Abstract: TH-PO674
Rescue of Suppressed Autophagy in Polycystic Kidney Disease
Session Information
- ADPKD: Genetic and Model Studies
October 25, 2018 | Location: Exhibit Hall, San Diego Convention Center
Abstract Time: 10:00 AM - 12:00 PM
Category: Genetic Diseases of the Kidney
- 1001 Genetic Diseases of the Kidney: Cystic
Authors
- Brown, Carolyn Nicole, UC Denver Anschutz Medical Campus, Aurora, Colorado, United States
- Holditch, Sara, UC Denver Anschutz Medical Campus, Aurora, Colorado, United States
- Serkova, Natalie J., University of Colorado Denver , Aurora, Colorado, United States
- Edelstein, Charles L., UC Denver Anschutz Medical Campus, Aurora, Colorado, United States
Background
Autophagy maintains proteostasis by sequestering damaged organelles and proteins into autophagosomes for delivery to the lysosome where cargo is degraded and recycled. Several treatments that have been shown to ameliorate PKD in mice are potent activators of autophagy. The aims of the study were to determine the effects of several autophagy inducers on autophagy, apoptosis, and proliferation markers in PKD kidneys and to determine whether kidney-specific autophagy knockout results in a cystic phenotype.
Methods
Mice were treated with 2-deoxyglucose (2DG), trehalose (TRE), or metformin (MET) followed by bafilomycin (BAF) to measure autophagic flux. p62 (autophagy-specific tag for degradation & marker of autophagy inhibition), LC3-II (marker of autophagosomes), Atg12-5 (marker of autophagosome elongation), cleaved caspase 3 (marker of apoptosis), and AMBRA1 (Activating molecule in Beclin1-regulated autophagy) that links autophagy to cell proliferation by promoting dephosphorylation and degradation of cMyc (transcription factor that activates pro-proliferative genes) were measured by immunoblot. Kidney-specific Atg7-/-mice received MRIs at 180D of age to precisely measure cyst volume and number.
Results
PKD kidneys had significantly reduced autophagic flux compared to WT, which was rescued by 2DG. Further, PKD kidneys had less Atg12-5 complex vs WT, an effect blocked by TRE. PKD kidneys had more cleaved caspase 3, a marker of apoptosis, compared to WT, which was decreased by MET, 2DG, and TRE. cMyc phosphorylation was greater in PKD compared to WT, an effect blocked by 2DG. Trehalose increased AMBRA expression in PKD kidneys (Table 1, *p<0.05). On MRI scan, mean number of cysts (0.5mm) in 180 day old Atg7-/- mice was 5±2 vs. 1±0.5 in WT (p<0.05). One Atg7-/- kidney was massively cystic.
Conclusion
In summary, 2DG restored autophagic flux in PKD kidneys. Autophagy inducers increased expression of autophagy-related proteins and decreased apoptosis and proliferation markers. Kidney-specific knockout of autophagy in 180 day old mice resulted in a cystic phenotype.
Table 1
| FLUX | Atg12-5 | p cMyc | AMBRA | CC3 | |
| WT VEH | 0.4±0.05* | 0.4±0.05* | 0.8±0.1* | 0.7±0.1 | 1.0±0.05* |
| PKD VEH | 0.1±0.03 | 0.2±0.02 | 1.3±0.1 | 0.7±0.1 | 1.6±0.09 |
| PKD 2DG | 0.3±0.06* | 0.2±0.08 | 0.9±0.1* | 1.3±0.1 | 1.0±0.15* |
| PKD TRE | 0.1±0.08 | 0.3±0.07 | 1.2±0.2 | 1.2±0.1* | 0.6±0.1* |
| PKD MET | 0.1±0.07 | 0.2±0.02 | 0.9±0.08* |
Funding
- Other U.S. Government Support