ASN's Mission

To create a world without kidney diseases, the ASN Alliance for Kidney Health elevates care by educating and informing, driving breakthroughs and innovation, and advocating for policies that create transformative changes in kidney medicine throughout the world.

learn more

Contact ASN

1401 H St, NW, Ste 900, Washington, DC 20005

email@asn-online.org

202-640-4660

The Latest on X

Kidney Week

Please note that you are viewing an archived section from 2020 and some content may be unavailable. To unlock all content for 2020, please visit the archives.

Abstract: PO0168

Endothelial Prolyl-Hydroxylase Domain Proteins Regulate Capillary Rarefaction Following Ischemic AKI and Reprogram Endothelial Metabolism

Session Information

  • AKI Mechanisms - 1
    October 22, 2020 | Location: On-Demand
    Abstract Time: 10:00 AM - 12:00 PM

Category: Acute Kidney Injury

  • 103 AKI: Mechanisms

Authors

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

Endothelial cell (EC) metabolism has emerged as a new regulator of EC behaviour, but its role in capillary rarefaction, a common feature of progressive renal disease, remains unknown. Because EC sense oxygen and metabolic alterations through Prolyl hydroxylases 1 to 3 (PHD1-3), here we wished to define the impact of PHD inactivation on post-ischemic kidney injury outcomes, while in vitro studies focused on metabolic consequences.

Methods

Following the induction of renal ischemia-reperfusion injury (IRI), concurrent deletion of PHD1,2,3 was achieved by the Cdh5(PAC)CreER inducible system. Analysis was performed at day 14 post-IRI. Furthermore, we examined the impact of DMOG, a PHD inhibitor on angiogenic capabilities and global metabolic profiles of endothelial cells.

Results

Post-ischemic kidneys of PHD1,2,3ECKO showed more fibrosis, as indicated by 68% increase in collagen area (P=0.005) and significant upregulation of profibrotic genes Loxl2, Tgf-β and Acta2 (n=6-8, P<0.5) compared to controls. Quantitative analysis of endomucin staining showed 50% decrease in peritubular capillary density, associated with reduced endothelial proliferation as indicated by Ki-67 immunostaining (n=4, p=0.005). Notably, biochemical inactivation of PHDs by DMOG reduced EC proliferation in MTT assay (P= 0.0001) while cell cycle analysis showed decrease of cells in S (~39%, n=3, p=0.0007) and G2/M phase (~24%, n=3, p=0.04). Furthermore, DMOG reduced EC migration (50%, n=3, p=0.005) and tube formation. LC-MS analysis showed a profound effect of DMOG in glycolytic, TCA cycle, lipid and, nucleotide metabolites. Specifically, EC treated with DMOG showed an increase in lactate (1.46-fold, p<0.05) and significant reductions in citrate (2.2 fold, p<0.001), alpha-ketoglutarate (2.5 fold, p<0.001), fumarate (1.4-fold, p<0.05) and malate (1.3 fold, p<0.01). Supplementation with citrate partially rescued the proliferation defect induced by DMOG, suggesting that PHDs may affect angiogenic responses through alterations in mitochondrial metabolism.

Conclusion

Post-ischemic endothelial inactivation of PHDs promotes peritubular capillary rarefaction and fibrosis following AKI, a response which could involve alterations in mitochondrial metabolism.

Funding

  • NIDDK Support