Nanopatterned Surfaces to Control Cell Fate

控制细胞命运的纳米图案表面

• 批准号: 7792209
• 负责人: KEVIN Edward HEALY
• 金额: $ 33.01万
• 依托单位: UNIVERSITY OF CALIFORNIA BERKELEY
• 依托单位国家: 美国
• 财政年份: 2009
• 资助国家: 美国
• 起止时间: 2009-04-01 至 2013-03-31
• 项目状态: 已结题
• 来源: https://reporter.nih.gov/project-details/7792209
• 关键词: Address; Adherent Culture; Adhesions; Animals; Area; Behavior Control; Biological; Cell Adhesion; Cell Culture Techniques; Cell Fate Control; Cell Shape; Cell Survival; Cells; Clinical; Collection; Conditioned Culture Media; Congestive Heart Failure; Culture Media; Development; Diabetes Mellitus; Disadvantaged; Disease; Embryo; Engineering; Environment; Exhibits; Extracellular Matrix Proteins; Focal Adhesions; Growth; Human; In Situ; Integrins; Ligands; Methods; Modeling; Morphology; Mus; Nuclear; Parkinson Disease; Patients; Peptides; Pharmaceutical Preparations; Phenotype; Production; Protein Biosynthesis; Protocols documentation; Regenerative Medicine; Reproducibility; Risk; Screening procedure; Shapes; Signal Transduction; Site; Source; Spinal cord injury; Stem cells; Surface; System; Technology; Testing; Tissue Engineering; Tissues; Treatment Efficacy; Zoonoses; base; cell type; chemotherapy; density; human embryonic stem cell; human embryonic stem cell line; large scale production; leukemia; nanopattern; neuronal cell body; novel; pathogen; physical state; public health relevance; self-renewal; stem; stem cell differentiation; stem cell population; transmission process

DESCRIPTION (provided by applicant): Human embryonic stem (hES) cells are being studied as potential source of cells for the treatment of many diseases (e.g. diabetes, spinal cord injury, Parkinson's, leukemia, congestive heart failure, etc.). These same cells are also being touted an ideal cell source for ex vivo tissue engineering or in situ regenerative medicine. The successful integration of hES cell into such therapies will hinge upon three critical steps: 1) stem cell expansion in number without differentiation (i.e., self-renewal); 2) directed differentiation into a specific cell type or collection of cell types; and, 3) cell survival and promotion of their functional integration into existing tissue. Precisely controlling each of these steps will be essential to maximize the hES cell's therapeutic efficacy. However, it is difficult to precisely control the behavior of hES cells, since environmental conditions for self-renewal and differentiation are poorly understood. We propose to develop a tunable completely synthetic surface and chemically defined media to control the self-renewal/expansion of hES cells. If hES cells can be derived and maintained within a completely synthetic environment, then it will be possible to eliminate pathogen transmission associated with animal-derived materials, provide a scalable basis for large-scale production of hES cells, and provide a precise base for further development to control hES cell differentiation. This application will develop materials to address the hypothesis that the contractile state of a hES cell, manifested by nuclear shape morphology via integrin engagement, regulates hES cell self-renewal. Our hypothesis is centered on a common mechanism by which cells respond differentially to either materials with variable moduli or materials that spatially confine a cell's shape via adhesion site distribution. We propose that a common mechanism that controls hES cell self-renewal and cell fate determination is the contractile state of the cell manifested by nuclear morphology, and integrin engagement and clustering. Thus, we wish to explore the spatial arrangement of cell adhesion domains (i.e., their size, number/cell body, and spatial arrangement) and assess their effect on the self-renewal of hES cells. We propose that altering the physical state of a pluripotent hES cell, via spatial clustering of its adhesions with a surface, will influence self-renewal and differentiation to a specific phenotype. The following specific aims are proposed. Specific Aim 1: To develop and characterize nanopatterned cell culture substrata where the size, peptide ligand density, number/cell body, and spatial arrangement of integrin-engaging domains will be varied to control cell and colony morphology. Specific Aim 2: To evaluate the nanopatterned substrata to support the long-term growth (5-10 passages) of human ES cells in chemically-defined media. PUBLIC HEALTH RELEVANCE: This application will focus specifically on engineering a tunable and well-defined environment presenting hES cells with a completely synthetic cell culture surface and chemically-defined media to promote self-renewal. The result will be a synthetic microenvironment that can both serve as a regenerative medicine technology platform for large scale hES cell expansion, as well as provide a novel and highly modular system for dissecting basic signaling mechanisms underlying hES cell self-renewal.

描述（由申请人提供）：正在研究人类胚胎（HES）细胞作为治疗许多疾病的潜在细胞来源（例如糖尿病，脊髓损伤，帕金森氏症，白血病，充血性心力衰竭等）。这些相同的细胞也被吹捧为离体组织工程或原位再生医学的理想细胞来源。将HES细胞成功整合到此类疗法中将取决于三个关键步骤：1）干细胞在没有分化的数量（即自我更新）的数量上扩展； 2）将分化为特定的细胞类型或细胞类型的集合； 3）细胞存活和促进其功能整合到现有组织中。精确控制这些步骤中的每个步骤对于最大化HES细胞的治疗功效至关重要。但是，很难精确控制HES细胞的行为，因为对自我更新和分化的环境条件知之甚少。我们建议开发一个完全合成的表面和化学定义的培养基，以控制HES细胞的自我更新/扩展。如果可以在完全合成的环境中得出并维持HES细胞，那么可以消除与动物衍生材料相关的病原体传播，为大规模生产HES细胞提供可扩展的基础，并为进一步的发展提供了控制HES细胞分化的精确基础。该应用将开发材料，以解决以下假设：HES细胞的收缩状态通过整合素的参与表现出核形状的形态，从而调节HES细胞自我更新。我们的假设以一种共同的机制为中心，通过该机制，细胞对具有可变模量的材料或通过粘附位点分布在空间上限制细胞形状的材料的材料有所不同。我们建议，控制HES细胞自我更新和细胞命运确定的一种共同机制是由核形态以及整合素的参与和聚类表现出的细胞的收缩状态。因此，我们希望探索细胞粘附域的空间排列（即它们的大小，数量/细胞体和空间排列），并评估其对HES细胞自我更新的影响。我们建议，通过其表面的空间聚类来改变多能HES细胞的物理状态，将影响自我更新和分化为特定表型。提出了以下特定目标。特定目的1：开发和表征纳米图案的细胞培养基质，其中大小，肽配体密度，数量/细胞体和整联蛋白启动结构域的空间排列将在控制细胞和菌落形态上有所不同。具体目的2：评估纳米图案的底层以支持化学定义培养基中人类ES细胞的长期生长（5-10段）。公共卫生相关性：该应用将专门针对一个可调且定义明确的环境，以完全合成的细胞培养表面和化学定义的培养基呈现HES细胞，以促进自我更新。结果将是一种合成的微环境，既可以用作大规模HES细胞扩展的再生医学技术平台，并且提供了一种新型且高度模块化的系统，用于剖析HES细胞自我更新的基本信号传导机制。