Laboratory of Cellular and Molecular Neurobiology-Stem Cells, Department of Neurobiology, Hellenic Pasteur Institute, Athens, Greece.
Methods Mol Biol. 2020;2155:23-39. doi: 10.1007/978-1-0716-0655-1_3.
Stem cell transplantation has attracted great interest for treatment of neurodegenerative diseases to provide neuroprotection, repair the lesioned neuronal network and restore functionality. Parkinson's disease (PD), in particular, has been a preferred target because motor disability that constitutes a core pathology of the disease is associated with local loss of dopaminergic neurons in a specific brain area, the substantia nigra pars compacta. These cells project to the striatum where they deliver the neurotransmitter dopamine that is involved in control of many aspects of motor behavior. Therefore, cell transplantation approaches in PD aim to replenish dopamine deficiency in the striatum. A major challenge in developing cell therapy approaches is the ability to generate large numbers of transplantable cells in a reliable and reproducible manner. In recent years the technological breakthrough of induced pluripotent stem cells (iPSCs) has demonstrated that this is possible at a preclinical level, accelerating clinical translation. A second important issue is to efficiently differentiate iPSCs into dopaminergic neuronal progenitors with restricted proliferation potential in order to avoid cellular overgrowth in vivo and minimize the risk of tumorigenesis. Here we describe an effective protocol that includes human iPSC differentiation to the dopaminergic lineage and enrichment in neuronal precursor cells expressing the polysialylated form of the neural cell adhesion molecule PSA-NCAM, through magnetically activated cell sorting. The resulting cells are transplanted and shown to survive, differentiate, and integrate within a striatal lesion model generated by unilateral 6-hydroxydopamine administration in mice of the NOD/SCID strain that supports xenografts.
干细胞移植在治疗神经退行性疾病方面引起了极大的兴趣，它可以提供神经保护、修复受损的神经元网络并恢复功能。帕金森病（PD）尤其成为了首选目标，因为构成该疾病核心病理学的运动障碍与特定脑区（黑质致密部）中多巴胺能神经元的局部丧失有关。这些细胞投射到纹状体，在那里它们释放参与控制运动行为许多方面的神经递质多巴胺。因此，PD 中的细胞移植方法旨在补充纹状体中多巴胺的缺乏。开发细胞治疗方法的一个主要挑战是能够以可靠和可重复的方式生成大量可移植细胞。近年来，诱导多能干细胞（iPSCs）的技术突破表明，这在临床前水平上是可行的，加速了临床转化。第二个重要问题是有效地将 iPSC 分化为多巴胺能神经元祖细胞，其增殖潜力受到限制，以避免体内细胞过度生长，并最大限度地降低致瘤风险。在这里，我们描述了一种有效的方案，包括人 iPSC 向多巴胺能谱系的分化，以及通过磁激活细胞分选富集表达神经细胞黏附分子 PSA-NCAM 多聚唾液酸化形式的神经元前体细胞。所得细胞被移植并显示在单侧 6-羟多巴胺处理的 NOD/SCID 小鼠产生的纹状体损伤模型中存活、分化和整合，该模型支持异种移植物。
