Neurorestoration in Parkinson’s disease by cell replacement and endogenous regeneration

Parkinson’s disease (PD) is characterised by a continuous and selective loss of dopaminergic neurons in the substantia nigra pars compacta with a subsequent reduction of the neurotransmitter dopamine. Thus, the prospect of replacing the missing or damaged dopaminergic cells is very attractive. Possible regenerative therapies include transplanting developing neural tissue or neural stem cells into the degenerated host brain and inducing proliferation of endogenous stem cells by pharmacological manipulations. Neural stem cells, with the capacity to self renew and produce the major cell types of the brain, exist in the developing and adult CNS. These cells can be generated and expanded in vitro while retaining the potential to differentiate into nervous tissue. However, one major problem is the control of growth and differentiation of these cells. This review discusses new data on stem cell technology in cell replacement strategies in PD as well as endogenous dopaminergic regeneration.     

Quantitative evaluation of MRI-based tracking of ferritin-labeled endogenous neural stem cell progeny in rodent brain

Endogenous neural stem cells have the potential to facilitate therapy for various neurodegenerative brain disorders. To increase our understanding of neural stem and progenitor cell biology in healthy and diseased brain, methods to label and visualize stem cells and their progeny in vivo are indispensable. Iron oxide particle based cell-labeling approaches enable cell tracking by MRI with high resolution and good soft tissue contrast in the brain. However, in addition to important concerns about unspecific labeling and low labeling efficiency, the dilution effect upon cell division is a major drawback for longitudinal follow-up of highly proliferating neural progenitor cells with MRI. Stable viral vector-mediated marking of endogenous stem cells and their progeny with a reporter gene for MRI could overcome these limitations. We stably and efficiently labeled endogenous neural stem/progenitor cells in the subventricular zone in situ by injecting a lentiviral vector expressing ferritin, a reporter for MRI. We developed an image analysis pipeline to quantify MRI signal changes at the level of the olfactory bulb as a result of migration of ferritin-labeled neuroblasts along the rostral migratory stream. We were able to detect ferritin-labeled endogenous neural stem cell progeny into the olfactory bulb of individual animals with ex vivo MRI at 30 weeks post injection, but could not demonstrate reliable in vivo detection and longitudinal tracking of neuroblast migration to the OB in individual animals. Therefore, although LV-mediated labeling of endogenous neural stem and progenitor cells resulted in efficient and stable ferritin-labeling of stem cell progeny in the OB, even with quantitative image analysis, sensitivity remains a limitation for in vivo applications.     

The future of cell therapies in the treatment of Parkinson's disease

Parkinson's disease (PD) is a common neurological disorder of the brain which has as a part of its core pathology the progressive degeneration of the dopaminergic nigrostriatal pathway. Therefore, cell therapies that aim to restore this degenerated dopaminergic network represent a promising strategy in helping to cure PD. In this review, the authors start by discussing the progress of research on the use of fetal ventral mesencephalic (VM) tissue in transplantation therapies in PD, both from the clinical and experimental perspectives. Then the issues pertinent to its adoption in the clinic are discussed, including the ethical and practical problems with its use, the varied composition of VM tissue that is implanted with the graft and how this may account for some of the problems seen in the clinical trials using this tissue, especially graft-induced dyskinesia. Finally other promising sources of tissue for PD cell therapy are described, including mesenchymal and embryonic stem cells, before concluding on what is the best approach to the cellular repair of the parkinsonian brain.     

The future of cell therapies in the treatment of Parkinson's disease

Parkinson's disease (PD) is a common neurological disorder of the brain which has as a part of its core pathology the progressive degeneration of the dopaminergic nigrostriatal pathway. Therefore, cell therapies that aim to restore this degenerated dopaminergic network represent a promising strategy in helping to cure PD. In this review, the authors start by discussing the progress of research on the use of fetal ventral mesencephalic (VM) tissue in transplantation therapies in PD, both from the clinical and experimental perspectives. Then the issues pertinent to its adoption in the clinic are discussed, including the ethical and practical problems with its use, the varied composition of VM tissue that is implanted with the graft and how this may account for some of the problems seen in the clinical trials using this tissue, especially graft-induced dyskinesia. Finally other promising sources of tissue for PD cell therapy are described, including mesenchymal and embryonic stem cells, before concluding on what is the best approach to the cellular repair of the parkinsonian brain.     

低氧环境下白细胞介素1β诱导中脑源性神经干细胞的分化

背景：在体外某些外来信号的调控和诱导下,神经干细胞可定向诱导分化成与受体部位细胞类型与构成相似的细胞群体,使其更容易掺入靶组织,并可满足移植所需的细胞数量,提高移植细胞的存活率。目的：验证白细胞介素1β在低氧环境下体外诱导中脑源性神经干细胞的分化。方法：分离孕12d胎鼠中脑组织,培养神经干细胞、传代并鉴定。将鉴定后传代培养的中脑源性神经干细胞按分组接种于含体积分数10%胎牛血清的DMEM/F12培养基和含体积分数10%胎牛血清的DMEM/F12＋白细胞介素1β培养基;分别置于常氧（体积分数21%O2）和低氧（体积分数3%O2）环境下诱导分化,诱导分化9-11d后行小鼠抗大鼠酪氨酸羟化酶免疫细胞化学染色,检测酪氨酸羟化酶及神经元特异性烯醇化酶阳性细胞率。结果与结论：与常氧组比较,低氧组尤其是低氧＋白细胞介素1β组中酪氨酸羟化酶阳性神经元的突起数目更多,且长度更为延长。孕12d胚鼠腹侧中脑组织可以在体外培养传代、并能诱导分化成多巴胺能神经元;在低氧环境或低氧＋白细胞介素1β诱导下,分化率均高于常氧组,其表型更成熟。说明在低氧或低氧和白细胞介素1β诱导下,可明显促进中脑源性干细胞分化为形态及功能成熟的多巴胺能神经元。     

低氧环境下胶质源性神经营养因子体外诱导中脑神经干细胞的分化

背景:有效的神经干细胞体外增殖与多巴胺能神经元的定向诱导分化是神经干细胞移植治疗帕金森病的关键所在。目的:观察低氧条件下胶质源性神经营养因子体外诱导中脑源性神经干细胞向多巴胺能神经元的分化。方法:体外分离培养孕12d胚鼠腹侧中脑组织,制成单细胞悬液,在含碱性成纤维细胞生长因子和B27的无血清培养基中培养并传代,分别置于常氧(体积分数21%O2)或低氧(体积分数3%O2)环境下增殖5~7d后,接种于含体积分数10%胎牛血清的DMEM/F12培养基,或含体积分数10%胎牛血清的DMEM/F12+1μg/L胶质源性神经营养因子。结果与结论:低氧环境下分化10~12d,中脑神经干细胞向多巴胺能神经元分化均高于常氧组,在胶质源性神经营养因子诱导下向多巴胺能神经元分化比例更高,表型更成熟。说明低氧环境下胶质源性神经营养因子可明显促进中脑神经干细胞分化为数量足够、形态及功能成熟的多巴胺能神经元。     

低氧环境下胶质源性神经营养因子体外诱导中脑神经干细胞的分化

背景：有效的神经干细胞体外增殖与多巴胺能神经元的定向诱导分化是神经干细胞移植治疗帕金森病的关键所在.目的：观察低氧条件下胶质源性神经营养因子体外诱导中脑源性神经干细胞向多巴胺能神经元的分化.方法：体外分离培养孕12 d胚鼠腹侧中脑组织,制成单细胞悬液,在含碱性成纤维细胞生长因子和B27的无血清培养基中培养并传代,分别置于常氧（体积分数21%O2）或低氧（体积分数3%O2）环境下增殖5~7 d后,接种于含体积分数10%胎牛血清的DMEM/F12培养基,或含体积分数10%胎牛血清的DMEM/F12＋1 μg/L胶质源性神经营养因子.结果与结论：低氧环境下分化10~12 d,中脑神经干细胞向多巴胺能神经元分化均高于常氧组,在胶质源性神经营养因子诱导下向多巴胺能神经元分化比例更高,表型更成熟.说明低氧环境下胶质源性神经营养因子可明显促进中脑神经干细胞分化为数量足够、形态及功能成熟的多巴胺能神经元.     

ES cell therapy for Parkinson's disease

Neural transplantation, as a treatment for advanced Parkinson's disease (PD), has been studied for more than a decade due to the potential replacement of degenerated dopaminergic (DA) neurons. Several open-label studies on implantation of fetal nigral neurons revealed improvement in motor functions. However, the benefits were incomplete in double-blind trials. Progressive neural or embryonic stem (ES) cell research has raised hopes of creating novel cell replacement therapies for PD. DA neurons have been efficiently produced from primate ES cells in astrocyte-conditioned medium. Transplantation of neuronal stem cells derived from primate ES cells into a primate model of PD restored striatal DA function, suggesting ES cells are suitable donor cells.     

止颤汤干预神经干细胞移植帕金森病大鼠脑内多巴胺及其代谢产物的变化

背景：如何促进脑内多巴胺含量的增加以及减少多巴胺的代谢,是治疗帕金森病的热点所在。目的：从多巴胺代谢途径角度观察止颤汤对神经干细胞移植帕金森病大鼠的脑黑质中多巴胺及其代谢产物含量的变化。方法：以大鼠脑立体定位和1-甲基-4-苯基-1,2,3,6-四氢吡啶建立帕金森病大鼠模型。应用高效液相色谱法测定帕金森病大鼠中脑多巴胺及其代谢产物的含量。结果与结论：止颤汤可以提高神经干细胞移植后帕金森病大鼠中脑多巴胺及其代谢产物双羟苯乙酸的含量,但对代谢产物高香草酸无明显影响。通过促进帕金森病大鼠干细胞移植后神经干细胞的存活,使之定向分化为多巴胺能神经元并分泌多巴胺,同时抑制多巴胺分解达到治疗作用。     

止颤汤干预神经干细胞移植帕金森病大鼠脑内多巴胺及其代谢产物的变化

背景:如何促进脑内多巴胺含量的增加以及减少多巴胺的代谢,是治疗帕金森病的热点所在.目的:从多巴胺代谢途径角度观察止颤汤对神经干细胞移植帕金森病大鼠的脑黑质中多巴胺及其代谢产物含量的变化.方法:以大鼠脑立体定位和1-甲基-4-苯基-1,2,3,6-四氢吡啶建立帕金森病大鼠模型.应用高效液相色谱法测定帕金森病大鼠中脑多巴胺及其代谢产物的含量.结果与结论:止颤汤可以提高神经干细胞移植后帕金森病大鼠中脑多巴胺及其代谢产物双羟苯乙酸的含量,但对代谢产物高香草酸无明显影响.通过促进帕金森病大鼠干细胞移植后神经干细胞的存活,使之定向分化为多巴胺能神经元并分泌多巴胺,同时抑制多巴胺分解达到治疗作用.     

神经干细胞转基因移植治疗帕金森病的可能与可行

背景：神经干细胞分化为何种类型的神经细胞,不仅取决于细胞自身基因调控,更受所处环境中外来信号的影响。目的：就神经干细胞生物学特性、分化以及转基因移植治疗帕金森病的研究进展进行综述。方法：由第一作者应用计算机检索PubMed数据库1992-01/2009-12及中国期刊网全文数据库2003-01/2010-12有关神经干细胞生物学特性、分化以及转基因移植治疗帕金森病的文章,英文检索词为＂neural stem cell（NSCs）,neural stem cell transplant,Parkinson’s disease（PD）＂,中文检索词为＂神经干细胞,干细胞移植,帕金森病＂。排除重复性研究及Meta分析,共保留30篇文献进行综述。结果与结论：转基因神经干细胞移植治疗帕金森病是目前公认的最有前景的治疗方案。在低氧条件下转基因培养的神经干细胞,能高效地诱导分化为多巴胺能神经元,为移植治疗帕金森病提供细胞源。但神经干细胞的生物学特性、诱导分化等还有待进一步分析。同时应用于人体治疗的生物安全性问题也逐渐引起人们的重视,如致瘤性等问题均在进一步探索中。     

鼠胚神经干细胞移植对帕金森模型大鼠的治疗作用

背景:干细胞移植是治疗帕金森的有潜力的方法之一。目的:观察神经干细胞纹状体移植对帕金森模型大鼠旋转行为及脑内多巴胺含量的影响。方法:采用6-羟基多巴胺定点注射毁损黑质纹状体的方法构建帕金森大鼠模型;向造模成功的大鼠纹状体内分别移植1×106(共计20μL)的第3代胚鼠神经干细胞或等量生理盐水。结果与结论:神经干细胞移植后,帕金森大鼠的旋转行为明显改善。干细胞移植后3周,免疫组化检测发现移植干细胞的帕金森大鼠脑黑质部位酪氨酸羟化酶阳性细胞数增多,纹状体内可见酪氨酸羟化酶阳性细胞;荧光显微镜下观察发现Hoechst33324d标记神经干细胞在移植针道附近最为密集,并向远隔部位迁徙。干细胞移植后8周,高效液相色谱检测显示移植干细胞的帕金森大鼠纹状体内多巴胺含量明显增高(P<0.01)。说明神经干细胞脑内移植能够减轻6-羟基多巴胺引起的大鼠中脑黑质多巴胺能神经元的损伤,改善大鼠的旋转行为。     

VEGF-expressing human umbilical cord mesenchymal stem cells, an improved therapy strategy for Parkinson's disease

The umbilical cord provides a rich source of primitive mesenchymal stem cells (human umbilical cord mesenchymal stem cells (HUMSCs)), which have the potential for transplantation-based treatments of Parkinson's Disease (PD). Our pervious study indicated that adenovirus-associated virus-mediated intrastriatal delivery of human vascular endothelial growth factor 165 (VEGF 165) conferred molecular protection to the dopaminergic system. As both VEGF and HUMSCs displayed limited neuroprotection, in this study we investigated whether HUMSCs combined with VEGF expression could offer enhanced neuroprotection. HUMSCs were modified by adenovirus-mediated VEGF gene transfer, and subsequently transplanted into rotenone-lesioned striatum of hemiparkinsonian rats. As a result, HUMSCs differentiated into dopaminergic neuron-like cells on the basis of neuron-specific enolase (NSE) (neuronal marker), glial fibrillary acidic protein (GFAP) (astrocyte marker), nestin (neural stem cell marker) and tyrosine hydroxylase (TH) (dopaminergic marker) expression. Further, VEGF expression significantly enhanced the dopaminergic differentiation of HUMSCs in vivo. HUMSC transplantation ameliorated apomorphine-evoked rotations and reduced the loss of dopaminergic neurons in the lesioned substantia nigra (SNc), which was enhanced significantly by VEGF expression in HUMSCs. These findings present the suitability of HUMSC as a vector for gene therapy and suggest that stem cell engineering with VEGF may improve the transplantation strategy for the treatment of PD.     

神经干细胞移植治疗帕金森病的实验研究

目的:神经干细胞的发现为神经系统退行性疾病的治疗,提供了新的方法。通过碱性成纤维细胞生长因子(basicfibroblastgrowthfactor,bFGF)反应性神经干细胞的移植,探讨神经干细胞在帕金森病(Parkinson'dis-ease,PD)治疗中的作用。方法:体外用bFGF作为丝裂原培养E16大鼠中脑神经干细胞,经BrdU标记后植入PD大鼠纹状体,了解PD大鼠旋转行为的改善情况。结果:神经干细胞移植组PD大鼠旋转行为改善明显。结论:bFGF反应神经干细胞移植对PD有明显的治疗作用。     

Adult neural stem cells and repair of the adult central nervous system

Neural stem cells are present not only in the developing nervous systems, but also in the adult central nervous system of mammals, including humans. The mature central nervous system has been traditionally regarded as an unfavorable environment for the regeneration of damaged axons of mature neurons and the generation of new neurons. In the adult central nervous system, however, newly generated neurons from adult neural stem cells in specific regions exhibit a striking ability to migrate, send out long axonal and dendritic projections, integrate into pre-existing neuronal circuits, and contribute to normal brain functions. Adult stem cells with potential neural capacity recently have been isolated from various neural and nonneural sources. Rapid advances in the stem cell biology have raised exciting possibilities of replacing damaged or lost neurons by activation of endogenous neural stem cells and/or transplantation of in vitro-expanded stem cells and/or their neuronal progeny. Before the full potential of adult stem cells can be realized for regenerative medicine, we need to identify the sources of stem cells, to understand mechanisms regulating their proliferation, fate specification, and, most importantly in the case of neuronal lineages, to characterize their functional properties. Equally important, we need to understand the neural development processes in the normal and diseased adult central nervous system environment, which is quite different from the embryonic central nervous system, where neural development has been traditionally investigated. Here we will review some recent progress of adult neural stem cell research that is applicable to developmental neurobiology and also has potential implications in clinical neuroscience.     

神经干细胞向多巴胺能神经元分化机制的研究进展

帕金森病是中脑黑质的多巴胺能神经元缺失引起的慢性中枢神经系统功能失调。目前,应用神经干细胞在体内外分化成多巴胺能神经元是帕金森病细胞替代治疗的重要途径。本文综合近十年来的研究进展,对神经干细胞向多巴胺能神经元分化过程中重要的分子机制、信号通路以及外界影响因素进行综述。     

Molecular imaging in Parkinson's disease

Molecular imaging techniques using PET or SPECT have provided major insights into not only objective diagnosis of Parkinson's disease(PD), but also understanding the pathophysiological process in the disease progression. At disease onset, a compensatory hyperactivity of dopa decarboxylase in the nigrostriatal and extrastriatal dopaminergic pathways and upregulation of postsynaptic D2 receptor have been demonstrated. In the advanced stage, an excessively earlier release of dopamine from the residual neurons has been shown, suggesting a relationship with motor complications. In terms of therapy of PD, functional images have provided some objective evidences for possible neuroprotective effect of dopamine agonists, survival of fetal dopaminergic tissue grafted into patient's putamen, an increase of dopamine release by BDNF focal infusion therapy, and functional modification by deep brain stimulation. In vivo imaging of gene expression under developing may be informative in the future gene therapy in PD.     

Engraftment of human nasal olfactory stem cells restores neuroplasticity in mice with hippocampal lesions

Stem cell-based therapy has been proposed as a potential means of treatment for a variety of brain disorders. Because ethical and technical issues have so far limited the clinical translation of research using embryonic/fetal cells and neural tissue, respectively, the search for alternative sources of therapeutic stem cells remains ongoing. Here, we report that upon transplantation into mice with chemically induced hippocampal lesions, human olfactory ecto-mesenchymal stem cells (OE-MSCs) - adult stem cells from human nasal olfactory lamina propria - migrated toward the sites of neural damage, where they differentiated into neurons. Additionally, transplanted OE-MSCs stimulated endogenous neurogenesis, restored synaptic transmission, and enhanced long-term potentiation. Mice that received transplanted OE-MSCs exhibited restoration of learning and memory on behavioral tests compared with lesioned, nontransplanted control mice. Similar results were obtained when OE-MSCs were injected into the cerebrospinal fluid. These data show that OE-MSCs can induce neurogenesis and contribute to restoration of hippocampal neuronal networks via trophic actions. They provide evidence that human olfactory tissue is a conceivable source of nervous system replacement cells. This stem cell subtype may be useful for a broad range of stem cell-related studies.     

SD胎鼠脑内WWOX表达与神经干细胞及神经元发育相关性初探

目的探讨胚胎期SD胎鼠脑内WWOX表达与神经干细胞及神经元的发育的相关性。方法通过免疫组织化学染色、尼氏染色及Western blot等方法,检测不同发育时段胎鼠脑内WWOX的表达及神经干细胞和神经元的发育特点。结果尼氏染色及免疫组织化学染色结果显示,E11-E13胎鼠脑发育仍处于神经管阶段,神经管上皮组成细胞多为神经干细胞及神经元前体细胞,WWOX表达呈下降趋势;E15天后,神经干细胞多已分化成神经元及神经胶质细胞等,神经干细胞迅速减少,神经元及神经胶质细胞等细胞成分在脑内迅速发育、增多,此期在SD胎鼠脑内新生的神经胶质细胞及脉络组织中的WWOX表达呈缓慢上升趋势,并维持至成年。结论在胎鼠脑发育过程中WWOX与神经干细胞的分化存在一定的相关性,但与神经元的发育没有直接的相关性。     

神经干细胞与脑梗死康复

近年来的研究证实,成年中枢神经系统内广泛分布着神经干细胞(NSC),海马齿状回颗粒下层和侧脑室下区是成体大脑内源性NSC存在的主要脑区。NSC正常情况下处于静息状态,当大脑受到损伤或出现某些病理性变化时会被激活,分化为成熟的神经细胞,修复受损的神经功能。可以通过某些手段(如康复训练、丰富环境、局部应用外源性神经生长因子)激活自身干细胞或移植干细胞(包括NSC移植和非NSC移植)治疗脑梗死。