The Cystogenic Effects of Ouabain in Autosomal Dominant Polycystic Kidney Disease (ADPKD) Require Cell Caveolae

Background: We have previously shown that the hormone ouabain is a circulating factor which can accelerate the progression of autosomal dominant polycystic kidney disease (ADPKD). At physiologic concentrations, ouabain increases cyst area and fibrosis in kidneys from ADPKD but not wildtype mice. These effects are mediated through ouabain binding to its specific receptor, Na,K-ATPase (NKA). In ADPKD, a fraction of the NKA has an abnormally high affinity for ouabain, leading to an over-activation of the NKA signal transduction cascade. Previous studies in cell lines suggest that ouabain’s effects are mediated by the NKA located in the cell caveolae. Here, we determined the role of caveolae in NKA-mediated and ouabain-induced effects in ADPKD using a new mouse model in which caveolar formation is disrupted on the background of ADPKD (Pkd1RC/RCCav1-/-). Methods: Mice homozygous for the knockout of caveolin-1 (Cav1), a main structural component of caveolae, were crossed with Pkd1RC/RC mice to obtain the Pkd1RC/RCCav1-/- model where caveolae no longer form in the kidney. Ouabain (0.3 mg/kg) or saline was injected intraperitoneally to Pkd1RC/RC or Pkd1RC/RCCav1-/- mice daily for 5 months. Kidneys from these mice were analyzed for cyst area, kidney weight to body weight ratio (%KW/BW) and fibrosis. Cell proliferation (using Ki67 as a marker) and the activation of downstream members of ouabain-NKA signaling (ERK and Akt) were measured. Results: The ablation of Cav1 and disruption of caveolae in the kidney was confirmed in Pkd1RC/RCCav1-/- mice by immunoblot and electron microscopy. Loss of caveolae abolished the high ouabain affinity component of NKA seen in Pkd1RC/RC mice, with all NKA having the same ouabain affinity as wildtype mice. After 5 months of ouabain administration, Pkd1RC/RCCav1-/- kidneys did not show the ouabain-induced increase in cyst number or fibrosis seen in Pkd1RC/RC mice. In addition, the ouabain-induced phosphorylation of ERK and AKT and the increase in cell proliferation in Pkd1RC/RC kidneys was blunted in the Pkd1RC/RCCav1-/- mice. Conclusion: Our results show that disruption of caveolae through deletion of caveolin-1 abolishes the abnormal high ouabain affinity fraction of NKA and the cystic effects of ouabain in ADPKD mice, providing in vivo evidence for the need of cell caveolae in the signaling function of NKA. Funding: NIDDK Support