Project Summary Human induced pluripotent stem cells (iPSCs) are unique in that they retain their ability to indefinitely self-renew while maintaining the capacity to self-organize and differentiate into both embryonic and extraembryonic lineages. iPSCs have emerged as a powerful tool to study human development, and disease, and have been integrated with tissue engineering approaches for regenerative medicine applications. To fulfill the promise of iPSC clinical utility, further investigation of the role of the stem cell niche in iPSC morphogenesis, lineage specification, and functional maturation is needed. While organoid approaches have revolutionized our ability to mimic organ-level function in a dish, they typically are comprised of cells from a single germ layer, missing critical cues shared by surrounding populations including the microvasculature and stroma. In addition, organoids are generated in ill-defined matrices such as Matrigel, which suffers from batch-to-batch variation, and limited tunability. To this end, we propose using micropatterned induced gastrulation models to better understand how paracrine and mechanical cues guide primitive stem cell fate. In addition, by leveraging assembloid technologies, synthetic extracellular matrix mimics, and dynamic microfluidic culture, we aim to better understand multi-germ layer interactions during tissue specification. Finally, we propose that improved iPSC derivatives can be used to better understand patient-specific differences in metabolic disorders. Collectively, we propose that using an integrative approach will permit iPSCs to be a powerful testbed for studying developmental biology and disease processes.