胎鼠神经干细胞培养及移植治疗Tourette综合征大鼠实验研究
|
摘要
研究背景
Tourette综合征(Tourette syndrome,TS)是儿童期最常见的精神运动障碍之一,表现为多发的运动性抽动和发声性抽动,近年来患病率有增加趋势。抽动是指刻板的、不随意的、无目的地重复性动作,该症状在整个TS的自然病程中可以持续多年,发作频率和强度不时地发生变化。TS伴发的行为症状复杂多样,如注意缺陷多动障碍、强迫障碍、自伤行为以及情绪障碍等,不同程度地干扰儿童的认知功能和发育,影响社会适应能力。多数情况下,抽动具有自限性,并可以通过行为矫正或药物治疗得到控制。尽管TS是一种良性的神经精神障碍,仍有接近半数的患者症状迁延,治疗困难,甚至延续到成人,导致终身疾患,严重影响生活质量。对一部分患者而言,抽动可导致终生的损害,约有5%的TS患者其症状是危及生命的,可称之为恶性TS。多种药物对恶性TS的治疗效果欠佳,因此近年来人们逐渐开始尝试用神经外科手术方法治疗恶性TS。深部脑刺激(deep brain stimulation,DBS)疗法对严重的难治性抽动症具有一定疗效,但昂贵的治疗费用以及某些手术并发症限制了DBS的应用。近年来在TS的治疗方面未见有重大的突破性进展,TS的治疗问题仍是本领域研究的重点和难点,因此,我们试图找出一条不同于传统对症治疗方法的途径,即从细胞水平进行治疗,尽可能从根本上替代和修复结构和功能受损伤的神经细胞,以期获得更好的治疗效果,探寻治疗TS,尤其是恶性难治性抽动症的新方法。
近年来,干细胞移植治疗中枢神经系统疾病的潜能引起人们极大的兴趣。神经干细胞(Neural stem cells,NSCs)是一种能够自我更新具有多种分化潜能的未成熟的前体细胞。动物实验表明,将神经干细胞注入动物脑内或血液内以后,细胞可以存活并迁移到受损伤的部位,整合到神经回路中,使动物的神经功能得到改善。神经干细胞移植为许多难以治疗的神经系统疾病提供了新的治疗途径,开拓了中枢神经系统疾病治疗的新视野,在生命科学研究领域中倍受重视。神经干细胞移植将成为多种中枢神经系统功能障碍及损伤后最有前途的神经组织替代性治疗策略。与传统的药物治疗相比,干细胞移植疗法具有其自身独特的优势,尤其适用于某些发病机制不清的神经系统疾病。关于TS的确切的发病机制,无论在分子水平还是细胞水平,至今仍然不是很清楚。神经影像学、神经生理学及尸检研究表明,TS是一种以基底神经节病变为基础的运动传导和调节障碍,病变部位有可能涉及皮质-纹状体-丘脑-皮质环路中的任一部分。
综上所述,我们推测如果将神经干细胞移植到TS病变部位,这些细胞有可能代替中枢神经系统中功能低下或受损的细胞发挥作用,重获内环境的稳态,从而减少刻板行为发作的频率及强度。关于神经干细胞治疗TS的研究还是空白,目前国内外尚未见到有关这方面研究的论文报道。因此,本研究试图考察神经干细胞移植能否改善TS大鼠的脑功能并减少其刻板行为,并对细胞移植治疗的作用机制进行初步地探讨。
目的
探讨胚胎大鼠大脑纹状体神经干细胞的分离、培养、标记与鉴定方法,观察神经干细胞增殖、传代和分化的规律;建立大鼠TS模型,采用立体定向脑内注射法,将神经干细胞移植到大鼠纹状体内,观察神经干细胞原位移植后移植细胞的成活、分化以及大鼠刻板行为改善情况。
方法
1、神经干细胞的取材、分离、培养和鉴定
本实验在无菌条件下取100只E13胚胎大鼠脑纹状体组织,解剖显微镜下用锐性刀片切割脑组织,吸管反复吹打机械分离制备单细胞悬液。调节单细胞悬液浓度,用DMEM/F12培养液调节稀释至细胞密度为5×10~5个/ml,接种到含有EGF(20ng/ml)和bFGF(20ng/ml)的DMEM/F12无血清培养基中,置于37℃、5%CO_2培养箱中培养。根据细胞生长情况每2～3天半量换液,低速离心未贴壁生长的细胞克隆,用新鲜培养液更换半量液体,吹打分散成单细胞及小细胞团,继续培养。原代培养5～6天后,形成数目较多、个体较大约含几十个甚至上百个细胞的“神经球”,采用机械吹打与胰蛋白酶消化相结合的方法及时进行传代培养。收集细胞,离心后弃上清液,加入1ml0.25%胰蛋白酶,同时用吸管反复轻柔吹打细胞悬液,避免产生气泡,将“神经球”重新分散制成单细胞悬液,当消化液略呈乳状时加入等体积的含10%胎牛血清的DMEM/F12培养基终止消化,调节细胞接种密度分瓶传代培养。分别用抗Nestin抗体、抗MAP2抗体、抗GFAP抗体,对培养的细胞进行干细胞特性和多分化潜能的免疫组织化学鉴定。取3～4代神经干细胞用于移植,移植前24小时将BrdU(10μg/ml)加入培养基中对细胞进行标记。
2、TS大鼠动物模型的建立与神经干细胞移植
抽取TS患者静脉血2.5ml,离心后分离血清,采用酶联免疫吸附实验(enzymelinked immunosorbent assay,ELISA)检测患者血清中的抗神经抗体滴度。从TS患者中选取8例OD值最高的患者血清用于大鼠纹状体微量灌注,以建立TS大鼠动物模型。32只Wistar大鼠被随机分为4组,每组8只:假模型组(微量注射正常儿童血清),TS组(微量注射TS患者血清),TS+PBS组(纹状体内被微量注射TS患者血清后再给予PBS做治疗对照),TS+NSCs移植组(纹状体内被微量注射TS患者血清后再进行NSC脑内移植治疗)。将大鼠麻醉后固定于立体定向仪上,无菌条件下将套管置入大鼠双侧纹状体内,套管置入位点为前囟前2.0mm、正中线左右4.0mm、颅下7.0mm。1周后通过微量渗透泵将血清以0.5μl/h的速度缓慢注入大鼠纹状体内,持续灌注72小时。血清灌注结束后,进行神经干细胞移植,在损伤位点原位注射神经干细胞悬液,每个注射位点接受2μl的细胞悬液,细胞密度为5×10~5个/μl。在移植后1天,7天,14天和21天评估大鼠刻板动作行为,在每12小时光循环结束前的最后30分钟观察大鼠的动作行为如啮咬(用牙齿咬笼子,嚼木屑,无目的咀嚼)、将前爪举到嘴边或面部、舔(不包括理毛)、理毛行为、摇头、摇动爪子、用后腿站立及阶段性的发声,将所观察到的动作行为计数的总和作为评估刻板运动的总分。
3、神经干细胞脑内移植后的存活和分化
神经干细胞移植后3周,处死大鼠,进行心脏灌流固定。快速灌注PBS150～200ml,冲洗脉管系统,随后换用4℃的4%多聚甲醛溶液继续灌注。灌流结束后将大鼠断头取脑,寻找双侧脑表面的标志针孔,以针孔为中心切取标志针孔周围脑组织;将脑组织浸入4%多聚甲醛固定。制备脑组织石蜡快,采取连续切片法,以移植部位为中心连续冠状切片。采用BCIP/NBT及AEC两种不同的显色系统,分别进行BrdU抗体+Nestin抗体、BrdU抗体+MAP2抗体、BrdU抗体+GFAP抗体双染免疫组化分析,观察神经干细胞移植到大鼠纹状体后在宿主脑内的存活及分化情况。
结果
1、神经干细胞的取材、分离、培养和鉴定
在含有EGF和bFGF的培养基中,大鼠纹状体NSCS开始增殖,细胞种植后24小时显微镜下观察可见大量分裂期的神经干细胞,细胞呈对称或不对称分裂,有的呈哑铃状,连续观察可见细胞分裂成两个完全独立的神经干细胞。随后神经干细胞进入快速增殖期,形成3～6个疏松连接的细胞团,随着时间的延长,细胞不断分裂,在48～72小时后形成含有10～20个细胞的克隆。神经球逐渐增大,细胞之间结合更加紧密,细胞光亮,折光性好,核、浆比例大。分离神经球后继续传代培养,细胞数量持续增加。神经干细胞在细胞分化液中培养,2小时后细胞贴壁,逐渐形成突起,随着培养时间延长,突起逐渐延长,显微镜下呈现神经元和胶质细胞形态,细胞染色证实细胞系巢蛋白Nestin阳性,经血清培养液诱导分化后部分呈MAP2阳性,部分呈GFAP阳性。
2、TS大鼠动物模型的建立与神经干细胞移植
基于抗神经抗体ELISA的OD值,从20例TS病人中选取8例OD值最高的病人的血清(0.904±0.177)用于大鼠纹状体微量灌注。同样从20例正常对照中选取8例OD值最低的血清(0.252±0.193)用于灌注。TS病人的血清中的抗神经抗体可致纹状体功能受损,纹状体内微量灌注TS血清后,成功诱导出大鼠的刻板行为。大鼠的刻板行为如啮咬、将前爪举到嘴边或面部、舔、摇头、摇动爪子、用后腿站立及阶段性的发声显著增加。灌注TS儿童血清的大鼠的刻板行为与灌注正常儿童血清的刻板行为之间差别显著,TS大鼠的啮咬、将前爪举到嘴边或面部、自咬、理毛、舔、摇头、摇动爪子、用后腿站立、阶段性的发声明显增加。在移植后1天、7天、14天和21天时监测四组大鼠的刻板行为。重复测量数据的方差分析表明组间差别(F=354.13,p＜0.0001)、时间差别(F=106.01,p＜0.0001)以及二者间的交互作用(F=21.38,p＜0.0001)差别均显著。对TS+NSCs和TS+PBS两组分析结果表明二者间其差别显著。注射神经干细胞的大鼠在14天和21天时的刻板行为明显减少,Tukey-Kramer post hoc test统计学分析表明在14天时即出现明显组间差别。移植组的刻板行为仍高于正常对照组(p＜0.0001),提示在3周内损伤并未得到完全恢复。
3、神经干细胞脑内移植后的存活和分化
在移植3周后对大鼠大脑进行免疫组织化学分析。本研究采用BrdU标记移植细胞,在纹状体内检测到细胞核着色为暗紫色的BrdU阳性细胞即为移植细胞。移植位点附近的BrdU阳性细胞分布较多,表明移植细胞在宿主脑内存活。部分移植细胞表达BrdU和MAP2,部分表达BrdU和GFAP,表明神经干细胞在移植部位生长并分化为神经元或星形胶质细胞。另外,仍有少数神经干细胞未分化,呈BrdU及Nestin双阳性。
结论
1、本实验成功分离和培养了来源于胚胎大鼠纹状体的NSCs,并证明它们具备了NSCs的基本特征:(1)表达神经干细胞标志性蛋白Nestin;(2)在培养中存活的NSCs可由单个细胞形成神经细胞球,表明具有自我更新、自我增殖能力;(3)在诱导分化条件下具有形成神经元、神经胶质细胞的能力。NSCs的自我增殖特性和多向分化潜能不因细胞传代而改变。
2、向大鼠纹状体内微量灌注含有高滴度抗神经抗体的TS患者血清,可以成功诱导出大鼠的刻板行为;纹状体内神经干细胞移植可减少TS大鼠的刻板行为。纹状体内注射神经干细胞可能对TS具有一定的治疗作用。
3、神经干细胞能够在TS大鼠脑内生长存活,部分神经干细胞分化成神经元及神经胶质细胞,可促进中枢神经系统的结构和功能的修复与重建。移植区未见异常肿瘤组织生成,无明显免疫排斥反应。
Background
Tourette syndrome (TS) is a neurobehavioural disorder occurring in children and is characterized by multiple motor and vocal tics. Tics are stereotypic, involuntary, purposeless and repetitive movements which persist for years in TS and wax and wane in frequency and intensity during their natural courses. Although TS is a benign neuropsychiatric disorder, nearly half of the cohort can not be free of tics even until 18 years of age. In most cases, tics are self-limited or can be treated by behavioural or pharmacological therapy. However, for some individuals, tics can cause lifelong impairment and about 5% of TS patients have life-threatening symptoms, which were defined malignant TS. These symptoms are intractable to conservative treatment and various attempts have been made to treat these patients through neurosurgical procedures. Recently, there has been increasing interest in deep brain stimulation (DBS) as a potential treatment for patients with severe, refractory tics. Nonetheless, the use of DBS for the treatment of TS is limited because of complications related to the surgical procedure and the infrequent hardware. Therefore, new therapy should be explored for those unfortunate patients who are significantly impaired.
During recent years, stem cell-based therapy has been proved promising as a potential treatment for many neurological disorders. Neural stem cells (NSCs) are considered a heterogeneous population of mitotically active, self-renewing, multipotent and immature progenitor cells. Animal experiments suggest that if NSCs are injected into the brain or even the bloodstream, the transplanted cells would survive and migrate to damaged portion of the nervous system to incorporate into working neural circuits, and the tested animals display significant functional improvement. Therefore, NSC replacement is thought to be a promising alternative therapeutic approach for treating neurological disorders. It may breakthrough some of the existing limitations of traditional pharmaceutical approaches. Most important of all, neural stem cell therapy is a good choice for the treatment of neural diseases whose exact pathogenesis are unclear.
The definitive pathophysiological mechanism of tics, at molecular and cellular level, is still unknown. However, structural and functional neuroimaging, neurophysiological, and post-mortem studies have shown the dysfunction of the basal ganglia and related cortico-striato-thalamo-cortical circuits, and the dopaminergic neuronal system. Taking these scholarship into consideration, we hypothesized that intrastriatal tansplantation of NSCs could replace disfunctioned or damaged cells in the central nervous system of TS, reduce the frequency and intensity of stereotypic behaviors and further help regain homeostasis.
Objectives
To investigate the methods of isolation, cultivation, purification, expanding, BrdU marking and identification of neural stem cells from striatum of fetal Wistar rat and observe the proliferation and differentiation of neural stem cells. Study the effect of neural stem cell transplantation on Wistar rats which were microinfused with the sera of TS patients. Observe the growth and differentiation of transplanted cells in the rat brain and the therapeutic effects of neural stem cells on the stereotypic behavior of TS rat.
Methods
NSCs were isolated from rat embryonic brain (E13). Embryonic striatum tissue was microdissected under a stereo microscope and minced into small pieces, and then mechanically triturated through a 26-gauge needle and dissociated to single cell suspension. Cells were cultured in serum-free basic medium DMEM: F12 supplemented with human recombinant epidermal growth factor (EGF) (20 ng/ml), fibroblast growth factor-basic (bFGF) (20 ng/ml), B27 (2%), penicillin-streptomycin 1% . Cells were incubated with 5% CO2 at 37℃. Within 2-3 days the cells grew as free floating neurospheres and half of the growth medium was replaced. Expanded neurospheres were collected by centrifugation and dissociated into single cells once every 5-6 days with 0.25% trypsin and mechanical trituration. NSCs were co-incubated with 5-bromodeoxyuridine (BrdU, 10μg/ml) for 24h prior to transplantation. Then BrdU-labeled NSCs (passage number 3-4) were harvested and enzymatically dissociated into single cells. Cells with higher than 85% viability were centrifuged and resuspended in basic medium supplemented with B27 at a density of 5×10~5 cells/μl and then stored on ice until grafting. To determine the phenotype of cultured cells, the cells were separately immuno-stained with anti-Nestin antibody, anti-MAP2 antibody or anti-GFAP antibody.
Blood samples were drawn from patients with TS and were sent for further enzyme-linked immunosorbent assays (ELISA) to a laboratory. The selection of sera for infusion was based on ELISA optical density readings against caudate homogenate, that is, we selected the highest serum OD readings for TS subjects and the lowest OD readings for the control infusion subgroup. A total of thirty-two Wistar rats were used in the study and randomly divided into four groups: sham (microinfused with normal sera), TS alone (microinfused with TS sera), TS plus PBS vehicle injection (TS+PBS) and TS plus NSCs grating (TS+NSCs) (n=32, i.e., n=8 for each group).
Rats were deeply anesthetized with chloral hydrate (400 mg/kg, i.p.) and placed in a stereotaxic apparatus with the incisor bar set at 3.5mm below the interaural line. Then using aseptic surgical technique, the skull was exposed and holes were drilled where appropriate and 28 gauge guide cannulae were implanted into bilateral striatum. Coordinates for cannulae placements were anterior-posterior 2.0mm from bregma and medial-lateral 4.0mm and dorsoventral -7.0mm from the skull. One week later, osmotic mini pump filled with PBS was connected to each cannula by a polyethylene tube loaded with 50μl of undiluted TS or control serum under sterile conditions. Sera were microinfused at a rate of 0.5μl /hour for 72 hours. The neural stem cell suspension (2μl) was injected bilaterally using a 5μl Hamilton syringe with a 26-gauge needle at the sera-infusion site after the pumps were removed. Rat movements were video and audio taped at the end of the rat's 12-hour light cycle for 30 min. Several categories of stereotypy including bites (teeth touching the cage, wood chips, vacuous chewing or other objects except the body), taffy pulling (raises of the forepaw to the mouth and face), self-gnawing, licking not associated with grooming, grooming, head shaking, paw shaking, rearing and episodic utterances (EU) were recorded. Stereotypic movements were recorded at 1 day, 7 days, 14 days and 21 days after transplantation.
Three weeks after transplantation, rats were given a lethal dose of chloride hydrate (600 mg/kg) and transcardially perfused with cold 0.9% NS followed by 4% paraformaldehyde (PFA) in 0.1 M PBS (pH 7.4, 4℃). Brains were removed from the cranium and post-fixed in PFA prior to sectioning. Rat brain sections were embedded in paraffin and 4-μm coronal sections were prepared. All the immunostaining processes were finished according to the instruction of histostain?-DS kit. Primary antibodies which included mouse monoclonal to GFAP—astrocyte marker, mouse monoclonal to MAP2—neuron marker and mouse monoclonal to Nestin—neural stem cell marker were added.
Results
Following the isolation, a close-to-single-cell suspension was obtained, which consisted of viable small, round cells. In suspension cultures containing EGF and bFGF, rat striatal NSCs began to proliferate and adherent clonal clusters of cells (10 - 20 cells/clone) started to appear after 48 - 72 h. The clusters divided rapidly and generated large colonies, i. e. neurospheres. These floating neurospheres displayed a spherical shape, and the cells forming the neurosphere were phase bright. These spheres were successfully passaged by dissociation and reculture. This procedure resulted in a rapid increase in cell number. The neural cells dissociated from E13 rat striatum had the capacity to proliferate and form neurospheres. The neurosphere could express Nestin, gernerate MAP2 or GFAP positive neural cells.
Eight children were selected from 20 patients with TS and their sera were used for rat striatal microinfusion. Similarly, eight specimens of sera were selected from 20 control samples. The selection of sera for infusion was based on ELISA optical density readings against caudate homogenate, that is, we selected the highest serum OD readings for TS subjects and the lowest OD readings for the control infusion subgroup. The mean±SD optical density readings in serum of 8 TS subjects selected for microinfusion was 0.904±0.177 and 0.252±0.193 for the 8 selected controls.
Serologic studies of children with TS have detected the existence of anti-neural antibodies, which can induce striatal dysfunction. In our study, stereotypies were successfully induced in rats by intrastriatal microinfusion of TS sera under noninflammatory conditions. After infusion of TS sera, stereotypic behaviors in rats increased significantly. Marked differences were observed in stereotypies of TS rats compared to control rats after microinfusion. TS rats exhibited significant increases in bites, taffy-pulling, gnawing, licking, head shaking, paw shaking, rearing and episodic utterance. All rats in four groups were observed at 1 day, 7 days, 14 days, and 21 days after their transplantations. Stereotypic behaviors of each rat were recorded and counted through each 30 minutes of observation period. The statistical analysis suggested that TS rats had increased stereotypic behaviors compared to normal rats (Sham). Using a repeated measurements analysis of variance (ANOVA), we found that the overall model had significant group (F=354.13, p<0.0001) and day (F= 106.01, p<0.0001) effects, as well as a (group×day) interaction (F=21.38, p<0.0001), indicating varying degrees of differences among the groups and across days. A model with just TS+NSCs and TS+PBS groups indicated significant differences between groups(F=36.63, p=0.0001) and a strong day effect(F=100.23, p<0.0001). Importantly, animals with NSCs grafts showed a significant decrease in stereotypic behaviors at 14 and 21 days. Tukey-Kramer post hoc test showed significant differences between TS+NSCs and TS+PBS groups beginning at 2 weeks post-transplantation. However, rats receiving NSC grafts (TS+NSCs) still had higher stereotypic behavior counts than sham controls (p<0.0001), which indicated that the animals' impairment did not fully recover in a short period of 3 weeks.
Histological analyses of recipient rat brains transplanted with NSCs or injected with PBS (vehicle) were given 3 weeks after transplantation. In order to trace the transplanted cells, we used BrdU labeling, so that the presence of the implanted NSCs within striatum of TS rats identifiable for the nuclei of the grafted cells were BrdU-positive and stained with purple. In the TS+NSCs rats, BrdU-positive cells were prevalent in the striatum sections near the injection site, suggesting the survival of the cells. Some of the grafted cells expressed both BrdU and MAP-2 and others expressed both BrdU and GFAP. These results suggested that NSCs used as grafts grew at the injection site and differentiated into neurons or astrocytes. Our results also indicated that a considerable portion of the transplanted cells had differentiated to mature neurons in vivo, which may lead to histological and functional reconstitution in the CNS. Besides, few BrdU and Nestin double positive cells were found, indicating undiferentiation of a small number of the grafted cells.
Conclusions
Neural stem cells were isolated from fetal rat striatum and were certificated as a heterogeneous population of mitotically active, self-renewing, multipotent, immature progenitor cells. These cells expressed the intermediate filament protein Nestin and grew as neurospheres in vitro. Neural stem cells had the capacity to differentiate into neurons and gilocytes..
In our study, stereotypies were successfully induced in rats by intrastriatal microinfusion of TS sera under noninflammatory conditions. Our results showed that intrastriatum transplanted rat NSCs relieved stereotypic behaviors. These results suggested the potential of using NSCs intrastriatum as a clinical treatment for TS. Neural stem cells survived in the brain of TS rat and part of them differentiated into neurons and gliocytes. These newly generated neurons from the transplanted NSCs might have played a role for the neural networks in the damaged areas. No signs of immunologic rejection or non-neural tissue growth were found in the host rat brain after transplantation.
Tourette综合征(Tourette syndrome,TS)是儿童期最常见的精神运动障碍之一,表现为多发的运动性抽动和发声性抽动,近年来患病率有增加趋势。抽动是指刻板的、不随意的、无目的地重复性动作,该症状在整个TS的自然病程中可以持续多年,发作频率和强度不时地发生变化。TS伴发的行为症状复杂多样,如注意缺陷多动障碍、强迫障碍、自伤行为以及情绪障碍等,不同程度地干扰儿童的认知功能和发育,影响社会适应能力。多数情况下,抽动具有自限性,并可以通过行为矫正或药物治疗得到控制。尽管TS是一种良性的神经精神障碍,仍有接近半数的患者症状迁延,治疗困难,甚至延续到成人,导致终身疾患,严重影响生活质量。对一部分患者而言,抽动可导致终生的损害,约有5%的TS患者其症状是危及生命的,可称之为恶性TS。多种药物对恶性TS的治疗效果欠佳,因此近年来人们逐渐开始尝试用神经外科手术方法治疗恶性TS。深部脑刺激(deep brain stimulation,DBS)疗法对严重的难治性抽动症具有一定疗效,但昂贵的治疗费用以及某些手术并发症限制了DBS的应用。近年来在TS的治疗方面未见有重大的突破性进展,TS的治疗问题仍是本领域研究的重点和难点,因此,我们试图找出一条不同于传统对症治疗方法的途径,即从细胞水平进行治疗,尽可能从根本上替代和修复结构和功能受损伤的神经细胞,以期获得更好的治疗效果,探寻治疗TS,尤其是恶性难治性抽动症的新方法。
近年来,干细胞移植治疗中枢神经系统疾病的潜能引起人们极大的兴趣。神经干细胞(Neural stem cells,NSCs)是一种能够自我更新具有多种分化潜能的未成熟的前体细胞。动物实验表明,将神经干细胞注入动物脑内或血液内以后,细胞可以存活并迁移到受损伤的部位,整合到神经回路中,使动物的神经功能得到改善。神经干细胞移植为许多难以治疗的神经系统疾病提供了新的治疗途径,开拓了中枢神经系统疾病治疗的新视野,在生命科学研究领域中倍受重视。神经干细胞移植将成为多种中枢神经系统功能障碍及损伤后最有前途的神经组织替代性治疗策略。与传统的药物治疗相比,干细胞移植疗法具有其自身独特的优势,尤其适用于某些发病机制不清的神经系统疾病。关于TS的确切的发病机制,无论在分子水平还是细胞水平,至今仍然不是很清楚。神经影像学、神经生理学及尸检研究表明,TS是一种以基底神经节病变为基础的运动传导和调节障碍,病变部位有可能涉及皮质-纹状体-丘脑-皮质环路中的任一部分。
综上所述,我们推测如果将神经干细胞移植到TS病变部位,这些细胞有可能代替中枢神经系统中功能低下或受损的细胞发挥作用,重获内环境的稳态,从而减少刻板行为发作的频率及强度。关于神经干细胞治疗TS的研究还是空白,目前国内外尚未见到有关这方面研究的论文报道。因此,本研究试图考察神经干细胞移植能否改善TS大鼠的脑功能并减少其刻板行为,并对细胞移植治疗的作用机制进行初步地探讨。
目的
探讨胚胎大鼠大脑纹状体神经干细胞的分离、培养、标记与鉴定方法,观察神经干细胞增殖、传代和分化的规律;建立大鼠TS模型,采用立体定向脑内注射法,将神经干细胞移植到大鼠纹状体内,观察神经干细胞原位移植后移植细胞的成活、分化以及大鼠刻板行为改善情况。
方法
1、神经干细胞的取材、分离、培养和鉴定
本实验在无菌条件下取100只E13胚胎大鼠脑纹状体组织,解剖显微镜下用锐性刀片切割脑组织,吸管反复吹打机械分离制备单细胞悬液。调节单细胞悬液浓度,用DMEM/F12培养液调节稀释至细胞密度为5×10~5个/ml,接种到含有EGF(20ng/ml)和bFGF(20ng/ml)的DMEM/F12无血清培养基中,置于37℃、5%CO_2培养箱中培养。根据细胞生长情况每2～3天半量换液,低速离心未贴壁生长的细胞克隆,用新鲜培养液更换半量液体,吹打分散成单细胞及小细胞团,继续培养。原代培养5～6天后,形成数目较多、个体较大约含几十个甚至上百个细胞的“神经球”,采用机械吹打与胰蛋白酶消化相结合的方法及时进行传代培养。收集细胞,离心后弃上清液,加入1ml0.25%胰蛋白酶,同时用吸管反复轻柔吹打细胞悬液,避免产生气泡,将“神经球”重新分散制成单细胞悬液,当消化液略呈乳状时加入等体积的含10%胎牛血清的DMEM/F12培养基终止消化,调节细胞接种密度分瓶传代培养。分别用抗Nestin抗体、抗MAP2抗体、抗GFAP抗体,对培养的细胞进行干细胞特性和多分化潜能的免疫组织化学鉴定。取3～4代神经干细胞用于移植,移植前24小时将BrdU(10μg/ml)加入培养基中对细胞进行标记。
2、TS大鼠动物模型的建立与神经干细胞移植
抽取TS患者静脉血2.5ml,离心后分离血清,采用酶联免疫吸附实验(enzymelinked immunosorbent assay,ELISA)检测患者血清中的抗神经抗体滴度。从TS患者中选取8例OD值最高的患者血清用于大鼠纹状体微量灌注,以建立TS大鼠动物模型。32只Wistar大鼠被随机分为4组,每组8只:假模型组(微量注射正常儿童血清),TS组(微量注射TS患者血清),TS+PBS组(纹状体内被微量注射TS患者血清后再给予PBS做治疗对照),TS+NSCs移植组(纹状体内被微量注射TS患者血清后再进行NSC脑内移植治疗)。将大鼠麻醉后固定于立体定向仪上,无菌条件下将套管置入大鼠双侧纹状体内,套管置入位点为前囟前2.0mm、正中线左右4.0mm、颅下7.0mm。1周后通过微量渗透泵将血清以0.5μl/h的速度缓慢注入大鼠纹状体内,持续灌注72小时。血清灌注结束后,进行神经干细胞移植,在损伤位点原位注射神经干细胞悬液,每个注射位点接受2μl的细胞悬液,细胞密度为5×10~5个/μl。在移植后1天,7天,14天和21天评估大鼠刻板动作行为,在每12小时光循环结束前的最后30分钟观察大鼠的动作行为如啮咬(用牙齿咬笼子,嚼木屑,无目的咀嚼)、将前爪举到嘴边或面部、舔(不包括理毛)、理毛行为、摇头、摇动爪子、用后腿站立及阶段性的发声,将所观察到的动作行为计数的总和作为评估刻板运动的总分。
3、神经干细胞脑内移植后的存活和分化
神经干细胞移植后3周,处死大鼠,进行心脏灌流固定。快速灌注PBS150～200ml,冲洗脉管系统,随后换用4℃的4%多聚甲醛溶液继续灌注。灌流结束后将大鼠断头取脑,寻找双侧脑表面的标志针孔,以针孔为中心切取标志针孔周围脑组织;将脑组织浸入4%多聚甲醛固定。制备脑组织石蜡快,采取连续切片法,以移植部位为中心连续冠状切片。采用BCIP/NBT及AEC两种不同的显色系统,分别进行BrdU抗体+Nestin抗体、BrdU抗体+MAP2抗体、BrdU抗体+GFAP抗体双染免疫组化分析,观察神经干细胞移植到大鼠纹状体后在宿主脑内的存活及分化情况。
结果
1、神经干细胞的取材、分离、培养和鉴定
在含有EGF和bFGF的培养基中,大鼠纹状体NSCS开始增殖,细胞种植后24小时显微镜下观察可见大量分裂期的神经干细胞,细胞呈对称或不对称分裂,有的呈哑铃状,连续观察可见细胞分裂成两个完全独立的神经干细胞。随后神经干细胞进入快速增殖期,形成3～6个疏松连接的细胞团,随着时间的延长,细胞不断分裂,在48～72小时后形成含有10～20个细胞的克隆。神经球逐渐增大,细胞之间结合更加紧密,细胞光亮,折光性好,核、浆比例大。分离神经球后继续传代培养,细胞数量持续增加。神经干细胞在细胞分化液中培养,2小时后细胞贴壁,逐渐形成突起,随着培养时间延长,突起逐渐延长,显微镜下呈现神经元和胶质细胞形态,细胞染色证实细胞系巢蛋白Nestin阳性,经血清培养液诱导分化后部分呈MAP2阳性,部分呈GFAP阳性。
2、TS大鼠动物模型的建立与神经干细胞移植
基于抗神经抗体ELISA的OD值,从20例TS病人中选取8例OD值最高的病人的血清(0.904±0.177)用于大鼠纹状体微量灌注。同样从20例正常对照中选取8例OD值最低的血清(0.252±0.193)用于灌注。TS病人的血清中的抗神经抗体可致纹状体功能受损,纹状体内微量灌注TS血清后,成功诱导出大鼠的刻板行为。大鼠的刻板行为如啮咬、将前爪举到嘴边或面部、舔、摇头、摇动爪子、用后腿站立及阶段性的发声显著增加。灌注TS儿童血清的大鼠的刻板行为与灌注正常儿童血清的刻板行为之间差别显著,TS大鼠的啮咬、将前爪举到嘴边或面部、自咬、理毛、舔、摇头、摇动爪子、用后腿站立、阶段性的发声明显增加。在移植后1天、7天、14天和21天时监测四组大鼠的刻板行为。重复测量数据的方差分析表明组间差别(F=354.13,p＜0.0001)、时间差别(F=106.01,p＜0.0001)以及二者间的交互作用(F=21.38,p＜0.0001)差别均显著。对TS+NSCs和TS+PBS两组分析结果表明二者间其差别显著。注射神经干细胞的大鼠在14天和21天时的刻板行为明显减少,Tukey-Kramer post hoc test统计学分析表明在14天时即出现明显组间差别。移植组的刻板行为仍高于正常对照组(p＜0.0001),提示在3周内损伤并未得到完全恢复。
3、神经干细胞脑内移植后的存活和分化
在移植3周后对大鼠大脑进行免疫组织化学分析。本研究采用BrdU标记移植细胞,在纹状体内检测到细胞核着色为暗紫色的BrdU阳性细胞即为移植细胞。移植位点附近的BrdU阳性细胞分布较多,表明移植细胞在宿主脑内存活。部分移植细胞表达BrdU和MAP2,部分表达BrdU和GFAP,表明神经干细胞在移植部位生长并分化为神经元或星形胶质细胞。另外,仍有少数神经干细胞未分化,呈BrdU及Nestin双阳性。
结论
1、本实验成功分离和培养了来源于胚胎大鼠纹状体的NSCs,并证明它们具备了NSCs的基本特征:(1)表达神经干细胞标志性蛋白Nestin;(2)在培养中存活的NSCs可由单个细胞形成神经细胞球,表明具有自我更新、自我增殖能力;(3)在诱导分化条件下具有形成神经元、神经胶质细胞的能力。NSCs的自我增殖特性和多向分化潜能不因细胞传代而改变。
2、向大鼠纹状体内微量灌注含有高滴度抗神经抗体的TS患者血清,可以成功诱导出大鼠的刻板行为;纹状体内神经干细胞移植可减少TS大鼠的刻板行为。纹状体内注射神经干细胞可能对TS具有一定的治疗作用。
3、神经干细胞能够在TS大鼠脑内生长存活,部分神经干细胞分化成神经元及神经胶质细胞,可促进中枢神经系统的结构和功能的修复与重建。移植区未见异常肿瘤组织生成,无明显免疫排斥反应。
Background
Tourette syndrome (TS) is a neurobehavioural disorder occurring in children and is characterized by multiple motor and vocal tics. Tics are stereotypic, involuntary, purposeless and repetitive movements which persist for years in TS and wax and wane in frequency and intensity during their natural courses. Although TS is a benign neuropsychiatric disorder, nearly half of the cohort can not be free of tics even until 18 years of age. In most cases, tics are self-limited or can be treated by behavioural or pharmacological therapy. However, for some individuals, tics can cause lifelong impairment and about 5% of TS patients have life-threatening symptoms, which were defined malignant TS. These symptoms are intractable to conservative treatment and various attempts have been made to treat these patients through neurosurgical procedures. Recently, there has been increasing interest in deep brain stimulation (DBS) as a potential treatment for patients with severe, refractory tics. Nonetheless, the use of DBS for the treatment of TS is limited because of complications related to the surgical procedure and the infrequent hardware. Therefore, new therapy should be explored for those unfortunate patients who are significantly impaired.
During recent years, stem cell-based therapy has been proved promising as a potential treatment for many neurological disorders. Neural stem cells (NSCs) are considered a heterogeneous population of mitotically active, self-renewing, multipotent and immature progenitor cells. Animal experiments suggest that if NSCs are injected into the brain or even the bloodstream, the transplanted cells would survive and migrate to damaged portion of the nervous system to incorporate into working neural circuits, and the tested animals display significant functional improvement. Therefore, NSC replacement is thought to be a promising alternative therapeutic approach for treating neurological disorders. It may breakthrough some of the existing limitations of traditional pharmaceutical approaches. Most important of all, neural stem cell therapy is a good choice for the treatment of neural diseases whose exact pathogenesis are unclear.
The definitive pathophysiological mechanism of tics, at molecular and cellular level, is still unknown. However, structural and functional neuroimaging, neurophysiological, and post-mortem studies have shown the dysfunction of the basal ganglia and related cortico-striato-thalamo-cortical circuits, and the dopaminergic neuronal system. Taking these scholarship into consideration, we hypothesized that intrastriatal tansplantation of NSCs could replace disfunctioned or damaged cells in the central nervous system of TS, reduce the frequency and intensity of stereotypic behaviors and further help regain homeostasis.
Objectives
To investigate the methods of isolation, cultivation, purification, expanding, BrdU marking and identification of neural stem cells from striatum of fetal Wistar rat and observe the proliferation and differentiation of neural stem cells. Study the effect of neural stem cell transplantation on Wistar rats which were microinfused with the sera of TS patients. Observe the growth and differentiation of transplanted cells in the rat brain and the therapeutic effects of neural stem cells on the stereotypic behavior of TS rat.
Methods
NSCs were isolated from rat embryonic brain (E13). Embryonic striatum tissue was microdissected under a stereo microscope and minced into small pieces, and then mechanically triturated through a 26-gauge needle and dissociated to single cell suspension. Cells were cultured in serum-free basic medium DMEM: F12 supplemented with human recombinant epidermal growth factor (EGF) (20 ng/ml), fibroblast growth factor-basic (bFGF) (20 ng/ml), B27 (2%), penicillin-streptomycin 1% . Cells were incubated with 5% CO2 at 37℃. Within 2-3 days the cells grew as free floating neurospheres and half of the growth medium was replaced. Expanded neurospheres were collected by centrifugation and dissociated into single cells once every 5-6 days with 0.25% trypsin and mechanical trituration. NSCs were co-incubated with 5-bromodeoxyuridine (BrdU, 10μg/ml) for 24h prior to transplantation. Then BrdU-labeled NSCs (passage number 3-4) were harvested and enzymatically dissociated into single cells. Cells with higher than 85% viability were centrifuged and resuspended in basic medium supplemented with B27 at a density of 5×10~5 cells/μl and then stored on ice until grafting. To determine the phenotype of cultured cells, the cells were separately immuno-stained with anti-Nestin antibody, anti-MAP2 antibody or anti-GFAP antibody.
Blood samples were drawn from patients with TS and were sent for further enzyme-linked immunosorbent assays (ELISA) to a laboratory. The selection of sera for infusion was based on ELISA optical density readings against caudate homogenate, that is, we selected the highest serum OD readings for TS subjects and the lowest OD readings for the control infusion subgroup. A total of thirty-two Wistar rats were used in the study and randomly divided into four groups: sham (microinfused with normal sera), TS alone (microinfused with TS sera), TS plus PBS vehicle injection (TS+PBS) and TS plus NSCs grating (TS+NSCs) (n=32, i.e., n=8 for each group).
Rats were deeply anesthetized with chloral hydrate (400 mg/kg, i.p.) and placed in a stereotaxic apparatus with the incisor bar set at 3.5mm below the interaural line. Then using aseptic surgical technique, the skull was exposed and holes were drilled where appropriate and 28 gauge guide cannulae were implanted into bilateral striatum. Coordinates for cannulae placements were anterior-posterior 2.0mm from bregma and medial-lateral 4.0mm and dorsoventral -7.0mm from the skull. One week later, osmotic mini pump filled with PBS was connected to each cannula by a polyethylene tube loaded with 50μl of undiluted TS or control serum under sterile conditions. Sera were microinfused at a rate of 0.5μl /hour for 72 hours. The neural stem cell suspension (2μl) was injected bilaterally using a 5μl Hamilton syringe with a 26-gauge needle at the sera-infusion site after the pumps were removed. Rat movements were video and audio taped at the end of the rat's 12-hour light cycle for 30 min. Several categories of stereotypy including bites (teeth touching the cage, wood chips, vacuous chewing or other objects except the body), taffy pulling (raises of the forepaw to the mouth and face), self-gnawing, licking not associated with grooming, grooming, head shaking, paw shaking, rearing and episodic utterances (EU) were recorded. Stereotypic movements were recorded at 1 day, 7 days, 14 days and 21 days after transplantation.
Three weeks after transplantation, rats were given a lethal dose of chloride hydrate (600 mg/kg) and transcardially perfused with cold 0.9% NS followed by 4% paraformaldehyde (PFA) in 0.1 M PBS (pH 7.4, 4℃). Brains were removed from the cranium and post-fixed in PFA prior to sectioning. Rat brain sections were embedded in paraffin and 4-μm coronal sections were prepared. All the immunostaining processes were finished according to the instruction of histostain?-DS kit. Primary antibodies which included mouse monoclonal to GFAP—astrocyte marker, mouse monoclonal to MAP2—neuron marker and mouse monoclonal to Nestin—neural stem cell marker were added.
Results
Following the isolation, a close-to-single-cell suspension was obtained, which consisted of viable small, round cells. In suspension cultures containing EGF and bFGF, rat striatal NSCs began to proliferate and adherent clonal clusters of cells (10 - 20 cells/clone) started to appear after 48 - 72 h. The clusters divided rapidly and generated large colonies, i. e. neurospheres. These floating neurospheres displayed a spherical shape, and the cells forming the neurosphere were phase bright. These spheres were successfully passaged by dissociation and reculture. This procedure resulted in a rapid increase in cell number. The neural cells dissociated from E13 rat striatum had the capacity to proliferate and form neurospheres. The neurosphere could express Nestin, gernerate MAP2 or GFAP positive neural cells.
Eight children were selected from 20 patients with TS and their sera were used for rat striatal microinfusion. Similarly, eight specimens of sera were selected from 20 control samples. The selection of sera for infusion was based on ELISA optical density readings against caudate homogenate, that is, we selected the highest serum OD readings for TS subjects and the lowest OD readings for the control infusion subgroup. The mean±SD optical density readings in serum of 8 TS subjects selected for microinfusion was 0.904±0.177 and 0.252±0.193 for the 8 selected controls.
Serologic studies of children with TS have detected the existence of anti-neural antibodies, which can induce striatal dysfunction. In our study, stereotypies were successfully induced in rats by intrastriatal microinfusion of TS sera under noninflammatory conditions. After infusion of TS sera, stereotypic behaviors in rats increased significantly. Marked differences were observed in stereotypies of TS rats compared to control rats after microinfusion. TS rats exhibited significant increases in bites, taffy-pulling, gnawing, licking, head shaking, paw shaking, rearing and episodic utterance. All rats in four groups were observed at 1 day, 7 days, 14 days, and 21 days after their transplantations. Stereotypic behaviors of each rat were recorded and counted through each 30 minutes of observation period. The statistical analysis suggested that TS rats had increased stereotypic behaviors compared to normal rats (Sham). Using a repeated measurements analysis of variance (ANOVA), we found that the overall model had significant group (F=354.13, p<0.0001) and day (F= 106.01, p<0.0001) effects, as well as a (group×day) interaction (F=21.38, p<0.0001), indicating varying degrees of differences among the groups and across days. A model with just TS+NSCs and TS+PBS groups indicated significant differences between groups(F=36.63, p=0.0001) and a strong day effect(F=100.23, p<0.0001). Importantly, animals with NSCs grafts showed a significant decrease in stereotypic behaviors at 14 and 21 days. Tukey-Kramer post hoc test showed significant differences between TS+NSCs and TS+PBS groups beginning at 2 weeks post-transplantation. However, rats receiving NSC grafts (TS+NSCs) still had higher stereotypic behavior counts than sham controls (p<0.0001), which indicated that the animals' impairment did not fully recover in a short period of 3 weeks.
Histological analyses of recipient rat brains transplanted with NSCs or injected with PBS (vehicle) were given 3 weeks after transplantation. In order to trace the transplanted cells, we used BrdU labeling, so that the presence of the implanted NSCs within striatum of TS rats identifiable for the nuclei of the grafted cells were BrdU-positive and stained with purple. In the TS+NSCs rats, BrdU-positive cells were prevalent in the striatum sections near the injection site, suggesting the survival of the cells. Some of the grafted cells expressed both BrdU and MAP-2 and others expressed both BrdU and GFAP. These results suggested that NSCs used as grafts grew at the injection site and differentiated into neurons or astrocytes. Our results also indicated that a considerable portion of the transplanted cells had differentiated to mature neurons in vivo, which may lead to histological and functional reconstitution in the CNS. Besides, few BrdU and Nestin double positive cells were found, indicating undiferentiation of a small number of the grafted cells.
Conclusions
Neural stem cells were isolated from fetal rat striatum and were certificated as a heterogeneous population of mitotically active, self-renewing, multipotent, immature progenitor cells. These cells expressed the intermediate filament protein Nestin and grew as neurospheres in vitro. Neural stem cells had the capacity to differentiate into neurons and gilocytes..
In our study, stereotypies were successfully induced in rats by intrastriatal microinfusion of TS sera under noninflammatory conditions. Our results showed that intrastriatum transplanted rat NSCs relieved stereotypic behaviors. These results suggested the potential of using NSCs intrastriatum as a clinical treatment for TS. Neural stem cells survived in the brain of TS rat and part of them differentiated into neurons and gliocytes. These newly generated neurons from the transplanted NSCs might have played a role for the neural networks in the damaged areas. No signs of immunologic rejection or non-neural tissue growth were found in the host rat brain after transplantation.
引文
1 Hogg RC, Chipperfield H, Whyte KA, Stafford MR, Hansen MA, et al. Functional maturation of isolated neural progenitor cells from the adult rat hippocampus. Eur JNeurosci 2004,19(9): 2410-20.
2 Reynolds BA, Tetzlaff W, Weiss S. A multipotent EGF-responsive striatal embryonic progenitor cell produces neurons and astrocytes. J Neurosci 1992, 12(11): 4565-74.
3 Reynolds BA, Weiss S. Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science 1992, 255(5052): 1707-10.
4 Richards LJ, Kilpatrick TJ, Bartlett PF. De novo generation of neuronal cells from the adult mouse brain. Proc Natl Acad Sci U S A1992, 89(18): 8591-5.
5 Shimazaki T. Biology and clinical application of neural stem cells. Horm Res 2003, Suppl 60(3): 1-9.
6 Sugaya K. Potential use of stem cells in neuroreplacement therapies for neurodegenerative diseases. Int Rev Cytol 2003,228: 1-30.
7 Lendahl U, Zimmeman LB, McKay RD. CNS stem cells express a new class of intermediate filament protein. Cell 1990, 60(4): 585
8 Dahlstrand J, Lardelli M, Lendahl U. Nestin mRNA expression correlates with the central nervous system progenitor cell state in many, but not all, regions of developing central nervous system. Brain Res Dev Brain Res 1995, 84(1): 109-29.
9 Johansson CB, Lothian C, Molin M, Okano H, Lendahl U. Nestin enhancer requirements for expression in normal and injured adult CNS. J Neurosci Res 2002, 69(6): 784-94.
10 Lothian C, Lendahl U. An evolutionary conserved region in the second intron of the human nestin gene directs gene expression to CNS progenitor cells and to early neural crest cells. Eur J Neurosci 1997,9(3): 452-62.
11 Akiyama Y, Honmou O, Kato T, Uede T, Hashi K, et al. Transplantation of clonal neural precursor cells derived from adult human brain establishes functional peripheral myelin in the rat spinal cord. Exp Neurol 2001, 167(1): 27-39.
12 Zigova T, Pencea V, Betarbet R, Wiegand SJ, Alexander C, et al. Neuronal progenitor cells of the neonatal subventricular zone differentiate and disperse following transplantation into the adult rat striatum. Cell Transplant 1998, 7(2): 137-56.
13 Lepore AC, Han SS, Tyler-Polsz CJ, Cai J, Rao MS, et al. Differential fate of multipotent and lineage-restricted neural precursors following transplantation into the adult CNS. Neuron Glia Biol 2004, 1(2): 113-126.
14 Martinez-Serrano A, Lundberg C, Horellou P, Fischer W, Bentlage C, et al. CNS-derived neural progenitor cells for gene transfer of nerve growth factor to the adult rat brain: complete rescue of axotomized cholinergic neurons after transplantation into the septum. J Neurosci 1995, 15(8): 5668-80.
15 Dwain I, Xiangpeng Y, Zeng Z, Patricia T, Joh SY. Neural stem cells—a promising potential therapy for brain tumors. Curr Stem Cell Res Ther 2006, 1(1): 79-84.
16 Cai J, Limke TL, Ginis I, Rao MS. Identifying and tracking neural stem cells. Blood Cells Mol Dis 2003, 31(1): 18-27.
17 Okano H, Yoshizaki T, Shimazaki T, Sawamoto K. Isolation and transplantation of dopaminergic neurons and neural stem cells. Parkinsonism Relat Disord 2002, 9(1): 23-8.
18 Ding S, Messam CA, Li P, Selzer ME, Dichter MA, et al. Murine brain progenitor cells have the ability to differentiate into functional neurons and integrate into the CNS. Cell Transplant 2006, 15(8-9): 699-710.
19 Taupin P. Therapeutic potential of adult neural stem cells. Recent Patents CNS Drug Discov 2006, 1(3): 299-303.
20 Li J, Imitola J, Snyder EY, Sidman RL. Neural stem cells rescue nervous purkinje neurons by restoring molecular homeostasis of tissue plasminogen activator and downstream targets. J Neurosci 2006,26(30): 7839-48.
21 Villa A, Rubio FJ, Navarro B, Bueno C, Martinez-Serrano A. Human neural stem cells in vitro. A focus on their isolation and perpetuation. Biomed Pharmacother 2001, 55(2): 91-5.
22 Tarasenko YI, Yu Y, Jordan PM, Bottenstein J, Wu P. Effect of growth factors on proliferation and phenotypic differentiation of human fetal neural stem cells. J Neurosci Res 2004, 78(5): 625-36.
23 Wang K, Wang JJ, Wang Y, He OH, Wang X, et al. Infusion of epidermal growth factor and basic fibroblast growth factor into the striatum of parkinsonian rats leads to in vitro proliferation and differentiation of adult neural progenitor cells. Neurosci Lett 2004, 364(3): 154-8.
24 Miltenyi S, Muller W, Weichel W, Radbruch A. High gradient magnetic cell separation with MACS. Cytometry 1990,11(2): 231-8.
25 Constantinescu SN. Stem cell generation and choice of fate: role of cytokines and cellular microenvironment. J Cell Mol Med 2000,4(4): 233-248.
26 Klassen HJ, Imfeld KL, Kirov II, Tai L, Gage FH, et al. Expression of cytokines by multipotent neural progenitor cells. Cytokine 2003, 22(3-4): 101-6
27 Abematsu M, Smith I, Nakashima K. Mechanisms of neural stem cell fate determination: extracellular cues and intracellular programs. Curr Stem Cell Res Ther 2006,1(2): 267-77.
28 Riaz SS, Theofilopoulos S, Jauniaux E, Stem GM, Bradford HF. The differentiation potential of human foetal neuronal progenitor cells in vitro. Brain Res Dev Brain Res 2004, 153(1): 39-51.
29 Erlandsson A, Larsson J, Forsberg-Nilsson K. Stem cell factor is a chemoattractant and a survival factor for CNS stem cells. Exp Cell Res 2004, 301(2): 201-10.
30 Benoit BO, Savarese T, Joly M, Engstrom CM, Pang L, et al. Neurotrophin channeling of neural progenitor cell differentiation. J Neurobiol 2001, 46(4): 265-80.
31 Hammang JP, Archer DR, Duncan ID. Myelination following transplantation of EGF-responsive neural stem cells into a myelin-deficient environment. Exp Neurol. 1997, 147(1): 84-95.
32 Lillien L, Raphael H. BMP and FGF regulate the development of EGF-responsive neural progenitor cells. Development 2000, 127(22): 4993-5005.
33 Kuhn HG, Winkler J, Kempermann G, Thal LJ, Gage FH. Epidermal growth factor and fibroblast growth factor-2 have different effects on neural progenitors in the adult rat brain. J Neurosci 1997, 17(15): 5820-9.
34 Santa-Olalla J, Covarrubias L. Basic fibroblast growth factor promotes epidermal growth factor responsiveness and survival of mesencephalic neural precursor cells. J Neurobiol 1999,40(1): 14-27.
35 Itoh T, Satou T, Dote K, Hashimoto S, Ito H. Effect of basic fibroblast growth factor on cultured rat neural stem cell in three-dimensional collagen gel. Neurol Res 2005, 27(4): 429-32.
36 Gobbel GT, Choi SJ, Beier S, Niranjan A. Long-term cultivation of multipotential neural stem cells from adult rat subependyma. Brain Res 2003, 980(2): 221-32.
37 Hinsch K, Zupanc GK. Isolation, cultivation, and differentiation of neural stem cells from adult fish brain. J Neurosci Methods 2006, 158(1): 75-88.
38 Zhang X, Li X, Wu J, Wang Z, Xu H, et al. Isolation, cultivation and identification of stem cells from cerebral cortex of mouse embryo. Zhonghua Yi Xue Za Zhi 2002, 82(12): 832-5.
39 Fu SL, Ma ZW, Yin L, Lu PH, Xu XM. Isolation and cultivation of neural stem cells from the embryonic rat brain and spinal cord. Sheng Li Xue Bao 2003, 55(3): 278-83.
40 Einstein O, Ben-Menachem-Tzidon O, Mizrachi-Kol R. Survival of neural precursor cells in growth factor-poor environment: implications for transplantation in chronic disease. Glia 2006, 53(4): 449-55.
41 Piper DR, Mujtaba T, Keyoung H, Roy NS, Goldman SA, et al. Identification and characterization of neuronal precursors and their progeny from human fetal tissue. J Neurosci Res 2001, 66(3): 356-68.
42 Bazan E, Alonso FJ, Redondo C, Lopez-Toledano MA, Alfaro JM, et al. In vitro and in vivo characterization of neural stem cells. Histol Histopathol 2004, 19(4): 1261-75.
43 Okano H, Kawahara H, Toriya M, Nakao K, Shibata S, et al. Function of RNA-binding protein Musashi-1 in stem cells.Exp Cell Res 2005,306(2):349-56.
1 Rampello L, Alvano A, Battaglia G, Bruno V, Raffaele R, et al. Tic disorders: from pathophysiology to treatment. J Neurol 2006, 253(1): 1-15.
2 Olson LL, Singer HS, Goodman WK, Maria BL. Tourette syndrome: diagnosis, strategies, therapies, pathogenesis, and future research directions. J Child Neurol 2006, 21(8): 630-41.
3 Stem JS, Burza S, Robertson MM. Gilles de la Tourette's syndrome and its impact in the UK. Postgrad Med J 2005, 81(951): 12-9.
4 Leckman JF, Zhang H, Vitale A, Lahnin F, Lynch K, et al. Course of tic severity in Tourette syndrome: the first two decades. Pediatrics 1998, 102(1): 14-9.
5 Riddle MA, Carlson J. Clinical psychopharmacology for Tourette syndrome and associated disorders. Adv Neurol 2001, 85: 343-54.
6 Deckersbach T, Rauch S, Buhlmann U, Wilhelm S. Habit reversal versus supportive psychotherapy in Tourette's disorder: a randomized controlled trial and predictors of treatment response. Behav Res Ther 2006,44(8): 1079-90.
7 Cheung MY, Shahed J, Jankovic J. Malignant Tourette syndrome. Mov Disord 2007, 22(12): 1743-50.
8 Maciunas RJ, Maddux BN, Riley DE, Whitney CM, Schoenberg MR, et al. Prospective randomized double-blind trial of bilateral thalamic deep brain stimulation in adults with Tourette syndrome. J Neurosurg 2007,107(5): 1004-14.
9 Hamani C, Moro E. Surgery for other movement disorders: dystonia, tics. Curr Opin Neurol 2007, 20(4): 470-6.
10 Neimat JS, Patil PG, Lozano AM. Novel surgical therapies for Tourette syndrome. J Child Neurol 2006,21(8): 715-8.
11 Anandan S, Wigg CL, Thomas CR, Coffey B. Psychosurgery for self-injurious behavior in Tourette's disorder. J Child Adolesc Psychopharmacol 2004, 14(4): 531-8.
12 Mink JW, Walkup J, Frey KA, Como P, Cath D, et al. Patient selection and assessment recommendations for deep brain stimulation in Tourette syndrome. Mov Disord 2006, 21(11): 1831-8.
13 Wang L, Martin DR, Baker HJ, Zinn KR, Kappes JC, et al. Neural progenitor cell transplantation and imaging in a large animal model. Neurosci Res 2007, 59(3): 327-40.
14 Jain M, Armstrong RJ, Elneil S, Barker RA. Transplanted human neural precursor cells migrate widely but show no lesion-specific tropism in the 6-hydroxydopamine rat model of Parkinson's disease. Cell Transplant 2006, 15(7): 579-93.
15 Cummings BJ, Uchida N, Tamaki SJ, Anderson AJ. Human neural stem cell differentiation following transplantation into spinal cord injured mice: association with recovery of locomotor function. Neurol Res 2006, 28(5): 474-81.
16 Lee ST, Park JE, Lee K, Kang L, Chu K, et al. Noninvasive method of immortalized neural stem-like cell transplantation in an experimental model of Huntington's disease. J Neurosci Methods 2006, 152(1-2): 250-4.
17 Lee ST, Chu K, Park JE, Lee K, Kang L. Intravenous administration of human neural stem cells induces functional recovery in Huntington's disease rat model. Neurosci Res 2005, 52(3): 243-9.
18 Peterson BS, Choi HA, Hao X, Amat JA, Zhu H, et al. Morphologic features of the amygdala and hippocampus in children and adults with Tourette syndrome. Arch Gen Psychiatry 2007, 64(11): 1281-91.
19 Muller N. Tourette's syndrome: clinical features, pathophysiology, and therapeutic approaches. Dialogues Clin Neurosci 2007, 9(2): 161-71.
20 Brovedani P, Masi G. Tourette's syndrome in children: neurological, neuropsychological and psychiatric issues. Panminerva Med 2000,42(2): 141-9.
21 Swerdlow NR, Sutherland AN. Preclinical models relevant to Tourette syndrome. Adv Neurol 2006, 99: 69-88.
22 Swerdlow NR, Sutherland AN. Using animal models to develop therapeutics for Tourette Syndrome. Pharmacol Ther 2005, 108(3): 281-93.
23 Sanberg PR, Emerich DF. Neural basis of behavior: animal models of human conditions. Brain Res Bull 1990, 25(3): 447-51.
24 Singer HS, Giuliano JD, Hansen BH, Hallett JJ, Laurino JP, et al. Antibodies against human putamen in children with Tourette syndrome. Neurology 1998, 50(6): 1618-24.
25 Hallett JJ, Harling-Berg CJ, Knopf PM, Stopa EG, Kiessling LS. Anti-striatal antibodies in Tourette syndrome cause neuronal dysfunction. J Neuroimmunol 2000,111(1-2): 195-202.
26 Taylor JR, Morshed SA, Parveen S, Mercadante MT, Scahill 1, et al. An animal model of Tourette's syndrome. Am J Psychiatry 2002, 159(4): 657-60.
27 Taupin P. Autologous transplantation in the central nervous system. Indian J Med Res 2006,124(6): 613-8.
28 Muller N, Riedel M, Straube A, Gunther W, Wilske B. Increased anti-streptococcal antibodies in patients with Tourette's syndrome. Psychiatry Res 2000, 94(1): 43-9.
29 Dale RC. Autoimmunity and the basal ganglia: new insights into old diseases. QJM 2003, 96(3): 183-91.
30 Hoekstra PJ, Anderson GM, Limburg PC, Korf J, Kallenberg CG, et al. Neurobiology and neuroimmunology of Tourette's syndrome: an update. Cell Mol Life Sci 2004, 61(7-8): 886-98.
31 Khalifa N, vonKnorring AL. Tourette syndrome and other tic disorders in a total population of children: clinical assessment and background. Acta Paediatr 2005, 94(11): 1608-14.
32 Burd L, Freeman RD, Klug MG, Kerbeshian J. Tourette Syndrome and learning disabilities. BMC Pediatr 2005, 5: 34.
33 Chang HL, Tu MJ, Wang HS. Tourette's syndrome: psychopathology in adolescents. Psychiatry Clin Neurosci 2004, 58(4): 353-8.
34 Channon S, Sinclair E, Waller D, Healey L, Robertson MM. Social cognition in Tourette's syndrome: intact theory of mind and impaired inhibitory functioning. J Autism Dev Disord 2004, 34(6): 669-77.
35 Goudriaan AE, Oosterlaan J, de Berrs E, van den Brink W. Neurocognitive functions in pathological gambling: a comparison with alcohol dependence, Tourette syndrome and normal controls. Addiction 2006, 101(4): 534-47.
36 Kano Y, Ohta M, Nagai Y, Pauls DL, Leckman JF. Obsessive-compulsive symptoms in parents of Tourette syndrome probands and autism spectrum disorder probands. Psychiatry Clin Neurosci 2004, 58(4): 348-52.
37 Stephens RJ, Sandor P. Aggressive behaviour in children with Tourette syndrome and comorbid attention-deficit hyperactivity disorder and obsessive-compulsive disorder. Can J Psychiatry 1999, 44(10): 1036-42.
38 Rizzo R, Gulisano M, Pavone P, Fogliani F, Robertson MM. Increased antistreptococcal antibody titers and anti-basal ganglia antibodies in patients with Tourette syndrome: controlled cross-sectional study. J Child Neurol 2006, 21(9): 747-53.
39 Loiselle CR, Wendlandt JT, Rohde CA, Singer HS. Antistreptococcal, neuronal, and nuclear antibodies in Tourette syndrome. Pediatr Neurol 2003, 28(2): 119-25.
40 Visvanathan K, Trifiletti RR, Altemus M, Zabriskie JB. Autoimmune mechanisms in movement disorders. Semin Pediatr Neurol 2000, 7(2): 103-7.
41 Muller N, Riedel M, Forderreuther S, Blendinger C, Abele-Hom M. Tourette's syndrome and mycoplasma pneumoniae infection. Am J Psychiatry 2000, 157(3): 481-2.
42 Perlmutter SJ, Leitman SF, Garvey MA, Hamburger S, Feldman E, et al. Therapeutic plasma exchange and intravenous immunoglobulin for obsessive-compulsive disorder and tic disorders in childhood. Lancet 1999, 354(9185): 1153-8.
43 Gilbert D. Treatment of children and adolescents with tics and Tourette syndrome. J Child Neurol 2006, 21(8): 690-700.
44 Kurlan R. Future and alternative therapies in Tourette syndrome. Adv Neurol 2006, 99: 248-53.
45 Bithell A, Williams BP. Neural stem cells and cell replacement therapy: making the right cells. Clin Sci (Lond) 2005,108(1): 13-22.
46 Cataudella T, Conti L, Cattaneo E. Neural stem and progenitor cells: choosing the right Shc. Prog Brain Res 2004, 146: 127-33.
47 Harris K, Singer HS. Tic disorders: neural circuits, neurochemistry, and neuroimmunology. J Child Neurol 2006,21(8): 678-89.
48 Albin RL. Neurobiology of basal ganglia and Tourette syndrome: striatal and dopamine function. Adv Neurol 2006, 99:99-106.
1 Galvin KA, Jones DG. Adult human neural stem cells for autologous cell replacement therapies for neurodegenerative disorders. NeuroRehabilitation 2006, 21(3): 255-65.
2 Kim HT, Kim IS, Lim SE, Lee IS, Park KI. Gene and cell replacement via neural stem cells. Yonsei Med J 2004, 45 Suppl: 32-40.
3 Ourednik J, Ourednik V, Lynch WP, Schachner M, Snyder EY. Neural stem cells display an inherent mechanism for rescuing dysfunctional neurons. Nat Biotechnol 2002,20(11): 1103-10. Epub 2002 Oct 15.
4 Lynch WP, Portis JL. Neural stem cells as tools for understanding retroviral neuropathogenesis. Virology 2000, 271(2): 227-33.
5 Yang H, Lu P, McKay HM, Bernot T, Keirstead H, et al. Endogenous neurogenesis replaces oligodendrocytes and astrocytes after primate spinal cord injury. J Neurosci 2006, 26(8): 2157-66
6 Lie DC, Song H, Colamarino SA, Ming GL, Gage FH. Neurogenesis in the adult brain: new strategies for central nervous system diseases. Annu Rev Pharmacol Toxicol 2004,44: 399-421.
7 Yasuhara T, matsukawa N, Hara K, Yu G, Xu L, et al. Transplantation of human neural stem cells exerts neuroprotection in a rat model of Parkinson's disease. J Neurosci 2006, 26(48): 12497-511.
8 Lee ST, Chu K, Park JE, Lee K, Kang L, et al. Intravenous administration of human neural stem cells induces functional recovery in Huntington's disease rat model. Neurosci Res 2005, 52(3): 243-9.
9 Karbanova J, Mokry J, kotingova L. Neural stem cells transplanted into intact brains as neurospheres form solid grafts composed of neurons, astrocytes and oligodendrocyte precursors. Biomed Pap Med Fac Univ Palacky Olomouc Czech Repub 2004,148(2): 217-20.
10 Lenka N. Derivation and characterization of neural cells from embryonic stem cells using nestin enhancer. Methods Mol Biol 2006, 330: 33-54.
11 Takagi Y, Nishimura M, Morizane A, Takahashi J, Nozaki K, et al. Survival and differentiation of neural progenitor cells derived from embryonic stem cells and transplanted into ischemic brain. J Neurosurg 2005,103(2): 304-10.
12 Aleksandrova MA, Podgomyi OV, Marei MV, Poltavtseva RA, Tsitrin EB, et al. Characteristics of human neural stem cells in vitro and after transplantation into rat brain. Bull Exp Biol Med 2005, 139(1): 114-20.
13 Zhu W, Mao Y, Zhao Y, Zhou LF, Wang Y, et al. Transplantation of vascular endothelial growth factor-transfected neural stem cells into the rat brain provides neuroprotection after transient focal cerebral ischemia. Neurosurgery 2005, 57(2): 325-33
14 Li T, Zheng J, Xie Y, Wang S, Zhang X, et al. Transplantable neural progenitor populations derived from rhesus monkey embryonic stem cells. Stem Cells 2005, 23(9): 1295-303.
15 Enzmann V, Howard RM, Yamauchi Y, Whittemore SR, Kaplan HJ. Enhanced induction of RPE lineage markers in pluripotent neural stem cells engrafted into the adult rat subretinal space. Invest Ophthalmol Vis Sci 2003, 44(12): 5417-22.
16 Yoshinaga T, Hashimoto E, Ukai W, Toki S, Saito S, et al. Neural stem cell transplantation in a model of fetal alcohol effects. J Neural Transm Suppl 2007, (72): 331-7.
17 Darsalia V, Kallur T, Kokaia Z. Survival, migration and neuronal differentiation of human fetal striatal and cortical neural stem cells grafted in stroke-damaged rat striatum. Eur J Neurosci 2007, 26(3): 605-14.
18 Yasuhara T, Matsukawa N, Hara K, Yu G, Xu L, et al. Transplantation of human neural stem cells exerts neuroprotection in a rat model of Parkinson's disease. J Neurosci 2006, 26(48): 12497-511
19 Heine W, Conant K, Griffin JW, Hoke A. Transplanted neural stem cells promote axonal regeneration through chronically denervated peripheral nerves. Exp Neurol 2004, 189(2): 231-40.
20 Riess P, Zhang C, Saatman KE, Laurer HL, Longhi LG, et al. Transplanted neural stem cells survive, differentiate, and improve neurological motor function after experimental traumatic brain injury. Neurosurgery 2002, 51(4): 1043-52.
21 Jeong SW, Chu K, Jung KH, Kim SU, Kim M, et al. Human neural stem cell transplantation promotes functional recovery in rats with experimental intracerebral hemorrhage. Stroke 2003, 34(9): 2258-63.
22 Enomoto M, Shinomiya K, Okabe S. Migration and differentiation of neural progenitor cells from two different regions of embryonic central nervous system after transplantation into the intact spinal cord. Eur J Neurosci 2003, 17(6): 1223-32.
23 Lee HJ, Kim KS, Kim EJ, Choi HB, Lee KH, et al. Brain transplantation of immortalized human neural stem cells promotes functional recovery in mouse intracerebral hemorrhage stroke model. Stem Cells 2007, 25(5): 1204-12.
24 Mueller D, Shamblott MJ, Fox HE, Gearhart JD, Martin LJ. Transplanted human embryonic germ cell-derived neural stem cells replace neurons and oligodendrocytes in the forebrain of neonatal mice with excitotoxic brain damage. J Neurosci Res 2005, 82(5): 592-608.
25 Niles LP, Armstrong KJ, Rincon Castro LM, Dao CV, Sharma R, et al. Neural stem cells express melatonin receptors and neurotrophic factors: colocalization of the MT1 receptor with neuronal and glial markers. BMC Neurosci 2004, 5(1): 41.
26 Lu P, Jones LL, Snyder EY, Tuszynski MH. Neural stem cells constitutively secrete neurotrophic factors and promote extensive host axonal growth after spinal cord injury. ExpNeurol 2003, 181(2): 115-29.
27 Ourednik J, Ourednik V, Lynch WP, Schachner M, Snyder EY. Neural stem cells display an inherent mechanism for rescuing dysfunctional neurons. Nat Biotechnol 2002, 20(11): 1103-10.
28 Yan J, Xu L, Welsh AM, Hatfield G, Hazel T, et al. Extensive neuronal differentiation of human neural stem cell grafts in adult rat spinal cord. PLoS Med 2007, 4(2): 39.
29 AnYH, Wang HY, Gao ZX, Wang ZC. Differentiation of rat neural stem cells and its relationship with environment. Biomed Environ Sci 2004,17(1): 1-7.
30 Jori FP, Galderisi U, Piegari E, Cipollaro M, Cascino A, et al. EGF-responsive rat neural stem cells: molecular follow-up of neuron and astrocyte differentiation in vitro. J Cell Physiol 2003, 195(2): 220-33.
31 Nakahara Y, Gage FH, Tuszynski MH. Grafts of fibroblasts genetically modified to secrete NGF, BDNF, NT-3, or basic FGF elicit differential responses in the adult spinal cord. Cell Transplant 1996, 5(2): 191-204.
32 Winkler C, Fricker RA, Gates MA, Olsson M, Hammang JP, et al. Incorporation and glial differentiation of mouse EGF-responsive neural progenitor cells after transplantation into the embryonic rat brain. Mol Cell Neurosci 1998, 11(3): 99-116.
33 Nikkhah G, Olsson M, Eberhard J. A micro transplantation approach for cell suspension grafting in the rat Parkinson model: A detailed account of methodology. Neuroscience 1994, 63(1): 57.
34 Muraoka K, Shingo T, Yasuhara T, Kameda M, Yuan W, et al. The high integration and differentiation potential of autologous neural stem cell transplantation compared with allogeneic transplantation in adult rat hippocampus. Exp Neurol 2006,199(2): 311-27.
35 Ehtesham M, Kabos P, Gutierrez MA, Chung NH, Griffith TS, et al. Induction of glioblastoma apoptosis using neural stem cell-mediated delivery of tumor necrosis factor-related apoptosis-inducing ligand. Cancer Res 2002, 62(24): 7170-4.
36 Mammolenti M, Gajavelli S, Tsoulfas P, Levy R. Absence of major histocompatibility complex class I on neural stem cells does not permit natural killer cell killing and prevents recognition by alloreactive cytotoxic T lymphocytes in vitro. Stem Cells 2004,22(6): 1101-10.
37 Lee EM, Kim JY, Cho BR, Chung WK, Yoon BW, et al. Down-regulation of MHC class I expression in human neuronal stem cells using viral stealth mechanism. Biochem Biophys Res Commun 2005, 326(4): 825-35.
38 Wei P, Liu J, Zhou HL, Han ZT, Wu QY, et al. Effects of engrafted neural stem cells derived from GFP transgenic mice in Parkinson's diseases rats. Neurosci Lett 2007, 419(1): 49-54.
39 Kon ND, Mills SA, Andrews CA, Miller LR, Cordell AR. Effect of desmopressin on vein graft patency in a microvascular model. Microsurgery 1991, 12(4): 288-91.
[1] Albin RL, Mink JW. Recent advances in Tourette syndrome research. Trends Neurosci 2006; 29: 175-82.
[2] Berthier ML, Campos VM, Kulisevsky J, Valero JA. Heroin and malignant coprolalia in Tourette's syndrome. J Neuropsychiatry Clin Neurosci 2003; 15: 116-7.
[3] Bjugstad KB, Redmond ED Jr, Teng YD, Elsworth JD, Roth RH, Blanchard BC, et al. Neural stem cells implanted into MPTP-treated monkeys increase the size of endogenous tyrosine hydroxylase-positive cells found in the striatum: a return to control measures. Cell Transplant 2005; 14: 183-92.
[4] Baizabal JM, Furlan-Magaril M, Santa-Olalla J, Covarrubias L. Neural stem cells in development and regenerative medicine. Arch Med Res 2003; 34: 572-88.
[5] Conti L, Retano E, Cattaneo E. Neural stem cell systems: diversities and properties after transplantation in animal models of diseases. Brain Pathol 2006; 16: 143-54.
[6] Diaz-Anzaldua A, Joober R, Riviere JB, Dion Y, Lesperance P, Richer F, et al. Tourette syndrome and dopaminergic genes: a family-based association study in the French Canadian founder population. Mol Psychiatry 2004; 9: 272-7.
[7] Fattapposta F, Restuccia R, Colonnese C, Labruna L, Garreffa G, Bianco F. Gilles de la Tourette syndrome and voluntary movement: a functional MRI study. Psychiatry Res 2005; 138: 269-72
[8] Gaughwin PM, Caldwell MA, Anderson JM, Schwiening CJ, Fawcett JW, Compston DA, et al. Astrocytes promote neurogenesis from oligodendrocyte precursor cells. Eur J Neurosci 2006; 23: 945-56.
[9] Hallett JJ, Harling-Berg CJ, Knopf PM, Stopa EG, Kiessling LS. Anti-striatal antibodies in Tourette syndrome cause neuronal dysfunction. J Neuroimmunol 2000; 111: 195-202.
[10] Hong JJ, Loiselle CR, Yoon DY, Lee O, Becker KG, Singer HS. Microarray analysis in Tourette syndrome postmortem putamen. J Neurol Sci 2004; 225: 57-64.
[11] Hu Z, Wei D, Johansson CB, Holmstrom N, Duan M, Frisen J, et al. Survival and neural differentiation of adult neural stem cells transplanted into the mature inner ear. Exp Cell Res 2005; 302: 40-7.
[12] Korochkin LI, Revishchi AV, Okhotin VE. Neural stem cells and their role in recovery processes in the nervous system. Neurosci Behav Physiol 2006; 36: 499-512.
[13] Leckman JF, Zhang H, Vitale A, Lahnin F, Lynch K, Bondi C, et al. Course of tic severity in Tourette syndrome: the first two decades. Pediatrics 1998; 102: 14-9.
[14] Loiselle CR, Lee O, Morgan TH, Singer HS. Striatal microinfusion of Tourette syndrome and PANDAS sera: failure to induce behavioral changes. Mov Disord 2004; 19: 390-6.
[15] Lee ST, Park JE, Lee K, Kang L, Chu K, Kim SU, et al. Noninvasive method of immortalized neural stem-like cell transplantation in an experimental model ofHuntington's disease. J Neurosci Methods 2006; 152: 250-4.
[16] Liang P, Jin LH, Liang T, Liu EZ, Zhao SG. Human neural stem cells promote corticospinal axons regeneration and synapse reformation in injured spinal cord of rats. Chin Med J (Engl) 2006; 119: 1331-8.
[17] Li J, Imitola J, Snyder EY, Sidman RL. Neural stem cells rescue nervous purkinje neurons by restoring molecular homeostasis of tissue plasminogen activator and downstream targets. J Neurosci 2006; 26: 7839-48.
[18] Mink JW, Walkup J, Frey KA, Como P, Cath D, Delong MR, et al. Patient selection and assessment recommendations for deep brain stimulation in Tourette syndrome. Mov Disord 2006; 21: 1831-8.
[19] Perlmutter SJ, Leitman SF, Garvey MA, Hamburger S, Feldman F, Leonard HL, et al. Therapeutic plasma exchange and intravenous immunoglobulin for obsessive-compulsive disorder and tic disorders in childhood. Lancet 1999; 354: 1153-8.
[20] Pallini R, Vitiani LR, Bez A, Casalbore P, Facchiano F, Di Giorgi Gerevini V, et al. Homologous transplantation of neural stem cells to the injured spinal cord of mice. Neurosurgery 2005; 57: 1014-25.
[21] Parati EA, Bez A, Ponti D, Sala S, Pozzi S, Pagano SF. Neural stem cells. Biological features and therapeutic potential in Parkinson's disease. J Neurosurg Sci 2003; 47: 8-17.
[22] Rampello L, Alvano A, Battaglia G, Bruno V, Raffaele R, Nicoletti F. Tic disorders: from pathophysiology to treatment. J Neurol 2006; 253: 1-15.
[23] Rizzo R, Gulisano M, Pavone P, fogliani F, Robertson MM. Increased antistreptococcal antibody titers and anti-basal ganglia antibodies in patients with Tourette syndrome: controlled cross-sectional study. J Child Neurol 2006; 21: 747-53.
[24] Singer HS, Loiselle CR, Lee O, Minzer K, Swedo S, Grus FH. Anti-basal ganglia antibodies in PANDAS. Mov Disord 2004; 19: 406-15.
[25] Singer HS, Mink JW, Loiselle CR, Burke KA, Ruchkina I, Morshed S, et al. Microinfusion of antineuronal antibodies into rodent striatum: failure to differentiate between elevated and low titers. J Neuroimmunol 2005; 163: 8-14.
[26] Swerdlow NR, Sutherland AN. Using animal models to develop therapeutics for Tourette Syndrome. Pharmacol Ther 2005; 108: 281-93.
[27] Taylor JR, Morshed SA, Parveen S, Mercadante MT, Scahill 1, Peterson BS, et al. An animal model of Tourette's syndrome. Am J Psychiatry 2002; 159: 657-60.
[28] Yeh CB, Wu CH, Tsung HC, Chen CW, Shyu JF, Leckman JF. Antineural antibody in patients with Tourette's syndrome and their family members. J Biomed Sci 2006; 13: 101-12.
1. Rampello L, Alvano A, Battaglia G, Bruno V, Raffaele R, Nicoletti F (2006): Tic disorders: from pathophysiology to treatment. J Neurol 253:1-15.
2. Leckman JF, Zhang H, Vitale A, Lahnin F, Lynch K, Bondi C, et al. (1998): Course of tic severity in Tourette syndrome: the first two decades. Pediatrics 102:14-19.
3. Berthier M, Campos VM, Kulisevsky J, Valero JA (2003): Heroin and malignant coprolalia in Tourette's syndrome. J Neuropsychiatry Clin Neurosci 15:116-117.
4. Mink JW, Walkup J, Frey KA, Como P, Cam D, Delong MR, et al. (2006): Patient selection and assessment recommendations for deep brain stimulation in Tourette syndrome. Mov Disord 21:1831-1838.
5. Kamishina H, Deng J, Oji T, Cheeseman JA, Clemmons RM (2006): Expression of neural markers on bone marrow-derived canine mesenchymal stem cells. Am J Vet Res 67:1921-1928.
6. Kim S, Honmou O, Kato K, Nonaka T, Houkin K, Hamada H, et al. (2006): Neural differentiation potential of peripheral blood- and bone-marrow-derived precursor cells. Brain Res 1123:27-33.
7. Pisati F, Bossolasco P, Meregalli M, Cova L, Belicchi M, Gavina M, et al. (2007): Induction of neurotrophin expression via human adult mesenchymal stem cells: implication for cell therapy in neurodegenerative diseases. Cell Transplan 16:41-55.
8. Mori K, Iwata J, Miyazaki M, Nakao Y, Maeda M (2005): Functional recovery of neuronal activity in rat whisker-barrel cortex sensory pathway from freezing injury after transplantation of adult bone marrow stromal cells. J Cereb Blood Flow Metab 25:887-898.
9. Bae JS, Han HS, Youn DH, Carter JE, Modo M, Schuchman EH, et al (2007): Bone marrow-derived mesenchymal stem cells promote neuronal networks with functional synaptic transmission after transplantation into mice with neurodegeneration. Stem Cells 25:1307-1316.
10. Albin RL, Mink JW (2006): Recent advances in Tourette syndrome research. Trends Neurosci 29:175-182.
11. Singer HS, Loiselle CR, Lee O, Minzer K, Swedo S, Grus FH (2004): Anti-basal ganglia antibodies in PANDAS. Mov Disord 19:406-415.
12. Hallett JJ, Harling-Berg CJ, Knopf PM, Stopa EG, Kiessling LS (2000): Anti-striatal antibodies in Tourette syndrome cause neuronal dysfunction. J Neuroimmuno, 111:195-202.
13. Taylor JR, Morshed SA, Parveen S, Mercadante MT, Scahill L, Peterson BS, et al. (2002): An animal model of Tourette's syndrome. Am J Psychiatry 159:657-660.
14. Rizzo R, Gulisano M, Pavone P, Fogliani F, Robertson MM (2006): Increased antistreptococcal antibody titers and anti-basal ganglia antibodies in patients with Tourette syndrome: controlled cross-sectional study. J Child Neurol 21:747-753.
15. Yeh CB, Wu CH, Tsung HC, Chen CW, Shyu JF, Leckman JF (2006): Antineural antibody in patients with Tourette's syndrome and their family members. J Biomed Sci 13:101-112.
16. Loiselle CR, Lee O, Morgan TH, Singer HS (2004): Striatal microinfusion of Tourette syndrome and PANDAS sera: failure to induce behavioral changes. Mov Disord 19:390-396.
17. Singer HS, Mink JW, Loiselle CR, Burke KA, Ruchkina I, Morshed S, et al. (2005): Microinfusion of antineuronal antibodies into rodent striatum: failure to differentiate between elevated and low titers. J Neuroimmunol 163:8-14.
18. Swerdlow NR, Sutherland AN (2005): Using animal models to develop therapeutics for Tourette Syndrome. Pharmacol Ther 108:281-293.
19. Perlmutter SJ, Leitman SF, Garvey MA, Hamburger S, Feldman F, Leonard HL, et al. (1999): Therapeutic plasma exchange and intravenous immunoglobulin for obsessive-compulsive disorder and tic disorders in childhood. Lancet 354:1153-1158.
20. Long X, Olszewski M, Huang W, Kletzel M (2005): Neural cell differentiation in vitro from adult human bone marrow mesenchymal stem cells. Stem Cells Dev 14:65-69.
21. Deng J, Petersen BE, Steindler DA, Jorgensen ML, Laywell ED (2006): Mesenchymal stem cells spontaneously express neural proteins in culture and are neurogenic after transplantation. Stem Cells 24:1054-1064.
22. Munoz-Elias G, Woodbury D, Black IB (2003): Marrow stromal cells, mitosis, and neuronal differentiation: stem cell and precursor functions. Stem Cells 21:437-448.
23. Chu Q, Wang Y, Fu X, Zhang S (2004): Mechanism of in vitro differentiation of bone marrow stromal cells into neuron-like cells. J Huazhong Univ Sci Technolog Med Sc 24:259-261.
24. Gaughwin PM, Caldwell MA, Anderson JM, Schwiening CJ, Fawcett JW, Compston DA, et al (2006): Astrocytes promote neurogenesis from oligodendrocyte precursor cells. Eur J Neurosc, 23:945-956.
25. Swanson RA, Farrell K, Stein BA (1997): Astrocyte energetics, function, and death under conditions of incomplete ischemia: a mechanism of glial death in the penumbra. Glia 21:142-153.
26. Zhong C, Qin Z, Zhong CJ, Wang Y, Shen XY (2003): Neuroprotective effects of bone marrow stromal cells on rat organotypic hippocampal slice culture model of cerebral ischemia. Neurosci Lett 342:93-96.
27. Cho KJ, Trzaska KA, Greco SJ, McArdle J, Wang FS, Ye JH, et al (2005): Neurons derived from human mesenchymal stem cells show synaptic transmission and can be induced to produce the neurotransmitter substance P by interleukin-1 alpha. Stem Cells 23:383-391.
2 Reynolds BA, Tetzlaff W, Weiss S. A multipotent EGF-responsive striatal embryonic progenitor cell produces neurons and astrocytes. J Neurosci 1992, 12(11): 4565-74.
3 Reynolds BA, Weiss S. Generation of neurons and astrocytes from isolated cells of the adult mammalian central nervous system. Science 1992, 255(5052): 1707-10.
4 Richards LJ, Kilpatrick TJ, Bartlett PF. De novo generation of neuronal cells from the adult mouse brain. Proc Natl Acad Sci U S A1992, 89(18): 8591-5.
5 Shimazaki T. Biology and clinical application of neural stem cells. Horm Res 2003, Suppl 60(3): 1-9.
6 Sugaya K. Potential use of stem cells in neuroreplacement therapies for neurodegenerative diseases. Int Rev Cytol 2003,228: 1-30.
7 Lendahl U, Zimmeman LB, McKay RD. CNS stem cells express a new class of intermediate filament protein. Cell 1990, 60(4): 585
8 Dahlstrand J, Lardelli M, Lendahl U. Nestin mRNA expression correlates with the central nervous system progenitor cell state in many, but not all, regions of developing central nervous system. Brain Res Dev Brain Res 1995, 84(1): 109-29.
9 Johansson CB, Lothian C, Molin M, Okano H, Lendahl U. Nestin enhancer requirements for expression in normal and injured adult CNS. J Neurosci Res 2002, 69(6): 784-94.
10 Lothian C, Lendahl U. An evolutionary conserved region in the second intron of the human nestin gene directs gene expression to CNS progenitor cells and to early neural crest cells. Eur J Neurosci 1997,9(3): 452-62.
11 Akiyama Y, Honmou O, Kato T, Uede T, Hashi K, et al. Transplantation of clonal neural precursor cells derived from adult human brain establishes functional peripheral myelin in the rat spinal cord. Exp Neurol 2001, 167(1): 27-39.
12 Zigova T, Pencea V, Betarbet R, Wiegand SJ, Alexander C, et al. Neuronal progenitor cells of the neonatal subventricular zone differentiate and disperse following transplantation into the adult rat striatum. Cell Transplant 1998, 7(2): 137-56.
13 Lepore AC, Han SS, Tyler-Polsz CJ, Cai J, Rao MS, et al. Differential fate of multipotent and lineage-restricted neural precursors following transplantation into the adult CNS. Neuron Glia Biol 2004, 1(2): 113-126.
14 Martinez-Serrano A, Lundberg C, Horellou P, Fischer W, Bentlage C, et al. CNS-derived neural progenitor cells for gene transfer of nerve growth factor to the adult rat brain: complete rescue of axotomized cholinergic neurons after transplantation into the septum. J Neurosci 1995, 15(8): 5668-80.
15 Dwain I, Xiangpeng Y, Zeng Z, Patricia T, Joh SY. Neural stem cells—a promising potential therapy for brain tumors. Curr Stem Cell Res Ther 2006, 1(1): 79-84.
16 Cai J, Limke TL, Ginis I, Rao MS. Identifying and tracking neural stem cells. Blood Cells Mol Dis 2003, 31(1): 18-27.
17 Okano H, Yoshizaki T, Shimazaki T, Sawamoto K. Isolation and transplantation of dopaminergic neurons and neural stem cells. Parkinsonism Relat Disord 2002, 9(1): 23-8.
18 Ding S, Messam CA, Li P, Selzer ME, Dichter MA, et al. Murine brain progenitor cells have the ability to differentiate into functional neurons and integrate into the CNS. Cell Transplant 2006, 15(8-9): 699-710.
19 Taupin P. Therapeutic potential of adult neural stem cells. Recent Patents CNS Drug Discov 2006, 1(3): 299-303.
20 Li J, Imitola J, Snyder EY, Sidman RL. Neural stem cells rescue nervous purkinje neurons by restoring molecular homeostasis of tissue plasminogen activator and downstream targets. J Neurosci 2006,26(30): 7839-48.
21 Villa A, Rubio FJ, Navarro B, Bueno C, Martinez-Serrano A. Human neural stem cells in vitro. A focus on their isolation and perpetuation. Biomed Pharmacother 2001, 55(2): 91-5.
22 Tarasenko YI, Yu Y, Jordan PM, Bottenstein J, Wu P. Effect of growth factors on proliferation and phenotypic differentiation of human fetal neural stem cells. J Neurosci Res 2004, 78(5): 625-36.
23 Wang K, Wang JJ, Wang Y, He OH, Wang X, et al. Infusion of epidermal growth factor and basic fibroblast growth factor into the striatum of parkinsonian rats leads to in vitro proliferation and differentiation of adult neural progenitor cells. Neurosci Lett 2004, 364(3): 154-8.
24 Miltenyi S, Muller W, Weichel W, Radbruch A. High gradient magnetic cell separation with MACS. Cytometry 1990,11(2): 231-8.
25 Constantinescu SN. Stem cell generation and choice of fate: role of cytokines and cellular microenvironment. J Cell Mol Med 2000,4(4): 233-248.
26 Klassen HJ, Imfeld KL, Kirov II, Tai L, Gage FH, et al. Expression of cytokines by multipotent neural progenitor cells. Cytokine 2003, 22(3-4): 101-6
27 Abematsu M, Smith I, Nakashima K. Mechanisms of neural stem cell fate determination: extracellular cues and intracellular programs. Curr Stem Cell Res Ther 2006,1(2): 267-77.
28 Riaz SS, Theofilopoulos S, Jauniaux E, Stem GM, Bradford HF. The differentiation potential of human foetal neuronal progenitor cells in vitro. Brain Res Dev Brain Res 2004, 153(1): 39-51.
29 Erlandsson A, Larsson J, Forsberg-Nilsson K. Stem cell factor is a chemoattractant and a survival factor for CNS stem cells. Exp Cell Res 2004, 301(2): 201-10.
30 Benoit BO, Savarese T, Joly M, Engstrom CM, Pang L, et al. Neurotrophin channeling of neural progenitor cell differentiation. J Neurobiol 2001, 46(4): 265-80.
31 Hammang JP, Archer DR, Duncan ID. Myelination following transplantation of EGF-responsive neural stem cells into a myelin-deficient environment. Exp Neurol. 1997, 147(1): 84-95.
32 Lillien L, Raphael H. BMP and FGF regulate the development of EGF-responsive neural progenitor cells. Development 2000, 127(22): 4993-5005.
33 Kuhn HG, Winkler J, Kempermann G, Thal LJ, Gage FH. Epidermal growth factor and fibroblast growth factor-2 have different effects on neural progenitors in the adult rat brain. J Neurosci 1997, 17(15): 5820-9.
34 Santa-Olalla J, Covarrubias L. Basic fibroblast growth factor promotes epidermal growth factor responsiveness and survival of mesencephalic neural precursor cells. J Neurobiol 1999,40(1): 14-27.
35 Itoh T, Satou T, Dote K, Hashimoto S, Ito H. Effect of basic fibroblast growth factor on cultured rat neural stem cell in three-dimensional collagen gel. Neurol Res 2005, 27(4): 429-32.
36 Gobbel GT, Choi SJ, Beier S, Niranjan A. Long-term cultivation of multipotential neural stem cells from adult rat subependyma. Brain Res 2003, 980(2): 221-32.
37 Hinsch K, Zupanc GK. Isolation, cultivation, and differentiation of neural stem cells from adult fish brain. J Neurosci Methods 2006, 158(1): 75-88.
38 Zhang X, Li X, Wu J, Wang Z, Xu H, et al. Isolation, cultivation and identification of stem cells from cerebral cortex of mouse embryo. Zhonghua Yi Xue Za Zhi 2002, 82(12): 832-5.
39 Fu SL, Ma ZW, Yin L, Lu PH, Xu XM. Isolation and cultivation of neural stem cells from the embryonic rat brain and spinal cord. Sheng Li Xue Bao 2003, 55(3): 278-83.
40 Einstein O, Ben-Menachem-Tzidon O, Mizrachi-Kol R. Survival of neural precursor cells in growth factor-poor environment: implications for transplantation in chronic disease. Glia 2006, 53(4): 449-55.
41 Piper DR, Mujtaba T, Keyoung H, Roy NS, Goldman SA, et al. Identification and characterization of neuronal precursors and their progeny from human fetal tissue. J Neurosci Res 2001, 66(3): 356-68.
42 Bazan E, Alonso FJ, Redondo C, Lopez-Toledano MA, Alfaro JM, et al. In vitro and in vivo characterization of neural stem cells. Histol Histopathol 2004, 19(4): 1261-75.
43 Okano H, Kawahara H, Toriya M, Nakao K, Shibata S, et al. Function of RNA-binding protein Musashi-1 in stem cells.Exp Cell Res 2005,306(2):349-56.
1 Rampello L, Alvano A, Battaglia G, Bruno V, Raffaele R, et al. Tic disorders: from pathophysiology to treatment. J Neurol 2006, 253(1): 1-15.
2 Olson LL, Singer HS, Goodman WK, Maria BL. Tourette syndrome: diagnosis, strategies, therapies, pathogenesis, and future research directions. J Child Neurol 2006, 21(8): 630-41.
3 Stem JS, Burza S, Robertson MM. Gilles de la Tourette's syndrome and its impact in the UK. Postgrad Med J 2005, 81(951): 12-9.
4 Leckman JF, Zhang H, Vitale A, Lahnin F, Lynch K, et al. Course of tic severity in Tourette syndrome: the first two decades. Pediatrics 1998, 102(1): 14-9.
5 Riddle MA, Carlson J. Clinical psychopharmacology for Tourette syndrome and associated disorders. Adv Neurol 2001, 85: 343-54.
6 Deckersbach T, Rauch S, Buhlmann U, Wilhelm S. Habit reversal versus supportive psychotherapy in Tourette's disorder: a randomized controlled trial and predictors of treatment response. Behav Res Ther 2006,44(8): 1079-90.
7 Cheung MY, Shahed J, Jankovic J. Malignant Tourette syndrome. Mov Disord 2007, 22(12): 1743-50.
8 Maciunas RJ, Maddux BN, Riley DE, Whitney CM, Schoenberg MR, et al. Prospective randomized double-blind trial of bilateral thalamic deep brain stimulation in adults with Tourette syndrome. J Neurosurg 2007,107(5): 1004-14.
9 Hamani C, Moro E. Surgery for other movement disorders: dystonia, tics. Curr Opin Neurol 2007, 20(4): 470-6.
10 Neimat JS, Patil PG, Lozano AM. Novel surgical therapies for Tourette syndrome. J Child Neurol 2006,21(8): 715-8.
11 Anandan S, Wigg CL, Thomas CR, Coffey B. Psychosurgery for self-injurious behavior in Tourette's disorder. J Child Adolesc Psychopharmacol 2004, 14(4): 531-8.
12 Mink JW, Walkup J, Frey KA, Como P, Cath D, et al. Patient selection and assessment recommendations for deep brain stimulation in Tourette syndrome. Mov Disord 2006, 21(11): 1831-8.
13 Wang L, Martin DR, Baker HJ, Zinn KR, Kappes JC, et al. Neural progenitor cell transplantation and imaging in a large animal model. Neurosci Res 2007, 59(3): 327-40.
14 Jain M, Armstrong RJ, Elneil S, Barker RA. Transplanted human neural precursor cells migrate widely but show no lesion-specific tropism in the 6-hydroxydopamine rat model of Parkinson's disease. Cell Transplant 2006, 15(7): 579-93.
15 Cummings BJ, Uchida N, Tamaki SJ, Anderson AJ. Human neural stem cell differentiation following transplantation into spinal cord injured mice: association with recovery of locomotor function. Neurol Res 2006, 28(5): 474-81.
16 Lee ST, Park JE, Lee K, Kang L, Chu K, et al. Noninvasive method of immortalized neural stem-like cell transplantation in an experimental model of Huntington's disease. J Neurosci Methods 2006, 152(1-2): 250-4.
17 Lee ST, Chu K, Park JE, Lee K, Kang L. Intravenous administration of human neural stem cells induces functional recovery in Huntington's disease rat model. Neurosci Res 2005, 52(3): 243-9.
18 Peterson BS, Choi HA, Hao X, Amat JA, Zhu H, et al. Morphologic features of the amygdala and hippocampus in children and adults with Tourette syndrome. Arch Gen Psychiatry 2007, 64(11): 1281-91.
19 Muller N. Tourette's syndrome: clinical features, pathophysiology, and therapeutic approaches. Dialogues Clin Neurosci 2007, 9(2): 161-71.
20 Brovedani P, Masi G. Tourette's syndrome in children: neurological, neuropsychological and psychiatric issues. Panminerva Med 2000,42(2): 141-9.
21 Swerdlow NR, Sutherland AN. Preclinical models relevant to Tourette syndrome. Adv Neurol 2006, 99: 69-88.
22 Swerdlow NR, Sutherland AN. Using animal models to develop therapeutics for Tourette Syndrome. Pharmacol Ther 2005, 108(3): 281-93.
23 Sanberg PR, Emerich DF. Neural basis of behavior: animal models of human conditions. Brain Res Bull 1990, 25(3): 447-51.
24 Singer HS, Giuliano JD, Hansen BH, Hallett JJ, Laurino JP, et al. Antibodies against human putamen in children with Tourette syndrome. Neurology 1998, 50(6): 1618-24.
25 Hallett JJ, Harling-Berg CJ, Knopf PM, Stopa EG, Kiessling LS. Anti-striatal antibodies in Tourette syndrome cause neuronal dysfunction. J Neuroimmunol 2000,111(1-2): 195-202.
26 Taylor JR, Morshed SA, Parveen S, Mercadante MT, Scahill 1, et al. An animal model of Tourette's syndrome. Am J Psychiatry 2002, 159(4): 657-60.
27 Taupin P. Autologous transplantation in the central nervous system. Indian J Med Res 2006,124(6): 613-8.
28 Muller N, Riedel M, Straube A, Gunther W, Wilske B. Increased anti-streptococcal antibodies in patients with Tourette's syndrome. Psychiatry Res 2000, 94(1): 43-9.
29 Dale RC. Autoimmunity and the basal ganglia: new insights into old diseases. QJM 2003, 96(3): 183-91.
30 Hoekstra PJ, Anderson GM, Limburg PC, Korf J, Kallenberg CG, et al. Neurobiology and neuroimmunology of Tourette's syndrome: an update. Cell Mol Life Sci 2004, 61(7-8): 886-98.
31 Khalifa N, vonKnorring AL. Tourette syndrome and other tic disorders in a total population of children: clinical assessment and background. Acta Paediatr 2005, 94(11): 1608-14.
32 Burd L, Freeman RD, Klug MG, Kerbeshian J. Tourette Syndrome and learning disabilities. BMC Pediatr 2005, 5: 34.
33 Chang HL, Tu MJ, Wang HS. Tourette's syndrome: psychopathology in adolescents. Psychiatry Clin Neurosci 2004, 58(4): 353-8.
34 Channon S, Sinclair E, Waller D, Healey L, Robertson MM. Social cognition in Tourette's syndrome: intact theory of mind and impaired inhibitory functioning. J Autism Dev Disord 2004, 34(6): 669-77.
35 Goudriaan AE, Oosterlaan J, de Berrs E, van den Brink W. Neurocognitive functions in pathological gambling: a comparison with alcohol dependence, Tourette syndrome and normal controls. Addiction 2006, 101(4): 534-47.
36 Kano Y, Ohta M, Nagai Y, Pauls DL, Leckman JF. Obsessive-compulsive symptoms in parents of Tourette syndrome probands and autism spectrum disorder probands. Psychiatry Clin Neurosci 2004, 58(4): 348-52.
37 Stephens RJ, Sandor P. Aggressive behaviour in children with Tourette syndrome and comorbid attention-deficit hyperactivity disorder and obsessive-compulsive disorder. Can J Psychiatry 1999, 44(10): 1036-42.
38 Rizzo R, Gulisano M, Pavone P, Fogliani F, Robertson MM. Increased antistreptococcal antibody titers and anti-basal ganglia antibodies in patients with Tourette syndrome: controlled cross-sectional study. J Child Neurol 2006, 21(9): 747-53.
39 Loiselle CR, Wendlandt JT, Rohde CA, Singer HS. Antistreptococcal, neuronal, and nuclear antibodies in Tourette syndrome. Pediatr Neurol 2003, 28(2): 119-25.
40 Visvanathan K, Trifiletti RR, Altemus M, Zabriskie JB. Autoimmune mechanisms in movement disorders. Semin Pediatr Neurol 2000, 7(2): 103-7.
41 Muller N, Riedel M, Forderreuther S, Blendinger C, Abele-Hom M. Tourette's syndrome and mycoplasma pneumoniae infection. Am J Psychiatry 2000, 157(3): 481-2.
42 Perlmutter SJ, Leitman SF, Garvey MA, Hamburger S, Feldman E, et al. Therapeutic plasma exchange and intravenous immunoglobulin for obsessive-compulsive disorder and tic disorders in childhood. Lancet 1999, 354(9185): 1153-8.
43 Gilbert D. Treatment of children and adolescents with tics and Tourette syndrome. J Child Neurol 2006, 21(8): 690-700.
44 Kurlan R. Future and alternative therapies in Tourette syndrome. Adv Neurol 2006, 99: 248-53.
45 Bithell A, Williams BP. Neural stem cells and cell replacement therapy: making the right cells. Clin Sci (Lond) 2005,108(1): 13-22.
46 Cataudella T, Conti L, Cattaneo E. Neural stem and progenitor cells: choosing the right Shc. Prog Brain Res 2004, 146: 127-33.
47 Harris K, Singer HS. Tic disorders: neural circuits, neurochemistry, and neuroimmunology. J Child Neurol 2006,21(8): 678-89.
48 Albin RL. Neurobiology of basal ganglia and Tourette syndrome: striatal and dopamine function. Adv Neurol 2006, 99:99-106.
1 Galvin KA, Jones DG. Adult human neural stem cells for autologous cell replacement therapies for neurodegenerative disorders. NeuroRehabilitation 2006, 21(3): 255-65.
2 Kim HT, Kim IS, Lim SE, Lee IS, Park KI. Gene and cell replacement via neural stem cells. Yonsei Med J 2004, 45 Suppl: 32-40.
3 Ourednik J, Ourednik V, Lynch WP, Schachner M, Snyder EY. Neural stem cells display an inherent mechanism for rescuing dysfunctional neurons. Nat Biotechnol 2002,20(11): 1103-10. Epub 2002 Oct 15.
4 Lynch WP, Portis JL. Neural stem cells as tools for understanding retroviral neuropathogenesis. Virology 2000, 271(2): 227-33.
5 Yang H, Lu P, McKay HM, Bernot T, Keirstead H, et al. Endogenous neurogenesis replaces oligodendrocytes and astrocytes after primate spinal cord injury. J Neurosci 2006, 26(8): 2157-66
6 Lie DC, Song H, Colamarino SA, Ming GL, Gage FH. Neurogenesis in the adult brain: new strategies for central nervous system diseases. Annu Rev Pharmacol Toxicol 2004,44: 399-421.
7 Yasuhara T, matsukawa N, Hara K, Yu G, Xu L, et al. Transplantation of human neural stem cells exerts neuroprotection in a rat model of Parkinson's disease. J Neurosci 2006, 26(48): 12497-511.
8 Lee ST, Chu K, Park JE, Lee K, Kang L, et al. Intravenous administration of human neural stem cells induces functional recovery in Huntington's disease rat model. Neurosci Res 2005, 52(3): 243-9.
9 Karbanova J, Mokry J, kotingova L. Neural stem cells transplanted into intact brains as neurospheres form solid grafts composed of neurons, astrocytes and oligodendrocyte precursors. Biomed Pap Med Fac Univ Palacky Olomouc Czech Repub 2004,148(2): 217-20.
10 Lenka N. Derivation and characterization of neural cells from embryonic stem cells using nestin enhancer. Methods Mol Biol 2006, 330: 33-54.
11 Takagi Y, Nishimura M, Morizane A, Takahashi J, Nozaki K, et al. Survival and differentiation of neural progenitor cells derived from embryonic stem cells and transplanted into ischemic brain. J Neurosurg 2005,103(2): 304-10.
12 Aleksandrova MA, Podgomyi OV, Marei MV, Poltavtseva RA, Tsitrin EB, et al. Characteristics of human neural stem cells in vitro and after transplantation into rat brain. Bull Exp Biol Med 2005, 139(1): 114-20.
13 Zhu W, Mao Y, Zhao Y, Zhou LF, Wang Y, et al. Transplantation of vascular endothelial growth factor-transfected neural stem cells into the rat brain provides neuroprotection after transient focal cerebral ischemia. Neurosurgery 2005, 57(2): 325-33
14 Li T, Zheng J, Xie Y, Wang S, Zhang X, et al. Transplantable neural progenitor populations derived from rhesus monkey embryonic stem cells. Stem Cells 2005, 23(9): 1295-303.
15 Enzmann V, Howard RM, Yamauchi Y, Whittemore SR, Kaplan HJ. Enhanced induction of RPE lineage markers in pluripotent neural stem cells engrafted into the adult rat subretinal space. Invest Ophthalmol Vis Sci 2003, 44(12): 5417-22.
16 Yoshinaga T, Hashimoto E, Ukai W, Toki S, Saito S, et al. Neural stem cell transplantation in a model of fetal alcohol effects. J Neural Transm Suppl 2007, (72): 331-7.
17 Darsalia V, Kallur T, Kokaia Z. Survival, migration and neuronal differentiation of human fetal striatal and cortical neural stem cells grafted in stroke-damaged rat striatum. Eur J Neurosci 2007, 26(3): 605-14.
18 Yasuhara T, Matsukawa N, Hara K, Yu G, Xu L, et al. Transplantation of human neural stem cells exerts neuroprotection in a rat model of Parkinson's disease. J Neurosci 2006, 26(48): 12497-511
19 Heine W, Conant K, Griffin JW, Hoke A. Transplanted neural stem cells promote axonal regeneration through chronically denervated peripheral nerves. Exp Neurol 2004, 189(2): 231-40.
20 Riess P, Zhang C, Saatman KE, Laurer HL, Longhi LG, et al. Transplanted neural stem cells survive, differentiate, and improve neurological motor function after experimental traumatic brain injury. Neurosurgery 2002, 51(4): 1043-52.
21 Jeong SW, Chu K, Jung KH, Kim SU, Kim M, et al. Human neural stem cell transplantation promotes functional recovery in rats with experimental intracerebral hemorrhage. Stroke 2003, 34(9): 2258-63.
22 Enomoto M, Shinomiya K, Okabe S. Migration and differentiation of neural progenitor cells from two different regions of embryonic central nervous system after transplantation into the intact spinal cord. Eur J Neurosci 2003, 17(6): 1223-32.
23 Lee HJ, Kim KS, Kim EJ, Choi HB, Lee KH, et al. Brain transplantation of immortalized human neural stem cells promotes functional recovery in mouse intracerebral hemorrhage stroke model. Stem Cells 2007, 25(5): 1204-12.
24 Mueller D, Shamblott MJ, Fox HE, Gearhart JD, Martin LJ. Transplanted human embryonic germ cell-derived neural stem cells replace neurons and oligodendrocytes in the forebrain of neonatal mice with excitotoxic brain damage. J Neurosci Res 2005, 82(5): 592-608.
25 Niles LP, Armstrong KJ, Rincon Castro LM, Dao CV, Sharma R, et al. Neural stem cells express melatonin receptors and neurotrophic factors: colocalization of the MT1 receptor with neuronal and glial markers. BMC Neurosci 2004, 5(1): 41.
26 Lu P, Jones LL, Snyder EY, Tuszynski MH. Neural stem cells constitutively secrete neurotrophic factors and promote extensive host axonal growth after spinal cord injury. ExpNeurol 2003, 181(2): 115-29.
27 Ourednik J, Ourednik V, Lynch WP, Schachner M, Snyder EY. Neural stem cells display an inherent mechanism for rescuing dysfunctional neurons. Nat Biotechnol 2002, 20(11): 1103-10.
28 Yan J, Xu L, Welsh AM, Hatfield G, Hazel T, et al. Extensive neuronal differentiation of human neural stem cell grafts in adult rat spinal cord. PLoS Med 2007, 4(2): 39.
29 AnYH, Wang HY, Gao ZX, Wang ZC. Differentiation of rat neural stem cells and its relationship with environment. Biomed Environ Sci 2004,17(1): 1-7.
30 Jori FP, Galderisi U, Piegari E, Cipollaro M, Cascino A, et al. EGF-responsive rat neural stem cells: molecular follow-up of neuron and astrocyte differentiation in vitro. J Cell Physiol 2003, 195(2): 220-33.
31 Nakahara Y, Gage FH, Tuszynski MH. Grafts of fibroblasts genetically modified to secrete NGF, BDNF, NT-3, or basic FGF elicit differential responses in the adult spinal cord. Cell Transplant 1996, 5(2): 191-204.
32 Winkler C, Fricker RA, Gates MA, Olsson M, Hammang JP, et al. Incorporation and glial differentiation of mouse EGF-responsive neural progenitor cells after transplantation into the embryonic rat brain. Mol Cell Neurosci 1998, 11(3): 99-116.
33 Nikkhah G, Olsson M, Eberhard J. A micro transplantation approach for cell suspension grafting in the rat Parkinson model: A detailed account of methodology. Neuroscience 1994, 63(1): 57.
34 Muraoka K, Shingo T, Yasuhara T, Kameda M, Yuan W, et al. The high integration and differentiation potential of autologous neural stem cell transplantation compared with allogeneic transplantation in adult rat hippocampus. Exp Neurol 2006,199(2): 311-27.
35 Ehtesham M, Kabos P, Gutierrez MA, Chung NH, Griffith TS, et al. Induction of glioblastoma apoptosis using neural stem cell-mediated delivery of tumor necrosis factor-related apoptosis-inducing ligand. Cancer Res 2002, 62(24): 7170-4.
36 Mammolenti M, Gajavelli S, Tsoulfas P, Levy R. Absence of major histocompatibility complex class I on neural stem cells does not permit natural killer cell killing and prevents recognition by alloreactive cytotoxic T lymphocytes in vitro. Stem Cells 2004,22(6): 1101-10.
37 Lee EM, Kim JY, Cho BR, Chung WK, Yoon BW, et al. Down-regulation of MHC class I expression in human neuronal stem cells using viral stealth mechanism. Biochem Biophys Res Commun 2005, 326(4): 825-35.
38 Wei P, Liu J, Zhou HL, Han ZT, Wu QY, et al. Effects of engrafted neural stem cells derived from GFP transgenic mice in Parkinson's diseases rats. Neurosci Lett 2007, 419(1): 49-54.
39 Kon ND, Mills SA, Andrews CA, Miller LR, Cordell AR. Effect of desmopressin on vein graft patency in a microvascular model. Microsurgery 1991, 12(4): 288-91.
[1] Albin RL, Mink JW. Recent advances in Tourette syndrome research. Trends Neurosci 2006; 29: 175-82.
[2] Berthier ML, Campos VM, Kulisevsky J, Valero JA. Heroin and malignant coprolalia in Tourette's syndrome. J Neuropsychiatry Clin Neurosci 2003; 15: 116-7.
[3] Bjugstad KB, Redmond ED Jr, Teng YD, Elsworth JD, Roth RH, Blanchard BC, et al. Neural stem cells implanted into MPTP-treated monkeys increase the size of endogenous tyrosine hydroxylase-positive cells found in the striatum: a return to control measures. Cell Transplant 2005; 14: 183-92.
[4] Baizabal JM, Furlan-Magaril M, Santa-Olalla J, Covarrubias L. Neural stem cells in development and regenerative medicine. Arch Med Res 2003; 34: 572-88.
[5] Conti L, Retano E, Cattaneo E. Neural stem cell systems: diversities and properties after transplantation in animal models of diseases. Brain Pathol 2006; 16: 143-54.
[6] Diaz-Anzaldua A, Joober R, Riviere JB, Dion Y, Lesperance P, Richer F, et al. Tourette syndrome and dopaminergic genes: a family-based association study in the French Canadian founder population. Mol Psychiatry 2004; 9: 272-7.
[7] Fattapposta F, Restuccia R, Colonnese C, Labruna L, Garreffa G, Bianco F. Gilles de la Tourette syndrome and voluntary movement: a functional MRI study. Psychiatry Res 2005; 138: 269-72
[8] Gaughwin PM, Caldwell MA, Anderson JM, Schwiening CJ, Fawcett JW, Compston DA, et al. Astrocytes promote neurogenesis from oligodendrocyte precursor cells. Eur J Neurosci 2006; 23: 945-56.
[9] Hallett JJ, Harling-Berg CJ, Knopf PM, Stopa EG, Kiessling LS. Anti-striatal antibodies in Tourette syndrome cause neuronal dysfunction. J Neuroimmunol 2000; 111: 195-202.
[10] Hong JJ, Loiselle CR, Yoon DY, Lee O, Becker KG, Singer HS. Microarray analysis in Tourette syndrome postmortem putamen. J Neurol Sci 2004; 225: 57-64.
[11] Hu Z, Wei D, Johansson CB, Holmstrom N, Duan M, Frisen J, et al. Survival and neural differentiation of adult neural stem cells transplanted into the mature inner ear. Exp Cell Res 2005; 302: 40-7.
[12] Korochkin LI, Revishchi AV, Okhotin VE. Neural stem cells and their role in recovery processes in the nervous system. Neurosci Behav Physiol 2006; 36: 499-512.
[13] Leckman JF, Zhang H, Vitale A, Lahnin F, Lynch K, Bondi C, et al. Course of tic severity in Tourette syndrome: the first two decades. Pediatrics 1998; 102: 14-9.
[14] Loiselle CR, Lee O, Morgan TH, Singer HS. Striatal microinfusion of Tourette syndrome and PANDAS sera: failure to induce behavioral changes. Mov Disord 2004; 19: 390-6.
[15] Lee ST, Park JE, Lee K, Kang L, Chu K, Kim SU, et al. Noninvasive method of immortalized neural stem-like cell transplantation in an experimental model ofHuntington's disease. J Neurosci Methods 2006; 152: 250-4.
[16] Liang P, Jin LH, Liang T, Liu EZ, Zhao SG. Human neural stem cells promote corticospinal axons regeneration and synapse reformation in injured spinal cord of rats. Chin Med J (Engl) 2006; 119: 1331-8.
[17] Li J, Imitola J, Snyder EY, Sidman RL. Neural stem cells rescue nervous purkinje neurons by restoring molecular homeostasis of tissue plasminogen activator and downstream targets. J Neurosci 2006; 26: 7839-48.
[18] Mink JW, Walkup J, Frey KA, Como P, Cath D, Delong MR, et al. Patient selection and assessment recommendations for deep brain stimulation in Tourette syndrome. Mov Disord 2006; 21: 1831-8.
[19] Perlmutter SJ, Leitman SF, Garvey MA, Hamburger S, Feldman F, Leonard HL, et al. Therapeutic plasma exchange and intravenous immunoglobulin for obsessive-compulsive disorder and tic disorders in childhood. Lancet 1999; 354: 1153-8.
[20] Pallini R, Vitiani LR, Bez A, Casalbore P, Facchiano F, Di Giorgi Gerevini V, et al. Homologous transplantation of neural stem cells to the injured spinal cord of mice. Neurosurgery 2005; 57: 1014-25.
[21] Parati EA, Bez A, Ponti D, Sala S, Pozzi S, Pagano SF. Neural stem cells. Biological features and therapeutic potential in Parkinson's disease. J Neurosurg Sci 2003; 47: 8-17.
[22] Rampello L, Alvano A, Battaglia G, Bruno V, Raffaele R, Nicoletti F. Tic disorders: from pathophysiology to treatment. J Neurol 2006; 253: 1-15.
[23] Rizzo R, Gulisano M, Pavone P, fogliani F, Robertson MM. Increased antistreptococcal antibody titers and anti-basal ganglia antibodies in patients with Tourette syndrome: controlled cross-sectional study. J Child Neurol 2006; 21: 747-53.
[24] Singer HS, Loiselle CR, Lee O, Minzer K, Swedo S, Grus FH. Anti-basal ganglia antibodies in PANDAS. Mov Disord 2004; 19: 406-15.
[25] Singer HS, Mink JW, Loiselle CR, Burke KA, Ruchkina I, Morshed S, et al. Microinfusion of antineuronal antibodies into rodent striatum: failure to differentiate between elevated and low titers. J Neuroimmunol 2005; 163: 8-14.
[26] Swerdlow NR, Sutherland AN. Using animal models to develop therapeutics for Tourette Syndrome. Pharmacol Ther 2005; 108: 281-93.
[27] Taylor JR, Morshed SA, Parveen S, Mercadante MT, Scahill 1, Peterson BS, et al. An animal model of Tourette's syndrome. Am J Psychiatry 2002; 159: 657-60.
[28] Yeh CB, Wu CH, Tsung HC, Chen CW, Shyu JF, Leckman JF. Antineural antibody in patients with Tourette's syndrome and their family members. J Biomed Sci 2006; 13: 101-12.
1. Rampello L, Alvano A, Battaglia G, Bruno V, Raffaele R, Nicoletti F (2006): Tic disorders: from pathophysiology to treatment. J Neurol 253:1-15.
2. Leckman JF, Zhang H, Vitale A, Lahnin F, Lynch K, Bondi C, et al. (1998): Course of tic severity in Tourette syndrome: the first two decades. Pediatrics 102:14-19.
3. Berthier M, Campos VM, Kulisevsky J, Valero JA (2003): Heroin and malignant coprolalia in Tourette's syndrome. J Neuropsychiatry Clin Neurosci 15:116-117.
4. Mink JW, Walkup J, Frey KA, Como P, Cam D, Delong MR, et al. (2006): Patient selection and assessment recommendations for deep brain stimulation in Tourette syndrome. Mov Disord 21:1831-1838.
5. Kamishina H, Deng J, Oji T, Cheeseman JA, Clemmons RM (2006): Expression of neural markers on bone marrow-derived canine mesenchymal stem cells. Am J Vet Res 67:1921-1928.
6. Kim S, Honmou O, Kato K, Nonaka T, Houkin K, Hamada H, et al. (2006): Neural differentiation potential of peripheral blood- and bone-marrow-derived precursor cells. Brain Res 1123:27-33.
7. Pisati F, Bossolasco P, Meregalli M, Cova L, Belicchi M, Gavina M, et al. (2007): Induction of neurotrophin expression via human adult mesenchymal stem cells: implication for cell therapy in neurodegenerative diseases. Cell Transplan 16:41-55.
8. Mori K, Iwata J, Miyazaki M, Nakao Y, Maeda M (2005): Functional recovery of neuronal activity in rat whisker-barrel cortex sensory pathway from freezing injury after transplantation of adult bone marrow stromal cells. J Cereb Blood Flow Metab 25:887-898.
9. Bae JS, Han HS, Youn DH, Carter JE, Modo M, Schuchman EH, et al (2007): Bone marrow-derived mesenchymal stem cells promote neuronal networks with functional synaptic transmission after transplantation into mice with neurodegeneration. Stem Cells 25:1307-1316.
10. Albin RL, Mink JW (2006): Recent advances in Tourette syndrome research. Trends Neurosci 29:175-182.
11. Singer HS, Loiselle CR, Lee O, Minzer K, Swedo S, Grus FH (2004): Anti-basal ganglia antibodies in PANDAS. Mov Disord 19:406-415.
12. Hallett JJ, Harling-Berg CJ, Knopf PM, Stopa EG, Kiessling LS (2000): Anti-striatal antibodies in Tourette syndrome cause neuronal dysfunction. J Neuroimmuno, 111:195-202.
13. Taylor JR, Morshed SA, Parveen S, Mercadante MT, Scahill L, Peterson BS, et al. (2002): An animal model of Tourette's syndrome. Am J Psychiatry 159:657-660.
14. Rizzo R, Gulisano M, Pavone P, Fogliani F, Robertson MM (2006): Increased antistreptococcal antibody titers and anti-basal ganglia antibodies in patients with Tourette syndrome: controlled cross-sectional study. J Child Neurol 21:747-753.
15. Yeh CB, Wu CH, Tsung HC, Chen CW, Shyu JF, Leckman JF (2006): Antineural antibody in patients with Tourette's syndrome and their family members. J Biomed Sci 13:101-112.
16. Loiselle CR, Lee O, Morgan TH, Singer HS (2004): Striatal microinfusion of Tourette syndrome and PANDAS sera: failure to induce behavioral changes. Mov Disord 19:390-396.
17. Singer HS, Mink JW, Loiselle CR, Burke KA, Ruchkina I, Morshed S, et al. (2005): Microinfusion of antineuronal antibodies into rodent striatum: failure to differentiate between elevated and low titers. J Neuroimmunol 163:8-14.
18. Swerdlow NR, Sutherland AN (2005): Using animal models to develop therapeutics for Tourette Syndrome. Pharmacol Ther 108:281-293.
19. Perlmutter SJ, Leitman SF, Garvey MA, Hamburger S, Feldman F, Leonard HL, et al. (1999): Therapeutic plasma exchange and intravenous immunoglobulin for obsessive-compulsive disorder and tic disorders in childhood. Lancet 354:1153-1158.
20. Long X, Olszewski M, Huang W, Kletzel M (2005): Neural cell differentiation in vitro from adult human bone marrow mesenchymal stem cells. Stem Cells Dev 14:65-69.
21. Deng J, Petersen BE, Steindler DA, Jorgensen ML, Laywell ED (2006): Mesenchymal stem cells spontaneously express neural proteins in culture and are neurogenic after transplantation. Stem Cells 24:1054-1064.
22. Munoz-Elias G, Woodbury D, Black IB (2003): Marrow stromal cells, mitosis, and neuronal differentiation: stem cell and precursor functions. Stem Cells 21:437-448.
23. Chu Q, Wang Y, Fu X, Zhang S (2004): Mechanism of in vitro differentiation of bone marrow stromal cells into neuron-like cells. J Huazhong Univ Sci Technolog Med Sc 24:259-261.
24. Gaughwin PM, Caldwell MA, Anderson JM, Schwiening CJ, Fawcett JW, Compston DA, et al (2006): Astrocytes promote neurogenesis from oligodendrocyte precursor cells. Eur J Neurosc, 23:945-956.
25. Swanson RA, Farrell K, Stein BA (1997): Astrocyte energetics, function, and death under conditions of incomplete ischemia: a mechanism of glial death in the penumbra. Glia 21:142-153.
26. Zhong C, Qin Z, Zhong CJ, Wang Y, Shen XY (2003): Neuroprotective effects of bone marrow stromal cells on rat organotypic hippocampal slice culture model of cerebral ischemia. Neurosci Lett 342:93-96.
27. Cho KJ, Trzaska KA, Greco SJ, McArdle J, Wang FS, Ye JH, et al (2005): Neurons derived from human mesenchymal stem cells show synaptic transmission and can be induced to produce the neurotransmitter substance P by interleukin-1 alpha. Stem Cells 23:383-391.