RHESUS ES CELLS: A MODEL TO STUDY PANCREAS DEVELOPMENT

恒河猴 ES 细胞：研究胰腺发育的模型

• 批准号: 6381948
• 负责人: Jon S Odorico
• 金额: $ 14.4万
• 依托单位: UNIVERSITY OF WISCONSIN-MADISON
• 依托单位国家: 美国
• 财政年份: 2000
• 资助国家: 美国
• 起止时间: 2000-05-15 至 2003-04-30
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/6381948
• 关键词: Macaca mulatta; animal tissue; biological models; cell differentiation; embryonic stem cell; gene expression; histogenesis; in situ hybridization; laboratory mouse; model design /development; northern blottings; pancreas; polymerase chain reaction

A comprehensive knowledge of the genetic programs and embryonic inductive events regulating human pancreas development would advance our understanding of congenital pancreatic malformations and aid in developing rational treatment strategies for diabetes mellitus. Significant insights to the mechanisms of pancreatic organogenesis have been gained through recent experiments in mice and chickens. However, critical differences in early developmental pathways between these species and humans places the direct applicability of these data to human development in question. Furthermore, functional studies in human development are restricted by the ethical implications of directly manipulating human embryos. Consequently, a more "human" model based on nonhuman primates would have broad application for studying human development. Recently, embryonic stem (ES) cell lines from nonhuman primates (rhesus macaque and common marmoset) have been derived and their potential for differentiation into derivatives of all three embryonic germ layers (ectoderm, mesoderm, and endoderm) in vivo has been characterized. Their capacity for lineage-restricted differentiation in vitro, however, has not been thoroughly tested to date. in contrast, neural, hematopoietic and muscle lineage differentiation of mouse ES cells in vitro has been extensively studied. Recently, the ability of mouse ES cells to undergo elements of endoderm and pancreatic differentiation in culture has also been demonstrated. Therefore, we will test the hypothesis that pancreatic endocrine and/or exocrine lineage specification can be recapitulated in rhesus ES cells induced to differentiate. The specific aims of this proposal are: l) To determine the expression pattern of endoderm related genes as well as pancreatic exocrine/endocrine-specific genes in rhesus ES cells differentiating in vitro, and 2) To determine the potential of rhesus ES cells for pancreatic lineage differentiation in ES cell derived tumors in immunocompromised mice. A combination of RT-PCR analysis, Northern hybridization, ribonuclease protection assay, in situ hybridization, and immunofluorescence studies will be used to characterize the pattern of pancreatic gene expression. These studies will provide a basis for the refinement of an in vitro primate model of human pancreatic development. Such a model system would have important implications for investigating the mechanisms controlling pancreatic differentiation and for developing new cell-based strategies for treating diabetes.

全面了解调控人类胰腺发育的遗传程序和胚胎诱导事件将促进我们对先天性胰腺畸形的理解，并有助于制定合理的糖尿病治疗策略。通过最近在小鼠和鸡中的实验，已经获得了对胰腺器官发生机制的重要见解。然而，这些物种和人类之间早期发育途径的关键差异使这些数据对人类发育的直接适用性受到质疑。此外，人类发育的功能研究受到直接操纵人类胚胎的伦理影响的限制。因此，基于非人类灵长类动物的更“人性化”的模型将在研究人类发展方面具有广泛的应用。最近，胚胎干（ES）细胞系，从非人灵长类动物（恒河猴和普通绒猴）已衍生和分化成所有三个胚胎胚层（外胚层，中胚层和内胚层）在体内的衍生物的潜力已被表征。然而，迄今为止，它们在体外进行谱系限制分化的能力还没有得到彻底的测试。相反，小鼠ES细胞在体外的神经、造血和肌肉谱系分化已被广泛研究。最近，小鼠ES细胞在培养中经历内胚层和胰腺分化的能力也已被证明。因此，我们将测试的假设，胰腺内分泌和/或外分泌谱系规范可以概括在恒河猴ES细胞诱导分化。本研究的具体目的是：1）确定内胚层相关基因以及胰腺外分泌/内分泌特异性基因在体外分化的恒河猴ES细胞中的表达模式; 2）确定恒河猴ES细胞在免疫受损小鼠ES细胞源性肿瘤中胰腺谱系分化的潜力。RT-PCR分析、北方杂交、核糖核酸酶保护试验、原位杂交和免疫荧光研究的组合将用于表征胰腺基因表达的模式。这些研究将为完善人类胰腺发育的体外灵长类动物模型提供基础。这样的模型系统对于研究控制胰腺分化的机制和开发治疗糖尿病的新的基于细胞的策略具有重要意义。