骨髓基质细胞与中药合用治疗缺血性脑损伤的相关问题

骨髓基质细胞具有高度的增殖能力和神经分化潜能，是修复中枢系统神经损伤的理想细胞，其作用途径可能是通过增殖、分化替代受损神经细胞并整合到原有的神经网络当中，并分泌细胞因子促进内源性神经干细胞的增殖与分化，以促进神经功能恢复，减少损伤区细胞凋亡等。中药治疗缺血性脑损伤疗效显著，而且可以促进骨髓基质细胞的增殖与神经分化，将中药与骨髓基质细胞联合治疗缺血性脑损伤具有很好的应用前景。目前亟需解决的是骨髓基质细胞的输入量、途径以及生物安全性，中药与骨髓基质细胞合用的时间窗、中药的剂型和浓度等问题。     

成体内源性神经干细胞的激活及活化策略

神经干细胞具有自我更新和多分化潜能，其应用为中枢神经系统神经再生和功能重建提供了全新的治疗思路。侧脑室外侧壁的室下带（subventricular zone，SVZ）和海马齿状回（dentate gyrus．DG）的颗粒下层是所有成年哺乳动物脑内存在的2个神经干细胞富集区，其他区域如脊髓、隔区、纹状体等也分离出了神经干细胞。目前应用神经干细胞修复中枢神经系统损伤已形成两种思路：移植外源性干细胞的“替代治疗”策略和激活内源性干细胞的“补充治疗”策略．而内源性神经干细胞具有来源稳定可靠、无伦理道德问题、无免疫排斥反应、无致瘤性等优势，因此如何充分调动内源性神经干细胞治疗神经系统疾病已成为当前新的研究热点。本文就内源性神经干细胞的激活及近年来的活化策略作一简述。[第一段]     

神经干细胞与神经再生的研究进展

神经干细胞是一类具有自我复制更新能力、高度增殖潜能和多向分化潜能的细胞,通过外源性或内源性途径的神经干细胞疗法为神经退行性病变及其他神经系统疾病的治疗提供了一种新的途径。     

神经干细胞在中枢神经系统损伤修复中的应用前景

神经干细胞是中枢神经系统中保持分裂和分化潜能的细胞。目前从神经干细胞的生物学特性、常规体外培养方法的建立 ,到神经干细胞的分化鉴定、电生理检测及定向诱导方法等方面的基础研究都取得了可喜成绩 ,这些成绩的取得为神经干细胞的临床应用奠定了坚实的实验基础。可以肯定 ,今后神经干细胞移植可以作为治疗神经系统损伤及疾病的一种方法。神经干细胞具有增殖和多向分化的潜能 ,在移植部位分裂增殖 ,并在局部微环境的作用下分化成相应的细胞来补充替代受损的细胞 ,恢复中枢神经系统的正常结构和功能。移植后产生的细胞也可以自主释放神经…     

神经干细胞的应用

神经于细胞存在于人类神经系统中，具有自我更新、活跃的增殖和分化能力、向脑内病变部位迁移以及分化成神经元、星形胶质细胞和少突胶质细胞的能力。近年来神经干细胞疗法已成为治疗多种疾病的新策略，其目的是替代、修复或加强受损的细胞或器官的生物学功能，是基因疗法的一种理想的靶细胞。应用主要集中以下几方面：①直接细胞移植进行替代治疗，神经干细胞作为细胞移植的来源，可以通过神经干细胞的体外移植或体内神经干细胞的激活，分化为神经元和胶质细胞，与已经存在的细胞结构整合到一起。②作为基因载体，携带治疗作用的报告基因进行移植，从而达到细胞替代和基因治疗的双重作用。③通过对牛长因子和细胞因子的研究，诱导自身的神经千细胞分化进行神经自我修复。可广泛应用于脑外伤、脑血管病后脑功能损伤、脑瘤及其他疾病的治疗。     

成体神经干细胞及其在神经修复中的应用

神经干细胞是一类具有多分化潜能的未分化细胞。在适宜的环境下可分化为神经元、神经胶质细胞和少突胶质细胞。神经干细胞的无限增殖性和多分化性使其成为细胞移植的良好供体。近年来，成年脑组织内也发现了具有多分化潜能的神经干细胞。成体神经干细胞的发现改变了以往中枢神经系统不可再生的理论，随着对成体神经干细胞研究的深入，通过诱导自体神经干细胞的体内增殖修复损伤的神经元，通过分离自体神经干细胞体外扩增后移植治疗等实验的探索为神经源性疾病的替代性治疗提供了新的思路。     

神经干细胞移植治疗缺血性脑卒中研究新进展

神经干细胞是一类具有自我复制更新能力、高度增殖潜能和多向分化潜能的细胞,通过外源性或内源性途径的神经干细胞疗法为缺血性脑卒中的治疗提供了一种新途径。现就神经干细胞及其治疗缺血性脑卒中的研究进展进行综述。     

脑脊液途径移植神经干细胞治疗中枢神经系统疾病的研究现状简

背景：目前报道的用于神经干细胞移植的途径主要有经局部病变部位途径、经血液循环途径及经脑脊液循环途径3种目的：综述经脑脊液途径移植神经干细胞或神经前体细胞的方式及其在治疗中枢神经系统疾病中的应用。 方法：检索Pubmed数据库和CHKD全文数据库2000至2009年相关文献,叙述脑脊液途径移植神经干细胞的方法、在动物实验和临床方面治疗中枢神经系统疾病的应用及治疗机制。结果与结论：脑脊液适宜神经干细胞的存活、增殖、分化,经脑脊液途径移植神经干细胞治疗中枢神经系统疾病是一种有效可行的方法。但因神经干细胞的来源问题及治疗的机制、最佳时间窗和数量、安全性等诸多问题,仍需更深更广的研究,从而为蛛网膜下腔注射神经干细胞治疗中枢神经系统疾病奠定坚实的基础。     

神经干细胞移植治疗研究进展

神经系统起源于胚胎神经管多能干细胞,而成人中枢神经系统内干细胞样前体细胞的发现为应用神经干细胞(NSCs)治疗各种神经系统疾病提供了前提条件。通过内源性NSCs增殖与迁移治疗各种神经元缺失疾病能力有限,而应用NSCs移植或有特定分化潜能的神经前体细胞移植治疗中枢神经系统损伤以及各种神经退行性疾病则可以促进患者神经功能恢复。目前,NSCs疗法所面临的主要问题是如何使NSCs定向分化替代特定功能的神经元,而NSCs疗法研究的热点是如何在分子水平调控NSCs的分化以及损伤区域NSCs的命运。     

神经干细胞脑内移植后的增殖、迁移与分化

成年哺乳动物中枢神经系统缺乏针对创伤反应产生新的细胞的能力 ,因此在受到各种损伤时 ,中枢神经的再生能力是非常有限的[1 ] 。神经干细胞能够自我更新 ,具有分化为中枢神经系统各种细胞包括神经元、星型胶质细胞和少突胶质细胞的能力[2 ] 。移植后的神经干细胞可以在受体脑内增殖、迁移和分化 ,与宿主的神经细胞形成新的神经环路 ,分泌特殊的递质和营养因子 ,在结构和功能上替代受损的神经元。因此 ,研究神经干细胞脑内移植后的增殖、迁移和分化对中枢神经系统疾病的细胞移植治疗具有重要的意义。本文就此研究进展综述如下。1　影响神经…     

神经干细胞植入阿尔茨海默病鼠脑后的效果观察

神经干细胞移植治疗阿尔茨海默病鼠包括细胞替代治疗和基因治疗,神经干细胞移植阿尔茨海默病鼠脑后其组织形态学与行为学效应均可以得到不同程度的修复和改善.细胞替代治疗中,神经干细胞与神经营养因子联合移植效应优于单纯的神经干细胞移植,但目前对神经干细胞体内分化机制的不确定导致了神经干细胞移植治疗的盲目性,同时对影响其疗效的各种可能因素也缺乏比较研究.神经干细胞基因治疗具有细胞替代和基因治疗的双重作用,但尚处于研究的初期阶段,仍以NGF,BDNF,GDNF等单一营养因子基因修饰的神经干细胞移植治疗为主,且转基因神经干细胞移植入阿尔茨海默病鼠脑后外源基因表达效率、促分化、功能修复情况以及安全性的研究还很缺乏.目前神经干细胞移植治疗阿尔茨海默病鼠脑后的疗效检测技术手段比较单一,免疫组化方法与活体示踪技术的结合、形态学指标与功能学指标的综合检测是疗效检测的发展趋势.     

Folate Deficiency Induces Neural Stem Cell Apoptosis by Increasing Homocysteine In Vitro

Cellular events for neural progenitor cells, such as proliferation and differentiation, are regulated by multiple intrinsic and extrinsic cell signals. Folate plays a central role in central nervous system development, so folate, as an extrinsic signal, may affect neural stem cell (NSC) proliferation and differentiation. In the present study, we investigated the effects of folate deficiency on the cell proliferation, cell apoptosis and homocysteine concentrations in NSCs. NSCs were isolated from fetal rats and identified as NSCs by their expression of immunoreactive nestin. Cell proliferation was quantitated by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. Apoptotic cells were detected and confirmed by flow cytometric analysis. We measured homocysteine concentrations in NSCs by high performance liquid chromatography and detected the expression of caspase-3 by western blot method. Folate deficiency not only decreased cell proliferation, but also increased the apoptotic rate of NSCs as demonstrated by the increased expression of early apoptotic markers such as caspase-3, compared to control group (p<0.05). Furthermore, There was a statistically significant increase in homocysteine concentration during folate deficiency in NSCs (p<0.05). These data suggest that folate affects the cell proliferation, apoptosis and homocysteine generation in NSC cells.     

Neural stem cells in the adult nervous system

The concept of the immutability of the nervous tissue has recently been replaced with the new idea that a continuous neurogenic turnover does occur in some limited areas of the central nervous system (CNS). At least two neurogenic regions of the adult mammalian CNS are involved in this process: the subventricular zone of the forebrain and the dentate gyrus of the hippocampus, which are considered to be a reservoir of new neural cells. Neural stem cells (NSCs) are multipotential progenitors that have self-renewal capability. While in vivo endogenous NSCs seem able to produce almost exclusively neurons, a single NSC in vitro is competent to generate neurons, astrocytes, and oligodendrocytes. NSCs lack a specific morphology and unambiguous surface markers that could allow their identification. For this reason, one of the major difficulties in identifying stem cells is that they are defined in terms of their functional capabilities, the determination of which might alter the cells' nature. The purpose of this review is to describe the characteristics of the NSCs of the adult mammalian CNS, their potentiality in terms of proliferation and differentiation capabilities, as well as their stability in long-term culture, all attributes that make them a good tool for tissue replacement therapies.     

神经干细胞在神经系统疾病中的应用

背景:神经干细胞的临床应用与基因治疗的有机结合可用于治疗多种神经系统遗传性和获得性疾病,但神经系统疾病多种多样,不同疾病神经组织内环境也不同,它们均影响神经干细胞的治疗效果.神经干细胞基因治疗中,如能同时转入多种外源性基因,让其相互作用及调节,就能更有效地促进神经组织不断再生.目的:文章就神经干细胞在神经系统疾病方面的应用进行综述.方法:应用计算机检索Medline数据库(2000-01/2009-08),以"Neural stem cells,Gene,Nervous system diseases"为检索词;应用计算机检索清华同方数据库(2000-01/2009-08),以"神经干细胞、基因、神经系统疾病"为检索词.结果与结论:计算机初检得到88篇英文文献,阅读标题和摘要进行初筛,排除因研究目的与此文无关的20篇,内容重复性的研究28篇,共保留40篇文献进行综述.神经干细胞作为一种新型的治疗手段,已用作供体细胞或转基因载体,对脑血管病、脑损伤性疾病、脊髓损伤、神经变性病、脑肿瘤、遗传代谢性疾病等治疗均取得了一定效果,但神经干细胞增殖、分化、迁移调控机制的基础性研究还有许多关键问题没有解决,不同疾病神经组织内的微环境亦影响着神经干细胞的治疗.     

离体实验条件下胞嘧啶脱氨酶基因修饰神经干细胞及其表达的观察

背景有关神经干细胞(neural stem cells,NSCs)治疗研究有明显增多趋势,已有许多研究认为NSCs能够增殖分化并在中枢神经系统内发生神经整合,NSCs在中枢神经系统中迁移的特性已成为近年来干细胞治疗神经系统疾病研究的热点.目的探讨胞嘧啶脱氨酶(CD)基因修饰神经干细胞及其基因表达.设计重复测量设计.单位解放军第三军医大学新桥医院神经外科、中南大学湘雅医院神经外科、Department of Urology,National Defense Medical College.材料新生第1天Wistar大鼠,由解放军第三军医大学实验动物中心提供.方法通过构建真核表达质粒pCMVCD,限制性内切酶消化鉴定后,采用Lipofectamine2000脂质体介导法转染新生大鼠室管膜下区NSCs,G418筛选阳性克隆,加入不同浓度的5-氟胞嘧啶(5-FC),MTT比色法测定NSCs的生存率.主要观察指标MTT比色法测定NSCs的生存率.结果本实验成功地培养并鉴定了神经干细胞,并将CD基因成功地转染了神经干细胞.基因转染使G418抗性细胞(NSCs/CD细胞)对5-FC高度敏感.未转染的NSCs对5-FC不敏感,IC50约为5 000 μmol/L,而转染基因后ICs0小于10 μmol/L.G418阳性NSCs对低浓度5-FC高度敏感.结论CD基因修饰神经干细胞的离体实验研究为干细胞治疗神经系统退行性病变及多种疾病研究提供依据.     

The therapeutic potential of adult neural stem cells

Neural stem cells (NSCs) are self-renewing, multipotent cells that generate the neuronal and glial cells of the nervous system. In mammals, contrary to long-held belief, neurogenesis occurs in the adult brain, and NSCs reside in the adult central nervous system. Thus, the brain may be amenable to repair following damage, and new avenues for cell-based therapy are being considered for the treatment of brain disease and injury, such as the stimulation of endogenous progenitor cells, the transplantation of adult-derived neural progenitor and stem cells, and, in particular, autologous cell transplantation. Although significant advances in this field have been made over the past decade, the adult NSC remains an elusive cell for study, and researchers are facing multiple challenges to the development of therapeutic applications from adult NSC research. Among these challenges are the identification and characterization of NSCs in vivo and in vitro, the understanding of the physiology of newly generated neuronal cells in the adult brain, the stimulation of endogenous progenitor cells to promote functional recovery, and the isolation and culture of homogenous populations of neural progenitor or stem cells from the adult brain for cell-based therapy.     

Neurotrophin 3 Transduction Augments Remyelinating and Immunomodulatory Capacity of Neural Stem Cells

Neural stem cells (NSCs) have therapeutic potential in experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS); however, to date, their use has resulted in only limited clinical and pathological improvement. To enhance their therapeutic capacity, in the present study, we transduced bone marrow–derived NSCs (BM-NSCs) with neurotrophin 3 (NT-3), a potent neurotrophic factor that is both neuroprotective and immunomodulatory. We found that BM-NSCs transduced with NT-3 reduced central nervous system (CNS) inflammation and neurological deficits in ongoing EAE significantly more than conventional NSC therapy, and, in addition, had the following advantages: (i) enhanced BM-NSC proliferation and differentiation into oligodendrocytes and neurons, as well as inhibited differentiation into astrocytes, thus promoting remyelination and neuronal repopulation, and reducing astrogliosis; (ii) enhanced anti-inflammatory capacity of BM-NSCs, thus more effectively suppressing CNS inflammation and accelerating remyelination; (iii) the easy accessibility of BM-NSCs provides another advantage over brain-derived NSCs for MS therapy; and (iv) a novel Tet-on system we used enables efficient control of NT-3 expression. Thus, our study provides a novel approach to break the vicious inflammation-demyelination cycle, and could pave the way to an easily accessible and highly effective therapy for CNS inflammatory demyelination.     

人脑神经干细胞体外培养中增殖、自我更新及多向分化潜能的观察

背景神经干细胞为神经系统损伤的修复开辟了新的途径,也是发现某些细胞因子和其作用机制的良好模型,神经干细胞的体外培养、增殖和诱导分化的探讨是这些研究的重要基础. 目的分离培养和鉴定人脑神经干细胞. 设计和方法采用无血清培养基分离和培养人胚胎脑皮质分离和培养神经干细胞,并应用免疫荧光技术进行细胞自我更新能力、巢蛋白表达和多向分化潜能的鉴定. 地点和材料实验在福建医科大学分子医学研究中心实施.实验材料为自然流产的孕龄 6～ 8周的人胚胎脑组织. 主要观察指标人脑神经干细胞在体外培养条件中的增殖、自我更新能力、巢蛋白表达和多向分化潜能. 结果人胚胎脑组织分离培养的细胞具有神经干细胞的特征,并可在体外大量增殖,经诱导可分化为神经元、星形胶质细胞和少突胶质细胞. 结论从人胚胎脑皮质成功分离培养出的神经干细胞,是研究神经干细胞诱导分化的良好模型.     

Robust In Vivo Transduction of Nervous System and Neural Stem Cells by Early Gestational Intra Amniotic Gene Transfer Using Lentiviral Vector

Presently, in vivo methods to efficiently and broadly transduce all major cell types throughout both the central (CNS) and peripheral adult nervous system (PNS) are lacking. In this study, we hypothesized that during early fetal development neural cell populations, including neural stem cells (NSCs), may be accessible for gene transfer via the open neural groove. To test this hypothesis, we injected lentiviral vectors encoding a green fluorescent protein (GFP) marker gene into the murine amniotic cavity at embryonic day 8. This method (i) efficiently and stably transduced the entire nervous system for at least 80% of the lifespan of the mice, (ii) transduced all major neural cell types, and (iii) transduced adult NSCs of the subventricular zone (SVZ) and subgranular zones (SGZs). This simple approach has broad applications for the study of gene function in nervous system development and adult NSCs and may have future clinical applications for treatment of genetic disorders of the nervous system.