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Ouabain Induces Endocytosis Of Plasmalemmal Na/K-Atpase In Llc-Pk1 Cells By A Clathrin-Dependent Mechanism, Jiang Liu, Riad Kesiry, Sankaridrug Periyasamy, Deppak Malhorta, Zijian Xie, Joseph Shapiro

Zijian Xie

Background: We have demonstrated that ouabain causes dose- and time-dependent decreases in ⁸⁶Rb uptake in porcine proximal tubular (LLC-PK1) cells. The present study addresses the molecular mechanisms involved in this process.

Methods: Studies were performed with cultured LLC-PK1 and Src family kinase deficient (SYF) cells.

Results: We found that 50 nmol/L ouabain applied to the basal, but not apical, aspect for 12 hours caused decreases in the plasmalemmal Na/K-ATPase. This loss of plasmalemmal Na/K-ATPase reverses completely within 12 to 24 hours after removal of ouabain. Ouabain also increased the Na/K-ATPase content in both early and late endosomes, activated phosphatidylinositol …

Ouabain-Induced Endocytosis Of The Plasmalemmal Na/K-Atpase In Llc-Pk1 Cells Requires Caveolin-1, Jiang Liu, Man Liang, Lijun Liu, Deepak Malhotra, Zijian Xie, Joseph Shapiro

Zijian Xie

Background: We have demonstrated that ouabain causes dose- and time-dependent decreases in 86Rb uptake in pig renal proximal tubule cell line (LLC-PK1) cells; and ouabain induces endocytosis of plasmalemmal Na/K-ATPase in LLC-PK1 cells in a clathrin-dependent pathway. Our data also suggest a role of endocytosis in both ouabain-induced signal transduction and proximal tubule sodium handling. The present study addresses the molecular mechanisms involved in this process. Methods: Studies were performed with cultured LLC-PK1 and a stable-expressed caveolin-1 knockdown LLC-PK1 cell line by SiRNA method. Results: In wild-type LLC-PK1 cells, depletion of cholesterol by methyl -cyclodextrin reduced ouabain-induced accumulation of Na/K-ATPase …