EVIDENCE HOW EPITHELIAL SODIUM CHANNEL PROMOTES STIFFNESS IN HUMAN AND MOUSE ENDOTHELIAL CELLS. (April 2021) - Shah, Vikash Kumar
Abstract : Objective: The epithelial sodium channels (ENaCs) in the kidney maintain body salt/water balance and contributes to blood pressure regulation and hypertension. Recent finding suggests that ENaCs in endothelial cells are potential players mediating endothelial cell stiffness (ECS). Increased ECS is an important determinant of hypertension that coincides with altered endothelial nitric oxide (NO) production and endothelial dysfunction. Herein, we aim to uncover the molecular mechanism how ENaC influences ECS. I hypothesize that F-actin changes as a potential target through which ENaC mediates its effect. Design and method: ENaC subunit protein levels in Human Umbilical Vein Endothelial Cells (HUVECs) and Mouse Myocardial Endothelial Cells (MyEnd) cells were quantified by Western blotting. Aldosterone was used to increase ENaC expression. Changes in ECS assessed with Atomic Force Microscopy (AFM).. If changes in F-actin cell cytoskeleton is correlated with ENaC expression and ECS, immunofluorescence microscopy was used and quantified. Results: Aldosterone was observed to significantly increase alpha-ENaC (both in human and mouse cells) and delta-ENaC (human specific) protein levels which were reversed by the additional incubation with amiloride (ENaC blocker). The elevated ENaC expression with aldosterone was associated with an increased Young's modulus that is indicative of elevated ECS observed in both human and mouse endothelial cells. The upregulated ENaC expression and increased stiffness was also translated into increased F-actin in response to aldosterone. Conclusions: Taken together, our study provides experimental evidence of increased ENaC protein level with increased F-actin cell cytoskeleton in endothelial cells that may promote ECS. This is a new mechanism through which ENaC mediate ECS and contributes to endothelial dysfunction and hypertension.