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Abstract: FR-PO382

The Regulatory Program of the Distalizing Mammalian Nephron

Session Information

Category: Development‚ Stem Cells‚ and Regenerative Medicine

  • 500 Development‚ Stem Cells‚ and Regenerative Medicine

Authors

  • Achieng, Mary Anne A., University of Southern California Keck School of Medicine, Los Angeles, California, United States
  • Schnell, Jack, University of Southern California Keck School of Medicine, Los Angeles, California, United States
  • Fausto, Connor, University of Southern California Keck School of Medicine, Los Angeles, California, United States
  • Lindstrom, Nils, University of Southern California Keck School of Medicine, Los Angeles, California, United States
Background

During development, nephron progenitors differentiate into over 24 cell types that become positioned along the nephron proximal-distal axis. To improve distal cell differentiation in the in vitro organoid model, we are delineating the developmental programs generating distal nephrons in vivo by analyzing scRNAseq data capturing human nephrogenesis. Comparisons of organoid scRNAseq data to the in vivo benchmark highlight dissimilarities between organoids and nephron cells, notably that organoids lack mature distal nephron markers. We present a scalable platform for synchronized nephrogenesis for studying patterning programs and forming functionally competent distal-like cells.

Methods

To correct patterning abnormalities in vitro, we generated a synchronized kidney organoid system for tuning differentiation of specific nephron cell fates. β-catenin signaling plays a role in expanding the distal domain and is required for distal tubule formation. Thus, we pulsed small molecule WNT agonists and quantified expression of early distal transcription factors to test whether β-catenin signaling plays a conserved role.

Results

Through early modulation of β-catenin signaling, we increased abundance of distal precursors and partially normalized patterning. Expression of transcription factors HNF1β and TFAP2A, which demarcate distal precursors in the comma- and S-shaped body of the mammalian nephron, is faithfully replicated in these treated organoids, with more elongated distal domains.

Conclusion

The synchronicity of our organoids enables manipulation of cohorts of nephron-like structures in a narrow developmental stage. In this system, β-catenin signaling manipulation in early organoids alters differentiation and morphogenesis, consistent with our model of β-catenin regulating positional identities along the nephron proximal-distal axis.

In CHIR-treated organoids, HNF1B+, TFAP2A+ distal domains are significantly increased compared to control, with more elongated tubules.