Ouabain-induced Internalization and Lysosomal Degradation of the Na+/K+-ATPase

Cherniavsky-Lev, Marina; Golani, Ofra; Karlish, Steven J. D.; Garty, Haim

doi:10.1074/jbc.m113.517003

Cited by 29 publications

(25 citation statements)

References 39 publications

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“…The knockdown of ␣1 in the work by Xie et al was correlated with an increase in basal Src activity and activation of consequent signal transduction pathways. Although our present work does not address this issue directly, our previously published study (4) showed no relationship between Src/phosphoinositide-3-kinase signaling pathways to ouabain-induced inhibition and subsequent internalization of Na ϩ -K ϩ -ATPase in our cellular system, as opposed to results described previously by Xie et al (11,(22)(23)(24).…”

Section: Discussioncontrasting

confidence: 99%

“…Biotinylation of cell surface proteins was done as reported previously (4). In brief, cell monolayers at ϳ90% confluency were incubated for 15 min at 4°C on a rocking platform with 1 mg/ml EZ-Link N-hydroxy-sulfo-succinimido-biotin (sulfo-NHS-SS-biotin; Pierce, Rockford, IL).…”

Section: Methodsmentioning

confidence: 99%

“…It is known that CGs, not only inhibit the Na ϩ -K ϩ -ATPase activity, but also induce internalization of its subunits (3,4,22). It was of interest to compare internalization of ␣1 and ␣2.…”

Section: C130 Isoforms Cardiac Glycoside Selectivity and Cell Deathmentioning

confidence: 99%

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Cardiac glycosides induced toxicity in human cells expressing α1-, α2-, or α3-isoforms of Na-K-ATPase

Lev

Karlish

Garty

2015

American Journal of Physiology-Cell Physiology

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Cherniavsky Lev M, Karlish SJ, Garty H. Cardiac glycosides induced toxicity in human cells expressing ␣1-, ␣2-, or ␣3-isoforms of Na-K-ATPase. Am J Physiol Cell Physiol 309: C126 -C135, 2015. First published May 20, 2015 doi:10.1152/ajpcell.00089.2015.-The Na ϩ -K ϩ -ATPase is specifically inhibited by cardiac glycosides, some of which may also function as endogenous mammalian hormones. Previous studies using Xenopus oocytes, yeast cells, or purified isoforms demonstrated that affinities of various cardiac glycosides for three isoforms of the Na ϩ -K ϩ -ATPase (␣1-␣3␤1) may differ, a finding with potential clinical implication. The present study investigates isoform selectivity and effects of cardiac glycosides on cultured mammalian cells under more physiological conditions. H1299 cells (non-small cell lung carcinoma) were engineered to express only one ␣-isoform (␣1, ␣2, or ␣3) by combining stable transfection of isoforms and silencing endogenous ␣1. [21153][21154][21155][21156][21157][21158][21159][21160][21161][21162] 2014). Relative ␣2:␣1 selectivity of digoxin vs. ouabain was also manifested by enhanced internalization of ␣2 in response to digoxin. Cellular proliferation assays of H1299 cells confirmed the patterns of ␣2:␣1 selectivity for ouabain, digoxin, and a synthetic derivative and reveal a crucial role of surface pump density on sensitivity to cardiac glycosides. Because cardiac glycosides are being considered as drugs for treatment of cancer, effects of ouabain on proliferation of 12 cancer and noncancer cell lines, with variable plasma membrane expression of ␣1, have been tested. These demonstrated that sensitivity to ouabain indeed depends linearly on the plasma membrane surface density of Na ϩ -K ϩ -ATPase irrespective of status, malignant or nonmalignant. cardiac glycosides; cell death; isoforms; Na-K-ATPase THE NA ϩ -K ϩ -ATPASE (the Na ϩ pump) is a ubiquitously expressed P-type ATPase that actively transports 3Na ϩ ions out of cells in exchange for 2K ϩ ions per one molecule of ATP hydrolyzed (17). The pump consists of a catalytic ␣-subunit, ␤-subunit that modulates activity and stabilizes the protein, and an auxiliary FXYD polypeptide. Four ␣-isoforms (␣1-␣4), three ␤-isoforms (␤1-␤3), and seven FXYD isoforms exist in mammalian organisms, resulting in considerable tissue-specific variability in pump structure and kinetics (21).As can be expected from plasma membrane protein with such a vital function, the Na ϩ -K ϩ -ATPase is regulated by a number of independent mechanisms (25). One such mode of regulation involves inhibition by a family of inhibitors termed cardiac glycosides (CGs). Most CGs are derived from plants.However, several so-called endogenous CGs, including ouabain, marinobufagenin, and digoxin, have been identified in mammalian tissues and are apparently synthesized by the adrenal gland (1). CGs such as digoxin are also clinically relevant specific inhibitors of the Na ϩ -K ϩ -ATPase, used classically to treat heart failure (9).Relatively limited information exists regarding th...

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Section: Discussioncontrasting

confidence: 99%

Section: Methodsmentioning

confidence: 99%

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Cardiac glycosides induced toxicity in human cells expressing α1-, α2-, or α3-isoforms of Na-K-ATPase

Lev

Karlish

Garty

2015

American Journal of Physiology-Cell Physiology

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“…2012; Cherniavsky-Lev et al. 2014). Therefore, the [ 3 H]-ouabain-binding assay may disclose a fraction of intracellular functional sodium pumps which leads to an overestimation of their surface density.…”

Section: Discussionmentioning

confidence: 99%

Airway surface liquid volume expansion induces rapid changes in amiloride-sensitive Na⁺transport across upper airway epithelium-Implications concerning the resolution of pulmonary edema

2015

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During airway inflammation, airway surface liquid volume (ASLV) expansion may result from the movement of plasma proteins and excess liquid into the airway lumen due to extravasation and elevation of subepithelial hydrostatic pressure. We previously demonstrated that elevation of submucosal hydrostatic pressure increases airway epithelium permeability resulting in ASLV expansion by 500 μL cm−2 h−1. Liquid reabsorption by healthy airway epithelium is regulated by active Na+ transport at a rate of 5 μL cm−2 h−1. Thus, during inflammation the airway epithelium may be submerged by a large volume of luminal liquid. Here, we have investigated the mechanism by which ASLV expansion alters active epithelial Na+ transport, and we have characterized the time course of the change. We used primary cultures of tracheal airway epithelium maintained under air interface (basal ASLV, depth is 7 ± 0.5 μm). To mimic airway flooding, ASLV was expanded to a depth of 5 mm. On switching from basal to expanded ASLV conditions, short-circuit current (Isc, a measure of total transepithelial active ion transport) declined by 90% with a half-time (t1/2) of 1 h. 24 h after the switch, there was no significant change in ATP concentration nor in the number of functional sodium pumps as revealed by [3H]-ouabain binding. However, amiloride-sensitive uptake of 22Na+ was reduced by 70% upon ASLV expansion. This process is reversible since after returning cells back to air interface, Isc recovered with a t1/2 of 5–10 h. These results may have important clinical implications concerning the development of Na+ channels activators and resolution of pulmonary edema.

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“…There are some data that are opposite with our model. It has been shown that c-Src activation was not involved in ouabain induced Na/K-ATPase endocytosis in non-small cell lung cancer cells [86] and no stable interactions were detected between purified recombinant Na/K-ATPase and purified human Src kinase [87]. We would like to point out that these observations are different from what we have reported using purified pig kidney Na/K-ATPase, LLC-PK1 cell, human HK-2, human dermal fibroblasts, renal fibroblast cell line, and rat cardiac fibroblasts cell line [84,88,89].…”

Section: Na/k-atpase – An Ion Pump a Signaling Receptor And A Sigmentioning

confidence: 99%

Sodium potassium adenosine triphosphatase (Na/K-ATPase) as a therapeutic target for uremic cardiomyopathy

Wang

Liu

Drummond

et al. 2017

Expert Opinion on Therapeutic Targets

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Introduction Clinically, patients with significant reductions in renal function present with cardiovascular dysfunction typically termed, uremic cardiomyopathy. It is a progressive series of cardiac pathophysiological changes, including left ventricular diastolic dysfunction and hypertrophy (LVH) which sometimes progress to left ventricular dilation (LVD) and systolic dysfunction in the setting of chronic kidney disease (CKD). Uremic cardiomyopathy is almost ubiquitous in patients afflicted with end stage renal disease (ESRD). Areas covered This article reviews recent epidemiology, pathophysiology of uremic cardiomyopathy and provide a board overview of Na/K-ATPase research with detailed discussion on the mechanisms of Na/K-ATPase/Src/ROS amplification loop. We also present clinical and preclinical evidences as well as molecular mechanism of this amplification loop in the development of uremic cardiomyopathy. A potential therapeutic peptide that targets on this loop is discussed. Expert opinion Current clinical treatment for uremic cardiomyopathy remains disappointing. Targeting the ROS amplification loop mediated by the Na/K-ATPase signaling function may provide a novel therapeutic target for uremic cardiomyopathy and related diseases. Additional studies of Na/K-ATPase and other strategies that regulate this loop will lead to new therapeutics.

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Ouabain-induced Internalization and Lysosomal Degradation of the Na+/K+-ATPase

Abstract: Background:The plasma membrane Na ϩ /K ϩ -ATPase undergoes internalization upon ouabain binding.

Cited by 29 publications

References 39 publications

Cardiac glycosides induced toxicity in human cells expressing α1-, α2-, or α3-isoforms of Na-K-ATPase

Cardiac glycosides induced toxicity in human cells expressing α1-, α2-, or α3-isoforms of Na-K-ATPase

Airway surface liquid volume expansion induces rapid changes in amiloride-sensitive Na⁺transport across upper airway epithelium-Implications concerning the resolution of pulmonary edema

Sodium potassium adenosine triphosphatase (Na/K-ATPase) as a therapeutic target for uremic cardiomyopathy

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Ouabain-induced Internalization and Lysosomal Degradation of the Na+/K+-ATPase

Abstract: Background:The plasma membrane Na ϩ /K ϩ -ATPase undergoes internalization upon ouabain binding.

Cited by 29 publications

References 39 publications

Cardiac glycosides induced toxicity in human cells expressing α1-, α2-, or α3-isoforms of Na-K-ATPase

Cardiac glycosides induced toxicity in human cells expressing α1-, α2-, or α3-isoforms of Na-K-ATPase

Airway surface liquid volume expansion induces rapid changes in amiloride-sensitive Na+transport across upper airway epithelium-Implications concerning the resolution of pulmonary edema

Sodium potassium adenosine triphosphatase (Na/K-ATPase) as a therapeutic target for uremic cardiomyopathy

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Airway surface liquid volume expansion induces rapid changes in amiloride-sensitive Na⁺transport across upper airway epithelium-Implications concerning the resolution of pulmonary edema