Nonhuman primate induced pluripotent stem cells in regenerative medicine

描述(由申请人提供)： 再生医学的目的是恢复因疾病或衰老而受损的器官的健康功能。一个主要问题是用于再生医学的细胞的来源。人们通常认为使用来自患者本人的细胞是可取的，因为这是为了避免需要给药来抑制移植细胞的免疫排斥反应。诱导多能干细胞(iPS细胞)的发现极大地扩展了在再生医学中使用患者特有细胞的可能性。IPS细胞可以来自任何体细胞，但具有胚胎干细胞的特性。就像胚胎细胞一样，它们可以用来生成再生医学中可能需要的任何身体细胞。这些研究的最终目标是从单个动物中提取iPS细胞，并在iPS细胞定向分化为特定细胞系后将其移植到供体动物体内。在这样的研究成为可能之前，需要进行适当的体外研究。一个主要问题涉及捐赠者年龄的潜在影响。大多数可能接受这种治疗的患者都是年长的，或者至少是成熟的成年人。非人灵长类动物(NHP)为iPS细胞及其分化后代的基础和临床前研究提供了一些优势，包括供体年龄的作用。这些优势包括与人类的遗传联系，以及类似发育的中枢神经系统。常见的绒猴(Callithrix Jacchus)作为一种小的、易于操作的NHP，已经被广泛地验证为生物医学模型。在这项研究中，我们解决了在NHP开始细胞移植研究之前必须回答的几个基本问题。我们问：(1)供体的年龄是否改变了来源的iPS细胞的特性，从而削弱了它们作为多能细胞的特性？在免疫缺陷小鼠体内，细胞是否正常分化，移植后细胞是否存活并进一步分化？我们将从三个年龄组的NHP中分离并鉴定iPS细胞。在多能性测试之后，我们将评估每个iPS细胞系分化为神经前体细胞的能力。为了提供正确分化的常规测试，我们将把神经干细胞移植到免疫缺陷小鼠的中枢神经系统中。(2)捐献者的年龄是否增加了存在代表畸胎瘤发展风险的分化耐受细胞亚群的可能性？我们将检查小鼠中枢神经系统中是否存在移植细胞的长期持续增殖。预计从长远来看，无论捐赠者的年龄如何，适当的分化将使增殖率降至微不足道的水平。(3)供者的年龄是否影响供者的免疫细胞对IPSC来源的细胞的预期识别为自体还是非自体？预计来自一个捐赠者的细胞在体外会与来自无关个人的免疫细胞发生反应，但不会与来自同一个人的免疫细胞发生反应；这种正常关系在IPSC来源的分化细胞中是否保持着正常的关系，而不考虑捐赠者的年龄？总而言之，这些研究将使未来再生医学的实施能够提出更有效的细胞疗法。如果研究发现与年龄有关的缺陷，将需要纠正此类缺陷的策略。然而，如果来自老年NHP捐赠者的iPS细胞在所有方面都与年轻捐赠者产生的iPS细胞功能相同，那么自体细胞治疗的体内测试将在NHP模型中进行。未来的研究将涉及将来自iPS细胞的分化细胞移植回捐赠者动物体内，以长期测试IPSC来源的细胞的治疗潜力和安全性。

DESCRIPTION (provided by applicant): The aims of regenerative medicine are to restore healthy function to organs damaged by disease or aging. A major issue is the source of cells to be used in regenerative medicine. It is often thought to be desirable to use cells derived from the patient himself/herself, because this is hypothesized to avoid the need to administer drugs to suppress immune rejection of the transplanted cells. The possibility of using patient- specific cells in regenerative medicine was greatly expanded by the discovery of induced pluripotent stem cells (iPS cells). iPS cells can be derived from any somatic cell, but have the properties of embryonic stem cells. Like embryonic cells, they can be used to generate any cell of the body that may be needed in regenerative medicine. The eventual goal of these studies is to derive iPS cells from individual animals and implant the cells into the donor animal following the directed differentiation of the iPS cells to specific cell lineages. Before such studies are possible, appropriate in vitro investigations are needed. A major question concerns the potential impact of donor age. Most patients who would be candidates for this form of therapy would be older, or, at least, mature adults. Nonhuman primates (NHPs) offer several advantages for basic and preclinical studies of iPS cells and their differentiated progeny, including the role of donor age. These advantages include genetic relatedness to humans, and similarly developed central nervous systems. The common marmoset (Callithrix jacchus), as a small, easily handled NHP, has been extensively validated as a biomedical model. In this research, we address several basic questions that must be answered before cell transplantation studies commence in NHPs. We ask: (1) Does the age of the donor change the properties of the derived iPS cells, impairing their properties as pluripotent cells? Does differentiation occur normally, and do the cells survive and undergo further differentiation following implantation in immunodeficient mice? We will derive and characterize iPS cells from NHPs of three age groups. Following tests of pluripotency, we will assess the ability of each iPS cell line to differentiate to neural progenitor cells (NPCs). To provide a robut test of proper differentiation, we will transplant NPCs into the CNS of immunodeficient mice. (2) Does the age of the donor increase the probability of the existence of a differentiation-resistant subpopulation of cells that could represent a risk for teratoma development? We will examine if there is long-term continued proliferation of transplanted cells in the mouse CNS. It is expected that appropriate differentiation will have reduced the rate of proliferation to negligible levels oer the long term, irrespective of donor age. (3) Does the age of the donor affect the expected recognition of the iPSC-derived cells as self versus nonself by immune cells of the donor? It is expected that the cells from one individual donor will elicit a reaction in vitro with immune cells from unrelated individuals, but not with immune cells from the same individual; is this normal relationship maintained in iPSC-derived differentiated cells, irrespective of donor age? Collectively, these studies will enable more effective cell therapy to be proposed in future implementations of regenerative medicine. If the studies reveal age-related defects, strategies to correct such defects will be required. However, if iPS cells generated from older NHP donors function equivalently to those from younger donors in all respects, in vivo tests of autologous cell therapy will proceed in the NHP model. Future studies would involve the transplantation of differentiated cells derived from iPS cells back into the donor animal, in long-term tests of the therapeutic potential and safety of iPSC-derived cells.