Chemically defined, plant-derived biomaterial platform for human cell culture

非技术性干细胞是人体内的细胞，与大多数细胞不同，它具有分化为许多不同类型细胞的潜力。由于干细胞在治疗人类疾病方面的潜力，干细胞一直是基础科学研究的重点领域。然而，精确控制干细胞分化为有用的治疗细胞仍然具有挑战性，更好地调控干细胞的行为，如黏附、增殖、分化和组织形成，可以加快人类干细胞的生物医学应用。天然和合成的生物材料可以作为控制细胞环境的“支架”，通常通过模仿细胞生长的自然环境来实现。植物在7亿多年的进化过程中形成了独特和多样化的材料特性，植物特性非常适合支持干细胞的生长。植物叶片生产的可伸缩性和可持续性质，以及增强的氧气扩散，广泛的形态和长度鳞片的多样性，以及它们的生物兼容性，都为它们成为广泛适用于人类干细胞制造的生物材料提供了强有力的理由。然而，植物来源的材料并不为人类细胞的正确附着和功能提供内在机制。通过这项工作开发的合成聚合物涂层将控制细胞-生物材料的附着，并向控制其命运的干细胞发出信号。通过这项工作开发的植物衍生生物材料可以交付给细胞生物学家和生物工程师，他们可以使用它们来探索关键的生物学问题或制造感兴趣的特定细胞。这项研究计划将通过开发跨学科学习模块来教育和激励农村高中学生和教师。该奖项将通过女性科学与工程领导力学院以及威斯康星大学麦迪逊分校本科生的研究经验，加强、留住和促进与代表不足的群体的接触。技术总结以脱细胞植物材料为基础的生物材料为生物分子的运输提供了复杂的相互连接的血管系统，并为细胞比对和图案配准提供了形态和长度尺度上的广泛多样性。本研究旨在开发一种脱细胞植物叶基生物材料，作为一种广泛适用于人类细胞培养的生物材料。这项研究将检验这样一种假设，即植物衍生材料表面涂有化学定义的聚合物薄膜，将支持细胞黏附、活性和排列，并促进干细胞向功能性腱细胞和肌细胞分化。这项工作的重点是开发方法学，以充分表征植物衍生的生物材料，并开发涂层化学，以修改叶表面，使其有利于人类细胞培养。这一理解将允许系统地询问表面形貌和可定制的表面化学对细胞排列和分化的影响。拟议研究的预期结果是：1)开发脱细胞过程，并对叶材料的化学成分、细胞毒性、形态、酶降解性、质量传输和机械性能进行综合表征，2)开发新型共聚涂层化学，以最小化生物分子的非特异性吸附，使粘附肽能够在纳米尺度上共定位，并定量地呈现肌肉和肌腱细胞外基质衍生多肽的组合，以及3)使用二次谐波产生显微镜将叶片微地形和表面化学与肌细胞和伸长细胞分化过程中的细胞排列相关联。这一奖项反映了NSF的法定使命，并通过使用基金会的智力优势和更广泛的影响审查标准进行评估，被认为值得支持。

NON-TECHNICAL SUMMARYStem cells are cells within our bodies that unlike most cells, have the potential to differentiate into many different types of cells. Stem cells have been an area of focused study in basic sciences due to their therapeutic potential in treating human disease. However, precise control over stem cell differentiation into useful therapeutic cells remains challenging, and better regulation of stem cell behaviors such as adhesion, proliferation, differentiation, and tissue formation could expedite biomedical applications of human stem cells. Natural and synthetic biomaterials can serve as a “scaffold” that controls the cell’s environment, often by mimicking the native environment in which the cells grow. Plants have developed unique and diverse material properties over more than 700 million years of evolution, and plant properties are ideally suited to support stem cell growth. The scalable, sustainable nature of plant leaf production, along with enhanced oxygen diffusion, wide diversity of morphology and length scales, and their biocompatibility, all make a strong case for their development as a broadly useful biomaterial for human stem cell manufacturing. However, plant-derived materials provide no intrinsic mechanism for proper attachment and function of human cells. The synthetic polymer coatings developed through this work will control cell-biomaterial attachment and present signals to stem cells that control their fate. The plant-derived biomaterials developed through this work can be delivered to cell biologists and bioengineers, who can use them to probe key biological questions or manufacture specific cells of interest. This research program will serve to educate and inspire rural high school students and teachers by the development of an interdisciplinary learning module. The award will enhance, retain, and promote engagement with underrepresented groups, through the Women in Science & Engineering Leadership Institute, and research experience for undergraduates at UW-Madison. TECHNICAL SUMMARYBiomaterials based on decellularized plant materials provide intricate interconnected vasculature for transport of biomolecules, as well as wide diversity in morphology and length scales for cell alignment and pattern registration. The research proposed here aims to develop a decellularized plant leaf-based biomaterial as a broadly useful and adaptable biomaterial for human cell culture. The research will test the hypothesis that plant-derived materials conformally coated with chemically-defined polymer films will support cell adhesion, viability and alignment, and also enhance stem cell differentiation toward functional tenocytes and myocytes. The work focuses on developing methodology to fully characterize plant-derived biomaterials and on developing coating chemistry to modify leaf surfaces to make them conducive for human cell culture. This understanding will allow systematic interrogation of the effect of surface topography and the customizable surface chemistry on the cellular alignment and differentiation. The expected outcomes of the proposed research are 1) the development of the decellularization process, and comprehensive characterization of chemical composition, cytotoxicity, morphology, enzymatic degradability, mass transport, and mechanical properties of the leaf material, 2) development of novel copolymer coating chemistry to minimize non-specific adsorption of biomolecules, enable the nanometer-scale co-localization of adhesion peptides, and quantitatively present a combination of muscle and tendon extra-cellular matrix derived peptides, and 3) correlating the leaf microtopography and surface chemistry with cell alignment during myogenic and tenogenic differentiation using second harmonic generation microscopy.This award reflects NSF's statutory mission and has been deemed worthy of support through evaluation using the Foundation's intellectual merit and broader impacts review criteria.