Integrating Collecting Systems via Nephron-Ureteric Bud Fusion in Kidney Organoids

Background: Kidney organoids are generated from human pluripotent stem cells (hPSCs) through directed differentiation of nephrogenic mesenchyme (NM) that subsequently epithelializes to form nephron-like structures. The nephrons lack collecting ducts (CDs) and instead terminate as blind-ended tubules, a limitation that has hampered their maturation into potentially functional tissue. Here we report an innovative, developmentally inspired system that addresses this deficiency through the incorporation and integration of hPSC-derived ureteric bud (UB) progenitor tissues, the embryologic building blocks of the kidney’s CDs and urinary collecting system. Methods: We established and optimized methods to grow kidney organoids comprising both NM and UB progenitor cells derived from hPSCs, and we used fluorescently labelled cells to trace the respective lineages. Both the early development and later differentiation were extensively characterized through wholemount staining, confocal imaging, and scRNA-seq. Results: Following recombination, NM rapidly and efficiently epithelialized into renal vesicle structures while the UB progenitors grew into extensive tubular networks. The resulting organoids contained both CDs and segmented nephrons, many of which were directly connected to the CDs via a connecting tubule-like anastomosis. The fusion process occurred during an early period of nephrogenesis, and it mimicked conserved steps by which the connection occurs in vivo. The ability to fuse with the UB was specific to only the distal-most nephron segments despite the greater abundance of proximal tubules, preserving normal polarity, and we identified pathways to manipulate nephron patterning and the frequency of fusion. Finally, the epithelial fusion also occurred in vivo following transplantation. Conclusion: By recapitulating developmental interactions, we established a novel organoid model with organized nephron-CD architecture and the first described system that recapitulates bona fide anastomoses between distal nephron and CD epithelia. This innovative system provides an unparalleled platform to interrogate the mechanisms of this important fusion event, and this approach is an important advance in kidney tissue engineering since generating a continuous drainage pathway from nephrons is an obligatory step toward production of functional renal tissue.