Neural Progenitor Grafting for Restorative Stroke Therapy

描述（由申请人提供）：本提案的总体目标是确定人胚胎干细胞（hESC）或诱导多能干细胞（iPSC）衍生的神经祖细胞（NPC）移植物改善实验性卒中后恢复的程度，并开始解决最终人类卒中治疗所需的关键安全性和有效性问题。hESC作为用于再生治疗的NPC来源提供了许多优点，例如容易获得的供应、巨大的分化潜力和易于遗传操作。iPSC提供了自体移植的额外优势，避免了对免疫抑制的需要。然而，对于这两种来源，理想的供体细胞类型和实现CNS再生所需的发育状态知之甚少。NPC移植是否能恢复中风后的功能，以及它可能恢复功能的机制也是未知的。我们已经开发了富集来源于hESC的特定NPC群体的方法，并且已经使用来源于人成纤维细胞的iPSC产生了NPC和多种神经元亚型。使用这些技术，我们提出测试以下假设：1）多能NPC（mpNPC）和神经元限制性前体（NRP）的纯化群体可以源自hESC或经由iPSC源自人类体细胞。这些群体在移植到完整或损伤的成年大鼠脑中后的迁移、分化和整合方面将不同;和2）在实验性中风后移植mpNPC或NRP将通过神经元替代或通过内源性NPC刺激修复直接增强功能恢复。具体目标1和2是使用基于启动子的报告基因或基于细胞表面抗原的选择来纯化和表征特定NPC群体。目的3是检查移植到成年大鼠脑的神经源性和非神经源性区域后这些NPC群体的行为，目的4是检查hESC和iPSC（人和大鼠）衍生的NPC移植物对实验性中风后恢复的影响。这些目标的进展将推进我们对移植因子如何影响hESC或iPSC衍生的NPC修复受损大脑和促进实验性中风后恢复的能力的认识，并将提供对这些疗法未开发的修复潜力和可能风险的深入了解。 公共卫生相关性：中风是一种常见的和潜在的破坏性神经系统疾病，没有经过证实的再生疗法。该提案旨在从人类胚胎干细胞或人类成人皮肤细胞的重编程中获得神经干细胞（NSC），并使用中风模型来确定用于脑修复性中风治疗的最佳NSC移植物。这一领域的进展为中风和其他脑损伤的新型细胞恢复疗法提供了进展。

DESCRIPTION (provided by applicant): The overall goals of this proposal are to determine the extent to which human embryonic stem cell (hESC)- or induced pluripotent stem cell (iPSC)-derived neural progenitor cell (NPC) grafts improve recovery after experimental stroke, and to begin addressing the critical safety and efficacy questions necessary for eventual human stroke therapy. hESCs offer many advantages as a source of NPCs for regenerative therapy, such as a readily available supply, vast differentiation potential and ease of genetic manipulation. iPSCs offer the added advantage of autologous grafting that obviates the need for immunosuppression. For either source, however, the ideal donor cell types and developmental states required to achieve CNS regeneration are poorly understood. Whether NPC grafting restores function after stroke and the mechanisms by which it might do so also are unknown. We have developed methods to enrich for specific NPC populations derived from hESCs, and have generated NPCs and multiple neuronal subtypes using iPSCs derived from human fibroblasts. Using these techniques, we propose to test the following hypotheses: 1) purified populations of multipotent NPCs (mpNPCs) and neuronal restricted precursors (NRPs) can be derived from hESCs or from human somatic cells via iPSCs. These populations will differ in their migration, differentiation and integration after transplantation into the intact or injured adult rat brain; and 2) grafting of mpNPCs or NRPs after experimental stroke will enhance functional recovery directly by neuronal replacement or by stimulating repair via endogenous NPCs. Specific Aims 1 and 2 are to purify and characterize specific NPC populations using promoter-based reporter or cell surface antigen-based selection. Aim 3 is to examine the behavior of these NPC populations after grafting into neurogenic and non-neurogenic regions of the adult rat brain, and Aim 4 is to examine the influence of hESC- and iPSC (human and rat)-derived NPC grafts on recovery after experimental stroke. Progress in these aims will advance our knowledge of how graft factors influence the capacity of hESC- or iPSC-derived NPCs to repair the injured brain and promote recovery after experimental stroke, and will provide insight into the untapped reparative potential and possible risks of these therapies. PUBLIC HEALTH RELEVANCE: Stroke is a common and potentially devastating neurological disorder with no proven regenerative therapies. This proposal aims to derive neural stem cells (NSCs) from human embryonic stem cells or reprogramming of human adult skin cells, and to use a stroke model to identify the optimal NSC grafts for brain reparative stroke therapy. Progress in this area offers advances toward novel cell-based restorative therapies for stroke and other brain insults.