mCD14 Expression in Human Monocytes Is Downregulated by Ouabain via Transactivation of Epithelial Growth Factor Receptor and Activation of p38 Mitogen-Activated Protein Kinase

Abstract

Background and Aims: The steroid ouabain is found in plasma and in many mammalian tissues, and is now considered as a hormone. In the immune system, ouabain regulates a number of lymphocyte functions, but little is known about its effects on monocyte function. Monocytes are important for adequate immune responses. The aim of this work was to analyze the effect of ouabain on mCD14 expression, a surface molecule involved in the response against Gram-negative bacteria and phagocytosis. Methods: Human peripheral blood mononuclear cells obtained from healthy donors were separated by density gradient centrifugation. Monocytes were separated by adherence and treated for 24 h with 100 nM ouabain. mCD14, CD1a and P-p38 expression was analyzed by flow cytometry. Inhibitors of cell-signaling pathways, i.e. SB202190, reduced glutathione, rottlerin, tyrphostin A23, genistein, chelerythrine chloride, PD98059, PP1 and Ly 294002, were used concomitantly with ouabain to observe their effect on mCD14 expression. Results: Ouabain induced a significant decrease in mCD14 expression. This feature was not related to receptor endocytosis or cell death. Furthermore, mCD14 downregulation did not reflect a shift in differentiation into dendritic cells because this hormone failed to induce CD1a expression. Amongst several inhibitors of cell-signaling pathways triggered by ouabain, only epidermal growth factor receptor (EGFR) and p38 mitogen-activated protein kinase (MAPK) inhibitors (tyrphostin A23 and SB202109) significantly reverted the effect of ouabain on mCD14 expression. Accordingly, the levels of P-p38 were increased on monocytes after ouabain treatment. However, incubation with epidermal growth factor did not alter mCD14 expression. Conclusion: These findings suggest that ouabain downregulates mCD14 expression on monocytes through EGFR transactivation and p38 MAPK activation.

Death of ouabain-treated renal epithelial cells: evidence for p38 MAPK-mediated Na i + /K i + -independent signaling

“…Thus, Alvardo-Kristensson and co-authors reported that p38 MAPK contributes to survival of human neutrophils by consequent phosphorylation and inhibition of caspase-8 and caspase-3 [32]. Valente and co-workers demonstrated that ouabain augments SB 202109-sensitive p38 phosphorylation in human peripheral blood mononuclear cells without any impact on cell survival [33]. In contrast, mouse and human macrophages apoptosis triggered by activation of P2X receptors and ROS is mediated by p38 activation [34,35].…”

Human cytomegalovirus induction of a unique signalsome during viral entry into monocytes mediates distinct functional changes: a strategy for viral dissemination

“…The activation of monocytic cell line surface EGFR and its downstream signaling cascades can stimulate the secretion of proinflammatory cytokines [70] and the acquisition of an antiapoptotic state [72]. Valente et al [73] found that the EGFRK activity of primary human monocytes mediated the up-regulation of CD14 by steroid ouabain. In accord with these observations, we have shown recently, for the first time, low-level EGFR expression on the surface of primary human peripheral blood monocytes [19], which has been confirmed subsequently [74].…”

Ouabain-induced alterations in ABCB1 of mesenteric lymph nodes and thymocytes of rats and mice

“…Certain studies have shown important implications of the ouabain hormone in cell proliferation, muscle contraction and cell survival in distinct cell types (32)(33)(34). In the immune system, ouabain modulates the expression of several important receptors in lymphocytes and monocytes (35,36), impairs lymphocyte proliferation induced by mitogens (36) and induces cytokine secretion in PBMC (37,38). In addition, the in vivo administration of ouabain has been shown to reduce the amount of mature B lymphocytes in the spleen and prevent the inflammation induced by several substances in mice (39,40).…”