显著增大,提示VEGF-C对NMDA受体产生协同作用；当给予VEGFR-2受体阻断剂Ki8751预处理后,这种协同效应被阻断,表明VEGF-C对NMDA受体的调节效应主要是由VEGFR-2受体介导的。
综上所述,本研究结果表明：第一、人MCD组织中VEGF-C及其受体VEGFR-2,VEGFR-3表达显著上调,且特征性地高表达于异构神经元；第二、MCD中VEGF-C下游通路中抗凋亡相关信号分子激活,有益地提示VEGF-C可能通过抗凋亡相关信号通路调节MCD内异构神经元的生物学行为；第三、VEGF-D在MCD中的表达上调；第四、VEGF-C通过VEGFR-2增强MCD大鼠异构神经元NMDA受体活性,易化异构神经元的兴奋性。总之,我们的结果提示VEGF-C信号通路参与了MCD的病理生理机制。
Malformations of cortical development (MCD) represents a well-recognized cause of intractable epilepsy, with a poor response to the currently available anti-epileptic drugs (AEDs). With the development of high-resolution imageology, the detection rate of MCD is continuously rising during the past decade in clinic.It has been demonstrated that approximately 40 percent of adult patients and more than 80 percent of pediatric with intractable epilepsy are associated with MCD.Thus, the pathogenesis of MCD has became a central concern in the field of intractable epilepsy. MCD encompasses a large spectrum of disorders related to abnormal cortical development, including tuberous sclerosis complex (TSC), focal cortical dysplasia (FCD), microgyria, congenital agyria and Sturge-Weber syndrome. Among these subsets, two of the most common components are TSC and FCD. Despite the fact that these subsets with varied genetic etiologies, anatomic abnormalities, and clinical manifestations, they often share similar histological features such as disorganization or lack of the normal six-layered cortical lamination structure, and the occurrence of malformed neurons (MNs) known as dysplastic neurons (DNs), giant neurons (GNs),balloon cells (BCs) and giant cells (GCs). Mounting clinical evidence has revealed that epilepsy readily occurs in patients with MNs, and appears drug resistance in the early stage, suggesting the dominant role of MNs in epileptogenesis of MCD. Until now, the pathogenesis of MNs remains unclear. A more popular view is that MNs is a malformation due to abnormal migration,mature and cell death during embryonic brain development. Moreover, recent pathological studies on surgical or autoptic human tissue suggest a role for perinatal and early postnatal brain injury in the formation of MNs. In spite of the fact that there are many reports on the subject of MCD, the exact mechanism of this disease has not yet been elucidated.
VEGF-C,a well-recognized regulator of lymphangiogenesis, binds to and activates the specific receptors VEGFR-2 and VEGFR-3, and the affinity of VEGFR-3 is much more than that of VEGFR-2. Previous studies show VEGF-C and VEGFR-3 are restrictly expresssed in lymphatic endothelial cells, and VEGFR-2 is expressed in both vascular endothelial cells and lymphatic endothelial cells.
Due to the deficiency of the lymphatic system in the brain, few studies have focused on the role of VEGF-C in the CNS. Until to 2006, an important study demonstrats that VEGF-C and VEGFR-3 are present in the neural progenitor cells of Xenopus laevis and mouse embryos. Furthermore, a lack of VEGF-C results in a severe defect in the proliferation of neuronal progenitor cells expressing VEGFR-3, suggesting that VEGF-C is a trophic factor in neuronal progenitor cells in the vertebrate brain. Recently, several studies revealed VEGF-C mediate multiple effects in CNS. For example,activation of the VEGF-C/VEGFR-3 signaling pathway was shown to mediate proliferation and chemotaxis in glial precursor cells. In addition, after a short period of brain ischemia, the innermost layer of granular cells in the hippocampal dental gyrus displayed increased levels of VEGFR-3 expression, these VEGFR-3 immunoreactive cells are also expressed PSA-NCAM, this might be related to neurogenesis in the hippocampus after cerebral ischemia. A recent study showed that the expression of VEGFR-3 increased in subventricular zone, and most of VEGFR-3 positive cells were neural progenitors, were highly proliferative. To our knowledge, there have been no reports about the roles of VEGF-C in in seizure disorders, especially those induced by MCD. In our preliminary experiment,we found that VEGF-C mRNA exhibited high levels in MCD. However, the detailed expression pattern of VEGF-C and it receptors in MCD, and the role of VEGF-C signal system in the epileptogenesis of MCD are unclear.
To address these concerns:First,we identified the distribution of VEGF-C system in MCD from patients with medically intractable epilepsy by means of RT-PCR, western blot and immunohistochemistry. Second,we tested the expression and activation of signaling molecule involved in VEGF-C pathway. Third, we investigated the expression and cellular distribution of VEGF-D,which is highly homologous with VEGF-C, in MCD. Fourth, to reveal the role of VEGF-C in the epileptogenesis of MCD, a MCD rat model was employed, and we investigated the effect of VEGF-C on NMDA receptor(NMDAR)of MNs by using whole-cell patch-clamp recordings technique on brain slice of MCD rat model. The results show as following:
Ⅰ. Expression and cellular distribution of VEGF-C and VEGF receptors 2 and 3 in MCD
1. RT-PCR and western blot analysis showed VEGF-C and its receptors mRNA and protein expression in total homogenates of normal control cortex (CTX); immunohistochemical experiments revealed immunostaining was mainly observed in pyramidal neurons.
2. There was a statistically significant increase of VEGF-C, VEGFR-2 and VEGFR-3 mRNA and protein levels in cortical tubers of TSC, in comparison with CTX. The immunostaining results demonstrated that the high-level expression of protein, including VEGF-C and its receptors, was mainly localized within GCs, DNs, and reactive astrocytes. The results of double-labeling immunofluorescence showed:①the co-localization of VEGF-C immunostaining with the neuronal marker NF-200 in majority of DNs and GCs. Intriguingly, we could not detect any co-immunostaining of VEGF-C with another neuronal marker, NeuN, in both DNs and GCs in all cases. Also, GFAP and CD68 immunostaining was not detected in any VEGF-C-positive DNs and GCs.②VEGFR-2 and VEGFR-3 shared a similar cellular distribution pattern in TSC, VEGFR-2 (VEGFR-3) immunoreactivity co-localized with NF-200 in the majority of DNs,the frequency of VEGFR-2 (VEGFR-3) and NF-200 co-expression in GCs was less than that in DNs. VEGFR-2 (VEGFR-3) and GFAP co-immunolabeling was not detected in any DNs and GCs, although prominent co-labeling of reactive astrocytes was noted in all of the TSC cases examined.
3. We found an obvious increase of VEGF-C and it receptors mRNA and protein expression in FCD specimens compared with CTX. The high-level expression of protein, including VEGF-C and its receptors, was mainly localized within DNs,GNs,GCs and reactive astrocytes. The results of double-labeling immunofluorescence showed:①GFAP and VEGF-C co-immunolabeling was not detected in any MNs. Interestingly,we detected co-immunostaining of VEGF-C with NeuN, which is usually recognized as a marker of mature neurons, in MNs.②VEGFR-3 and GFAP/NF-200 co-immunolabeling was not observed in any MNs,suggesting a complex phenotype of MNs.
Ⅱ. The expression and activation of signaling molecule involved in VEGF-C pathway
1. In comparison with CTX, RT-PCR analysis showed a increase of Akt mRNA levels in MCD specimens. Immunoblots revealed a upward trend in the three types of Akt subsets and the most obvious change was observed in the expression of Akt1.These findings suggested the Akt pathway involved in the pathogenesis of MCD.
2. A similar expression levels of Bad was detected in both CTX and MCD specimens, but we detected a significant increase of phosphorylated Bad (P-Bad) in MCD.
3. We observed representative immunoblot double-bands of ERK1/2 (42 kDa and 44 kDa) in homogenates from MCD and CTX, and the ERK1/2 protein level in MCD was much high than that in CTX.
Ⅲ. Expression and cellular distribution of VEGF-D in MCD
1. We detected the VEGF-D mRNA and protein expression in total homogenates of CTX; immunohistochemical results revealed a weak immunostaining was restricted to pyramidal neurons.
2. VEGF-D mRNA and protein levels were significantly increased in MCD compared with CTX. The immunostaining results demonstrated:①In TSC specimens, the high-level expression of VEGF-D protein was mainly localized within most of DNs,GCs, and reactive astrocytes; confocal images showed no co-localization of VEGF-D with NeuN, NF-200, GFAP and CD68 in MNs;②In FCD subtype, we found the VEGF-D was highly expressed in DNs,GNs,BCs and reactive astrocytes; the results of double-labeling showed no co-localization of VEGF-D with NF-200 and GFAP in these MNs;
V.The modulatory effects of VEGF-C on NMDAR function in MNs from MCD rats model.
1. We employed a well-established MCD rat model by in utero irradiation, and In vitro brain slices were obtained from these rats.The results of patch clamp recording showed:①With step-wise depolarization, MNs of MCD responded initially with action potentials (APs),but the shape of APs varied, the spike frequence adaption (SFA) was damaged,even disappeared,suggesting a immature characteristic in electrophysiology.②We successfully observed evoked-EPSC in MNs, and the amplitude and area of evoked-EPSC of MNs were clearly increased,suggesting a complex response comprising multiple inward currents;③The current ratio of NMDAR/AMPAR was increased in MNs compared with pyramidal neurons from normal control, mainly induced by the increase of NMDAR-mediated current, revealing a upregulation of the function of NMDAR.
2. The immunostaining results showed there was a significant increase of VEGF-C and VEGFR-2 expression in MNs from MCD rats; the expression level of VEGFR-3 was also statistically increased.
3. 100μmol/L NMDA could evoke the inward. After treatment with 100 ng/mL VEGF-C,100μmol/L NMDA caused a pronounced increase of transmembrane current in MNs, the data indicated a synergistic action between VEGF-C and NMDAR in MNs from MCD rats. We also found that Ki8751 (1 nmol/L), a VEGFR-2 inhibitor, abolished the upregulative effects of VEGF-C on NMDA receptor channel, this finding illuminated that the potentiation effects of VEGF-C on NMDAR is mediated by VEGFR-2.
In summary, in the present study, we found an obvious increase of VEGF-C and it receptors mRNA and protein expression in MCD specimens compared with the control sample. The high-level expression of protein, including VEGF-C and its receptors, was mainly localized within MNs(DNs,GNs,GCs and BCs). Subsequently, we observed the activation of signaling molecule Akt, Bad, and ERK in MCD, which suggested the VEGF-C miediated anti-apoptotic mechanism might involve in the pathogenesis of MCD. Moreover, our data showed that NMDA receptor-mediated current is markedly potentiated by VEGF-C in the MNs from MCD rats, which suggests that a synergistic action between VEGF-C and glutamate systems exists in MCD. Altogether, these findings illuminate the VEGF-C system involves in the pathogenesis of MCD.
引文
1. Wong M. Mechanisms of epileptogenesis in tuberous sclerosis complex and related malformations of cortical development with abnormal glioneuronal proliferation. Epilepsia 2008; 49:8-21.
2. Madan N, Grant PE. New directions in clinical imaging of cortical dysplasias. Epilepsia 2009; 50 Suppl 9:9-18.
3. Mathew T, Srikanth SG, Satishchandra P. Malformations of cortical development (MCDs) and epilepsy:experience from a tertiary care center in south India. Seizure 2010; 19:147-152.
4. Najm IM, Tilelli CQ, Oghlakian R. Pathophysiological mechanisms of focal cortical dysplasia:a critical review of human tissue studies and animal models. Epilepsia 2007; 48 Suppl 2:21-32.
5. Andres M, Andre VM, Nguyen S, Salamon N, Cepeda C, Levine MS et al. Human cortical dysplasia and epilepsy:an ontogenetic hypothesis based on volumetric MRI and NeuN neuronal density and size measurements. Cereb Cortex 2005; 15:194-210.
6. Storkebaum E, Carmeliet P. VEGF:a critical player in neurodegeneration.J Clin Invest 2004;113:14-18.
7. Partanen TA, Arola J, Saaristo A, Jussila L, Ora A, Miettinen M et al. VEGF-C and VEGF-D expression in neuroendocrine cells and their receptor, VEGFR-3, in fenestrated blood vessels in human tissues. FASEB J 2000; 14:2087-2096.
8. Le Bras B, Barallobre MJ, Homman-Ludiye J, Ny A, Wyns S, Tammela T et al. VEGF-C is a trophic factor for neural progenitors in the vertebrate embryonic brain. Nat Neurosci 2006;9:340-348.
9. Shin YJ, Choi JS, Choi JY, Cha JH, Chun MH, Lee MY. Enhanced expression of vascular endothelial growth factor receptor-3 in the subventricular zone of stroke-lesioned rats. Neurosci Lett 2010; 469:194-198.
10. Choi JS, Shin YJ, Lee JY, Yun H, Cha JH, Choi JY et al. Expression of vascular endothelial growth factor receptor-3 mRNA in the rat developing forebrain and retina. J Comp Neurol 2010; 518:1064-1081.
11. Shin YJ, Choi JS, Lee JY, Choi JY, Cha JH, Chun MH et al. Differential regulation of vascular endothelial growth factor-C and its receptor in the rat hippocampus following transient forebrain ischemia. Acta Neuropathol 2008; 116:517-527.
12. Kranich S, Hattermann K, Specht A, Lucius R, Mentlein R. VEGFR-3/Flt-4 mediates proliferation and chemotaxis in glial precursor cells. Neurochem Int 2009; 55:747-753.
13. Palmini A, Najm I, Avanzini G, Babb T, Guerrini R, Foldvary-Schaefer N et al. Terminology and classification of the cortical dysplasias. Neurology 2004; 62:S2-8.
14. Napolioni V, Moavero R, Curatolo P. Recent advances in neurobiology of Tuberous Sclerosis Complex. Brain Dev 2009; 31:104-113.
15. Liu S, An N, Yang H, Yang M, Hou Z, Liu L et al. Pediatric intractable epilepsy syndromes:reason for early surgical intervention. Brain Dev 2007; 29:69-78.
16. Grau SJ, Trillsch F, Herms J, Thon N, Nelson PJ, Tonn JC et al. Expression of VEGFR3 in glioma endothelium correlates with tumor grade. J Neurooncol 2007; 82: 141-150.
17. Boer K, Troost D, Spliet WG, van Rijen PC, Gorter JA, Aronica E. Cellular distribution of vascular endothelial growth factor A (VEGFA) and B (VEGFB) and VEGF receptors 1 and 2 in focal cortical dysplasia type ⅡB. Acta Neuropathol 2008; 115:683-696.
18. Croll SD, Goodman JH, Scharfman HE. Vascular endothelial growth factor (VEGF) in seizures:a double-edged sword. Adv Exp Med Biol 2004; 548:57-68.
19. Gollmer JC, Ladoux A, Gioanni J, Paquis P, Dubreuil A, Chatel M et al. Expression of vascular endothelial growth factor-b in human astrocytoma. Neuro Oncol 2000; 2: 80-86.
20. Huber H, Eggert A, Janss AJ, Wiewrodt R, Zhao H, Sutton LN et al. Angiogenic profile of childhood primitive neuroectodermal brain tumours/medulloblastomas. Eur J Cancer 2001; 37:2064-2072.
21. Jenny B, Harrison JA, Baetens D, Tille JC, Burkhardt K, Mottaz H et al. Expression and localization of VEGF-C and VEGFR-3 in glioblastomas and haemangioblastomas. J Pathol 2006; 209:34-43.
22. Schonberger A, Niehusmann P, Urbach H, Majores M, Grote A, Holthausen H et al. Increased frequency of distinct TSC2 allelic variants in focal cortical dysplasias with balloon cells and mineralization. Neuropathology 2009; 29:559-565.
23. Crino PB. Focal brain malformations:a spectrum of disorders along the mTOR cascade. Novartis Found Symp 2007; 288:260-272; discussion 272-281.
24. Brugarolas JB, Vazquez F, Reddy A, Sellers WR, Kaelin WG, Jr. TSC2 regulates VEGF through mTOR-dependent and-independent pathways. Cancer Cell 2003; 4:147-158.
25. El-Hashemite N, Walker V, Zhang H, Kwiatkowski DJ. Loss of Tscl or Tsc2 induces vascular endothelial growth factor production through mammalian target of rapamycin. Cancer Res 2003; 63:5173-5177.
26. Mahimainathan L, Ghosh-Choudhury N, Venkatesan B, Das F, Mandal CC, Dey N et al. TSC2 deficiency increases pten via HIF1alpha. J Biol Chem 2009.
27. Simiantonaki N, Jayasinghe C, Michel-Schmidt R, Peters K, Hermanns MI, Kirkpatrick CJ. Hypoxia-induced epithelial VEGF-C/VEGFR-3 upregulation in carcinoma cell lines. Int J Oncol 2008; 32:585-592.
28. Meierkord H, Holtkamp M. Status epilepticus:an independent outcome predictor after cerebral anoxia. Neurology 2008; 70:2015-2016; author reply 2015-2016.
29. Boer K, Jansen F, Nellist M, Redeker S, van den Ouweland AM, Spliet WG et al. Inflammatory processes in cortical tubers and subependymal giant cell tumors of tuberous sclerosis complex. Epilepsy Res 2008; 78:7-21.
30. Rodgers KM, Hutchinson MR, Northcutt A, Maier SF, Watkins LR, Barth DS. The cortical innate immune response increases local neuronal excitability leading to seizures. Brain 2009; 132:2478-2486.
31. Ristimaki A, Narko K, Enholm B, Joukov V, Alitalo K. Proinflammatory cytokines regulate expression of the lymphatic endothelial mitogen vascular endothelial growth factor-C. J Biol Chem 1998; 273:8413-8418.
32. Watari K, Nakao S, Fotovati A, Basaki Y, Hosoi F, Bereczky B et al. Role of macrophages in inflammatory lymphangiogenesis:Enhanced production of vascular endothelial growth factor C and D through NF-kappaB activation. Biochem Biophys Res Commun 2008; 377:826-831.
33. Tang RF, Wang SX, Zhang FR, Peng L, Xiao Y, Zhang M. Interleukin-1alpha,6 regulate the secretion of vascular endothelial growth factor A, C in pancreatic cancer. Hepatobiliary Pancreat Dis Int 2005; 4:460-463.
34. Ravizza T, Boer K, Redeker S, Spliet WG, van Rijen PC, Troost D et al. The IL-1beta system in epilepsy-associated malformations of cortical development. Neurobiol Dis 2006; 24:128-143.
35. Pajusola K, Aprelikova O, Korhonen J, Kaipainen A, Pertovaara L, Alitalo R et al. FLT4 receptor tyrosine kinase contains seven immunoglobulin-like loops and is expressed in multiple human tissues and cell lines. Cancer Res 1992; 52:5738-5743.
36. Aprelikova O, Pajusola K, Partanen J, Armstrong E, Alitalo R, Bailey SK et al. FLT4, a novel class Ⅲ receptor tyrosine kinase in chromosome 5q33-qter. Cancer Res 1992; 52: 746-748.
37. Brockington A, Wharton SB, Fernando M, Gelsthorpe CH, Baxter L, Ince PG et al. Expression of vascular endothelial growth factor and its receptors in the central nervous system in amyotrophic lateral sclerosis. J Neuropathol Exp Neurol 2006; 65:26-36.
38. Hlobilkova A, Ehrmann J, Knizetova P, Krejci V, Kalita O, Kolar Z. Analysis of VEGF, Flt-1, Flk-1, nestin and MMP-9 in relation to astrocytoma pathogenesis and progression. Neoplasma 2009; 56:284-290.
39. Krum JM, Mani N, Rosenstein JM. Roles of the endogenous VEGF receptors flt-1 and flk-1 in astroglial and vascular remodeling after brain injury. Exp Neurol 2008; 212: 108-117.
40. Raab S, Plate KH. Different networks, common growth factors:shared growth factors and receptors of the vascular and the nervous system. Acta Neuropathol 2007; 113: 607-626.
41. Pietsch T, Valter MM, Wolf HK, von Deimling A, Huang HJ, Cavenee WK et al. Expression and distribution of vascular endothelial growth factor protein in human brain tumors. Acta Neuropathol 1997; 93:109-117.
42. Rigau V, Morin M, Rousset MC, de Bock F, Lebrun A, Coubes P et al. Angiogenesis is associated with blood-brain barrier permeability in temporal lobe epilepsy. Brain 2007; 130:1942-1956.
43. Miyata H, Chiang AC, Vinters HV. Insulin signaling pathways in cortical dysplasia and TSC-tubers:tissue microarray analysis. Ann Neurol 2004; 56:510-519.
44. Boer K, Lucassen PJ, Spliet WG, Vreugdenhil E, van Rijen PC, Troost D et al. Doublecortin-like (DCL) expression in focal cortical dysplasia and cortical tubers. Epilepsia 2009.
45. Qu M, Aronica E, Boer K, Fallmar D, Kumlien E, Nister M et al. DLG3/SAP102 protein expression in malformations of cortical development:a study of human epileptic cortex by tissue microarray. Epilepsy Res 2009; 84:33-41.
46. Sugiura C, Miyata H, Ueda M, Ohama E, Vinters HV, Ohno K. Immunohistochemical expression of fibroblast growth factor (FGF)-2 in epilepsy-associated malformations of cortical development (MCDs). Neuropathology 2008; 28:372-381.
47. Toering ST, Boer K, de Groot M, Troost D, Heimans JJ, Spliet WG et al. Expression patterns of synaptic vesicle protein 2A in focal cortical dysplasia and TSC-cortical tubers. Epilepsia 2009; 50:1409-1418.
48. Susan D. Croll,, Richard M. Ransohoff, Ning Caia, Qing Zhanga, Francis J. Martina, Tao Weib, Lora J. Kasselmana, Jennifer Kintnera, Andrew J. Murphya, George D. Yancopoulosa and Stanley J. Wieganda. VEGF-mediated inflammation precedes angiogenesis in adult brain. Experimental Neurology 2004; 187:388-402.
49. Vezzani A. VEGF and seizures:cross-talk between endothelial and neuronal environments. Epilepsy Curr 2005;5:72-74.
50. Vezzani A, Granata T. Brain inflammation in epilepsy:experimental and clinical evidence. Epilepsia 2005;46:1724-1743.
51. van Vliet EA, da Costa Araujo S, Redeker S, van Schaik R, Aronica E, Gorter JA. Blood-brain barrier leakage may lead to progression of temporal lobe epilepsy. Brain 2007;130:521-534.
52. Maurer MH, Tripps WK, Feldmann RE, Jr., Kuschinsky W. Expression of vascular endothelial growth factor and its receptors in rat neural stem cells. Neurosci Lett 2003; 344:165-168.
53. Salven P, Mustjoki S, Alitalo R, Alitalo K, Rafii S. VEGFR-3 and CD133 identify a population of CD34+ lymphatic/vascular endothelial precursor cells. Blood 2003;101: 168-172.
54. Ma YH, Mentlein R, Knerlich F, Kruse ML, Mehdorn HM, Held-Feindt J. Expression of stem cell markers in human astrocytomas of different WHO grades. J Neurooncol 2008; 86:31-45.
55. Crino PB, Trojanowski JQ, Dichter MA, Eberwine J. Embryonic neuronal markers in tuberous sclerosis:single-cell molecular pathology. Proc Natl Acad Sci U S A 1996; 93: 14152-14157.
56. Jozwiak J, Jozwiak S, Skopinski P. Immunohistochemical and microscopic studies on giant cells in tuberous sclerosis. Histol Histopathol 2005; 20:1321-1326.
57. Fauser S, Becker A, Schulze-Bonhage A, Hildebrandt M, Tuxhorn I, Pannek HW et al. CD34-immunoreactive balloon cells in cortical malformations. Acta Neuropathol 2004; 108:272-278.
58. Boer K, Lucassen PJ, Spliet WG, Vreugdenhil E, van Rijen PC, Troost D et al. Doublecortin-like (DCL) expression in focal cortical dysplasia and cortical tubers. Epilepsia 2009; 50:2629-2637.
59. Kranich S, Hattermann K, Specht A, Lucius R, Mentlein R. VEGFR-3/Flt-4 mediates proliferation and chemotaxis in glial precursor cells. Neurochem Int 2009.
1. Chamberlain WA, Prayson RA. Focal cortical dysplasia type II (malformations of cortical development) aberrantly expresses apoptotic proteins. Appl Immunohistochem Mol Morphol 2008; 16:471-476.
2. Ruch C, Skiniotis G, Steinmetz MO, Walz T, Ballmer-Hofer K. Structure of a VEGF-VEGF receptor complex determined by electron microscopy. Nat Struct Mol Biol 2007; 14:249-250.
3. Chien MH, Ku CC, Johansson G, Chen MW, Hsiao M, Su JL et al. Vascular endothelial growth factor-C (VEGF-C) promotes angiogenesis by induction of COX-2 in leukemic cells via the VEGF-R3/JNK/AP-1 pathway. Carcinogenesis 2009; 30:2005-2013.
4. Abid MR, Guo S, Minami T, Spokes KC, Ueki K, Skurk C et al. Vascular endothelial growth factor activates PI3K/Akt/forkhead signaling in endothelial cells. Arterioscler Thromb Vasc Biol 2004; 24:294-300.
5. Andres M, Andre VM, Nguyen S, Salamon N, Cepeda C, Levine MS et al. Human cortical dysplasia and epilepsy:an ontogenetic hypothesis based on volumetric MRI and NeuN neuronal density and size measurements. Cereb Cortex 2005; 15:194-210.
6. Najm IM, Tilelli CQ, Oghlakian R. Pathophysiological mechanisms of focal cortical dysplasia:a critical review of human tissue studies and animal models. Epilepsia 2007; 48 Suppl2:21-32.
7. Sen A, Thom M, Martinian L, Harding B, Cross JH, Nikolic M et al. Pathological tau tangles localize to focal cortical dysplasia in older patients. Epilepsia 2007; 48: 1447-1454.
8. Cepeda C, Andre VM, Levine MS, Salamon N, Miyata H, Vinters HV et al. Epileptogenesis in pediatric cortical dysplasia:the dysmature cerebral developmental hypothesis. Epilepsy Behav 2006; 9:219-235.
9. Muller-Deile J, Worthmann K, Saleem M, Tossidou I, Haller H, Schiffer M. The balance of autocrine VEGF-A and VEGF-C determines podocyte survival. Am J Physiol Renal Physiol 2009; 297:F1656-1667.
10. Kirkin V, Thiele W, Baumann P, Mazitschek R, Rohde K, Fellbrich G et al. MAZ51, an indolinone that inhibits endothelial cell and tumor cell growth in vitro, suppresses tumor growth in vivo. Int J Cancer 2004; 112:986-993.
11. Duronio V. The life of a cell:apoptosis regulation by the PI3K/PKB pathway. Biochem J 2008; 415:333-344.
12. Le Page C, Koumakpayi IH, Alam-Fahmy M, Mes-Masson AM, Saad F. Expression and localisation of Akt-1, Akt-2 and Akt-3 correlate with clinical outcome of prostate cancer patients. Br J Cancer 2006; 94:1906-1912.
13. Price J, Zaidi AK, Bohensky J, Srinivas V, Shapiro IM, Ali H. Akt-1 mediates survival of chondrocytes from endoplasmic reticulum-induced stress. J Cell Physiol; 222: 502-508.
14. Chipuk JE, Moldoveanu T, Llambi F, Parsons MJ, Green DR. The BCL-2 family reunion. Mol Cell; 37:299-310.
15. Patel MP, Masood A, Patel PS, Chanan-Khan AA. Targeting the Bcl-2. Curr Opin Oncol 2009; 21:516-523.
16. Zhu Y, Hoell P, Ahlemeyer B, Krieglstein J. PTEN:a crucial mediator of mitochondria-dependent apoptosis. Apoptosis 2006; 11:197-207.
17. Narasimhan P, Liu J, Song YS, Massengale JL, Chan PH. VEGF Stimulates the ERK 1/2 signaling pathway and apoptosis in cerebral endothelial cells after ischemic conditions. Stroke 2009; 40:1467-1473.
18. Salameh A, Galvagni F, Bardelli M, Bussolino F, Oliviero S. Direct recruitment of CRK and GRB2 to VEGFR-3 induces proliferation, migration, and survival of endothelial cells through the activation of ERK, AKT, and JNK pathways. Blood 2005; 106: 3423-3431.
19. Girault JA, Valjent E, Caboche J, Herve D. ERK2:a logical AND gate critical for drug-induced plasticity? Curr Opin Pharmacol 2007; 7:77-85.
20. Suzuki M, Satoh A, Ide H, Tamura K. Transgenic Xenopus with prxl limb enhancer reveals crucial contribution of MEK/ERK and PI3K/AKT pathways in blastema formation during limb regeneration. Dev Biol 2007; 304:675-686.
21. Kilic E, Kilic U, Wang Y, Bassetti CL, Marti HH, Hermann DM. The phosphatidylinositol-3 kinase/Akt pathway mediates VEGF's neuroprotective activity and induces blood brain barrier permeability after focal cerebral ischemia. FASEB J 2006; 20:1185-1187.
22. Frebel K, Wiese S. Signalling molecules essential for neuronal survival and differentiation. Biochem Soc Trans 2006; 34:1287-1290.
23. Hu P, Han Z, Couvillon AD, Ex ton JH. Critical role of endogenous Akt/IAPs and MEK1/ERK pathways in counteracting endoplasmic reticulum stress-induced cell death. J Biol Chem 2004; 279:49420-49429.
24. Dai R, Chen R, Li H. Cross-talk between PI3K/Akt and MEK/ERK pathways mediates endoplasmic reticulum stress-induced cell cycle progression and cell death in human hepatocellular carcinoma cells. Int J Oncol 2009; 34:1749-1757.
25. Numakawa T, Suzuki S, Kumamaru E, Adachi N, Richards M, Kunugi H. BDNF function and intracellular signaling in neurons. Histol Histopathol; 25:237-258.
26. Miyata H, Chiang AC, Vinters HV. Insulin signaling pathways in cortical dysplasia and TSC-tubers:tissue microarray analysis. Ann Neurol 2004; 56:510-519.
27. Friedrichs N, Kuchler J, Endl E, Koch A, Czerwitzki J, Wurst P et al. Insulin-like growth factor-1 receptor acts as a growth regulator in synovial sarcoma. J Pathol 2008; 216:428-439.
28. Staniszewska I, Sariyer IK, Lecht S, Brown MC, Walsh EM, Tuszynski GP et al. Integrin alpha9 betal is a receptor for nerve growth factor and other neurotrophins. J Cell Sci 2008;121:504-513.
1. Orlandini M, Marconcini L, Ferruzzi R, Oliviero S. Identification of a c-fos-induced gene that is related to the platelet-derived growth factor/vascular endothelial growth factor family. Proc Natl Acad Sci U S A 1996; 93:11675-11680.
2. Achen MG, Jeltsch M, Kukk E, Makinen T, Vitali A, Wilks AF et al. Vascular endothelial growth factor D (VEGF-D) is a ligand for the tyrosine kinases VEGF receptor 2 (Flkl) and VEGF receptor 3 (Flt4). Proc Natl Acad Sci U S A 1998; 95: 548-553.
3. Jeltsch M, Kaipainen A, Joukov V, Meng X, Lakso M, Rauvala H et al. Hyperplasia of lymphatic vessels in VEGF-C transgenic mice. Science 1997; 276:1423-1425.
4. Baldwin ME, Halford MM, Roufail S, Williams RA, Hibbs ML, Grail D et al. Vascular endothelial growth factor D is dispensable for development of the lymphatic system. Mol Cell Biol 2005; 25:2441-2449.
5. Bates DO, Harper SJ. Regulation of vascular permeability by vascular endothelial growth factors. Vascul Pharmacol 2002; 39:225-237.
6. Achen MG, Williams RA, Minekus MP, Thornton GE, Stenvers K, Rogers PA et al. Localization of vascular endothelial growth factor-D in malignant melanoma suggests a role in tumour angiogenesis. J Pathol 2001; 193:147-154.
7. Jenny B, Harrison JA, Baetens D, Tille JC, Burkhardt K, Mottaz H et al. Expression and localization of VEGF-C and VEGFR-3 in glioblastomas and haemangioblastomas. J Pathol 2006; 209:34-43.
8. Grau SJ, Trillsch F, Herms J, Thon N, Nelson PJ, Tonn JC et al. Expression of VEGFR3 in glioma endothelium correlates with tumor grade. J Neurooncol 2007; 82:141-150.
9. Gollmer JC, Ladoux A, Gioanni J, Paquis P, Dubreuil A, Chatel M et al. Expression of vascular endothelial growth factor-b in human astrocytoma. Neuro Oncol 2000; 2: 80-86.
10. Huber H, Eggert A, Janss AJ, Wiewrodt R, Zhao H, Sutton LN et al. Angiogenic profile of childhood primitive neuroectodermal brain tumours/medulloblastomas. Eur J Cancer 2001; 37:2064-2072.
11. Shin YJ, Choi JS, Choi JY, Cha JH, Chun MH, Lee MY. Enhanced expression of vascular endothelial growth factor receptor-3 in the subventricular zone of stroke-lesioned rats. Neurosci Lett 2010; 469:194-198.
12. Shin YJ, Choi JS, Lee JY, Choi JY, Cha JH, Chun MH et al. Differential regulation of vascular endothelial growth factor-C and its receptor in the rat hippocampus following transient forebrain ischemia. Acta Neuropathol 2008; 116:517-527.
13. Debinski W, Slagle-Webb B, Achen MG, Stacker SA, Tulchinsky E, Gillespie GY et al. VEGF-D is an X-linked/AP-1 regulated putative onco-angiogen in human glioblastoma multiforme. Mol Med 2001; 7:598-608.
14. Takahashi H, Shibuya M. The vascular endothelial growth factor (VEGF)/VEGF receptor system and its role under physiological and pathological conditions. Clin Sci (Lond) 2005; 109:227-241.
15. Von Marschall Z, Scholz A, Stacker SA, Achen MG, Jackson DG, Alves F et al. Vascular endothelial growth factor-D induces lymphangiogenesis and lymphatic metastasis in models of ductal pancreatic cancer. Int J Oncol 2005; 27:669-679.
16. Akahane M, Akahane T, Matheny SL, Shah A, Okajima E, Thorgeirsson UP. Vascular endothelial growth factor-D is a survival factor for human breast carcinoma cells. Int J Cancer 2006; 118:841-849.
17. Ishii H, Yazawa T, Sato H, Suzuki T, Ikeda M, Hayashi Y et al. Enhancement of pleural dissemination and lymph node metastasis of intrathoracic lung cancer cells by vascular endothelial growth factors (VEGFs). Lung Cancer 2004; 45:325-337.
18. Sie M, Wagemakers M, Molema G, Mooij JJ, de Bont ES, den Dunnen WF. The angiopoietin 1/angiopoietin 2 balance as a prognostic marker in primary glioblastoma multiforme. J Neurosurg 2009; 110:147-155.
19. Currie MJ, Hanrahan V, Gunningham SP, Morrin HR, Frampton C, Han C et al. Expression of vascular endothelial growth factor D is associated with hypoxia inducible factor (HIF-1 alpha) and the HIF-1 alpha target gene DEC1, but not lymph node metastasis in primary human breast carcinomas. J Clin Pathol 2004; 57:829-834.
20. Maxwell PH. Hypoxia-inducible factor as a physiological regulator. Exp Physiol 2005; 90:791-797.
21.Maurer MH, Tripps WK, Feldmann RE, Jr., Kuschinsky W. Expression of vascular endothelial growth factor and its receptors in rat neural stem cells. Neurosci Lett 2003; 344:165-168.
1. Cepeda C, Hurst RS, Flores-Hernandez J, Hernandez-Echeagaray E, Klapstein GJ, Boylan MK et al. Morphological and electrophysiological characterization of abnormal cell types in pediatric cortical dysplasia. J Neurosci Res 2003; 72:472-486.
2. Cepeda C, Andre VM, Vinters HV, Levine MS, Mathern GW. Are cytomegalic neurons and balloon cells generators of epileptic activity in pediatric cortical dysplasia? Epilepsia 2005; 46 Suppl 5:82-88.
3. Calcagnotto ME, Paredes MF, Tihan T, Barbaro NM, Baraban SC. Dysfunction of synaptic inhibition in epilepsy associated with focal cortical dysplasia. J Neurosci 2005; 25:9649-9657.
4. Lai MC, Lui CC, Yang SN, Wang JY, Huang LT. Epileptogenesis is increased in rats with neonatal isolation and early-life seizure and ameliorated by MK-801:a long-term MRI and histological study. Pediatr Res 2009; 66:441-447.
5. Prasad A, Williamson JM, Bertram EH. Phenobarbital and MK-801, but not phenytoin, improve the long-term outcome of status epilepticus. Ann Neurol 2002; 51:175-181.
6. Mikuni N, Babb TL, Ying Z, Najm I, Nishiyama K, Wylie C et al. NMDA-receptors 1 and 2A/B coassembly increased in human epileptic focal cortical dysplasia. Epilepsia 1999; 40:1683-1687.
7. Talos DM, Kwiatkowski DJ, Cordero K, Black PM, Jensen FE. Cell-specific alterations of glutamate receptor expression in tuberous sclerosis complex cortical tubers. Ann Neurol 2008; 63:454-465.
8. Yamanouchi H. Activated remodeling and N-methyl-D-aspartate (NMDA) receptors in cortical dysplasia.J Child Neurol 2005; 20:303-307.
9. Moddel G, Jacobson B, Ying Z, Janigro D, Bingaman W, Gonzalez-Martinez J et al. The NMDA receptor NR2B subunit contributes to epileptogenesis in human cortical dysplasia. Brain Res 2005; 1046:10-23.
10. Bandyopadhyay S, Hablitz JJ. NR2B antagonists restrict spatiotemporal spread of activity in a rat model of cortical dysplasia. Epilepsy Res 2006; 72:127-139.
11. Wang YT, Yu XM, Salter MW. Ca(2+)-independent reduction of N-methyl-D-aspartate channel activity by protein tyrosine phosphatase. Proc Natl Acad Sci U S A 1996; 93: 1721-1725.
12. Wang YT, Salter MW. Regulation of NMDA receptors by tyrosine kinases and phosphatases. Nature 1994; 369:233-235.
13. Leonard AS, Hell JW. Cyclic AMP-dependent protein kinase and protein kinase C phosphorylate N-methyl-D-aspartate receptors at different sites. J Biol Chem 1997; 272: 12107-12115.
14. Lu WY, Xiong ZG, Lei S, Orser BA, Dudek E, Browning MD et al. G-protein-coupled receptors act via protein kinase C and Src to regulate NMDA receptors. Nat Neurosci 1999; 2:331-338.
15. Borgland SL, Taha SA, Sarti F, Fields HL, Bonci A. Orexin A in the VTA is critical for the induction of synaptic plasticity and behavioral sensitization to cocaine. Neuron 2006; 49:589-601.
16. Ren K, Dubner R. Pain facilitation and activity-dependent plasticity in pain modulatory circuitry:role of BDNF-TrkB signaling and NMDA receptors. Mol Neurobiol 2007; 35: 224-235.
17. Roper SN. In utero irradiation of rats as a model of human cerebrocortical dysgenesis:a review. Epilepsy Res 1998; 32:63-74.
18. Jenny B, Harrison JA, Baetens D, Tille JC, Burkhardt K, Mottaz H et al. Expression and localization of VEGF-C and VEGFR-3 in glioblastomas and haemangioblastomas. J Pathol 2006; 209:34-43.
19. Grau SJ, Trillsch F, Herms J, Thon N, Nelson PJ, Tonn JC et al. Expression of VEGFR3 in glioma endothelium correlates with tumor grade. J Neurooncol 2007; 82:141-150.
20. Le Bras B, Barallobre MJ, Homman-Ludiye J, Ny A, Wyns S, Tammela T et al. VEGF-C is a trophic factor for neural progenitors in the vertebrate embryonic brain. Nat Neurosci 2006; 9:340-348.
21. Shin YJ, Choi JS, Lee JY, Choi JY, Cha JH, Chun MH et al. Differential regulation of vascular endothelial growth factor-C and its receptor in the rat hippocampus following transient forebrain ischemia. Acta Neuropathol 2008; 116:517-527.
22. Jin K, Mao XO, Greenberg DA. Vascular endothelial growth factor stimulates neurite outgrowth from cerebral cortical neurons via Rho kinase signaling. J Neurobiol 2006; 66:236-242.
23. Kim HY, Choi JS, Cha JH, Choi JY, Lee MY. Expression of vascular endothelial growth factor receptors Flt-1 and Flk-1 in embryonic rat forebrain. Neurosci Lett 2007; 425: 131-135.
24. Storkebaum E, Carmeliet P. VEGF:a critical player in neurodegeneration. J Clin Invest 2004; 113:14-18.
25. Cahana A, Escamez T, Nowakowski RS, Hayes NL, Giacobini M, von Holst A et al. Targeted mutagenesis of Lis1 disrupts cortical development and LIS1 homodimerization. Proc Natl Acad Sci U S A 2001; 98:6429-6434.
26. Schottler F, Couture D, Rao A, Kahn H, Lee KS. Subcortical connections of normotopic and heterotopic neurons in sensory and motor cortices of the tish mutant rat. J Comp Neurol 1998; 395:29-42.
27. Battaglia G, Becker AJ, LoTurco J, Represa A, Baraban SC, Roper SN et al. Basic mechanisms of MCD in animal models. Epileptic Disord 2009; 11:206-214.
28.Bai J, Ramos RL, Ackman JB, Thomas AM, Lee RV, LoTurco JJ. RNAi reveals doublecortin is required for radial migration in rat neocortex. Nat Neurosci 2003; 6: 1277-1283.
29. Redecker C, Luhmann HJ, Hagemann G, Fritschy JM, Witte OW. Differential downregulation of GABAA receptor subunits in widespread brain regions in the freeze-lesion model of focal cortical malformations. J Neurosci 2000; 20:5045-5053.
30. Hasling TA, Gierdalski M, Jablonska B, Juliano SL. A radialization factor in normal cortical plate restores disorganized radial glia and disrupted migration in a model of cortical dysplasia. Eur J Neurosci 2003; 17:467-480.
31.王文,徐天乐.NMDA受体通道的结构与功能.生物化学与生物物理进展1997；24：321-326.
32. Kellinghaus C, Kunieda T, Ying Z, Pan A, Luders HO, Najm IM. Severity of histopathologic abnormalities and in vivo epileptogenicity in the in utero radiation model of rats is dose dependent. Epilepsia 2004; 45:583-591.
33.Finardi A, Gardoni F, Bassanini S, Lasio G, Cossu M, Tassi L et al. NMDA receptor composition differs among anatomically diverse malformations of cortical development. J Neuropathol Exp Neurol 2006; 65:883-893.
34. Voronin LL. Intrasynaptic ephaptic feedback in central synapses. Neurosci Behav Physiol 2000; 30:575-585.
35. Roper SN, Eisenschenk S, King MA. Reduced density of parvalbumin-and calbindin D28-immunoreactive neurons in experimental cortical dysplasia. Epilepsy Res 1999; 37: 63-71.
36. Zhu WJ, Roper SN. Reduced inhibition in an animal model of cortical dysplasia. J Neurosci 2000; 20:8925-8931.
37. Chen BS, Roche KW. Regulation of NMDA receptors by phosphorylation. Neuropharmacology 2007; 53:362-368.
38. Mikuni N, Nishiyama K, Babb TL, Ying Z, Najm I, Okamoto T et al. Decreased calmodulin-NR1 co-assembly as a mechanism for focal epilepsy in cortical dysplasia. Neuroreport 1999; 10:1609-1612.
39. Babb TL, Ying Z, Hadam J, Penrod C. Glutamate receptor mechanisms in human epileptic dysplastic cortex. Epilepsy Res 1998; 32:24-33.
40. Najm IM, Ying Z, Babb T, Mohamed A, Hadam J, LaPresto E et al. Epileptogenicity correlated with increased N-methyl-D-aspartate receptor subunit NR2A/B in human focal cortical dysplasia. Epilepsia 2000; 41:971-976.
41.DeFazio RA, Hablitz JJ. Alterations in NMDA receptors in a rat model of cortical dysplasia. J Neurophysiol 2000; 83:315-321.
42. Shin YJ, Choi JS, Choi JY, Cha JH, Chun MH, Lee MY. Enhanced expression of vascular endothelial growth factor receptor-3 in the subventricular zone of stroke-lesioned rats. Neurosci Lett 2010; 469:194-198.
43. Sondell M, Kanje M. Postnatal expression of VEGF and its receptor flk-1 in peripheral ganglia. Neuroreport 2001; 12:105-108.
44. Yang SZ, Zhang LM, Huang YL, Sun FY Distribution of Flk-1 and Flt-1 receptors in neonatal and adult rat brains. Anat Rec A Discov Mol Cell Evol Biol 2003; 274: 851-856.
45. Greengard P. The neurobiology of slow synaptic transmission. Science 2001; 294: 1024-1030.
46. Liu L, Wong TP, Pozza MF, Lingenhoehl K, Wang Y, Sheng M et al. Role of NMDA receptor subtypes in governing the direction of hippocampal synaptic plasticity. Science 2004; 304:1021-1024.
47. Yuen EY, Jiang Q, Chen P, Gu Z, Feng J, Yan Z. Serotonin 5-HT1A receptors regulate NMDA receptor channels through a microtubule-dependent mechanism. J Neurosci 2005; 25:5488-5501.
48. Xu JY, Zheng P, Shen DH, Yang SZ, Zhang LM, Huang YL et al. Vascular endothelial growth factor inhibits outward delayed-rectifier potassium currents in acutely isolated hippocampal neurons. Neuroscience 2003; 118:59-67.
49. Qiu MH, Zhang R, Sun FY. Enhancement of ischemia-induced tyrosine phosphorylation of Kvl.2 by vascular endothelial growth factor via activation of phosphatidylinositol 3-kinase. J Neurochem 2003; 87:1509-1517.
50. Ma YY, Li KY, Wang JJ, Huang YL, Huang Y, Sun FY. Vascular endothelial growth factor acutely reduces calcium influx via inhibition of the Ca2+ channels in rat hippocampal neurons. J Neurosci Res 2009; 87:393-402.
51. Kim BW, Choi M, Kim YS, Park H, Lee HR, Yun CO et al. Vascular endothelial growth factor (VEGF) signaling regulates hippocampal neurons by elevation of intracellular calcium and activation of calcium/calmodulin protein kinase Ⅱ and mammalian target of rapamycin. Cell Signal 2008; 20:714-725.
52. Daniel P M, Croll SD, Scharfman HE. Depression of Synaptic Transmission by Vascular Endothelial Growth Factor in Adult Rat Hippocampus and Evidence for Increased Efficacy after Chronic Seizures. J Neurosci 2005; 25:8889-8897.
1. Palmini A, Najm I, Avanzini G, Babb T, Guerrini R, Foldvary-Schaefer N et al. Terminology and classification of the cortical dysplasias. Neurology 2004; 62:S2-8.
2. Mischel PS, Nguyen LP, Vinters HV. Cerebral cortical dysplasia associated with pediatric epilepsy. Review of neuropathologic features and proposal for a grading system. J Neuropathol Exp Neurol 1995; 54:137-153.
3. Sisodiya SM. Malformations of cortical development:burdens and insights from important causes of human epilepsy. Lancet Neurol 2004; 3:29-38.
4. Kuzniecky RI, Barkovich AJ. Pathogenesis and pathology of focal malformations of cortical development and epilepsy. J Clin Neurophysiol 1996; 13:468-480.
5. Aydin A, Cakmakci H, Kovanlikaya A, Dirik E. Sturge-Weber syndrome without facial nevus. Pediatr Neurol 2000; 22:400-402.
6. Guerrini R, Canapicchi R, Dobyns WB. Epilepsy and malformations of the cerebral cortex. Neurologia 1999; 14 Suppl 3:32-47.
7. Freedom RM, Lee KJ, MacDonald C, Taylor G. Selected aspects of cardiac tumors in infancy and childhood. Pediatr Cardiol 2000; 21:299-316.
8. Najm IM, Tilelli CQ, Oghlakian R. Pathophysiological mechanisms of focal cortical dysplasia:a critical review of human tissue studies and animal models. Epilepsia 2007; 48 Suppl 2.21-32.
9. Cepeda C, Andre VM, Vinters HV, Levine MS, Mathern GW. Are cytomegalic neurons and balloon cells generators of epileptic activity in pediatric cortical dysplasia? Epilepsia 2005; 46 Suppl 5:82-88.
10. Andres M, Andre VM, Nguyen S, Salamon N, Cepeda C, Levine MS et al. Human cortical dysplasia and epilepsy:an ontogenetic hypothesis based on volumetric MRI and NeuN neuronal density and size measurements. Cereb Cortex 2005; 15:194-210.
11. Cepeda C, Andre VM, Levine MS, Salamon N, Miyata H, Vinters HV et al. Epileptogenesis in pediatric cortical dysplasia:the dysmature cerebral developmental hypothesis. Epilepsy Behav 2006; 9:219-235.
12. Thom M, Martinian L, Sen A, Cross JH, Harding BN, Sisodiya SM. Cortical neuronal densities and lamination in focal cortical dysplasia. Acta Neuropathol 2005; 110: 383-392.
13. Shin YJ, Choi JS, Choi JY, Cha JH, Chun MH, Lee MY. Enhanced expression of vascular endothelial growth factor receptor-3 in the subventricular zone of stroke-lesioned rats. Neurosci Lett 2010; 469:194-198.
14. Lamparello P, Baybis M, Pollard J, Hol EM, Eisenstat DD, Aronica E et al. Developmental lineage of cell types in cortical dysplasia with balloon cells. Brain 2007; 130:2267-2276.
15. Boer K, Lucassen PJ, Spliet WG, Vreugdenhil E, van Rijen PC, Troost D et al. Doublecortin-like (DCL) expression in focal cortical dysplasia and cortical tubers. Epilepsia 2009; 50:2629-2637.
16. Crino PB, Trojanowski JQ, Dichter MA, Eberwine J. Embryonic neuronal markers in tuberous sclerosis:single-cell molecular pathology. Proc Natl Acad Sci U S A 1996; 93: 14152-14157.
17. Jozwiak J, Jozwiak S, Skopinski P. Immunohistochemical and microscopic studies on giant cells in tuberous sclerosis. Histol Histopathol 2005; 20:1321-1326.
18. Fauser S, Becker A, Schulze-Bonhage A, Hildebrandt M, Tuxhorn I, Pannek HW et al. CD34-immunoreactive balloon cells in cortical malformations. Acta Neuropathol 2004; 108:272-278.
19. Chamberlain WA, Prayson RA. Focal cortical dysplasia type Ⅱ (malformations of cortical development) aberrantly expresses apoptotic proteins. Appl Immunohistochem Mol Morphol 2008; 16:471-476.
20. Mathew T, Srikanth SG, Satishchandra P. Malformations of cortical development (MCDs) and epilepsy:experience from a tertiary care center in south India. Seizure 2010; 19:147-152.
21. Ying Z, Gonzalez-Martinez J, Tilelli C, Bingaman W, Najm I. Expression of neural stem cell surface marker CD133 in balloon cells of human focal cortical dysplasia. Epilepsia 2005; 46:1716-1723.
22. Sen A, Thom M, Martinian L, Harding B, Cross JH, Nikolic M et al. Pathological tau tangles localize to focal cortical dysplasia in older patients. Epilepsia 2007; 48: 1447-1454.
23. Chipuk JE, Moldoveanu T, Llambi F, Parsons MJ, Green DR. The BCL-2 family reunion. Mol Cell; 37:299-310.
24. Boer K, Troost D, Spliet WG, van Rijen PC, Gorter JA, Aronica E. Cellular distribution of vascular endothelial growth factor A (VEGFA) and B (VEGFB) and VEGF receptors 1 and 2 in focal cortical dysplasia type ⅡB. Acta Neuropathol 2008; 115:683-696.
25. Carelli S, Lesma E, Paratore S, Grande V, Zadra G, Bosari S et al. Survivin expression in tuberous sclerosis complex cells. Mol Med 2007; 13:166-177.
26. Wong M. Animal models of focal cortical dysplasia and tuberous sclerosis complex: recent progress toward clinical applications. Epilepsia 2009; 50 Suppl 9:34-44.
1. Mullen RJ, Buck CR, Smith AM. NeuN, a neuronal specific nuclear protein in vertebrates. Development 1992; 116:201-211.
2. Lind D, Franken S, Kappler J, Jankowski J, Schilling K. Characterization of the neuronal marker NeuN as a multiply phosphorylated antigen with discrete subcellular localization. J Neurosci Res 2005; 79:295-302.
3. Kim KK, Adelstein RS, Kawamoto S. Identification of neuronal nuclei (NeuN) as Fox-3, a new member of the Fox-1 gene family of splicing factors. J Biol Chem 2009; 284: 31052-31061.
4. Wolf HK, Buslei R, Schmidt-Kastner R, Schmidt-Kastner PK, Pietsch T, Wiestler OD et al. NeuN:a useful neuronal marker for diagnostic histopathology. J Histochem Cytochem 1996; 44:1167-1171.
5. Van Nassauw L, Wu M, De Jonge F, Adriaensen D, Timmermans JP. Cytoplasmic, but not nuclear, expression of the neuronal nuclei (NeuN) antibody is an exclusive feature of Dogiel type II neurons in the guinea-pig gastrointestinal tract. Histochem Cell Biol 2005; 124:369-377.
6. Kumar SS, Buckmaster PS. Neuron-specific nuclear antigen NeuN is not detectable in gerbil subtantia nigra pars reticulata. Brain Res 2007; 1142:54-60.
7. Voelker CC, Garin N, Taylor JS, Gahwiler BH, Hornung JP, Molnar Z. Selective neurofilament (SMI-32, FNP-7 and N200) expression in subpopulations of layer V pyramidal neurons in vivo and in vitro. Cereb Cortex 2004; 14:1276-1286.
8. Sasaki S, Maruyama S. Immunocytochemical and ultrastructural studies of the motor cortex in amyotrophic lateral sclerosis. Acta Neuropathol 1994; 87:578-585.
9. Toering ST, Boer K, de Groot M, Troost D, Heimans JJ, Spliet WG et al. Expression patterns of synaptic vesicle protein 2A in focal cortical dysplasia and TSC-cortical tubers. Epilepsia 2009; 50:1409-1418.
10. Magavi SS, Macklis JD. Immunocytochemical analysis of neuronal differentiation. Methods Mol Biol 2008; 438:345-352.
11.徐虹.,陈耀文.,韩太真.,沈建新.,霍霞.激光共聚焦扫描观察脑片及其神经元内RNA与DNA变化的技术.激光生物学报 2000；9：15-161.
12. Brazelton TR, Rossi FM, Keshet GI, Blau HM. From marrow to brain:expression of neuronal phenotypes in adult mice. Science 2000; 290:1775-1779.
13. Goetz AK, Scheffler B, Chen HX, Wang S, Suslov O, Xiang H et al. Temporally restricted substrate interactions direct fate and specification of neural precursors derived from embryonic stem cells. Proc Natl Acad Sci U S A 2006; 103:11063-11068.
14. Damianov A, Black DL. Autoregulation of Fox protein expression to produce dominant negative splicing factors. RNA 2010; 16:405-416.
15. Ponthier JL, Schluepen C, Chen W, Lersch RA, Gee SL, Hou VC et al. Fox-2 splicing factor binds to a conserved intron motif to promote inclusion of protein 4.1R alternative exon 16. J Biol Chem 2006; 281:12468-12474.
16. Yeo G, Holste D, Kreiman G, Burge CB. Variation in alternative splicing across human tissues. Genome Biol 2004; 5:R74.
17. Lee CJ, Irizarry K. Alternative splicing in the nervous system:an emerging source of diversity and regulation. Biol Psychiatry 2003; 54:771-776.
18. Preusser M, Laggner U, Haberler C, Heinzl H, Budka H, Hainfellner JA. Comparative analysis of NeuN immunoreactivity in primary brain tumours:conclusions for rational use in diagnostic histopathology. Histopathology 2006; 48:438-444.
19. Edgar MA, Rosenblum MK. The differential diagnosis of central nervous system tumors: a critical examination of some recent immunohistochemical applications. Arch Pathol Lab Med 2008; 132:500-509.
20. Soylemezoglu F, Onder S, Tezel GG, Berker M. Neuronal nuclear antigen (NeuN):a new tool in the diagnosis of central neurocytoma. Pathol Res Pract 2003; 199:463-468.
21.Shen CC, Yang YC, Chiao MT, Cheng WY, Ko JL, Tsuei YS. Characterization of Endogenous Neural Progenitor Cells after Experimental Ischemic Stroke. Curr Neurovasc Res.
22. Li Y, Chopp M, Chen J, Wang L, Gautam SC, Xu YX et al. Intrastriatal transplantation of bone marrow nonhematopoietic cells improves functional recovery after stroke in adult mice. J Cereb Blood Flow Metab 2000; 20:1311-1319.
23. Toda H, Takahashi J, Iwakami N, Kimura T, Hoki S, Mozumi-Kitamura K et al. Grafting neural stem cells improved the impaired spatial recognition in ischemic rats. Neurosci Lett 2001; 316:9-12.
24. Davoli MA, Fourtounis J, Tam J, Xanthoudakis S, Nicholson D, Robertson GS et al. Immunohistochemical and biochemical assessment of caspase-3 activation and DNA fragmentation following transient focal ischemia in the rat. Neuroscience 2002; 115: 125-136.
25. Sugawara T, Lewen A, Noshita N, Gasche Y, Chan PH. Effects of global ischemia duration on neuronal, astroglial, oligodendroglial, and microglial reactions in the vulnerable hippocampal CA1 subregion in rats. J Neurotrauma 2002; 19:85-98.
26. Unal-Cevik I, Kilinc M, Gursoy-Ozdemir Y, Gurer G, Dalkara T. Loss of NeuN immunoreactivity after cerebral ischemia does not indicate neuronal cell loss:a cautionary note. Brain Res 2004; 1015:169-174.
27. Baquet ZC, Bickford PC, Jones KR. Brain-derived neurotrophic factor is required for the establishment of the proper number of dopaminergic neurons in the substantia nigra pars compacta. J Neurosci 2005; 25:6251-6259.
28. Novikova L, Garris BL, Garris DR, Lau YS. Early signs of neuronal apoptosis in the substantia nigra pars compacta of the progressive neurodegenerative mouse 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/probenecid model of Parkinson's disease. Neuroscience 2006; 140:67-76.
29. Zhu C, Vourc'h P, Fernagut PO, Fleming SM, Lacan S, Dicarlo CD et al. Variable effects of chronic subcutaneous administration of rotenone on striatal histology. J Comp Neurol 2004; 478:418-426.
30. Cannon JR, Greenamyre JT. NeuN is not a reliable marker of dopamine neurons in rat substantia nigra. Neurosci Lett 2009; 464:14-17.
31. McPhail LT, McBride CB, McGraw J, Steeves JD, Tetzlaff W. Axotomy abolishes NeuN expression in facial but not rubrospinal neurons. Exp Neurol 2004; 185:182-190.
32. Collombet JM, Masqueliez C, Four E, Burckhart MF, Bernabe D, Baubichon D et al. Early reduction of NeuN antigenicity induced by soman poisoning in mice can be used to predict delayed neuronal degeneration in the hippocampus. Neurosci Lett 2006; 398: 337-342.
33. Wu KL, Li YQ, Tabassum A, Lu WY, Aubert I, Wong CS. Loss of Neuronal Protein Expression in Mouse Hippocampus After Irradiation. J Neuropathol Exp Neurol 2010; 69:272-280.
34. Long X, Olszewski M, Huang W, Kletzel M. Neural cell differentiation in vitro from adult human bone marrow mesenchymal stem cells. Stem Cells Dev 2005; 14:65-69.
35.Safford KM, Safford SD, Gimble JM, Shetty AK, Rice HE. Characterization of neuronal/glial differentiation of murine adipose-derived adult stromal cells. Exp Neurol 2004; 187:319-328.
36. Darlington PJ, Goldman JS, Cui QL, Antel JP, Kennedy TE. Widespread immunoreactivity for neuronal nuclei in cultured human and rodent astrocytes. J Neurochem 2008; 104:1201-1209.
37. Wong M. Mechanisms of epileptogenesis in tuberous sclerosis complex and related malformations of cortical development with abnormal glioneuronal proliferation. Epilepsia 2008; 49:8-21.
38. Palmini A, Najm I, Avanzini G, Babb T, Guerrini R, Foldvary-Schaefer N et al. Terminology and classification of the cortical dysplasias. Neurology 2004; 62:S2-8.
39. Lee A, Maldonado M, Baybis M, Walsh CA, Scheithauer B, Yeung R et al. Markers of cellular proliferation are expressed in cortical tubers. Ann Neurol 2003; 53:668-673.
40. Talos DM, Kwiatkowski DJ, Cordero K, Black PM, Jensen FE. Cell-specific alterations of glutamate receptor expression in tuberous sclerosis complex cortical tubers. Ann Neurol 2008; 63:454-465.
41. Thom M, Martinian L, Sen A, Cross JH, Harding BN, Sisodiya SM. Cortical neuronal densities and lamination in focal cortical dysplasia. Acta Neuropathol 2005;110: 383-392.
42. Garbelli R, Meroni A, Magnaghi G, Beolchi MS, Ferrario A, Tassi L et al. Architectural (Type IA) focal cortical dysplasia and parvalbumin immunostaining in temporal lobe epilepsy. Epilepsia 2006; 47:1074-1078.
43. Gonzalez-Martinez JA, Moddel G, Ying Z, Prayson RA, Bingaman WE, Najm IM. Neuronal nitric oxide synthase expression in resected epileptic dysplastic neocortex. J Neurosurg 2009; 110:343-349.
44. Portiansky EL, Barbeito CG, Gimeno EJ, Zuccolilli GO, Goya RG. Loss of NeuN immunoreactivity in rat spinal cord neurons during aging. Exp Neurol 2006; 202: 519-521.
2. Madan N, Grant PE. New directions in clinical imaging of cortical dysplasias. Epilepsia 2009; 50 Suppl 9:9-18.
3. Mathew T, Srikanth SG, Satishchandra P. Malformations of cortical development (MCDs) and epilepsy:experience from a tertiary care center in south India. Seizure 2010; 19:147-152.
4. Najm IM, Tilelli CQ, Oghlakian R. Pathophysiological mechanisms of focal cortical dysplasia:a critical review of human tissue studies and animal models. Epilepsia 2007; 48 Suppl 2:21-32.
5. Andres M, Andre VM, Nguyen S, Salamon N, Cepeda C, Levine MS et al. Human cortical dysplasia and epilepsy:an ontogenetic hypothesis based on volumetric MRI and NeuN neuronal density and size measurements. Cereb Cortex 2005; 15:194-210.
6. Storkebaum E, Carmeliet P. VEGF:a critical player in neurodegeneration.J Clin Invest 2004;113:14-18.
7. Partanen TA, Arola J, Saaristo A, Jussila L, Ora A, Miettinen M et al. VEGF-C and VEGF-D expression in neuroendocrine cells and their receptor, VEGFR-3, in fenestrated blood vessels in human tissues. FASEB J 2000; 14:2087-2096.
8. Le Bras B, Barallobre MJ, Homman-Ludiye J, Ny A, Wyns S, Tammela T et al. VEGF-C is a trophic factor for neural progenitors in the vertebrate embryonic brain. Nat Neurosci 2006;9:340-348.
9. Shin YJ, Choi JS, Choi JY, Cha JH, Chun MH, Lee MY. Enhanced expression of vascular endothelial growth factor receptor-3 in the subventricular zone of stroke-lesioned rats. Neurosci Lett 2010; 469:194-198.
10. Choi JS, Shin YJ, Lee JY, Yun H, Cha JH, Choi JY et al. Expression of vascular endothelial growth factor receptor-3 mRNA in the rat developing forebrain and retina. J Comp Neurol 2010; 518:1064-1081.
11. Shin YJ, Choi JS, Lee JY, Choi JY, Cha JH, Chun MH et al. Differential regulation of vascular endothelial growth factor-C and its receptor in the rat hippocampus following transient forebrain ischemia. Acta Neuropathol 2008; 116:517-527.
12. Kranich S, Hattermann K, Specht A, Lucius R, Mentlein R. VEGFR-3/Flt-4 mediates proliferation and chemotaxis in glial precursor cells. Neurochem Int 2009; 55:747-753.
13. Palmini A, Najm I, Avanzini G, Babb T, Guerrini R, Foldvary-Schaefer N et al. Terminology and classification of the cortical dysplasias. Neurology 2004; 62:S2-8.
14. Napolioni V, Moavero R, Curatolo P. Recent advances in neurobiology of Tuberous Sclerosis Complex. Brain Dev 2009; 31:104-113.
15. Liu S, An N, Yang H, Yang M, Hou Z, Liu L et al. Pediatric intractable epilepsy syndromes:reason for early surgical intervention. Brain Dev 2007; 29:69-78.
16. Grau SJ, Trillsch F, Herms J, Thon N, Nelson PJ, Tonn JC et al. Expression of VEGFR3 in glioma endothelium correlates with tumor grade. J Neurooncol 2007; 82: 141-150.
17. Boer K, Troost D, Spliet WG, van Rijen PC, Gorter JA, Aronica E. Cellular distribution of vascular endothelial growth factor A (VEGFA) and B (VEGFB) and VEGF receptors 1 and 2 in focal cortical dysplasia type ⅡB. Acta Neuropathol 2008; 115:683-696.
18. Croll SD, Goodman JH, Scharfman HE. Vascular endothelial growth factor (VEGF) in seizures:a double-edged sword. Adv Exp Med Biol 2004; 548:57-68.
19. Gollmer JC, Ladoux A, Gioanni J, Paquis P, Dubreuil A, Chatel M et al. Expression of vascular endothelial growth factor-b in human astrocytoma. Neuro Oncol 2000; 2: 80-86.
20. Huber H, Eggert A, Janss AJ, Wiewrodt R, Zhao H, Sutton LN et al. Angiogenic profile of childhood primitive neuroectodermal brain tumours/medulloblastomas. Eur J Cancer 2001; 37:2064-2072.
21. Jenny B, Harrison JA, Baetens D, Tille JC, Burkhardt K, Mottaz H et al. Expression and localization of VEGF-C and VEGFR-3 in glioblastomas and haemangioblastomas. J Pathol 2006; 209:34-43.
22. Schonberger A, Niehusmann P, Urbach H, Majores M, Grote A, Holthausen H et al. Increased frequency of distinct TSC2 allelic variants in focal cortical dysplasias with balloon cells and mineralization. Neuropathology 2009; 29:559-565.
23. Crino PB. Focal brain malformations:a spectrum of disorders along the mTOR cascade. Novartis Found Symp 2007; 288:260-272; discussion 272-281.
24. Brugarolas JB, Vazquez F, Reddy A, Sellers WR, Kaelin WG, Jr. TSC2 regulates VEGF through mTOR-dependent and-independent pathways. Cancer Cell 2003; 4:147-158.
25. El-Hashemite N, Walker V, Zhang H, Kwiatkowski DJ. Loss of Tscl or Tsc2 induces vascular endothelial growth factor production through mammalian target of rapamycin. Cancer Res 2003; 63:5173-5177.
26. Mahimainathan L, Ghosh-Choudhury N, Venkatesan B, Das F, Mandal CC, Dey N et al. TSC2 deficiency increases pten via HIF1alpha. J Biol Chem 2009.
27. Simiantonaki N, Jayasinghe C, Michel-Schmidt R, Peters K, Hermanns MI, Kirkpatrick CJ. Hypoxia-induced epithelial VEGF-C/VEGFR-3 upregulation in carcinoma cell lines. Int J Oncol 2008; 32:585-592.
28. Meierkord H, Holtkamp M. Status epilepticus:an independent outcome predictor after cerebral anoxia. Neurology 2008; 70:2015-2016; author reply 2015-2016.
29. Boer K, Jansen F, Nellist M, Redeker S, van den Ouweland AM, Spliet WG et al. Inflammatory processes in cortical tubers and subependymal giant cell tumors of tuberous sclerosis complex. Epilepsy Res 2008; 78:7-21.
30. Rodgers KM, Hutchinson MR, Northcutt A, Maier SF, Watkins LR, Barth DS. The cortical innate immune response increases local neuronal excitability leading to seizures. Brain 2009; 132:2478-2486.
31. Ristimaki A, Narko K, Enholm B, Joukov V, Alitalo K. Proinflammatory cytokines regulate expression of the lymphatic endothelial mitogen vascular endothelial growth factor-C. J Biol Chem 1998; 273:8413-8418.
32. Watari K, Nakao S, Fotovati A, Basaki Y, Hosoi F, Bereczky B et al. Role of macrophages in inflammatory lymphangiogenesis:Enhanced production of vascular endothelial growth factor C and D through NF-kappaB activation. Biochem Biophys Res Commun 2008; 377:826-831.
33. Tang RF, Wang SX, Zhang FR, Peng L, Xiao Y, Zhang M. Interleukin-1alpha,6 regulate the secretion of vascular endothelial growth factor A, C in pancreatic cancer. Hepatobiliary Pancreat Dis Int 2005; 4:460-463.
34. Ravizza T, Boer K, Redeker S, Spliet WG, van Rijen PC, Troost D et al. The IL-1beta system in epilepsy-associated malformations of cortical development. Neurobiol Dis 2006; 24:128-143.
35. Pajusola K, Aprelikova O, Korhonen J, Kaipainen A, Pertovaara L, Alitalo R et al. FLT4 receptor tyrosine kinase contains seven immunoglobulin-like loops and is expressed in multiple human tissues and cell lines. Cancer Res 1992; 52:5738-5743.
36. Aprelikova O, Pajusola K, Partanen J, Armstrong E, Alitalo R, Bailey SK et al. FLT4, a novel class Ⅲ receptor tyrosine kinase in chromosome 5q33-qter. Cancer Res 1992; 52: 746-748.
37. Brockington A, Wharton SB, Fernando M, Gelsthorpe CH, Baxter L, Ince PG et al. Expression of vascular endothelial growth factor and its receptors in the central nervous system in amyotrophic lateral sclerosis. J Neuropathol Exp Neurol 2006; 65:26-36.
38. Hlobilkova A, Ehrmann J, Knizetova P, Krejci V, Kalita O, Kolar Z. Analysis of VEGF, Flt-1, Flk-1, nestin and MMP-9 in relation to astrocytoma pathogenesis and progression. Neoplasma 2009; 56:284-290.
39. Krum JM, Mani N, Rosenstein JM. Roles of the endogenous VEGF receptors flt-1 and flk-1 in astroglial and vascular remodeling after brain injury. Exp Neurol 2008; 212: 108-117.
40. Raab S, Plate KH. Different networks, common growth factors:shared growth factors and receptors of the vascular and the nervous system. Acta Neuropathol 2007; 113: 607-626.
41. Pietsch T, Valter MM, Wolf HK, von Deimling A, Huang HJ, Cavenee WK et al. Expression and distribution of vascular endothelial growth factor protein in human brain tumors. Acta Neuropathol 1997; 93:109-117.
42. Rigau V, Morin M, Rousset MC, de Bock F, Lebrun A, Coubes P et al. Angiogenesis is associated with blood-brain barrier permeability in temporal lobe epilepsy. Brain 2007; 130:1942-1956.
43. Miyata H, Chiang AC, Vinters HV. Insulin signaling pathways in cortical dysplasia and TSC-tubers:tissue microarray analysis. Ann Neurol 2004; 56:510-519.
44. Boer K, Lucassen PJ, Spliet WG, Vreugdenhil E, van Rijen PC, Troost D et al. Doublecortin-like (DCL) expression in focal cortical dysplasia and cortical tubers. Epilepsia 2009.
45. Qu M, Aronica E, Boer K, Fallmar D, Kumlien E, Nister M et al. DLG3/SAP102 protein expression in malformations of cortical development:a study of human epileptic cortex by tissue microarray. Epilepsy Res 2009; 84:33-41.
46. Sugiura C, Miyata H, Ueda M, Ohama E, Vinters HV, Ohno K. Immunohistochemical expression of fibroblast growth factor (FGF)-2 in epilepsy-associated malformations of cortical development (MCDs). Neuropathology 2008; 28:372-381.
47. Toering ST, Boer K, de Groot M, Troost D, Heimans JJ, Spliet WG et al. Expression patterns of synaptic vesicle protein 2A in focal cortical dysplasia and TSC-cortical tubers. Epilepsia 2009; 50:1409-1418.
48. Susan D. Croll,, Richard M. Ransohoff, Ning Caia, Qing Zhanga, Francis J. Martina, Tao Weib, Lora J. Kasselmana, Jennifer Kintnera, Andrew J. Murphya, George D. Yancopoulosa and Stanley J. Wieganda. VEGF-mediated inflammation precedes angiogenesis in adult brain. Experimental Neurology 2004; 187:388-402.
49. Vezzani A. VEGF and seizures:cross-talk between endothelial and neuronal environments. Epilepsy Curr 2005;5:72-74.
50. Vezzani A, Granata T. Brain inflammation in epilepsy:experimental and clinical evidence. Epilepsia 2005;46:1724-1743.
51. van Vliet EA, da Costa Araujo S, Redeker S, van Schaik R, Aronica E, Gorter JA. Blood-brain barrier leakage may lead to progression of temporal lobe epilepsy. Brain 2007;130:521-534.
52. Maurer MH, Tripps WK, Feldmann RE, Jr., Kuschinsky W. Expression of vascular endothelial growth factor and its receptors in rat neural stem cells. Neurosci Lett 2003; 344:165-168.
53. Salven P, Mustjoki S, Alitalo R, Alitalo K, Rafii S. VEGFR-3 and CD133 identify a population of CD34+ lymphatic/vascular endothelial precursor cells. Blood 2003;101: 168-172.
54. Ma YH, Mentlein R, Knerlich F, Kruse ML, Mehdorn HM, Held-Feindt J. Expression of stem cell markers in human astrocytomas of different WHO grades. J Neurooncol 2008; 86:31-45.
55. Crino PB, Trojanowski JQ, Dichter MA, Eberwine J. Embryonic neuronal markers in tuberous sclerosis:single-cell molecular pathology. Proc Natl Acad Sci U S A 1996; 93: 14152-14157.
56. Jozwiak J, Jozwiak S, Skopinski P. Immunohistochemical and microscopic studies on giant cells in tuberous sclerosis. Histol Histopathol 2005; 20:1321-1326.
57. Fauser S, Becker A, Schulze-Bonhage A, Hildebrandt M, Tuxhorn I, Pannek HW et al. CD34-immunoreactive balloon cells in cortical malformations. Acta Neuropathol 2004; 108:272-278.
58. Boer K, Lucassen PJ, Spliet WG, Vreugdenhil E, van Rijen PC, Troost D et al. Doublecortin-like (DCL) expression in focal cortical dysplasia and cortical tubers. Epilepsia 2009; 50:2629-2637.
59. Kranich S, Hattermann K, Specht A, Lucius R, Mentlein R. VEGFR-3/Flt-4 mediates proliferation and chemotaxis in glial precursor cells. Neurochem Int 2009.
1. Chamberlain WA, Prayson RA. Focal cortical dysplasia type II (malformations of cortical development) aberrantly expresses apoptotic proteins. Appl Immunohistochem Mol Morphol 2008; 16:471-476.
2. Ruch C, Skiniotis G, Steinmetz MO, Walz T, Ballmer-Hofer K. Structure of a VEGF-VEGF receptor complex determined by electron microscopy. Nat Struct Mol Biol 2007; 14:249-250.
3. Chien MH, Ku CC, Johansson G, Chen MW, Hsiao M, Su JL et al. Vascular endothelial growth factor-C (VEGF-C) promotes angiogenesis by induction of COX-2 in leukemic cells via the VEGF-R3/JNK/AP-1 pathway. Carcinogenesis 2009; 30:2005-2013.
4. Abid MR, Guo S, Minami T, Spokes KC, Ueki K, Skurk C et al. Vascular endothelial growth factor activates PI3K/Akt/forkhead signaling in endothelial cells. Arterioscler Thromb Vasc Biol 2004; 24:294-300.
5. Andres M, Andre VM, Nguyen S, Salamon N, Cepeda C, Levine MS et al. Human cortical dysplasia and epilepsy:an ontogenetic hypothesis based on volumetric MRI and NeuN neuronal density and size measurements. Cereb Cortex 2005; 15:194-210.
6. Najm IM, Tilelli CQ, Oghlakian R. Pathophysiological mechanisms of focal cortical dysplasia:a critical review of human tissue studies and animal models. Epilepsia 2007; 48 Suppl2:21-32.
7. Sen A, Thom M, Martinian L, Harding B, Cross JH, Nikolic M et al. Pathological tau tangles localize to focal cortical dysplasia in older patients. Epilepsia 2007; 48: 1447-1454.
8. Cepeda C, Andre VM, Levine MS, Salamon N, Miyata H, Vinters HV et al. Epileptogenesis in pediatric cortical dysplasia:the dysmature cerebral developmental hypothesis. Epilepsy Behav 2006; 9:219-235.
9. Muller-Deile J, Worthmann K, Saleem M, Tossidou I, Haller H, Schiffer M. The balance of autocrine VEGF-A and VEGF-C determines podocyte survival. Am J Physiol Renal Physiol 2009; 297:F1656-1667.
10. Kirkin V, Thiele W, Baumann P, Mazitschek R, Rohde K, Fellbrich G et al. MAZ51, an indolinone that inhibits endothelial cell and tumor cell growth in vitro, suppresses tumor growth in vivo. Int J Cancer 2004; 112:986-993.
11. Duronio V. The life of a cell:apoptosis regulation by the PI3K/PKB pathway. Biochem J 2008; 415:333-344.
12. Le Page C, Koumakpayi IH, Alam-Fahmy M, Mes-Masson AM, Saad F. Expression and localisation of Akt-1, Akt-2 and Akt-3 correlate with clinical outcome of prostate cancer patients. Br J Cancer 2006; 94:1906-1912.
13. Price J, Zaidi AK, Bohensky J, Srinivas V, Shapiro IM, Ali H. Akt-1 mediates survival of chondrocytes from endoplasmic reticulum-induced stress. J Cell Physiol; 222: 502-508.
14. Chipuk JE, Moldoveanu T, Llambi F, Parsons MJ, Green DR. The BCL-2 family reunion. Mol Cell; 37:299-310.
15. Patel MP, Masood A, Patel PS, Chanan-Khan AA. Targeting the Bcl-2. Curr Opin Oncol 2009; 21:516-523.
16. Zhu Y, Hoell P, Ahlemeyer B, Krieglstein J. PTEN:a crucial mediator of mitochondria-dependent apoptosis. Apoptosis 2006; 11:197-207.
17. Narasimhan P, Liu J, Song YS, Massengale JL, Chan PH. VEGF Stimulates the ERK 1/2 signaling pathway and apoptosis in cerebral endothelial cells after ischemic conditions. Stroke 2009; 40:1467-1473.
18. Salameh A, Galvagni F, Bardelli M, Bussolino F, Oliviero S. Direct recruitment of CRK and GRB2 to VEGFR-3 induces proliferation, migration, and survival of endothelial cells through the activation of ERK, AKT, and JNK pathways. Blood 2005; 106: 3423-3431.
19. Girault JA, Valjent E, Caboche J, Herve D. ERK2:a logical AND gate critical for drug-induced plasticity? Curr Opin Pharmacol 2007; 7:77-85.
20. Suzuki M, Satoh A, Ide H, Tamura K. Transgenic Xenopus with prxl limb enhancer reveals crucial contribution of MEK/ERK and PI3K/AKT pathways in blastema formation during limb regeneration. Dev Biol 2007; 304:675-686.
21. Kilic E, Kilic U, Wang Y, Bassetti CL, Marti HH, Hermann DM. The phosphatidylinositol-3 kinase/Akt pathway mediates VEGF's neuroprotective activity and induces blood brain barrier permeability after focal cerebral ischemia. FASEB J 2006; 20:1185-1187.
22. Frebel K, Wiese S. Signalling molecules essential for neuronal survival and differentiation. Biochem Soc Trans 2006; 34:1287-1290.
23. Hu P, Han Z, Couvillon AD, Ex ton JH. Critical role of endogenous Akt/IAPs and MEK1/ERK pathways in counteracting endoplasmic reticulum stress-induced cell death. J Biol Chem 2004; 279:49420-49429.
24. Dai R, Chen R, Li H. Cross-talk between PI3K/Akt and MEK/ERK pathways mediates endoplasmic reticulum stress-induced cell cycle progression and cell death in human hepatocellular carcinoma cells. Int J Oncol 2009; 34:1749-1757.
25. Numakawa T, Suzuki S, Kumamaru E, Adachi N, Richards M, Kunugi H. BDNF function and intracellular signaling in neurons. Histol Histopathol; 25:237-258.
26. Miyata H, Chiang AC, Vinters HV. Insulin signaling pathways in cortical dysplasia and TSC-tubers:tissue microarray analysis. Ann Neurol 2004; 56:510-519.
27. Friedrichs N, Kuchler J, Endl E, Koch A, Czerwitzki J, Wurst P et al. Insulin-like growth factor-1 receptor acts as a growth regulator in synovial sarcoma. J Pathol 2008; 216:428-439.
28. Staniszewska I, Sariyer IK, Lecht S, Brown MC, Walsh EM, Tuszynski GP et al. Integrin alpha9 betal is a receptor for nerve growth factor and other neurotrophins. J Cell Sci 2008;121:504-513.
1. Orlandini M, Marconcini L, Ferruzzi R, Oliviero S. Identification of a c-fos-induced gene that is related to the platelet-derived growth factor/vascular endothelial growth factor family. Proc Natl Acad Sci U S A 1996; 93:11675-11680.
2. Achen MG, Jeltsch M, Kukk E, Makinen T, Vitali A, Wilks AF et al. Vascular endothelial growth factor D (VEGF-D) is a ligand for the tyrosine kinases VEGF receptor 2 (Flkl) and VEGF receptor 3 (Flt4). Proc Natl Acad Sci U S A 1998; 95: 548-553.
3. Jeltsch M, Kaipainen A, Joukov V, Meng X, Lakso M, Rauvala H et al. Hyperplasia of lymphatic vessels in VEGF-C transgenic mice. Science 1997; 276:1423-1425.
4. Baldwin ME, Halford MM, Roufail S, Williams RA, Hibbs ML, Grail D et al. Vascular endothelial growth factor D is dispensable for development of the lymphatic system. Mol Cell Biol 2005; 25:2441-2449.
5. Bates DO, Harper SJ. Regulation of vascular permeability by vascular endothelial growth factors. Vascul Pharmacol 2002; 39:225-237.
6. Achen MG, Williams RA, Minekus MP, Thornton GE, Stenvers K, Rogers PA et al. Localization of vascular endothelial growth factor-D in malignant melanoma suggests a role in tumour angiogenesis. J Pathol 2001; 193:147-154.
7. Jenny B, Harrison JA, Baetens D, Tille JC, Burkhardt K, Mottaz H et al. Expression and localization of VEGF-C and VEGFR-3 in glioblastomas and haemangioblastomas. J Pathol 2006; 209:34-43.
8. Grau SJ, Trillsch F, Herms J, Thon N, Nelson PJ, Tonn JC et al. Expression of VEGFR3 in glioma endothelium correlates with tumor grade. J Neurooncol 2007; 82:141-150.
9. Gollmer JC, Ladoux A, Gioanni J, Paquis P, Dubreuil A, Chatel M et al. Expression of vascular endothelial growth factor-b in human astrocytoma. Neuro Oncol 2000; 2: 80-86.
10. Huber H, Eggert A, Janss AJ, Wiewrodt R, Zhao H, Sutton LN et al. Angiogenic profile of childhood primitive neuroectodermal brain tumours/medulloblastomas. Eur J Cancer 2001; 37:2064-2072.
11. Shin YJ, Choi JS, Choi JY, Cha JH, Chun MH, Lee MY. Enhanced expression of vascular endothelial growth factor receptor-3 in the subventricular zone of stroke-lesioned rats. Neurosci Lett 2010; 469:194-198.
12. Shin YJ, Choi JS, Lee JY, Choi JY, Cha JH, Chun MH et al. Differential regulation of vascular endothelial growth factor-C and its receptor in the rat hippocampus following transient forebrain ischemia. Acta Neuropathol 2008; 116:517-527.
13. Debinski W, Slagle-Webb B, Achen MG, Stacker SA, Tulchinsky E, Gillespie GY et al. VEGF-D is an X-linked/AP-1 regulated putative onco-angiogen in human glioblastoma multiforme. Mol Med 2001; 7:598-608.
14. Takahashi H, Shibuya M. The vascular endothelial growth factor (VEGF)/VEGF receptor system and its role under physiological and pathological conditions. Clin Sci (Lond) 2005; 109:227-241.
15. Von Marschall Z, Scholz A, Stacker SA, Achen MG, Jackson DG, Alves F et al. Vascular endothelial growth factor-D induces lymphangiogenesis and lymphatic metastasis in models of ductal pancreatic cancer. Int J Oncol 2005; 27:669-679.
16. Akahane M, Akahane T, Matheny SL, Shah A, Okajima E, Thorgeirsson UP. Vascular endothelial growth factor-D is a survival factor for human breast carcinoma cells. Int J Cancer 2006; 118:841-849.
17. Ishii H, Yazawa T, Sato H, Suzuki T, Ikeda M, Hayashi Y et al. Enhancement of pleural dissemination and lymph node metastasis of intrathoracic lung cancer cells by vascular endothelial growth factors (VEGFs). Lung Cancer 2004; 45:325-337.
18. Sie M, Wagemakers M, Molema G, Mooij JJ, de Bont ES, den Dunnen WF. The angiopoietin 1/angiopoietin 2 balance as a prognostic marker in primary glioblastoma multiforme. J Neurosurg 2009; 110:147-155.
19. Currie MJ, Hanrahan V, Gunningham SP, Morrin HR, Frampton C, Han C et al. Expression of vascular endothelial growth factor D is associated with hypoxia inducible factor (HIF-1 alpha) and the HIF-1 alpha target gene DEC1, but not lymph node metastasis in primary human breast carcinomas. J Clin Pathol 2004; 57:829-834.
20. Maxwell PH. Hypoxia-inducible factor as a physiological regulator. Exp Physiol 2005; 90:791-797.
21.Maurer MH, Tripps WK, Feldmann RE, Jr., Kuschinsky W. Expression of vascular endothelial growth factor and its receptors in rat neural stem cells. Neurosci Lett 2003; 344:165-168.
1. Cepeda C, Hurst RS, Flores-Hernandez J, Hernandez-Echeagaray E, Klapstein GJ, Boylan MK et al. Morphological and electrophysiological characterization of abnormal cell types in pediatric cortical dysplasia. J Neurosci Res 2003; 72:472-486.
2. Cepeda C, Andre VM, Vinters HV, Levine MS, Mathern GW. Are cytomegalic neurons and balloon cells generators of epileptic activity in pediatric cortical dysplasia? Epilepsia 2005; 46 Suppl 5:82-88.
3. Calcagnotto ME, Paredes MF, Tihan T, Barbaro NM, Baraban SC. Dysfunction of synaptic inhibition in epilepsy associated with focal cortical dysplasia. J Neurosci 2005; 25:9649-9657.
4. Lai MC, Lui CC, Yang SN, Wang JY, Huang LT. Epileptogenesis is increased in rats with neonatal isolation and early-life seizure and ameliorated by MK-801:a long-term MRI and histological study. Pediatr Res 2009; 66:441-447.
5. Prasad A, Williamson JM, Bertram EH. Phenobarbital and MK-801, but not phenytoin, improve the long-term outcome of status epilepticus. Ann Neurol 2002; 51:175-181.
6. Mikuni N, Babb TL, Ying Z, Najm I, Nishiyama K, Wylie C et al. NMDA-receptors 1 and 2A/B coassembly increased in human epileptic focal cortical dysplasia. Epilepsia 1999; 40:1683-1687.
7. Talos DM, Kwiatkowski DJ, Cordero K, Black PM, Jensen FE. Cell-specific alterations of glutamate receptor expression in tuberous sclerosis complex cortical tubers. Ann Neurol 2008; 63:454-465.
8. Yamanouchi H. Activated remodeling and N-methyl-D-aspartate (NMDA) receptors in cortical dysplasia.J Child Neurol 2005; 20:303-307.
9. Moddel G, Jacobson B, Ying Z, Janigro D, Bingaman W, Gonzalez-Martinez J et al. The NMDA receptor NR2B subunit contributes to epileptogenesis in human cortical dysplasia. Brain Res 2005; 1046:10-23.
10. Bandyopadhyay S, Hablitz JJ. NR2B antagonists restrict spatiotemporal spread of activity in a rat model of cortical dysplasia. Epilepsy Res 2006; 72:127-139.
11. Wang YT, Yu XM, Salter MW. Ca(2+)-independent reduction of N-methyl-D-aspartate channel activity by protein tyrosine phosphatase. Proc Natl Acad Sci U S A 1996; 93: 1721-1725.
12. Wang YT, Salter MW. Regulation of NMDA receptors by tyrosine kinases and phosphatases. Nature 1994; 369:233-235.
13. Leonard AS, Hell JW. Cyclic AMP-dependent protein kinase and protein kinase C phosphorylate N-methyl-D-aspartate receptors at different sites. J Biol Chem 1997; 272: 12107-12115.
14. Lu WY, Xiong ZG, Lei S, Orser BA, Dudek E, Browning MD et al. G-protein-coupled receptors act via protein kinase C and Src to regulate NMDA receptors. Nat Neurosci 1999; 2:331-338.
15. Borgland SL, Taha SA, Sarti F, Fields HL, Bonci A. Orexin A in the VTA is critical for the induction of synaptic plasticity and behavioral sensitization to cocaine. Neuron 2006; 49:589-601.
16. Ren K, Dubner R. Pain facilitation and activity-dependent plasticity in pain modulatory circuitry:role of BDNF-TrkB signaling and NMDA receptors. Mol Neurobiol 2007; 35: 224-235.
17. Roper SN. In utero irradiation of rats as a model of human cerebrocortical dysgenesis:a review. Epilepsy Res 1998; 32:63-74.
18. Jenny B, Harrison JA, Baetens D, Tille JC, Burkhardt K, Mottaz H et al. Expression and localization of VEGF-C and VEGFR-3 in glioblastomas and haemangioblastomas. J Pathol 2006; 209:34-43.
19. Grau SJ, Trillsch F, Herms J, Thon N, Nelson PJ, Tonn JC et al. Expression of VEGFR3 in glioma endothelium correlates with tumor grade. J Neurooncol 2007; 82:141-150.
20. Le Bras B, Barallobre MJ, Homman-Ludiye J, Ny A, Wyns S, Tammela T et al. VEGF-C is a trophic factor for neural progenitors in the vertebrate embryonic brain. Nat Neurosci 2006; 9:340-348.
21. Shin YJ, Choi JS, Lee JY, Choi JY, Cha JH, Chun MH et al. Differential regulation of vascular endothelial growth factor-C and its receptor in the rat hippocampus following transient forebrain ischemia. Acta Neuropathol 2008; 116:517-527.
22. Jin K, Mao XO, Greenberg DA. Vascular endothelial growth factor stimulates neurite outgrowth from cerebral cortical neurons via Rho kinase signaling. J Neurobiol 2006; 66:236-242.
23. Kim HY, Choi JS, Cha JH, Choi JY, Lee MY. Expression of vascular endothelial growth factor receptors Flt-1 and Flk-1 in embryonic rat forebrain. Neurosci Lett 2007; 425: 131-135.
24. Storkebaum E, Carmeliet P. VEGF:a critical player in neurodegeneration. J Clin Invest 2004; 113:14-18.
25. Cahana A, Escamez T, Nowakowski RS, Hayes NL, Giacobini M, von Holst A et al. Targeted mutagenesis of Lis1 disrupts cortical development and LIS1 homodimerization. Proc Natl Acad Sci U S A 2001; 98:6429-6434.
26. Schottler F, Couture D, Rao A, Kahn H, Lee KS. Subcortical connections of normotopic and heterotopic neurons in sensory and motor cortices of the tish mutant rat. J Comp Neurol 1998; 395:29-42.
27. Battaglia G, Becker AJ, LoTurco J, Represa A, Baraban SC, Roper SN et al. Basic mechanisms of MCD in animal models. Epileptic Disord 2009; 11:206-214.
28.Bai J, Ramos RL, Ackman JB, Thomas AM, Lee RV, LoTurco JJ. RNAi reveals doublecortin is required for radial migration in rat neocortex. Nat Neurosci 2003; 6: 1277-1283.
29. Redecker C, Luhmann HJ, Hagemann G, Fritschy JM, Witte OW. Differential downregulation of GABAA receptor subunits in widespread brain regions in the freeze-lesion model of focal cortical malformations. J Neurosci 2000; 20:5045-5053.
30. Hasling TA, Gierdalski M, Jablonska B, Juliano SL. A radialization factor in normal cortical plate restores disorganized radial glia and disrupted migration in a model of cortical dysplasia. Eur J Neurosci 2003; 17:467-480.
31.王文,徐天乐.NMDA受体通道的结构与功能.生物化学与生物物理进展1997；24：321-326.
32. Kellinghaus C, Kunieda T, Ying Z, Pan A, Luders HO, Najm IM. Severity of histopathologic abnormalities and in vivo epileptogenicity in the in utero radiation model of rats is dose dependent. Epilepsia 2004; 45:583-591.
33.Finardi A, Gardoni F, Bassanini S, Lasio G, Cossu M, Tassi L et al. NMDA receptor composition differs among anatomically diverse malformations of cortical development. J Neuropathol Exp Neurol 2006; 65:883-893.
34. Voronin LL. Intrasynaptic ephaptic feedback in central synapses. Neurosci Behav Physiol 2000; 30:575-585.
35. Roper SN, Eisenschenk S, King MA. Reduced density of parvalbumin-and calbindin D28-immunoreactive neurons in experimental cortical dysplasia. Epilepsy Res 1999; 37: 63-71.
36. Zhu WJ, Roper SN. Reduced inhibition in an animal model of cortical dysplasia. J Neurosci 2000; 20:8925-8931.
37. Chen BS, Roche KW. Regulation of NMDA receptors by phosphorylation. Neuropharmacology 2007; 53:362-368.
38. Mikuni N, Nishiyama K, Babb TL, Ying Z, Najm I, Okamoto T et al. Decreased calmodulin-NR1 co-assembly as a mechanism for focal epilepsy in cortical dysplasia. Neuroreport 1999; 10:1609-1612.
39. Babb TL, Ying Z, Hadam J, Penrod C. Glutamate receptor mechanisms in human epileptic dysplastic cortex. Epilepsy Res 1998; 32:24-33.
40. Najm IM, Ying Z, Babb T, Mohamed A, Hadam J, LaPresto E et al. Epileptogenicity correlated with increased N-methyl-D-aspartate receptor subunit NR2A/B in human focal cortical dysplasia. Epilepsia 2000; 41:971-976.
41.DeFazio RA, Hablitz JJ. Alterations in NMDA receptors in a rat model of cortical dysplasia. J Neurophysiol 2000; 83:315-321.
42. Shin YJ, Choi JS, Choi JY, Cha JH, Chun MH, Lee MY. Enhanced expression of vascular endothelial growth factor receptor-3 in the subventricular zone of stroke-lesioned rats. Neurosci Lett 2010; 469:194-198.
43. Sondell M, Kanje M. Postnatal expression of VEGF and its receptor flk-1 in peripheral ganglia. Neuroreport 2001; 12:105-108.
44. Yang SZ, Zhang LM, Huang YL, Sun FY Distribution of Flk-1 and Flt-1 receptors in neonatal and adult rat brains. Anat Rec A Discov Mol Cell Evol Biol 2003; 274: 851-856.
45. Greengard P. The neurobiology of slow synaptic transmission. Science 2001; 294: 1024-1030.
46. Liu L, Wong TP, Pozza MF, Lingenhoehl K, Wang Y, Sheng M et al. Role of NMDA receptor subtypes in governing the direction of hippocampal synaptic plasticity. Science 2004; 304:1021-1024.
47. Yuen EY, Jiang Q, Chen P, Gu Z, Feng J, Yan Z. Serotonin 5-HT1A receptors regulate NMDA receptor channels through a microtubule-dependent mechanism. J Neurosci 2005; 25:5488-5501.
48. Xu JY, Zheng P, Shen DH, Yang SZ, Zhang LM, Huang YL et al. Vascular endothelial growth factor inhibits outward delayed-rectifier potassium currents in acutely isolated hippocampal neurons. Neuroscience 2003; 118:59-67.
49. Qiu MH, Zhang R, Sun FY. Enhancement of ischemia-induced tyrosine phosphorylation of Kvl.2 by vascular endothelial growth factor via activation of phosphatidylinositol 3-kinase. J Neurochem 2003; 87:1509-1517.
50. Ma YY, Li KY, Wang JJ, Huang YL, Huang Y, Sun FY. Vascular endothelial growth factor acutely reduces calcium influx via inhibition of the Ca2+ channels in rat hippocampal neurons. J Neurosci Res 2009; 87:393-402.
51. Kim BW, Choi M, Kim YS, Park H, Lee HR, Yun CO et al. Vascular endothelial growth factor (VEGF) signaling regulates hippocampal neurons by elevation of intracellular calcium and activation of calcium/calmodulin protein kinase Ⅱ and mammalian target of rapamycin. Cell Signal 2008; 20:714-725.
52. Daniel P M, Croll SD, Scharfman HE. Depression of Synaptic Transmission by Vascular Endothelial Growth Factor in Adult Rat Hippocampus and Evidence for Increased Efficacy after Chronic Seizures. J Neurosci 2005; 25:8889-8897.
1. Palmini A, Najm I, Avanzini G, Babb T, Guerrini R, Foldvary-Schaefer N et al. Terminology and classification of the cortical dysplasias. Neurology 2004; 62:S2-8.
2. Mischel PS, Nguyen LP, Vinters HV. Cerebral cortical dysplasia associated with pediatric epilepsy. Review of neuropathologic features and proposal for a grading system. J Neuropathol Exp Neurol 1995; 54:137-153.
3. Sisodiya SM. Malformations of cortical development:burdens and insights from important causes of human epilepsy. Lancet Neurol 2004; 3:29-38.
4. Kuzniecky RI, Barkovich AJ. Pathogenesis and pathology of focal malformations of cortical development and epilepsy. J Clin Neurophysiol 1996; 13:468-480.
5. Aydin A, Cakmakci H, Kovanlikaya A, Dirik E. Sturge-Weber syndrome without facial nevus. Pediatr Neurol 2000; 22:400-402.
6. Guerrini R, Canapicchi R, Dobyns WB. Epilepsy and malformations of the cerebral cortex. Neurologia 1999; 14 Suppl 3:32-47.
7. Freedom RM, Lee KJ, MacDonald C, Taylor G. Selected aspects of cardiac tumors in infancy and childhood. Pediatr Cardiol 2000; 21:299-316.
8. Najm IM, Tilelli CQ, Oghlakian R. Pathophysiological mechanisms of focal cortical dysplasia:a critical review of human tissue studies and animal models. Epilepsia 2007; 48 Suppl 2.21-32.
9. Cepeda C, Andre VM, Vinters HV, Levine MS, Mathern GW. Are cytomegalic neurons and balloon cells generators of epileptic activity in pediatric cortical dysplasia? Epilepsia 2005; 46 Suppl 5:82-88.
10. Andres M, Andre VM, Nguyen S, Salamon N, Cepeda C, Levine MS et al. Human cortical dysplasia and epilepsy:an ontogenetic hypothesis based on volumetric MRI and NeuN neuronal density and size measurements. Cereb Cortex 2005; 15:194-210.
11. Cepeda C, Andre VM, Levine MS, Salamon N, Miyata H, Vinters HV et al. Epileptogenesis in pediatric cortical dysplasia:the dysmature cerebral developmental hypothesis. Epilepsy Behav 2006; 9:219-235.
12. Thom M, Martinian L, Sen A, Cross JH, Harding BN, Sisodiya SM. Cortical neuronal densities and lamination in focal cortical dysplasia. Acta Neuropathol 2005; 110: 383-392.
13. Shin YJ, Choi JS, Choi JY, Cha JH, Chun MH, Lee MY. Enhanced expression of vascular endothelial growth factor receptor-3 in the subventricular zone of stroke-lesioned rats. Neurosci Lett 2010; 469:194-198.
14. Lamparello P, Baybis M, Pollard J, Hol EM, Eisenstat DD, Aronica E et al. Developmental lineage of cell types in cortical dysplasia with balloon cells. Brain 2007; 130:2267-2276.
15. Boer K, Lucassen PJ, Spliet WG, Vreugdenhil E, van Rijen PC, Troost D et al. Doublecortin-like (DCL) expression in focal cortical dysplasia and cortical tubers. Epilepsia 2009; 50:2629-2637.
16. Crino PB, Trojanowski JQ, Dichter MA, Eberwine J. Embryonic neuronal markers in tuberous sclerosis:single-cell molecular pathology. Proc Natl Acad Sci U S A 1996; 93: 14152-14157.
17. Jozwiak J, Jozwiak S, Skopinski P. Immunohistochemical and microscopic studies on giant cells in tuberous sclerosis. Histol Histopathol 2005; 20:1321-1326.
18. Fauser S, Becker A, Schulze-Bonhage A, Hildebrandt M, Tuxhorn I, Pannek HW et al. CD34-immunoreactive balloon cells in cortical malformations. Acta Neuropathol 2004; 108:272-278.
19. Chamberlain WA, Prayson RA. Focal cortical dysplasia type Ⅱ (malformations of cortical development) aberrantly expresses apoptotic proteins. Appl Immunohistochem Mol Morphol 2008; 16:471-476.
20. Mathew T, Srikanth SG, Satishchandra P. Malformations of cortical development (MCDs) and epilepsy:experience from a tertiary care center in south India. Seizure 2010; 19:147-152.
21. Ying Z, Gonzalez-Martinez J, Tilelli C, Bingaman W, Najm I. Expression of neural stem cell surface marker CD133 in balloon cells of human focal cortical dysplasia. Epilepsia 2005; 46:1716-1723.
22. Sen A, Thom M, Martinian L, Harding B, Cross JH, Nikolic M et al. Pathological tau tangles localize to focal cortical dysplasia in older patients. Epilepsia 2007; 48: 1447-1454.
23. Chipuk JE, Moldoveanu T, Llambi F, Parsons MJ, Green DR. The BCL-2 family reunion. Mol Cell; 37:299-310.
24. Boer K, Troost D, Spliet WG, van Rijen PC, Gorter JA, Aronica E. Cellular distribution of vascular endothelial growth factor A (VEGFA) and B (VEGFB) and VEGF receptors 1 and 2 in focal cortical dysplasia type ⅡB. Acta Neuropathol 2008; 115:683-696.
25. Carelli S, Lesma E, Paratore S, Grande V, Zadra G, Bosari S et al. Survivin expression in tuberous sclerosis complex cells. Mol Med 2007; 13:166-177.
26. Wong M. Animal models of focal cortical dysplasia and tuberous sclerosis complex: recent progress toward clinical applications. Epilepsia 2009; 50 Suppl 9:34-44.
1. Mullen RJ, Buck CR, Smith AM. NeuN, a neuronal specific nuclear protein in vertebrates. Development 1992; 116:201-211.
2. Lind D, Franken S, Kappler J, Jankowski J, Schilling K. Characterization of the neuronal marker NeuN as a multiply phosphorylated antigen with discrete subcellular localization. J Neurosci Res 2005; 79:295-302.
3. Kim KK, Adelstein RS, Kawamoto S. Identification of neuronal nuclei (NeuN) as Fox-3, a new member of the Fox-1 gene family of splicing factors. J Biol Chem 2009; 284: 31052-31061.
4. Wolf HK, Buslei R, Schmidt-Kastner R, Schmidt-Kastner PK, Pietsch T, Wiestler OD et al. NeuN:a useful neuronal marker for diagnostic histopathology. J Histochem Cytochem 1996; 44:1167-1171.
5. Van Nassauw L, Wu M, De Jonge F, Adriaensen D, Timmermans JP. Cytoplasmic, but not nuclear, expression of the neuronal nuclei (NeuN) antibody is an exclusive feature of Dogiel type II neurons in the guinea-pig gastrointestinal tract. Histochem Cell Biol 2005; 124:369-377.
6. Kumar SS, Buckmaster PS. Neuron-specific nuclear antigen NeuN is not detectable in gerbil subtantia nigra pars reticulata. Brain Res 2007; 1142:54-60.
7. Voelker CC, Garin N, Taylor JS, Gahwiler BH, Hornung JP, Molnar Z. Selective neurofilament (SMI-32, FNP-7 and N200) expression in subpopulations of layer V pyramidal neurons in vivo and in vitro. Cereb Cortex 2004; 14:1276-1286.
8. Sasaki S, Maruyama S. Immunocytochemical and ultrastructural studies of the motor cortex in amyotrophic lateral sclerosis. Acta Neuropathol 1994; 87:578-585.
9. Toering ST, Boer K, de Groot M, Troost D, Heimans JJ, Spliet WG et al. Expression patterns of synaptic vesicle protein 2A in focal cortical dysplasia and TSC-cortical tubers. Epilepsia 2009; 50:1409-1418.
10. Magavi SS, Macklis JD. Immunocytochemical analysis of neuronal differentiation. Methods Mol Biol 2008; 438:345-352.
11.徐虹.,陈耀文.,韩太真.,沈建新.,霍霞.激光共聚焦扫描观察脑片及其神经元内RNA与DNA变化的技术.激光生物学报 2000；9：15-161.
12. Brazelton TR, Rossi FM, Keshet GI, Blau HM. From marrow to brain:expression of neuronal phenotypes in adult mice. Science 2000; 290:1775-1779.
13. Goetz AK, Scheffler B, Chen HX, Wang S, Suslov O, Xiang H et al. Temporally restricted substrate interactions direct fate and specification of neural precursors derived from embryonic stem cells. Proc Natl Acad Sci U S A 2006; 103:11063-11068.
14. Damianov A, Black DL. Autoregulation of Fox protein expression to produce dominant negative splicing factors. RNA 2010; 16:405-416.
15. Ponthier JL, Schluepen C, Chen W, Lersch RA, Gee SL, Hou VC et al. Fox-2 splicing factor binds to a conserved intron motif to promote inclusion of protein 4.1R alternative exon 16. J Biol Chem 2006; 281:12468-12474.
16. Yeo G, Holste D, Kreiman G, Burge CB. Variation in alternative splicing across human tissues. Genome Biol 2004; 5:R74.
17. Lee CJ, Irizarry K. Alternative splicing in the nervous system:an emerging source of diversity and regulation. Biol Psychiatry 2003; 54:771-776.
18. Preusser M, Laggner U, Haberler C, Heinzl H, Budka H, Hainfellner JA. Comparative analysis of NeuN immunoreactivity in primary brain tumours:conclusions for rational use in diagnostic histopathology. Histopathology 2006; 48:438-444.
19. Edgar MA, Rosenblum MK. The differential diagnosis of central nervous system tumors: a critical examination of some recent immunohistochemical applications. Arch Pathol Lab Med 2008; 132:500-509.
20. Soylemezoglu F, Onder S, Tezel GG, Berker M. Neuronal nuclear antigen (NeuN):a new tool in the diagnosis of central neurocytoma. Pathol Res Pract 2003; 199:463-468.
21.Shen CC, Yang YC, Chiao MT, Cheng WY, Ko JL, Tsuei YS. Characterization of Endogenous Neural Progenitor Cells after Experimental Ischemic Stroke. Curr Neurovasc Res.
22. Li Y, Chopp M, Chen J, Wang L, Gautam SC, Xu YX et al. Intrastriatal transplantation of bone marrow nonhematopoietic cells improves functional recovery after stroke in adult mice. J Cereb Blood Flow Metab 2000; 20:1311-1319.
23. Toda H, Takahashi J, Iwakami N, Kimura T, Hoki S, Mozumi-Kitamura K et al. Grafting neural stem cells improved the impaired spatial recognition in ischemic rats. Neurosci Lett 2001; 316:9-12.
24. Davoli MA, Fourtounis J, Tam J, Xanthoudakis S, Nicholson D, Robertson GS et al. Immunohistochemical and biochemical assessment of caspase-3 activation and DNA fragmentation following transient focal ischemia in the rat. Neuroscience 2002; 115: 125-136.
25. Sugawara T, Lewen A, Noshita N, Gasche Y, Chan PH. Effects of global ischemia duration on neuronal, astroglial, oligodendroglial, and microglial reactions in the vulnerable hippocampal CA1 subregion in rats. J Neurotrauma 2002; 19:85-98.
26. Unal-Cevik I, Kilinc M, Gursoy-Ozdemir Y, Gurer G, Dalkara T. Loss of NeuN immunoreactivity after cerebral ischemia does not indicate neuronal cell loss:a cautionary note. Brain Res 2004; 1015:169-174.
27. Baquet ZC, Bickford PC, Jones KR. Brain-derived neurotrophic factor is required for the establishment of the proper number of dopaminergic neurons in the substantia nigra pars compacta. J Neurosci 2005; 25:6251-6259.
28. Novikova L, Garris BL, Garris DR, Lau YS. Early signs of neuronal apoptosis in the substantia nigra pars compacta of the progressive neurodegenerative mouse 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine/probenecid model of Parkinson's disease. Neuroscience 2006; 140:67-76.
29. Zhu C, Vourc'h P, Fernagut PO, Fleming SM, Lacan S, Dicarlo CD et al. Variable effects of chronic subcutaneous administration of rotenone on striatal histology. J Comp Neurol 2004; 478:418-426.
30. Cannon JR, Greenamyre JT. NeuN is not a reliable marker of dopamine neurons in rat substantia nigra. Neurosci Lett 2009; 464:14-17.
31. McPhail LT, McBride CB, McGraw J, Steeves JD, Tetzlaff W. Axotomy abolishes NeuN expression in facial but not rubrospinal neurons. Exp Neurol 2004; 185:182-190.
32. Collombet JM, Masqueliez C, Four E, Burckhart MF, Bernabe D, Baubichon D et al. Early reduction of NeuN antigenicity induced by soman poisoning in mice can be used to predict delayed neuronal degeneration in the hippocampus. Neurosci Lett 2006; 398: 337-342.
33. Wu KL, Li YQ, Tabassum A, Lu WY, Aubert I, Wong CS. Loss of Neuronal Protein Expression in Mouse Hippocampus After Irradiation. J Neuropathol Exp Neurol 2010; 69:272-280.
34. Long X, Olszewski M, Huang W, Kletzel M. Neural cell differentiation in vitro from adult human bone marrow mesenchymal stem cells. Stem Cells Dev 2005; 14:65-69.
35.Safford KM, Safford SD, Gimble JM, Shetty AK, Rice HE. Characterization of neuronal/glial differentiation of murine adipose-derived adult stromal cells. Exp Neurol 2004; 187:319-328.
36. Darlington PJ, Goldman JS, Cui QL, Antel JP, Kennedy TE. Widespread immunoreactivity for neuronal nuclei in cultured human and rodent astrocytes. J Neurochem 2008; 104:1201-1209.
37. Wong M. Mechanisms of epileptogenesis in tuberous sclerosis complex and related malformations of cortical development with abnormal glioneuronal proliferation. Epilepsia 2008; 49:8-21.
38. Palmini A, Najm I, Avanzini G, Babb T, Guerrini R, Foldvary-Schaefer N et al. Terminology and classification of the cortical dysplasias. Neurology 2004; 62:S2-8.
39. Lee A, Maldonado M, Baybis M, Walsh CA, Scheithauer B, Yeung R et al. Markers of cellular proliferation are expressed in cortical tubers. Ann Neurol 2003; 53:668-673.
40. Talos DM, Kwiatkowski DJ, Cordero K, Black PM, Jensen FE. Cell-specific alterations of glutamate receptor expression in tuberous sclerosis complex cortical tubers. Ann Neurol 2008; 63:454-465.
41. Thom M, Martinian L, Sen A, Cross JH, Harding BN, Sisodiya SM. Cortical neuronal densities and lamination in focal cortical dysplasia. Acta Neuropathol 2005;110: 383-392.
42. Garbelli R, Meroni A, Magnaghi G, Beolchi MS, Ferrario A, Tassi L et al. Architectural (Type IA) focal cortical dysplasia and parvalbumin immunostaining in temporal lobe epilepsy. Epilepsia 2006; 47:1074-1078.
43. Gonzalez-Martinez JA, Moddel G, Ying Z, Prayson RA, Bingaman WE, Najm IM. Neuronal nitric oxide synthase expression in resected epileptic dysplastic neocortex. J Neurosurg 2009; 110:343-349.
44. Portiansky EL, Barbeito CG, Gimeno EJ, Zuccolilli GO, Goya RG. Loss of NeuN immunoreactivity in rat spinal cord neurons during aging. Exp Neurol 2006; 202: 519-521.