Abstract
With-No lysine Kinases (WNKs) have been newly implicated in alveolar fluid clearance (AFC). Epithelial sodium channels (ENaCs) serve a vital role in AFC. The potential protective effect of WNK4 in acute respiratory distress syndrome (ARDS), mediated by ENaC-associated AFC was investigated in the study. A model of lipopolysaccharide (LPS)-induced ARDS was established in C57BL/6 mice. WNK4, Sterile 20-related proline-alanine-rich kinase (SPAK), small interfering RNA (siRNA)-WNK4 or siRNA-SPAK were transfected into mouse lung or primary alveolar epithelial type II (ATII) cells. AFC, bronchoalveolar lavage fluid and lung histomorphology were determined. The expression of ENaC was determined to investigate the regulation of AFC by WNK4-SPAK signaling pathway. Activation of WNK4-SPAK signaling improved lung injury and survival rate, with enhanced AFC and reduced pulmonary edema via the upregulation of ENaC in ARDS. In primary rat ATII cells, gene-silencing by siRNA transfection reduced ENaC expression and the level of WNK4-associated SPAK phosphorylation. Immunoprecipitation revealed that the level of neural precursor cell-expressed developmentally downregulated gene 4 (Nedd4-2) binding to ENaC was decreased as a result of WNK4-SPAK signaling. The present study demonstrated that the WNK4/SPAK pathway improved AFC during LPS-induced ARDS, which is mainly dependent on the upregulation of ENaC with Nedd4-2-mediated ubiquitination.