Pluripotent stem cell-derived organoids can recapitulate significant features of organ development in vitro. We hypothesized that organoids thus created can be made substantially more relevant by mimicking aspects of in utero gestation, leading to higher physiological and anatomical resemblance to their in vivo counterparts. This dissertation introduces developmentally inspired methodologies to differentiate early human heart organoids into patterned heart tube organoids in a reproducible and high-throughput fashion by self-organization. Our strategy is based on controlled and stepwise exposure to crucial metabolic and hormonal factors eliciting important molecular, cellular, morphological, and functional changes in heart organoids over a 10-day period consistent with human cardiac development. Single-cell transcriptomics revealed the emergence of all cell types present in the developing heart tube, and past this stage. Structurally, heart organoids developed distinct atrial and ventricular chambers, a polarized proepicardial organ and anterior-posterior patterning due to endogenous retinoic acid signaling, mimicking the developmental processes found in the primitive heart post heart tube formation. Moreover, we showed the importance of our system for disease modeling by exploring the effects of ondansetron, a drug administered to pregnant women and associated with congenital heart defects, on embryonic heart development. Taken together, our results constitute a significant technical advance in human synthetic development and provide a powerful tool for cardiac disease modeling and pharmacological studies.
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