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Abstract: PO1547

A Fluidic Model of ARPKD Using Vascularized Kidney Organoids Identifies HIF-1 as a Potential Therapeutic Target

Session Information

Category: Genetic Diseases of the Kidneys

  • 1001 Genetic Diseases of the Kidneys: Cystic

Authors

  • Hiratsuka, Ken, Massachusetts General Hospital/Harvard Medical School, Boston, Massachusetts, United States
  • Miyoshi, Tomoya, Massachusetts General Hospital/Harvard Medical School, Boston, Massachusetts, United States
  • Kroll, Katharina T., Wyss Institute/Harvard University, Boston, Massachusetts, United States
  • Gupta, Navin R., Massachusetts General Hospital/Harvard Medical School, Boston, Massachusetts, United States
  • Valerius, M. Todd, Brigham and Women's Hospital/Harvard Medical School, Boston, Massachusetts, United States
  • Matsumoto, Takuya, Massachusetts General Hospital/Harvard Medical School, Boston, Massachusetts, United States
  • Lewis, Jennifer A., Wyss Institute/Harvard University, Boston, Massachusetts, United States
  • Morizane, Ryuji, Massachusetts General Hospital/Harvard Medical School, Boston, Massachusetts, United States
Background

We have recently reported a method to generate vascularized kidney organoids using fluidic chips. Vascularized kidney organoids derived from PKHD1 mutants demonstrated clinically relevant phenotypes that recapitulate cystogenesis in distal nephrons unlike static models with forskolin which induces cysts in proximal tubules. Here, we utilized this new PKD model to elucidate pathomechanisms and identified potential therapeutic targets for ARPKD patients.

Methods

PKHD1-mutant hPSCs were generated by CRISPR/Cas9 genome editing and differentiated into kidney organoids by following our reported protocol. Cystogenesis was stimulated by either fluidic flow on fluidic chips or forskolin in static culture. Cystic phenotypes were quantitatively determined by immunostaining. Gene expression was evaluated by 3D-gene microarray, and signal pathways were assessed by Metacore. Based on signal pathway results, candidate compounds were tested, and phenotypic improvement was evaluated by measuring tubular/cyst diameters using whole-mount immunostaining.

Results

Fluidic flow altered 407 signal pathways in PKHD1-/- organoids when compared to PKHD1+/- organoids while 63 pathways were changed by forskolin treatment in conventional static culture. In those pathways, 32 were involved in both flow- and forskolin-induced signal changes. In the common 32 pathways, HIF-1 pathway was top ranked with lowest p value of 3.71×10-14, suggested as a potential pathomechanim of ARPKD. To validate the result, we treated vascularized kidney organoids with HIF-1 inhibitor from day 16, the earliest stage of nephron differentiation. The distal nephron diameter was increased from 36.2 ± 7.7 μm (n=96) to 54.4 ± 21.8 μm (n=59) by fluidic flow (p=8.65×10-12), which was decreased to 46.1 ± 16.8 μm (n=158) by a HIF-1 inhibitor (p=0.004).

Conclusion

We identified HIF-1 as a potential therapeutic target for ARPKD patients. PKD organoids using an organ-on-a-chip platform might serve as a better model to elucidate disease developing mechanism and discover disease-specific new therapeutic targets in vitro.

Funding

  • Other NIH Support