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Abstract: SA-PO090

Loss of Pax2 and Pax8 in Proximal Tubules Protects Against Failed Repair After Ischemic AKI

Session Information

  • AKI: Mechanisms - III
    November 05, 2022 | Location: Exhibit Hall, Orange County Convention Center‚ West Building
    Abstract Time: 10:00 AM - 12:00 PM

Category: Acute Kidney Injury

  • 103 AKI: Mechanisms

Authors

  • Beamish, Jeffrey A., University of Michigan, Ann Arbor, Michigan, United States
  • Alsuraimi, Anas, University of Michigan, Ann Arbor, Michigan, United States
  • McElliott, Madison C., University of Michigan, Ann Arbor, Michigan, United States
  • Soofi, Abdul A., University of Michigan, Ann Arbor, Michigan, United States
  • Dressler, Greg R., University of Michigan, Ann Arbor, Michigan, United States
Background

Pax2 and Pax8 are two highly homologous transcription factors with critical and overlapping functions in kidney development and physiology. Both proteins are also upregulated following acute kidney injury (AKI), but their function in injury and repair is unknown. Our aim was to determine if selective deletion of Pax2 and Pax8 in proximal tubules affects sensitivity to injury and recovery after AKI.

Methods

Pax double mutant mice were generated carrying conditional floxed Pax2 and Pax8 alleles, the proximal-tubule-selective phosphoenolpyruvate carboxykinase (PEPCK) Cre driver, and a Cre-activated green fluorescent protein (GFP) reporter. Control mice included PEPCK-Cre and GFP reporter with wild type Pax2 and Pax8. 12-week-old male mice were subjected to unilateral ischemia-reperfusion injury (uIRI). Animals were sacrificed at various times after uIRI. Kidney tissue was analyzed by immunofluorescence (IF). Areas of failed repair 14 d post uIRI were marked by Kim1 or Vcam1 expression. Stained sections were scanned then quantified using U-net image segmentation.

Results

At baseline, mutants showed no difference in histology or kidney function. Gene deletions in proximal tubules were confirmed by IF and PCR. Mutant animals showed significantly fewer Vcam1+ tubules (46.6±33.8 vs 94.1±38.4 per mm2, N=14-15, p=0.002) and Kim1+ tubules (31.0±28.0 vs 56.3±27.5 per mm2, N=14-15, p=0.017). In mutant animals, we found that areas of failed repair contained significantly fewer GFP+ mutant cells compared with control animals where GFP marks Cre activity without Pax2 and Pax8 deletion (35.9±5.0% vs 46.4±6.7% of Vcam1+ cells, N=14-15, p<0.0001).

Conclusion

Surprisingly, mice with proximal-tubule-selective loss of Pax2 and Pax8 were less sensitive to IRI, with fewer Vcam1+ cells indicative of failed repair. As Vcam1+ cells may represent a population of survivors that do not fully recover their proximal tubule phenotype after injury, the data suggest that Pax mutant cells may be less likely to survive the initial injury and are rapidly cleared such that fewer stressed or damaged cells remain. Alternatively, Pax mutant cells may be inherently more resistant to ischemic injury. Mechanisms and experiments that distinguish these possibilities will be discussed.

Funding

  • NIDDK Support