Glucocorticoid receptor is indispensable for physiological responses to aldosterone in epithelial Na+ channel induction via the mineralocorticoid receptor in a human colonic cell line

Tumor necrosis factor (TNF) is known to activate the epithelial Na+ channel (ENaC) in A549 cells. A549 cells are widely used model for ENaC research. The role of δ-ENaC subunit in TNF-induced activation has not been studied. In this study we hypothesized that δ-ENaC plays a major role in TNF-induced activation of ENaC channel in A549 cells which are widely used model for ENaC research. We used CRISPR/Cas 9 approach to knock down (KD) the δ-ENaC in A549 cells. Western blot and immunofluorescence assays were performed to analyze efficacy of δ-ENaC protein KD. Whole-cell patch clamp technique was used to analyze the TNF-induced activation of ENaC. Overexpression of wild type δ-ENaC in the δ-ENaC KD of A549 cells restored the TNF-induced activation of whole-cell Na+ current. Neither N-linked glycosylation sites nor carboxyl terminus domain of δ-ENaC was necessary for the TNF-induced activation of whole-cell Na+ current in δ-ENaC KD of A549 cells. Our data demonstrated that in A549 cells the δ-ENaC plays a major role in TNF-induced activation of ENaC.

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The glucocorticoid receptor: cause of or cure for obesity?

AJP Endocrinology and Metabolism ◽

10.1152/ajpendo.00478.2015 ◽

2016 ◽

Vol 310 (4) ◽

pp. E249-E257 ◽

Cited By ~ 55

Author(s):

Kezia John ◽

Joseph S. Marino ◽

Edwin R. Sanchez ◽

Terry D. Hinds

Keyword(s):

Lipid Metabolism ◽

Glucocorticoid Receptor ◽

Chronic Exposure ◽

Mineralocorticoid Receptor ◽

Acute Treatment ◽

Glucocorticoid Hormones ◽

Future Studies ◽

Obesity And Diabetes ◽

Metabolic Conditions ◽

Insufficient Knowledge

Glucocorticoid hormones (GCs) are important regulators of lipid metabolism, promoting lipolysis with acute treatment but lipogenesis with chronic exposure. Conventional wisdom posits that these disparate outcomes are mediated by the classical glucocorticoid receptor GRα. There is insufficient knowledge of the GC receptors (GRα and GRβ) in metabolic conditions such as obesity and diabetes. We present acute models of GC exposure that induce lipolysis, such as exercise, as well as chronic-excess models that cause obesity and lipid accumulation in the liver, such as hepatic steatosis. Alternative mechanisms are then proposed for the lipogenic actions of GCs, including induction of GC resistance by the GRβ isoform, and promotion of lipogenesis by GC activation of the mineralocorticoid receptor (MR). Finally, the potential involvement of chaperone proteins in the regulation of adipogenesis is considered. This reevaluation may prove useful to future studies on the steroidal basis of adipogenesis and obesity.

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Acute effects of aldosterone on the epithelial Na channel in rat kidney

AJP Renal Physiology ◽

10.1152/ajprenal.00585.2014 ◽

2015 ◽

Vol 308 (6) ◽

pp. F572-F578 ◽

Cited By ~ 26

Author(s):

Gustavo Frindt ◽

Lawrence G. Palmer

Keyword(s):

Rat Kidney ◽

Surface Protein ◽

Early Response ◽

Surface Expression ◽

Na Channel ◽

Channel Activity ◽

Acute Effects ◽

Chronic Stimulation ◽

Epithelial Na Channels ◽

Epithelial Na Channel

The acute effects of aldosterone administration on epithelial Na channels (ENaC) in rat kidney were examined using electrophysiology and immunodetection. Animals received a single injection of aldosterone (20 μg/kg body wt), which reduced Na excretion over the next 3 h. Channel activity was assessed in principal cells of cortical collecting ducts as amiloride-sensitive whole cell clamp current ( INa). INa averaged 100 pA/cell, 20–30% of that reported for the same preparation under conditions of chronic stimulation. INa was negligible in control animals that did not receive hormone. The acute physiological response correlated with changes in ENaC processing and trafficking. These effects included increases in the cleaved forms of α-ENaC and γ-ENaC, assessed by Western blot, and increases in the surface expression of β-ENaC and γ-ENaC measured after surface protein biotinylation. These changes were qualitatively and quantitatively similar to those of chronic stimulation. This suggests that altered trafficking to or from the apical membrane is an early response to the hormone and that later increases in channel activity require stimulation of channels residing at the surface.

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