Ouabain exacerbates activation-induced cell death in human peripheral blood lymphocytes

Mabel B Esteves,Ottília R Affonso-Mitidieri,Vivian M Rumjanek,Luis F Marques-Santos

doi:10.1590/s0001-37652005000200008

Abstract

Lymphocytes activated by mitogenic lectins display changes in transmembrane potential, an elevation in the cytoplasmic Ca2+ concentrations, proliferation and/or activation induced cell death. Low concentrations of ouabain (an inhibitor of Na+,K+-ATPase) suppress mitogen-induced proliferation and increases cell death. To understand the mechanisms involved, a number of parameters were analyzed using fluorescent probes and flow cytometry. The addition of 100 nM ouabain to cultures of peripheral blood lymphocytes activated with 5 microg/ml phytohemagglutinin (PHA) did not modify the increased expression of the Fas receptor or its ligand FasL induced by the mitogen. However, treatment with ouabain potentiated apoptosis induced by an anti-Fas agonist antibody. A synergy between ouabain and PHA was also observed with regard to plasma membrane depolarization. PHA per se did not induce dissipation of mitochondrial membrane potential but when cells were also exposed to ouabain a marked depolarization could be observed, and this was a late event. It is possible that the inhibitory effect of ouabain on activated peripheral blood lymphocytes involves the potentiation of some of the steps of the apoptotic process and reflects an exacerbation of the mechanism of activation-induced cell death.

Highlights

Ouabain is a cardiotonic steroid that has been widely used, as well as other cardiac glycosides, in the clinic
The molecules Fas and FasL are expressed in PHAactivated lymphocytes independent of their exposure to ouabain
In the Jurkat cell line, ouabain potentiates Fas-induced apoptosis and the death program triggered by this molecule (Bortner et al 2001, Nobel et al 2000)

Summary

Introduction

Ouabain is a cardiotonic steroid that has been widely used, as well as other cardiac glycosides, in the clinic. The known mode of action of ouabain is through the inhibition of the Na+,K+ pump. The pump uses the energy of hydrolysis of one ATP molecule to transport to the Quastel and Kaplan (1968) were the first to describe the inhibition by ouabain of lymphocyte proliferation induced by PHA. A number of workers, including ourselves, has described the inhibitory effects of ouabain on lymphocyte proliferation induced by various stimuli such as mitogens (Szamel et al 1981, Moraes et al 1989, Olej et al 1998), phorbol ester (Olej et al 1994, Brodie et al 1995), CD3 (Szamel et al 1995), Interleukin-2 (Redondo et al 1986) and calcium ionophore (Jensen et al 1977).

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