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Kidney Week

Abstract: TH-PO022

Pannexin 1 Channels Regulate Mitochondrial Function, Autophagy, and Cell Survival During Kidney Ischemia-Reperfusion Injury in Mice

Session Information

Category: Acute Kidney Injury

  • 103 AKI: Mechanisms

Authors

  • Poudel, Nabin, University of Virginia, Charlottesville, Virginia, United States
  • Jankowski, Jakub, University of Virginia, Charlottesville, Virginia, United States
  • Morioka, Sho, University of Virginia, Charlottesville, Virginia, United States
  • Schinderle, Colleen, University of Virginia, Charlottesville, Virginia, United States
  • Zheng, Shuqiu, University of Virginia, Charlottesville, Virginia, United States
  • Rosin, Diane L., University of Virginia, Charlottesville, Virginia, United States
  • Okusa, Mark D., University of Virginia, Charlottesville, Virginia, United States
Background

Pannexin 1 (Panx1) channels are membrane associated non-selective channels that are activated by mechanical/physiological stimuli during injury and serve as a conduit for release of small molecules, including ATP. We have previously shown that pharmacological inhibition or genetic deletion of Panx1 in mice is protective against renal ischemia reperfusion injury (IRI), and Panx1 deficiency in murine proximal tubule-derived (TKPTS) cells results in reduced extracellular ATP and concomitant increase in intracellular ATP (PMID: 29866797). While the effects of extracellular ATP released from cells via Panx1 during injury have been extensively studied, the physiological role of Panx1 in cellular homeostasis is unknown.

Methods

Mice were subjected to IRI (26 mins of ischemia and 24 hrs of reperfusion) to assess plasma creatinine and kidney ATP levels. Control and Panx1 deficient TKPTS cells were subjected to hypoxia reperfusion (HR). Mitochondrial biogenesis and autophagy were assessed using real-time PCR and western blotting. Mitochondrial membrane potential was assessed by flow cytometry. Mitochondrial respiration was assessed using an Agilent Seahorse® assay. Cyclic AMP levels were measured using cAMP biosensor. For ATP depletion studies, cells were pretreated with 100 nM oligomycin prior to HR.

Results

Panx1-/- mice have higher kidney ATP levels 24 hours after IRI than control mice. Panx1-/-mice have higher levels of p62 in kidneys. In vitro findings show that Panx1 deficient cells retain more intracellular ATP after HR and have better survivability. Panx1 deficient cells have reduced mRNA expression of Pgc1a and Tfam, increased intensity of Mitotracker Red CMXRos® staining, and reduced cAMP-dependent signaling.

Conclusion

Our findings demonstrate that Panx1 deficiency leads to increased intracellular ATP, reduced cAMP signaling, reduced autophagy, increased mitochondrial function, and better cell survival during hypoxia. We conclude that deficiency of Panx1 leads to improved mitochondrial health and increased tubule cell survival during hypoxia resulting in protection during IRI. The development of selective pharmacological inhibitors of Panx1 could provide a novel approach to the treatment of acute kidney injury by maintaining mitochondrial health during stress and injury.

Funding

  • NIDDK Support