EAGER: Mechanics-Guided Multicellular Self-Organization

在多细胞组织的发育过程中，形状和图案的形成以相当标准的方式发生。均质细胞群被触发自主地自组织成结构化的多细胞组织。然而，指导这种组织的信号仍然是一个谜，这些过程并不总是在实验室培养环境中正确或完全地发生。这个探索性研究（EAGER）项目的早期概念资助的目标是开发一种微流体系统，在这种系统中，培养环境可以精确控制，这样干细胞的组织发育就可以定向到可复制的结构。正确的组织发育需要生化和机械信号，通过这个系统可以精确地控制这些信号。然后，这些信号在最初同质的细胞组中触发基因表达，使它们开始发育路径。微流体系统还将与实时成像兼容，以支持未来对驱动细胞组织和组织发育机制的高级研究。提高对三维组织发育成正确组织结构的理解将显著加强发育生物学的相关研究，包括先天性畸形，以及组织工程和再生医学。教育和推广计划包括让高中生（优先考虑女性和代表性不足的少数族裔）进入实验室进行为期4-8周的研究体验，以及让本科生参与研究。这些项目还将为高级研究生和博士后提供获得指导经验的机会。与干细胞生物技术和再生医学相关的一门新课程也正在开发中，这门课程将直接利用支持这项研究的科学进展。这个跨学科项目涉及干细胞生物学、发育生物学、信号转导、机械生物学和微流体学。本研究的目的是开发一种微流控装置，并利用它实现人多能干细胞的外胚层囊肿（柱状组织）和羊膜囊肿（鳞状组织）的可控合成。多室微流体系统将改变细胞负载和化学诱导因子。细胞播种将通过共聚焦成像进行评估。实时成像和免疫染色将用于监测囊肿形成的形态发生动力学，包括腔形成（腔形成）以及细胞形状的变化。细胞命运将通过外胚层和羊膜谱系标记的染色来确认，上皮化的动态将在不同的时间点进行检查。该奖项反映了美国国家科学基金会的法定使命，并通过使用基金会的知识价值和更广泛的影响审查标准进行评估，被认为值得支持。

During the development of multicellular tissues, the formation of shape and pattern occur in fairly standard ways. Homogeneous cell populations are triggered to autonomously self-organize into structured, multicellular tissues. However, the signals that guide this organization remain a mystery, and these processes do not consistently occur properly or completely in an a laboratory culture environment. The goal of this EArly-concept Grant for Exploratory Research (EAGER) project is to develop a microfluidic system in which the culture environment can be precisely controlled so that tissue development from stem cells can be directed towards reproducible structures. Correct tissue development requires both biochemical and mechanical signals, which will be precisely controllable through this system. These signals then trigger gene expression within initially homogeneous sets of cells so that they begin a developmental path. The microfluidic system will also be compatible with live imaging, to support future advanced studies into mechanisms driving cellular organization and tissue development. Improved understanding of 3D tissue development into correctly organized structures will significantly enhance research related to developmental biology, including congenital malformations, as well as tissue engineering and regenerative medicine. Educational and outreach plans include integrating high school students, with priority given to women and underrepresented minorities, into the lab for a 4-8 week research experience as well as engaging undergraduates in research. These projects will also provide an opportunity for senior graduate students and post-doctoral fellows to gain mentorship experience. A new course is also being developed related to stem cell biotechnology and regenerative medicine, which will directly draw on the advances in science that support this research. This cross-disciplinary project involves stem cell biology, developmental biology, signal transduction, mechanobiology, and microfluidics. The objectives of this study are to develop a microfluidic device and use it to achieve controllable synthesis of both the epiblast cyst (a columnar tissue) and the amniotic cyst (a squamous tissue) from human pluripotent stem cells. The multi-chambered, microfluidic system will vary both cell loading and chemical induction factors. Cell seeding will be assessed with confocal imaging. Live imaging and immunostaining will be used to monitor mofphogenetic dynamics of the cyst formation, including lumenogenesis (cavity formation) as well as changes in cell shape. Cell fate will be confirmed by staining for epiblast and amniotic lineage markers, and the dynamics of epithelialization will be examined at varying time points.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.