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Abstract: SA-OR20

Differential Role of Endothelial Prolyl-Hydroxylase 1, 2, 3 in AKI

Session Information

Category: Acute Kidney Injury

  • 103 AKI: Mechanisms

Authors

  • Tiwari, Ratnakar, Northwestern University Feinberg School of Medicine, Chicago, Illinois, United States
  • An, Si Young, Northwestern University Feinberg School of Medicine, Chicago, Illinois, United States
  • Kapitsinou, Pinelopi P., Northwestern University Feinberg School of Medicine, Chicago, Illinois, United States
Background

Recently, we showed that pre-ischemic inhibition of endothelial cell (EC)-HIF Prolyl Hydroxylase 2 (Phd2) protects against kidney ischemia-reperfusion injury (IRI). However, the role of post-ischemic inactivation of EC-Phd2 in kidney repair remains unclear. Further, recent single-cell RNA sequencing (scRNA-seq) data suggest a role for other EC-Phd isoforms (EC-Phd1 and EC-Phd3) in oxygen sensing. Here, we wished to address the role of post-ischemic inactivation of EC-Phd1, EC-Phd2, and EC-Phd3 in kidney repair.

Methods

Post-ischemic inactivation of EC-Phd1 (EC-Phd1KO), Phd2 (EC-Phd2KO), and Phd3 (EC-Phd3KO) was achieved by the Cdh5(PAC)CreER inducible system. To avoid compensatory effects between Phds, we generated mice with concurrent deletion of EC-Phd1, 2, and 3 (EC-Phd123KO) and induced recombination after IRI. Analysis was performed on day 14 post-IRI.

Results

Post-ischemic inactivation of EC-Phd1 or EC-Phd2 failed to protect kidneys based on mRNA expression of kidney injury molecule 1 (Kim1) and profibrotic genes lysyl oxidase-like 2 (Loxl2), transforming growth factor-beta 1 (Tgfβ1), and smooth muscle actin (Acta2) and histopathological analysis (n=7-8). Surprisingly, the inactivation of EC-Phd3 following IRI exacerbated kidney damage and fibrosis as indicated by increased expression of Kim1, Tgf-β, and Acta2 and deposition of collagen (n=6-8; p< 0.05). Likewise, post-ischemic concurrent deletion of EC-Phd123 increased kidney damage and fibrosis assessed by histopathological analysis and increased expression of profibrotic genes and collagen deposition (n=7-8, p<0.05), respectively, compared to Cre-controls. These changes were associated with significant worsening of renal function assessed by blood urea nitrogen level and transdermal GFR measurements (n=7, p<0.05). scRNA-seq data of the EC-Phd123KO post-ischemic kidneys showed significant transcriptional alterations in the EC cluster compared to Cre- controls with prominent changes in metabolic genes. Significant transcriptional changes were also observed in tubular, fibroblast, and inflammatory cell clusters between the two genotypes.

Conclusion

Post-ischemic concurrent inactivation of Phd1, 2, and 3 significantly impaired renal function, induced fibrosis which was mainly driven by EC-Phd3 inactivation. We delineated a critical role for EC-Phd3 in post-ischemic AKI repair.

Funding

  • NIDDK Support