Kidney Week

Abstract: FR-PO071

Pannexin1 Regulates Intracellular ATP in a Cisplatin Model of AKI

Session Information

• AKI: Tubules, Metabolism, New Models
  October 26, 2018 | Location: Exhibit Hall, San Diego Convention Center
  Abstract Time: 10:00 AM - 12:00 PM

Category: Acute Kidney Injury

• 103 AKI: Mechanisms

Authors

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

Jakub Jankowski,

• Yao, Junlan, University of Virginia, Charlottesville, Virginia, United States

Junlan Yao,

• Zheng, Shuqiu, University of Virginia, Charlottesville, Virginia, United States

Shuqiu Zheng,

• Bajwa, Amandeep, University of Virginia, Charlottesville, Virginia, United States

Amandeep Bajwa,

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

Diane L. Rosin,

• Isakson, Brant, University of Virginia School of Medicine, Charlottesville, Virginia, United States

Brant Isakson,

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

Kodi S. Ravichandran,

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

Mark D. Okusa,

Background

Pannexin1 (Panx1) is a membrane channel that can release ATP upon injury, thereby exacerbating inflammation. We reported that Panx1 deficiency protects kidneys from ischemia-reperfusion injury (IRI). We found significant differences in kidney ATP between Panx1 KO and control kidneys after injury and hypothesized that retention of ATP is a universal protective mechanism in AKI, including the cisplatin model.

Methods

Global Panx1 KO and WT mice (n=4) were subjected to 26min bilateral kidney ischemia and 24h of reperfusion. Global (n=8) and proximal tubule (PT, n=4) Panx1 KO mice and appropriate controls (n=6 and 4, respectively) were given cisplatin (20 mg/kg, ip). Injury was assessed 72 h later by plasma creatinine (PCr), blood urea nitrogen (BUN) and weight loss. Whole kidney ATP was obtained by chloroform extraction of kidney homogenates. CRISPR/Cas9 was used to generate stable Panx1 KO in murine proximal tubule cells (TKPTS). Cells were treated with 20 µM cisplatin overnight or 20 µM antimycin A for 1 hour to induce injury. Intracellular ATP concentration was measured using luminescent assay.

Results

At baseline, global Panx1 KO had less ATP (normalized to tissue weight) in whole kidney homogenates than WT mice (n=8 and 9; 0.59 vs. 1.00; p<0.05). Following IRI kidney ATP was higher (2.9 vs. 1.0; p<0.05) and PCr was lower (0.48 vs. 1.45 mg/dL; p<0.01) in Panx1 KO than in WT mice. After cisplatin, BUN was higher in Panx1 KO than WT mice (81.74 vs. 58.46 mg/dL; p=0.07) but there was no difference in kidney ATP (0.89 vs. 0.92; p=0.879). Similarly, after cisplatin, BUN (113.8 vs. 61.49 mg/dL; p<0.05) and body weight loss (21.76 vs. 17.37%; p<0.05) was higher in PT-specific Panx1 KO than WT mice. Kidney ATP was not greater in PT-Panx1 KO than WT mice (1.31 vs. 1.00; p=0.34) after cisplatin treatment. Cultured Panx1 KO cells retained more ATP than controls after cisplatin (54 vs. 36%, p<0.0001) or antimycin A (66 vs. 49%, p<0.001) treatment.

Conclusion

These results show that Panx1 influences ATP balance in cisplatin injury setting. PT-specific Panx1 deletion is sufficient to exacerbate injury, contrary to in vitro observations that Panx1 knockout tubules retain more ATP after antimycin A or cisplatin stimulation. The complex role of intracellular ATP homeostasis in AKI will require additional investigation.

Funding

• NIDDK Support