EAGER: Mechanics-Guided Multicellular Self-Organization

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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.