Optimization of the Chemical-Physical Environments for Stem Cell Differentiation

描述（由申请人提供）：多能人胚胎干细胞（hESC）可以在体外分化为多种细胞类型，包括心血管细胞（心肌细胞、血管内皮细胞和平滑肌细胞）。体外控制干细胞生长和分化的能力对于成功应用分化细胞进行基于细胞的治疗至关重要。致力于所需谱系的适当细胞类型与心血管系统的相关元件一起可用于治疗心血管疾病。 所提出的关于确定控制hESC命运的最佳条件的研究使用了一种由全面的化学-物理微环境组成的新型中高通量微阵列平台。这些包括（1）固定化的生长因子（GF）和细胞外基质蛋白（ECM），（2）ECM的机械性质，和（3）作用于细胞的外部机械力。目前的知识表明，这些参数中的每一个（即，GF、ECMP、基质刚性以及机械负荷）在调节干细胞命运中起作用，但单独作用时效率较低。由于干细胞经历了多个微环境因素的影响，这些因素在发育阶段会发生变化，因此有必要研究多因素复杂性的组合效应。在拟议的研究中，我们将研究ECM蛋白（ECMP）和GF对hESCs（联邦批准的WA 01和WA 09细胞系）分化的组合影响。我们已经设计了一种水凝胶系统来控制基质的刚性，覆盖了在不同组织中遇到的范围，用于我们在干细胞培养中的ECMP阵列。我们还将把我们实验室开发的流动和拉伸装置结合到微阵列系统中，以评估外部机械力在ECMP/GF/Rigidity平台上调节hESC细胞命运中的作用。这种新的组合微阵列系统允许全面测试的化学-物理微环境的最佳条件的选择hESC的生长和分化。使用这种最佳选择的hESC转化为心血管组织工程将为显着推进人工血管，血管生成补丁以及心力衰竭细胞替代的发展提供机会，这反过来将改善心血管疾病患者的医疗保健和我们公民的福祉。 公共卫生相关性：多能人类胚胎干细胞可以分化成多种细胞类型，包括心血管系统中的细胞。具有相关血管细胞网络的适当细胞类型可用于治疗心脏、肺和血液疾病。该项目旨在开发一种新的系统方法来理解，定义并最终控制干细胞分化的过程，最终目标是开发治疗心血管疾病的再生医学工具。

DESCRIPTION (provided by applicant): Pluripotent human embryonic stem cells (hESCs) can be differentiated in vitro into multiple cell types, including cardiovascular cells (cardiomyocytes, vascular endothelial and smooth muscle cells). The ability to control the growth and differentiation of stem cells in vitro is essential for the successful application of differentiated cells for cell-based therapies. The appropriate cell types committed to a desired lineage, together with the relevant elements of the cardiovascular system, can be used to treat cardiovascular diseases. The proposed research on the identification of the optimal condition for controlling hESC fate uses a novel medium-high throughput microarray platform composed of comprehensive chemical-physical microenvironments. These include (1) immobilized growth factors (GFs) and extracellular matrix proteins (ECMPs), (2) mechanical properties of the ECM, and (3) external mechanical forces acting on the cells. Current knowledge indicates that each of these parameters (i.e., GFs, ECMPs, substrate rigidities, as well as mechanical loadings) plays roles in regulating stem cell fate, but the efficiency is low when acting alone. Since stem cells experience the influence of multiple microenvironmental factors which change during the developmental stages, it is essential to examine the combinatory effects of multi-factorial complexities of the niches, as proposed in the current research. In the proposed research, we will investigate the combinatorial effects of ECM proteins (ECMPs) and GFs on the differentiation of hESCs (Federally approved WA01 and WA09 cell lines). We have designed a hydrogel system to control the rigidities of matrices, covering a range encountered in different tissues, for our ECMP array in stem cell culture. We will also incorporate the flow and stretch devices developed in our lab into the microarray system to assess the roles of external mechanical forces in regulating the cell fate of hESCs on the ECMP/GF/Rigidity platform. This novel combinatory microarray system allows the comprehensive testing of the chemical-physical microenvironment for the choice of optimal conditions for hESC growth and differentiation. The use of such optimally chosen hESCs for translation to cardiovascular tissue engineering will provide the opportunity to significantly advance the development of artificial vessels, angiogenesis patches, as well as cell replacement for heart failure, which will in turn improve the healthcare of patients with cardiovascular diseases and the wellbeing of our citizens. PUBLIC HEALTH RELEVANCE: Pluripotent human embryonic stem cells can be differentiated into multiple cell types, including cells in the cardiovascular systems. The appropriate cell types with the associated networks of vascular cells can be used to treat heart, lung, and blood diseases. This project is aiming at developing a novel systematic approach to understand, define, and ultimately control the process of stem cell differentiation, with the ultimate goal of developing tools of regenerative medicine to treat cardiovascular diseases.