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Abstract: FR-PO355

Tuning Kidney Organoids to Generate High-Fidelity Proximal Tubule Cells

Session Information

Category: Development‚ Stem Cells‚ and Regenerative Medicine

  • 500 Development‚ Stem Cells‚ and Regenerative Medicine

Authors

  • Schnell, Jack, University of Southern California Department of Stem Cell Biology and Regenerative Medicine, Los Angeles, California, United States
  • Achieng, Mary Anne A., University of Southern California Department of Stem Cell Biology and Regenerative Medicine, Los Angeles, California, United States
  • Lindstrom, Nils, University of Southern California Department of Stem Cell Biology and Regenerative Medicine, Los Angeles, California, United States
Background

The most abundant cell in the kidney, proximal tubule (PT) cells, are poorly replicated across 3D kidney organoid models. PTs perform the bulk of renal reabsorption via the work of solute carriers and transporters for nutrients, toxins, and substrates. Defects to these cells often manifest as urinary wasting of nutrients. Studies have suggested role for transcription factor Hnf4a in PT maturation by driving the expression of transporters that impart physiological function to the PT, such as Urat1. Yet, recapitulating PT maturation in kidney organoids remains a challenge for regenerative therapeutics.

Methods

We undertook a stepwise approach that mimics early proximal precursor populations. HNF4A+ PTs within organoids were increased through a modified protocol for the directed differentiation of a human induced pluripotent stem cell HNF4A reporter line. This was done by transiently treating organoids with a small-molecule inhibitor of PI3K signaling. RNA-sequencing and immunofluorescent validation were performed to assess organoids for PT maturation genes.

Results

Treated organoids underwent increases in the abundance of HNF4A+ PTs. RNA-sequencing of organoids suggests increases in the urate and organic ion transporter URAT1, the glucose transporter SGLT6, amino acid transporters SLC3A1 and EAAC1, and others (Fig. 1). These data suggest enhanced PT maturation and better recapitulation of normal physiology compared to control samples.

Conclusion

These data demonstrate that organoid nephron maturation can be enhanced by modifying differentiation. Promoting proximal precursors within kidney organoids increases the abundance of HNF4A+ PTs at later stages of differentiation. These organoids exhibit increased expression of vital kidney transporters, suggesting that tuning early patterning can improve organoid functional potential. As such, these studies inform in vitro nephrogenesis strategies building regenerative therapeutics.

Percentage of total read counts from day 18 bulk RNA-seq of whole-organoid samples (n = 2).

Funding

  • Other NIH Support