Integrating Biomaterials and Biophotonics to Assess How ECM Mechanics Regulate Cell Function in 3-D

ID: MPS/DMR/BMAT(7623) 0805164 PI: Putnam， Andrew ORG：加州大学欧文分校标题：整合生物材料和生物光子学来评估ECM力学如何调节3- din3细胞功能知识价值：细胞外基质（ECM）为细胞提供了化学和机械线索，并且提高了对这些线索如何在3-D中控制细胞功能的理解对于设计仿生材料作为组织工程的形态发生指南至关重要。在该提案中，PI计划利用基于聚乙二醇和纤维蛋白原（peg -纤维蛋白原）的独特生物合成混合水凝胶来明确测试局部底物机械特性影响细胞表型的3d假设。这一假设将使用血管平滑肌细胞（SMCs）作为生理学相关的模型细胞系统来解决，不仅基于PI对这种细胞类型的记录经验，而且基于这些细胞在体外和体内对静态和动态机械应力的众所周知的反应能力。初步数据表明，peg -纤维蛋白原水凝胶在相对较长的培养周期内支持SMC的粘附、扩散、活力和分化标志物的三维表达。这种材料还提供了一种方法，可以独立于粘附配体密度和蛋白水解敏感性来可预测地调整整体机械性能，这是使用天然生物聚合物（如胶原蛋白、纤维蛋白）无法实现的。PI还建议开发新的方法来测量peg -纤维蛋白原凝胶的局部机械特性，利用co-PI的光学镊子专业知识来研究机械转导机制，以解决细胞-物质相互作用领域的一个主要未解问题。以下三个目标构成了拟议的研究：(1)利用peg纤维蛋白原设计、表征和开发新型生物合成混合水凝胶，并证明在保持纤维蛋白原浓度不变的情况下，它们的整体机械性能可以被预测地操纵。(2)询问peg -纤维蛋白原凝胶的局部弹性和粘弹性特性，将测量值与上一个目标中的散装测量值进行比较。(3)在peg -纤维蛋白原ECM类似物的背景下，评估凝胶力学性能对三维培养SMCs（合成到收缩）表型转换的影响，并量化这些细胞材料结构的体积和局部力学性能如何随时间变化。更广泛的影响：完成本提案的目标需要整合来自生物材料、生物光子学、细胞和分子生物学的新兴原理。它将有助于开发材料系统和方法的近期研究目标，以解决有关ECM化学和力学的基本问题。这样的系统和方法将对定义组织工程中有用的生物材料的生物力学设计参数产生更广泛的影响，并允许在未来的提案中解决与发育生物学、心血管生理学、伤口愈合和肿瘤发生相关的基本力学问题。为了促进这里开发的方法的传播，它们将被整合到UCI校园贝克曼激光研究所的美国国立卫生研究院生物医学技术资源中心的激光微束和医学项目中。除了在项目上培训研究生外，pi还与加州少数民族参与联盟（CAMP）和UCI校园的数学、工程和科学成就（MESA）项目建立了工作关系，让少数民族本科生和高中生参与该项目。

ID: MPS/DMR/BMAT(7623) 0805164 PI: Putnam, Andrew ORG: University of California-IrvineTitle: Integrating Biomaterials and Biophotonics to Assess How ECM Mechanics Regulate Cell Function in 3-DINTELLECTUAL MERIT: The extracellular matrix (ECM) provides both chemical and mechanical cues to cells, and an improved understanding of how these cues govern cell function in 3-D is critically important to design biomimetic materials as morphogenetic guides for tissue engineering. In this proposal the PI plans to utilize a unique biosynthetic hybrid hydrogel based on poly(ethylene glycol) and fibrinogen (PEG-fibrinogen) to explicitly test the hypothesis that local substrate mechanical properties influence cell phenotype in 3-D. This hypothesis will be addressed using vascular smooth muscle cells (SMCs) as a physiologically relevant model cell system, based not only on the PI's documented experience with this cell type but also these cells' well-known ability to respond to static and dynamic mechanical stresses both in vitro and in vivo. Preliminary data demonstrate that PEG-fibrinogen hydrogels support SMC adhesion, spreading, viability, and the expression of differentiation markers in 3-D over relatively long culture periods. This material also provides the means to predictably tune bulk mechanical properties independently from adhesion ligand density and proteolytic sensitivity, something which cannot be achieved using native biopolymers (e.g., collagen, fibrin). The PI also proposes to develop novel methodology to measure the local mechanical properties of PEG-fibrinogen gels, leveraging the co-PI's expertise with optical tweezers to investigate mechanotransduction mechanisms to address a major unanswered question in the area of cell-material interactions. The following three objectives constitute the proposed study: (1) Engineer, characterize, and develop novel biosynthetic hybrid hydrogels using PEG-fibrinogen and demonstrate that their bulk mechanical properties can be predictably manipulated while holding the concentration of fibrinogen constant. (2) Interrogate the local elastic and viscoelastic properties of the PEG-fibrinogen gels, comparing the measured values to the bulk measurements in the previous objective. (3) Assess the impact of gel mechanical properties on the phenotypic switch of SMCs (synthetic to contractile) cultured in 3-D in the context of this PEG-fibrinogen ECM analog, and quantify how the bulk and local mechanical properties of these cell-material constructs change over time.BROADER IMPACTS: Completion of the objectives of this proposal requires an integration of emerging principles from biomaterials, biophotonics, and cell and molecular biology. It will contribute to the near-term research goal of developing material systems and methods to address fundamental questions regarding ECM chemistry and mechanics. Such systems and methods will have a much broader impact defining biomechanical design parameters for biomaterials useful in tissue engineering, and allow fundamental mechanics questions relevant for developmental biology, cardiovascular physiology, wound healing, and tumorigenesis to be addressed in future proposals. To facilitate dissemination of the methods developed here they will be integrated into the Laser Microbeam and Medical Program, an NIH Biomedical Technology Resource Center at the Beckman Laser Institute on the UCI campus. In addition to training graduate students on the project, the PIs have established a working relationship with the California Alliance for Minority Participation (CAMP) and the Mathematics, Engineering, and Science Achievement (MESA) programs on the UCI campus to involve minority undergraduate and high school students in the program.