Osteoblast Lineage Progression from Embryonic Stem Cells

描述(申请人提供)：这项建议旨在通过提高我们对局部微环境在人类胚胎干细胞分化中的作用的理解，专门回应再生牙科医学中的RFA。我们将在RFA中概述的另一个关键概念来补充体外方法，即在体内设计的生物材料支架上研究HES细胞分化。我们研究的长期目标是开始了解指定HES细胞成骨细胞分化的信号。基于我们团队的研究兴趣和干细胞生物学知识的主要空白，我们将评估核心假设，即人胚胎干细胞向成骨细胞的谱系进化可以通过局部微环境的信号来控制。我们将首先确定滋养层和/或促成骨生长因子在体外引导HES细胞向成骨细胞样细胞方向发展的影响。在第二个目标中，我们计划使用谱系特异的转基因驱动超生命标记基因来分离、操纵和跟踪细胞的命运，以便将它们作为一个同源群体进行分离和研究。一旦确定了促进HES细胞分化的特定组织培养条件，我们将使用更具生理学相关性的模型系统，通过控制设计的生物材料上的微环境来研究体内人ES细胞的成骨细胞分化。在我们的第三个目标中，我们将通过将未分化的HES细胞和成骨细胞系选择的细胞移植到确定的体内位置来确定局部微环境信号如何在体内调节ES细胞的命运。 这些研究将进一步加深我们对正常发育过程中谱系分离生物学的理解，并为未来组织再生策略的发展提供必要的信息。

DESCRIPTION (provided by applicant): This proposal is designed to specifically respond to the RFA in Regenerative Dental Medicine by improving our understanding of the role of the local microenvironment on human embryonic stem (ES) cell differentiation. We will compliment in vitro approaches with another key concept outlined in the RFA, that is, the study of hES cell differentiation on designed biomaterial scaffolds in vivo. The long term goal of our research is to begin to understand the signals that specify osteoblast differentiation of hES cells. Based on the research interests of our team and the major gaps in knowledge of stem cell biology, we will evaluate the central hypothesis that the lineage progression of human ESCs to osteoblasts can be controlled by signaling from the local microenvironment. We will first determine the influence of the feeder layer and/or pro-osteogenie growth factors in directing lineage progression of hES cells to osteoblast-like cells in vitro. In the second aim, we plan to use lineage specific transgenes driving supravital marker genes to isolate, manipulate and follow the fate of cells so that they can be isolated and studied as a homogeneous population. Once specific tissue culture conditions are identified that promote bone cell differentiation from the hES cells, we will use a more physiologically relevant model system to study osteoblast differentiation of human ES cells in vivo by controlling the microenvironment on designed biomaterials. In our third aim we will determine how local microenvironmental cues regulate the fate of ES cells in vivo by transplanting both undifferentiated hES cells and osteoblast lineage-selected cells to defined in vivo locations. These studies will further our understanding of the biology of lineage segregation during normal development and provide essential information for the development of future tissue regeneration strategies.