Derivation and Characterization of Baboon ES Cells

描述（由申请人提供）： 干细胞及其作为治疗人类复杂、衰弱性疾病或病症的新方法的潜力，引发了人们对这一领域研究的前所未有的兴趣。考虑到以细胞为基础的新型治疗方法对复杂疾病和残疾的潜力，这种关注是当之无愧的，迄今为止，其他方法未能提供足够的解决方案。此外，干细胞表现出许多对细胞功能至关重要的基本生物学过程，包括自我更新和/或分化。这些相同的过程是正常器官和组织的发育、分化和功能的基础，并且这些过程中的异常是许多严重疾病的根源，包括癌症、神经退行性疾病、影响心脏和血液系统的疾病以及许多其他使人衰弱的病症。然而，用人类胚胎干（ES）细胞进行实验的能力目前受到联邦法规的限制，并且即使在没有这样的法规的情况下，在任何尝试将基于干细胞的治疗方法应用于人类之前，仍然迫切需要在相关动物模型中研究ES细胞的生物学、功能和可操作性。在这个应用程序中，我们建议启动实验，开发狒狒作为一个新的模型系统的灵长类动物胚胎干细胞的研究。狒狒作为非人灵长类系统的模型，为ES细胞生物学研究和与人类高度相关的应用提供了许多潜在的优势。因此，在这项R21申请中，我们提出了三个目标- 1）从通过辅助生殖技术体外产生的狒狒胚胎中衍生至少5个新的ES细胞系，2）在细胞水平上表征每个狒狒ES细胞系，包括生长特征、染色体组型、自我更新能力、多能细胞特征性标志物的表达、在培养物中进行定向分化的能力，和在体内经历自发分化的能力，和3）在分子水平上表征每种狒狒ES细胞系，包括与未分化ES细胞中多能性相关的基因的表达，在体外诱导定向分化时多能性基因表达的抑制，以及两个示例性印迹基因的亲本特异性单等位基因表达和等位基因特异性差异DNA甲基化（H19和Snrpn）在未分化和分化的ES细胞中的表达。

DESCRIPTION (provided by applicant): Stem cells and the potential they hold for novel approaches to treating complex, debilitating diseases or conditions in humans have sparked unprecedented interest in research in this area. This attention is well deserved given the potential for novel cell-based therapeutic approaches to complex diseases and disabilities for which other approaches have, to date, failed to offer sufficient solutions. In addition, stem cells demonstrate a number of basic biological processes that are central to cellular function, including self-renewal and/or differentiation. These same processes underlie the development, differentiation and function of normal organs and tissues, and abnormalities in these processes are at the root of many severe diseases including cancer, neurodegenerative diseases, diseases affecting the heart and blood systems, and many other debilitating conditions. However, the ability to conduct experiments with human embryonic stem (ES) cells is currently limited by federal regulations, and even in the absence of such regulations, there remains a compelling need for research on the biology, function, and manipulability of ES cells in a relevant animal model prior to any attempt to apply stem cell-based therapeutic approaches to humans. In this application we propose to initiate experiments to develop the baboon as a new model system for studies of primate embryonic stem cells. Baboons offer a number of potential advantages as a model nonhuman primate system for studies of ES cell biology and applications that will be highly relevant to humans. Thus, in this R21 application, we propose three aims - 1) To derive at least 5 new ES cell lines from baboon embryos produced in vitro by assisted reproductive technologies, 2) To characterize each baboon ES cell line at the cellular level, including growth characteristics, karyotype, ability to self-renew, expression of markers characteristic of pluripotent cells, ability to undergo directed differentiation in culture, and ability to undergo spontaneous differentiation in vivo, and 3) To characterize each baboon ES cell line at the molecular level, including expression of genes associated with pluripotency in undifferentiated ES cells, suppression of pluripotency gene expression upon induction of directed differentiation in vitro, and both parent-of-originspecific monoallelic expression and allele-specific differential DNA methylation of two exemplary imprinted genes (H19 and Snrpn) in both undifferentiated and differentiated ES cells.