Kidney Week

Abstract: PO0348

Regulation of Mitochondrial Metabolism in T-Regulatory Cells by Programmed Cell Death Protein 1 in AKI

Session Information

• AKI: Mechanisms of Injury
  November 04, 2021 | Location: On-Demand, Virtual Only
  Abstract Time: 10:00 AM - 12:00 PM

Category: Acute Kidney Injury

• 103 AKI: Mechanisms

Authors

• Dogan, Murat, University of Virginia School of Medicine, Charlottesville, Virginia, United States

Murat Dogan, MD, PhD

• Sabapathy, Vikram, University of Virginia School of Medicine, Charlottesville, Virginia, United States

Vikram Sabapathy,

• Venkatadri, Rajkumar, University of Virginia School of Medicine, Charlottesville, Virginia, United States

Rajkumar Venkatadri,

• O'Neill, Christopher, University of Virginia School of Medicine, Charlottesville, Virginia, United States

Christopher O'Neill,

• Xavier, Sandhya, University of Virginia School of Medicine, Charlottesville, Virginia, United States

Sandhya Xavier,

• Portilla, Didier, University of Virginia School of Medicine, Charlottesville, Virginia, United States

Didier Portilla,

• Sharma, Rahul, University of Virginia School of Medicine, Charlottesville, Virginia, United States

Rahul Sharma,

Background

The T-regulatory cells (Tregs) are important for suppressing inflammation and for resolution of injury during acute kidney injury (AKI). Absence or blocking of programmed cell death 1 (PD-1) mitigates the Treg-mediated protection in AKI. The mechanisms, however remain unknown. Here we test the hypothesis that PD-1 regulates mitochondrial metabolism of Tregs during AKI.

Methods

Bilateral renal ischemia reperfusion injury (IRI) was used to investigate the role of PD-1 function in Tregs. Adoptive transfer of splenic CD4⁺CD25⁺ Tregs isolated either from wild-type (WT) or PD-1-/- Foxp3-GFP mice was performed into recipient mice 1h or 24h prior to IRI. Renal structure and function was assessed by plasma creatinine, Kim1 and Ngalexpression, histopathology and flow cytometry. Mitochondrial metabolism of isolated Tregs was assessed ex vivo using Seahorse Metabolic Flux Analyzer and staining with mitochondrial dyes (TMRE and Mito-ROS) for mitochondrial membrane potential and ROS production respectively, as well as by RT-PCR for genes related to mitochondrial dynamics. Scanning Electron microscopy (SEM) was performed on isolated Tregs to image mitochondrial morphology followed by ImageJ^TM analyses.

Results

Contrary to the WT, adoptive transfer of PD-1^-/- Tregs was unable to protect the recipient mice from IRI-induced AKI. The oxygen consumption rate (OCR), a measure of oxidative phosphorylation, was significantly reduced in PD-1^-/- Tregs at baseline and under maximal respiration as compared to WT. PD-1^-/- Tregs also had reduced mitochondrial mass, lower mitochondrial membrane potential, and greater mitochondrial ROS production than WT Tregs. Further, the expression of Pgc1a, Nrf1, Brf2, Tfam, Drp1, Mfn1, Mfn2 and Mff was also significantly reduced in the PD-1^-/- Tregs. Importantly, compared to Tregs from age and sex-matched WT mice and as measured using SEM and ImageJ^TM, the mitochondria in Tregs from PD-1^-/- mice were fewer in number and had lower average pixel area.

Conclusion

The data suggest that PD-1 regulates mitochondrial function and dynamics of Tregs and this is critical for protection from AKI and other inflammatory diseases.

Funding

• NIDDK Support