Differentiation of human pluripotent stem cells into dopaminergic neurons: security risk for heterogeneity

doi:10.3969/j.issn.2095-4344.1532

Abstract

Abstract:

BACKGROUND: The transplantation of human pluripotent stem cell-derived nerve cells for the treatment of Parkinson’s disease has made great progress and the relevant research is at the subclinical stage. However, there are still many problems to be solved. As one of the existing problems, heterogeneity of differentiation can cause tumor formation and dyskinesia in the host after transplantation, which brings potential and unpredictable risks to patients.
OBJECTIVE: To summarize the current status of human pluripotent stem cells differentiated into dopaminergic neurons, the principle and process of differentiation design, the induction process and the research status of heterogeneous differentiation, laying a theoretical foundation for the clinical transformation of transplantation therapy.
METHODS: The keywords were “iPSC AND Parkinson’s disease, induced pluripotent stem cells AND Parkinson’s disease, ES cells AND Parkinson’s disease, embryonic stem cells AND Parkinson’s disease, pluripotent stem cells AND Parkinson’s Disease” in English and Chinese, respectively. Relevant articles published from 1980 to 2018 were retrieved by the first author in the PubMed, CNKI, and WanFang. Literature addressing the treatment of Parkinson’s disease with induced pluripotent stem cell-derived neurons in recent years was reviewed, and finally 46 articles were retained.
RESULTS AND CONCLUSION: A variety of induction differentiation protocols can be used to induce the differentiation of pluripotent stem cells into A9 dopaminergic neurons. Cell transplantation can promote the behavioral and limb function recovery of different Parkinson's disease models. However, in addition to the differentiation of A9 dopaminergic neurons, the current differentiation protocol products include different neurons such as A10 dopaminergic neurons and serotonergic neurons. There is no differentiation scheme that can achieve uniform differentiation. Further optimizing the differentiation conditions of human pluripotent stem cells in vitro and homogenizing differentiation into A9 dopaminergic neurons may further improve the behavioral performance of Parkinson’s disease model animals and promote the clinical transformation of the cell therapy.

中国组织工程研究杂志出版内容重点：干细胞；骨髓干细胞；造血干细胞；脂肪干细胞；肿瘤干细胞；胚胎干细胞；脐带脐血干细胞；干细胞诱导；干细胞分化；组织工程

Key words: Parkinson Disease, Multipotent Stem Cells, Dopamine, Neurons, Tissue Engineering

CLC Number:

R459.9

Chen Li, Hu Lan, Peng Yanan, Yang Liu, Shen Hui, Wang Tan, Zhao Zhenqiang. Differentiation of human pluripotent stem cells into dopaminergic neurons: security risk for heterogeneity[J]. Chinese Journal of Tissue Engineering Research, 2019, 23(1): 118-124.

Figures/Tables 1

2.1 帕金森病的细胞替代疗法现状胚胎腹侧中脑组织中含有多巴胺能神经祖细胞，其移植治疗帕金森病患者的临床研究已经开展了30多年。尽管临床结局好坏不一，但这些临床研究已证明细胞替代疗法的确是一种非常有希望的治疗策略，成为持续的研究热点[10]。然而由于伦理问题，难以得到大量的胎儿中脑组织，存在移植物质量参差不齐，难以标准化等问题，胚胎腹侧中脑细胞移植很难大规模开展。人多能干细胞(human pluripotent stem cells，hPSCs)包括人胚胎干细胞(human embryonic stem cells，hESCs)和人诱导多能干细胞(human induced pluripotent stem cells，hiPSCs)，均具有自我更新和分化成多种人类细胞类型(包括中脑多巴胺能神经元)的潜能[11]。因此，人多能干细胞为细胞替代疗法的开展提供了有价值的资源[12-13]。通常情况下，人多能干细胞在体内分化的不可控性，并不适合直接移植。此外，神经干细胞倾向于分化为中间神经元类型和星形胶质细胞，体外很难产生足够数量的多巴胺能神经元，因此从治疗的安全性和有效性综合考虑，移植已经定向的多巴胺能祖细胞更具可行性[14]。Kikuchi小组[15]的实验显示无饲养层和无血清条件下由人诱导多能干细胞分化衍生的多巴胺能神经元，在1-甲基-4-苯基-1，2，3，6-四氢吡啶(1-Methyl-4-phenyl-1，2，3，6-tetrahydropyridine，MPTP)损伤的帕金森病猴体内存活6个月。该报道支持人诱导多能干细胞对未来临床试验的治疗潜力。Kirkeby小组[16]则使用优化的方案分化人胚胎干细胞，将衍生的多巴胺能神经元移植入纹状体内，能改善6-羟基多巴胺诱导的半帕金森病大鼠运动功能。同时多个研究小组证实，通过特定的诱导方案可将人多能干细胞分化为A9多巴胺能神经元[17-19]。移植后的多巴胺能神经元可以在动物模型脑内存活、神经支配，促进功能修复[20-21]，而没有产生肿瘤[22]。2017年中国科学院周琪院士主持的“人胚胎干细胞来源的神经祖细胞治疗帕金森病”的临床前期研究在郑州大学第一附属医院正式开展，表明干细胞替代疗法已经十分接近临床。 2.2 当前人多能干细胞分化为中脑多巴胺能神经元诱导方案的设计基本原理及机制 2.2.1 中脑多巴胺能神经元发育的前后模式和背腹模式在胚胎发育过程中，发育模式和细胞命运特异性分化受到局部分泌的形态发生素和生长因子的调节。神经的形成是一个发展的过程。它从外胚层开始，神经上皮层或神经板折叠形成神经管，大脑和脊髓从该神经管发育而来。神经管沿着延喙尾轴和背腹轴生长，此发育模式在时间和空间上受沿前后模式和背腹模式轴分布的形态发生素浓度梯度的协调控制。此过程中，细胞分化由形态发生因子和邻近细胞分泌的信号分子诱导[23]。影响前后模式的形态发生素包括成纤维细胞生长因子、WNT和视黄酸，经典WNT/ β-catenin信号通路在其中扮演了关键角色。影响背腹模式的成分包括WNT、骨形态发生蛋白和音猬因子，SHH信号通路起到了决定性作用。沿前后模式和背腹模式轴分布的形态发生素浓度梯度限定了转录密码和特定区域神经前体细胞的身份，见图2。形态发生素浓度轻微的变化就可以引起细胞命运的转变[24]。这两种模式是体外方案中调控WNT信号通路和SHH信号通路可以决定中脑神经前体表型的理论依据。"

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