Surface-Directed Differentiation of Human Mesenchymal Stem Cells on Orthogonal Peptide Concentration Gradient Surfaces

该奖项由阿克伦大学材料研究部生物材料项目颁发，旨在支持旨在量化影响人类间充质干细胞细胞分化途径的配体-受体相互作用的浓度依赖性和空间性质的研究。再生医学的未来发展将需要使用不止一种肽或生物活性分子来驱动干细胞进入明确的特异性分化群体。虽然许多研究都集中在离散浓度下的个体分子相互作用上，但细胞外基质衍生的成骨生长肽和骨形态发生蛋白的短肽片段单独或组合的浓度依赖性的完整相互作用组仍然难以捉摸。梯度方法还提供了一种将细胞整合素相互作用引起的局部信号与可溶性旁分泌效应解耦的方法。一旦确定，这些信息将推动高阶生物活性和仿生材料的设计，用于合成材料与生物系统接触的几种应用。该提案的目的概述了一种方法来制造和表征单向和正交肽浓度（10-200 pm/cm2为单向和10-100 pm/cm2为正交）梯度。利用表面光谱、免疫组织化学、自动荧光显微镜和实时聚合酶链反应的多维表征方法，该项目将捕获影响细胞增殖、谱系承诺和群体异质性的浓度阈值和协同信号事件。多学科研究生态系统将为材料化学、化学生物学和再生医学前沿的各级研究人员（高中、本科、研究生和博士后）提供培训机会。与当地一所高中合作，现实世界的研究活动将被纳入高年级生物课程中与年龄相适应的模块，并将用于教授物理和化学之间的基本概念和关系。推动再生医学进入主流应用的主要障碍之一是可靠、安全的细胞来源问题。在短期内，用于手术植入的干细胞必须来自需要替换部分的人（自体来源）。这一限制将需要术前分离干细胞，然后进行体外（体外）扩增方案，以产生足够的干细胞用于支架播种。现有的合成材料不足以满足这些协议。本提案概述了一种方法，以确定最佳组合和浓度的小仿生肽产生大量明确定义的干细胞群。自体方法不存在各种人群提出的大多数安全和伦理问题。如果成功，确定的浓度将对我们理解肽的个体和组合如何影响细胞分化产生深远的影响，并进一步提高用于研究和临床调查的明确定义的干细胞群体的数量和质量。多学科的研究环境为各级（高中、本科、研究生和博士后）的培训提供了机会。该项目包括培训当地一所学校的学生、本科生和高中生。适合年龄的教育模块将被设计为使用先进的研究课题在不同层次上教授高中生物课程的基本原理。鼓励这个年龄段的学生了解工程原理的重要性以及它们与促进人类健康的关系是至关重要的。该项目预计将支持国家努力增加工程学生管道的数量和多样性，这需要一个人深入并为代表性不足/少数民族学生群体提供独特的研究机会，鼓励追求科学和工程职业。

This award by the Biomaterials program in the Division of Materials Research to University of Akron is in support of research that seeks to quantify the concentration dependence and spatial nature of ligand-receptor interactions, which influence cell differentiation pathways in human mesenchymal stem cells. Future advances in regenerative medicine will require the use of more than one peptide or bioactive molecule to drive stem cells into well-defined specifically differentiated populations. While many investigations have focused on individual molecular interactions at discrete concentrations, a complete interactome of the concentration dependence of extra cellular matrix-derived, osteogenic growth peptide and short peptide fragments of bone morphogenic proteins singly and in combination has remained elusive. The gradient approach also affords a method to decouple local signaling that occurs due to cell-integrin interactions from soluble paracrine effects. When identified, this information will drive the design of higher-order bioactive and biomimetic materials for use in several applications where synthetic materials contact biological systems. The aims of the proposal outline an approach to fabricate and characterize both unidirectional and orthogonal peptide concentration (10-200 pm/cm2 for unidirectional and 10-100 pm/cm2 for orthogonal) gradients. Using a multidimensional characterization approach with surface spectroscopy, immunohistochemistry, automated fluorescence microscopy and real-time polymerase chain reaction, the project will capture the concentration thresholds and synergistic signaling events that influence cell proliferation, lineage commitment and population heterogeneities. The multidisciplinary research ecosystem will offer training opportunities to researchers at all levels (high school, undergraduate, graduate and postdoctoral) at the cutting edge of materials chemistry, chemical biology and regenerative medicine. In partnership with a local high school, real world research activities will be incorporated into age-appropriate modules into upper level biology courses and will be used to teach fundamental concepts and relationships between both physics and chemistry.One of the major remaining barriers to advancing regenerative medicine into mainstream applications is the issue of reliable, safe cell sources. In the near term, stem cells for surgical implantation must come from the person needing the replacement part (autologous sourcing). This limitation will require a pre-surgical isolation of stem cells followed by an expansion protocol ex vivo (outside the body) to generate enough stem cells for the scaffold seeding. The existing synthetic materials are insufficient for these protocols. This proposal outlines an approach to identify the optimal combinations and concentrations of small biomimetic peptides to yield large quantities of well-defined stem cell populations. The autologous approach is free of most safety and ethical concerns that have been raised by various populations. If successful, the identified concentrations will have far reaching impact on our understanding of how individual and combinations of peptides influence cell differentiation and furthermore improve the quantity and quality of well-defined stem cell populations available for research and clinical investigations. The multidisciplinary research environment offers opportunities for training at all levels (high school, undergraduate, graduate and postdoctoral). The project involves training students, undergraduate, and high school students from a local school. The age appropriate educational modules will be designed to teach fundamental principles to high school biology courses at various levels using advanced research topics. It is critical to encourage students at this age to understand the importance of engineering principles and how they relate to advancing human health. This project is expected to support the nation's efforts to increase the numbers and diversity of the engineering student pipeline that require one to reach down and providing unique research opportunities to underrepresented/minority students populations which encourage the pursuit of science and engineering careers.