Recent studies have demonstrated the tissue-specific effect of Na+/K+ pump inhibition by
ouabain and other
cardiac glycosides on cell viability. The vascular endothelium is an initial target of
cardiac glycosides employed for the management of
congestive heart failure as well as circulating endogenous
ouabain-like substances (EOLS), the production of which is augmented in volume-expanded
hypertension. This study examined the role of the Na+/K+ pump in the survival of cultured porcine aortic endothelial cells (PAEC). Complete Na+/K+ pump inhibition with
ouabain led to PAEC death, indicated by cell detachment and decreased staining with
3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT). Based on cell swelling and resistance to
benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (
z-VAD.fmk) a pan-
caspase inhibitor, this type of cell death was classified as
necrosis. In contrast to
ouabain, Na+/K+ pump inhibition in K+-free medium did not affect PAEC viability and sharply attenuated apoptosis triggered by 3H decay-induced DNA damage.
Necrosis evoked by
ouabain was preserved after dissipation of the transmembrane gradient of K+ and Na+, whereas dissipation of the Na+ gradient abolished the antiapoptotic action of K+-free medium. Comparative analysis of these results and modulation of intracellular Na+ and K+ content by the above-listed stimuli showed that interaction of
ouabain with Na+/K+-
ATPase triggered
necrosis independently of inhibition of Na+/K+ pump-mediated ion fluxes and inversion of the [Na+]i/[K+]i ratio, whereas protection against apoptosis under Na+/K+ pump inhibition in K+-depleted medium was mediated by [Na+]i elevation. The role of Na+/K+ pump-mediated regulation of endothelial cell survival and vascular remodelling seen in
hypertension should be investigated further in context of EOLS and chronic treatment with digitalis.