UCSF

Stem cells within the early embryo have the incredible capacity to self-organize and form higher order structures that provide the basis for large scale tissue morphogenesis. Understanding how embryonic signals are shaped, perceived, and transduced by cells early in embryonic development is critical for progress in the field of tissue engineering, so that we are able to recapitulate these cues in vitro and grow cells and tissue constructs outside of the body for high-throughput and animal-free drug/toxicological testing and cell/tissue replacement and therapy. However, efforts to study signaling and stem cell specification early in human development have largely been hindered by significant morphological differences in embryonic development of model organisms and ethical limitations regarding human embryo research. Thus, there is a need for tractable in vitro models which closely resemble native embryonic tissue, in both structure and behavior. Selecting an appropriate in vitro model that is tailored to hypothesis testing is critical for deriving conclusions about in vivo development. In this dissertation, I present a review on engineering co-emergence in organoid models, along with two studies which use novel 2D and 3D pluripotent stem cell models to study cell decision and symmetry breaking events during two different stages of early human embryonic development. In Chapter 1, I discuss the current state of organoid engineering, and define stringent criteria for organoid models. I discuss how primitive patterns and asymmetries integrate with endogenous cellular response systems to create autonomous and highly complex patterns that serve as a blueprint for higher order structure formation. In Chapter 2, I introduce cell structure as a key parameter which defines cell migration, organization, and specification. Specifically, I discuss how silencing of the cell-cell junction protein, CDH1, in a subpopulation of pluripotent stem cells within an embryoid body mimics zona pellucida hatching prior to gastrulation and prompts robust segregation and organization of CDH1- cells, along with their spontaneous differentiation to extraembryonic lineages. In Chapter 3, I focus on another cell-cell junction protein, ZO1, whose role is to both partition the plasma membrane into apical and basolateral functional domains, and assemble transmembrane tight junction proteins which prevent paracellular diffusion of macromolecules between the apical- and basolateral-facing lumens. Specifically, I demonstrate that cellular polarity, maintained by ZO1 in pluripotent stem cells, is a key regulator of morphogen signaling and pattern formation in a 2D peri-gastrulation model. In Chapter 4, I discuss the implications of these studies for signaling, specification, and migration of stem cells during embryonic development.