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Kidney Week

Abstract: PO0621

Zebrafish Kidney Regeneration as a Model for Engraftment of Stem Cell-Derived Kidney Replacement Tissue

Session Information

Category: Development, Stem Cells, and Regenerative Medicine

  • 500 Development, Stem Cells, and Regenerative Medicine

Authors

  • Kamei, Caramai Nanae, Mount Desert Island Biological Laboratory, Bar Harbor, Maine, United States
  • Hughes, Samuel M., Mount Desert Island Biological Laboratory, Bar Harbor, Maine, United States
  • Wolf, Amber M., Mount Desert Island Biological Laboratory, Bar Harbor, Maine, United States
  • Schenk, Heiko Joachim, Medizinische Hochschule Hannover, Hannover, Niedersachsen, Germany
  • Drummond, Iain A., Mount Desert Island Biological Laboratory, Bar Harbor, Maine, United States
Background

In vivo engraftment of iPS cell derived mammalian kidney organoids is a major goal for kidney regenerative medicine. A major challenge in engraftment is in establishing patent tubule conduits between organoid graft and host tubules to allow fluid filtration and excretion. Stem cell-derived nephrons are continuously made during zebrafish kidney growth and regeneration that "plumb into" the pre-existing collecting system, making the zebrafish a viable model of kidney tissue engraftment. Using the zebrafish adult kidney to model synchronous fusion, we investigated the role of growth factor signaling pathways in this process.

Methods

Tg(Lhx1a:eGFP) expression labels distal invading ends of new nephrons. Tg(TCFLef-miniP:dGFP) Wnt reporter expression was used to reveal high Wnt signaling domains in new nephrons. The Wnt inhibitors IWR1 and IWP2 were applied to injured adult zebrafish to test requirements for Wnt signaling. Homozygous adult Crispr/Cas9 indel mutants in fzd9b and wnt9b were generated.

Results

We find that new nephron aggregates are patterned by canonical Wnt signaling. Cells with high canonical Wnt signaling form a single cell thick dome within cell aggregates and polarize to form rosettes with an apical constriction predicting the site of future tubule lumen. Tg(Lhx1a:eGFP) marks cells at the distal end of the new nephron which extend invasive processes or invadopodia into the underlying tubular epithelium. Short term inhibition of Wnt signaling using IWR1 and IWP2 inhibits invadopodia formation and blocks tubule interconnection events. Adult homozygous fzd9b mutants exhibit ectopic distal cell proliferation and a failure of convergent extension in new nephrons after injury while wnt9b mutants produce fewer new nephrogenic aggregates. A quantitative RT-PCR screen of candidate genes highly upregulated in both zebrafish nephron progenitors after injury and human cancer metastasis implicates invadopodia markers mmp14a/b, cortactin, tks5, as well as cdh11, c-jun, and id-1 in the invasion and interconnection process.

Conclusion

Wnt signaling is required for kidney tubule invasion and engraftment and correlates with expression of multiple genes associated with metastatic cell invasiveness. Manipulation of Wnt signaling is an opportunity to engineer kidney tubule interconnections.

Funding

  • NIDDK Support