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Abstract: SA-PO334

Inflammatory Cytokine Signaling in New Nephron Formation After AKI in Adult Zebrafish

Session Information

Category: Development, Stem Cells, and Regenerative Medicine

  • 600 Development, Stem Cells, and Regenerative Medicine

Authors

  • Schenk, Heiko Joachim, Medizinische Hochschule Hannover, Hannover, Niedersachsen, Germany
  • Sampson, William, Mount Desert Island Biological Laboratory, Salsbury Cove, Maine, United States
  • Kamei, Caramai Nanae, Mount Desert Island Biological Laboratory, Salsbury Cove, Maine, United States
  • Drummond, Iain A., Mount Desert Island Biological Laboratory, Salsbury Cove, Maine, United States
Background

Adult progenitor cells in the mesonephric zebrafish kidneys are required during neo-nephrogenesis replacing injured tubules by forming new nephrons. Single-cell RNA transcriptomes of adult kidney progenitor cells point to components of NF-kB and inflammatory cytokine receptors that may initiate stem cell-based nephrogenesis. Here, we present evidence that gentamicin induces inflammation-associated injury, which potentially stimulates stem cell-based nephrogenesis. The stimulatory effect on the progenitor cells to form new nephrons may be recapitulated by LPS injection without prior injury.

Methods

Adult zebrafish were injected i.p. with gentamicin or LPS at day 0. NF-kB signaling was determined 4 days post-injection (dpi) by NF-kB:GFP detection of the NF-kB reporter line Tg(NF-kB:EGFP) and NF-kB-associated gene expression using qRTPCR. Requirement of NF-kB signaling during regeneration was evaluated by pharmacological NF-kB inhibition. Bulk RNAseq from positive selected GFP+ and mcherry+ single cells by FACS was performed from kidneys 7 dpi by gentamicin injection using Tg(lhx1a:EGFP;cdh17:mCherry) fish.

Results

Gentamicin-induced kidney injury leads to increased tubular NF-kB reporter expression at 4 dpi and is associated with an upregulation of NF-kB target gene expression detected by qRTPCR. Additionally, gentamicin administration upregulated mRNA expression of GH receptors at 7 dpi, along with kidney progenitor markers osr1 and eya4. This was accompanied by an increased formation of new nephron aggregates, as indicated by elevated expression of Tg(lhx1a:GFP). Pharmacological inhibition of NF-kB signaling resulted in reduced expression of kidney progenitor markers, while LPS injection induced an upregulation of these markers. Bulk RNAseq analysis of positive-selected GFP+ Lhx1a+ cells at 7 dpi with gentamicin confirmed the induction of cytokine receptors in kidney progenitor cells.

Conclusion

Multiple pathways may converge on adult kidney stem cells to activate the formation of new nephrons. We propose that gentamicin-induced acute kidney injury (AKI) triggers inflammation, which, in turn, activates stem cells in the distal tubules. However, it remains unclear at this stage whether cytokine stimulation, such as that induced by LPS in the absence of injury, is sufficient to induce neo-nephrogenesis.

Funding

  • NIDDK Support