Kidney Week

Abstract: TH-OR017

Kidney PANX1 Releases ATP to Mediate Ischemia-Reperfusion Injury

Session Information

• Cellular Responses to Kidney Injury
  November 02, 2017 | Location: Room 395, Morial Convention Center
  Abstract Time: 05:42 PM - 05:54 PM

Category: Acute Kidney Injury

• 001 AKI: Basic

Authors

• Jankowski, Jakub, University of Virginia, Charlottesville, Virginia, United States

Jakub Jankowski, MS

• Perry, Heather M., University of Virginia, Charlottesville, Virginia, United States

Heather M. Perry,

• Huang, Liping, University of Virginia, Charlottesville, Virginia, United States

Liping Huang,

• Rosin, Diane L., University of Virginia, Charlottesville, Virginia, United States

Diane L. Rosin,

• Medina, Christopher B., University of Virginia, Charlottesville, Virginia, United States

Christopher B. Medina,

• Isakson, Brant, University of Virginia, Charlotesville, Virginia, United States

Brant Isakson,

• Ravichandran, Kodi S., University of Virginia, Charlottesville, Virginia, United States

Kodi S. Ravichandran,

• Okusa, Mark D., University of Virginia, Charlottesville, Virginia, United States

Mark D. Okusa,

Background

Extracellular ATP, a DAMP molecule, is deleterious in a number of kidney disease models. There is little data on its source and impact in AKI. We hypothesize that ATP is released from injured kidney cells by transmembrane pannexin1 (PANX1) channels and mediates ischemia-reperfusion injury (IRI). Earlier we reported that global PANX1 KO mice are protected against kidney IRI. We hypothesize that PANX1 expression on specific cell types in the kidney can contribute to injury by different mechanisms.

Methods

Proximal tubule (PT) and endothelial cell (EC) specific PANX1 KO mice (PepckCrePanx1^f/f and VECadCrePanx1^f/f, n=7 and 9 respectively) and appropriate controls were subjected to 26m bilateral kidney IRI or sham operation and 24h of reperfusion. To generate bone marrow chimeras global PANX1 KO and control mice (n=18 and 20) were lethally irradiated and 1x10⁷ donor bone marrow cells were administered i.v. and after 9 weeks mice were subjected to IRI. Kidney function and injury were assessed by plasma creatinine (PCr) and stereological quantification of acute tubular necrosis (ATN). Markers of kidney injury were quantified by real-time PCR of whole kidney lysates. Murine proximal tubule cell line (TKPTS) was transfected using CRISPR/Cas9 to create stable PANX1 deficiency. Injury after in vitro hypoxia/reoxygenation (H/R) was assessed by fluorescent ATP release assay and qPCR.

Results

PANX1 KO mice receiving PANX1 KO bone marrow (KO→KO) had lower PCr levels compared to WT→WT group after injury (0.57 vs.1.67; p<0.0001). KO→WT chimeras had increased level of plasma creatinine compared to WT→KO (1.67 vs. 0.38; p<0.0001), suggesting importance of parenchymal PANX1 deficiency in mediating tissue protection. The increase in PCr in WT mice subjected to IRI was attenuated in both PT (1.73 vs. 0.28; p<0.0001) and EC specific PANX1KO (1.3 vs. 0.16; p<0.0001). Histological injury scores were also lower in both PT (28.7% vs 87.8%; p<0.01) and EC PANX1KO (22.5% vs. 89.3%; p<0.001) compared to their respective controls. In TKPTS cells subjected to H/R medium ATP content and TNFa expression correlated positively with Panx1 expression.

Conclusion

These results show that loss of PANX1 from both PT and endothelium protects mouse kidneys from IRI. Targeting parenchymal PANX1 may lead to new therapeutic agents in the treatment of AKI.

Funding

• NIDDK Support