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Kidney Week

Abstract: PO1712

Targeting mTOR Signaling Improved Kidney Function in APOL1 Risk Variant Mice with Chronic Exposure to Inflammatory Stimuli

Session Information

Category: Glomerular Diseases

  • 1204 Podocyte Biology

Authors

  • Shah, Shrijal, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States
  • Tattersfield, Calum, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States
  • Demeritt, David, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States
  • Donovan, Olivia, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States
  • Schaller, Lena B., Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States
  • Mccarthy, Gizelle, Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States
  • Pollak, Martin R., Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States
  • Friedman, David J., Beth Israel Deaconess Medical Center, Boston, Massachusetts, United States
Background

Inheriting two copies of APOL1 risk variants significantly increases the likelihood of developing chronic kidney disease in African Americans. Many different mechanisms of disease have been proposed using cell-based model systems. Here we try to understand risk variant pathophysiology using a transgenic mouse model.

Methods

Transgenic mice with a single copy of human APOL1 G0 and G2 were generated using bacterial artificial chromosome (BAC) on a FVB background. Mice were injected with interferon gamma (IFN-g) plasmid via hydrodynamic tail vein injections to induce APOL1 expression. Since mTOR activation has been frequently observed in podocytes during FSGS, we blocked the pathway using rapamycin to see if it could improve disease outcomes. Mice were injected with 2mg/kg of rapamycin every other day (3 days/week, intraperitoneally) for a total of 2 weeks. Paraffin embedded tissue sections were used for immunohistochemistry and Periodic acid–Schiff (PAS) staining. Glomerular isolations were performed using Dynabeads. APOL1 oligomerization was assessed using blue native PAGE.

Results

7 Days after IFN-g plasmid injection, the podocytes of G2 mice stained positive for phospho-S6 ribosomal protein indicating mTOR activation accompanied by proteinuria. Blocking mTOR activation using rapamycin reversed ribosomal protein S6 phosphorylation, reduced proteinuria, and improved tissue histology as seen by PAS staining. We hypothesized that rapamycin might be activating autophagy and clearing APOL1 oligomers, but found no evidence for this mechanism of rescue. Instead, we were able to replicate the rescue we had observed with rapamycin using a cell cycle inhibitor, suggesting that rapamycin might be rescuing G2 phenotype by inhibiting podocyte cell cycle entry downstream of risk variant mediated injury.

Conclusion

Persistent expression of APOL1 risk variants pushes podocytes into cell cycle entry. Inhibiting mTOR signaling and subsequent cell cycle entry alleviated injury.

Funding

  • Other NIH Support