Configuration of Microprinted Stem Cell Colonies in Heterocellular Niches Regulates Neural Differentiation

智力优势：干细胞在替换受损细胞和恢复患者失去的组织功能方面具有很大的前景。实现这一潜力需要有能力设计良好定义的微环境，并确定诱导干细胞向特定谱系分化的因素。这一建议源于PI最近的工作，该工作发现，在异细胞干细胞支持细胞微环境中，干细胞集落的大小和间距协同调节神经分化表型。研究这一现象的一个主要挑战是很难在支持细胞层上直接产生大小和间距确定的干细胞集落。提出的工作将使用一种新的高通量细胞打印微技术来解决这个问题，该技术可以在空间和时间上控制干细胞在支持细胞上的直接和无接触定位。提出的工作的中心假设是，干细胞集落的大小和空间组织创造了调节神经分化效率的内源性分化诱导信号分子的梯度。这一假设将通过设计菌落的大小和间距来验证，从而系统地操纵细胞分泌的内源性因子的信号阈值。主要目的是获得控制这一事件的分子事件的机制理解，并提出一种增强胚胎干细胞（ESCs）和诱导多能干细胞（iPSCs）神经分化的方法。对于在传统培养中分化效率较低的iPSCs来说，这将是一个显著的改进。该研究的成功将是实现干细胞生成的神经细胞转移治疗神经退行性疾病的重要一步。这个项目具有变革性，因为这里开发的方法将广泛适用于高通量筛选干细胞在人细胞和异细胞生态位中对各种谱系的命运承诺。实现这种微技术的便利性，只需要廉价的材料和现成的设备，使其完全为研究界所接受。更广泛的影响：基于胚胎和诱导多能干细胞的使用开发新的细胞替代治疗方法，将对治疗包括目前缺乏有效治疗的神经退行性疾病在内的广泛疾病具有重大的公共卫生益处。提出的微技术方法工程干细胞龛将确定特定的因素，加强神经细胞的分化超出目前可实现的，因此将具有重大的社会影响。生态位工程的方法也将通过提供一种有用的技术来研究干细胞在异细胞生态位中向各种细胞类型的分化，并对组织修复和再生医学产生长期影响，从而广泛地造福于研究界。除了潜在的医学影响外，拟议的项目还将通过几种机制改善本科STEM教育。首先，本科生将成为项目不可或缺的一部分，执行、分析和展示研究成果。根据pi的经验，从事研究的本科生，包括女性和少数族裔，继续进入STEM研究生领域（从事研究的本科生中有35/51是女性或代表性不足的少数族裔，22/32的毕业生继续进入STEM领域的研究生院（主要是BME））。其次，本工作将整合本科二年级计算机课程的几个方面。学生将接触到模拟扩散的基本方程以及对问题的解释。然后，他们将在课堂上使用MATLAB®研究模型的解决方案。最后，该项目将通过为阿克伦大学夏令营的中学生和高中生开发一个模块，增加妇女和代表性不足的少数民族对STEM的参与。本模块将允许学生与博士后和研究生一起工作，学习易于使用的微技术基础知识，并设计和创建细胞打印。因此，这些学生将接触到生物医学工程的先进技术和研究。

PI: Hossein TavanaProposal ID 1264562Intellectual merit: Stem cells hold a great promise for replacing damaged cells and recovering lost tissue function in patients. Realizing this potential requires the ability to engineer well-defined microenvironments and determine the factors that induce differentiation of stem cells toward specific lineages. This proposal stems from PI's recent works in which it was found that size and interspacing of stem cell colonies in heterocellular stem cell-support cell microenvironments synergistically regulate neural differentiation phenotypes. A major challenge to investigating this phenomenon is the difficulty of creating stem cell colonies of defined size and interspacing directly on a support cell layer. The proposed work will tackle this problem using a new high throughput cell printing microtechnology that enables spatial and temporal control over direct and contact-free positioning of stem cells on support cells. The central hypothesis of the proposed work is that size and spatial organization of stem cell colonies creates gradients of endogenous differentiation-inducing signaling molecules that regulate neural differentiation efficiency. This hypothesis will be tested by engineering the size and interspacing of colonies and thus, systematically manipulating signaling thresholds of cell-secreted endogenous factors. The main objectives are to gain a mechanistic understanding of molecular events that govern this event and propose an approach for enhanced neural differentiation of embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs). This will be a marked improvement for iPSCs that differentiate with low efficiency in traditional cultures. Success of this proposal will be a step towards realizing transition of stem cell-generated neural cells for treatment of neurodegenerative diseases. This project is transformative as the methodology developed here will be broadly applicable to high throughput screening of fate commitment of stem cells to various lineages in homo- and hetero-cellular niches. The ease of implementing this microtechnology, which only requires inexpensive materials and off-the-shelf equipment, makes it fully accessible to the research community. Broader impact: Developing new cell replacement therapy approaches based on the use of embryonic and induced pluripotent stem cells will have significant public health benefits for treating a wide range of disorders including neurodegenerative diseases that currently lack effective treatment. The proposed microtechnological approach to engineering stem cell niches will identify specific factors that enhance differentiation to neural cells beyond that currently achievable, and therefore will have significant societal impact. The approach to niche engineering will also broadly benefit the research community by offering a useful technology to investigate stem cell differentiation to various cell types in heterocellular niches with long-term impact in tissue repair and regenerative medicine. In addition to the potential medical impacts, the proposed project will improve undergraduate STEM education through several mechanisms. First, undergraduates will be integral to the project, performing, analyzing, and presenting research. It is the PIs' experience that undergraduate students, including women and minorities, who perform research continue on into STEM graduate fields (35/51 undergraduates performing research were females or underrepresented minorities and 22/32 who have graduated continue on to graduate school in STEM fields (primarily BME)). Second, this work will be integrated several aspects of a sophomore level undergraduate computing course. Students will be exposed to the fundamental equations for modeling diffusion as well as an explanation of the problem. They will then investigate solutions for the model in the class using MATLAB®. Finally, this project will increase participation of women and underrepresented minorities in STEM by development of a module for middle and high school students in summer camps at The University of Akron. This module will allow the students to work with the postdoctoral fellow and the graduate student, learn basics of easy-to-use microtechnology, and design and create a print of cells. Therefore, these students will be exposed to advanced technology and research in biomedical engineering.