Mechanosensitive hair cells in the cochlea are responsible for hearing but are vulnerable to damage by genetic mutations and environmental insults. The paucity of human cochlear tissues makes it difficult to study cochlear hair cells. Organoids offer a compelling platform to study scarce tissues in vitro; however, derivation of cochlear cell types has proven non-trivial. Here, using 3D cultures of human pluripotent stem cells, we sought to replicate key differentiation cues of cochlear specification. We found that timed modulations of Sonic Hedgehog and WNT signaling promote ventral gene expression in otic progenitors. Ventralized otic progenitors subsequently give rise to elaborately patterned epithelia containing hair cells with morphology, marker expression, and functional properties consistent with both outer and inner hair cells in the cochlea. These results suggest that early morphogenic cues are sufficient to drive cochlear induction and establish an unprecedented system to model the human auditory organ.

Accumulating evidence demonstrates important roles for metabolism in cell fate determination. However, it is a challenge to assess metabolism at a spatial resolution that acknowledges both heterogeneity and cellular dynamics in its tissue microenvironment. Using a multi-omics platform to study cell-type-specific dynamics in metabolism in complex tissues, we describe the metabolic trajectories during nephrogenesis in the developing human kidney. Exploiting in situ analysis of isotopic labeling, a shift from glycolysis toward fatty acid β-oxidation was observed during the differentiation from the renal vesicle toward the S-shaped body and the proximal tubules. In addition, we show that hiPSC-derived kidney organoids are characterized by a metabolic immature phenotype that fails to use mitochondrial long-chain fatty acids for energy metabolism. Furthermore, supplementation of butyrate enhances tubular epithelial differentiation and maturation in cultured kidney organoids. Our findings highlight the relevance of understanding metabolic trajectories to efficiently guide stem cell differentiation.

Pluripotent stem cells (PSCs) could provide a powerful system to model development of the human esophagus, whose distinct tissue organization compared to rodent esophagus suggests that developmental mechanisms may not be conserved between species. We therefore established an efficient protocol for generating esophageal progenitor cells (EPCs) from human PSCs. We found that inhibition of TGF-ß and BMP signaling is required for sequential specification of EPCs, which can be further purified using cell-surface markers. These EPCs resemble their human fetal counterparts and can recapitulate normal development of esophageal stratified squamous epithelium during in vitro 3D cultures and in vivo. Importantly, combining hPSC differentiation strategies with mouse genetics elucidated a critical role for Notch signaling in the formation of this epithelium. These studies therefore not only provide an efficient approach to generate EPCs, but also offer a model system to study the regulatory mechanisms underlying development of the human esophagus.