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Abstract: PO1214

Autophagy Inhibition Ameliorates Polycystic Kidney Disease

Session Information

Category: Genetic Diseases of the Kidneys

  • 1001 Genetic Diseases of the Kidneys: Cystic

Authors

  • Atwood, Daniel, University of Colorado Denver School of Medicine, Aurora, Colorado, United States
  • Pokhrel, Deepak, University of Colorado Denver School of Medicine, Aurora, Colorado, United States
  • Dwivedi, Nidhi, University of Colorado Denver School of Medicine, Aurora, Colorado, United States
  • Hopp, Katharina, University of Colorado Denver School of Medicine, Aurora, Colorado, United States
  • Edelstein, Charles L., University of Colorado Denver School of Medicine, Aurora, Colorado, United States
Background

We published that there is a decrease in autophagy proteins in Pkd1RC/RC mouse kidneys. Our study aimed to determine the mechanistic role of suppressed autophagy in causing cyst growth using pharmacological and genetic autophagy inhibition.

Methods

Male Pkd1RC/RC (RC) mice were treated with 2-Deoxyglucose (2DG) or Chloroquine (CHLQ) from 50-120d of age. Kidney specific Pkd1, Atg7 double knockout mice were generated by Ksp1.3 Cre-lox recombination. Relative densitometry units (RDU) were determined on immunoblot. Autophagic flux was measured by the change in LC3-II (autophagosomes) -/+ Bafilomycin (Baf).

Results

Autophagic flux was present in wild type (WT) and 120 d old RC but suppressed in 150 d old RC kidneys. LC3-II (RDU) -/+ Baf was 0.1 vs 0.7 in WT (p<0.01), 0.6 vs 1.0 in 120 d old (p<0.05) and 2.4 vs 2.1 (NS) in 150 d old RC. 2DG resulted in a decrease in ATG12-5 complex and suppressed autophagic flux in RC kidneys. LC3-II (RDU) -/+ Baf was 0.5 vs 0.8 in VEH (p<0.05), 0.7 vs 0.7 in 2DG (NS). 2DG significantly reduced cyst growth and improved kidney function. Cystic index (%), count -/+ 2DG: 7.7 vs 3.7 (p<0.01), 211 vs 161 (p<0.05). BUN (mg/dL) -/+ 2DG: 35 vs 27 (p<0.01). Next, RC mice were treated with CHLQ, a specific autophagy inhibitor. CHLQ resulted in suppressed autophagic flux, less PKD and improved kidney function in RC mice. LC3-II (RDU) -/+ Baf was 1.2 vs 1.2 (NS) in CHLQ treated kidneys. Cyst index (%), Cyst no -/+ CHLQ in RC mice was 15.5 vs 7 (p<0.07), 231 vs 105 (p<0.05). BUN and creatinine (by HPLC) (mg/dL) -/+ CHLQ: 41 vs 26 (p<0.05), 2.9 vs 2.3 mg/dL (p<0.05). Next, autophagy was inhibited in PKD kidneys by generating double Pkd1 Atg7 KO mice. The 2 kidney/BW (%) was improved in Pkd1 Atg7 KO vs. single Pkd1 KO mice (32 vs 39 p<0.05). Atg7 KO kidneys had a massive increase in p62 indicating a build-up of autophagic cargo. p62 in WT vs Atg7 KO (RDU) 0.1 vs 1.8 p<0.001. Interestingly Atg7 KO kidneys were filled with tertiary lymphoid organs (TLO): large condensed infiltrates of T and B cells. pAMPKT172 was increased in Atg7 KO kidneys. AMPK activation is known to reduce PKD.

Conclusion

Both 2DG and CHLQ suppressed autophagic flux in RC kidneys and resulted in less PKD and improved kidney function. Double Pkd1 Atg7 KO mice had significantly lower kidney weight than single Pkd1 KO mice. Both pharmacological and genetic autophagy inhibition resulted in less PKD.

Funding

  • Veterans Affairs Support