Histamine increases sphingosine kinase-1 expression and activity in the human arterial endothelial cell line EA.hy 926 by a PKC-α-dependent mechanism

• Published in: Biochimica et biophysica acta Vol. 1761; no. 3; pp. 367 - 376
• Main Authors: Huwiler, Andrea, Döll, Frauke, Ren, Shuyu, Klawitter, Sabine, Greening, Anna, Römer, Isolde, Bubnova, Svetlana, Reinsberg, Luise, Pfeilschifter, Josef
• Format: Journal Article
• Language: English
• Published: Netherlands Elsevier B.V 01.03.2006
• Subjects: Arteries - cytology; Cell Line; Cell Movement - physiology; Cycloheximide - metabolism; Endothelial cell; Endothelial Cells - cytology; Endothelial Cells - metabolism; Enzyme Activation; Enzyme Inhibitors - metabolism; Histamine; Histamine - metabolism; Humans; Isoenzymes - genetics; Isoenzymes - metabolism; Migration; Phorbol ester; Phosphotransferases (Alcohol Group Acceptor) - genetics; Phosphotransferases (Alcohol Group Acceptor) - metabolism; PKC isoenzyme; Protein Kinase C-alpha - metabolism; Protein Synthesis Inhibitors - metabolism; RNA, Small Interfering - genetics; RNA, Small Interfering - metabolism; Signal Transduction - physiology; Sphingosine kinase-1; Tetradecanoylphorbol Acetate - metabolism; PBS; Endothelial cell; TPA; Phorbol ester; Migration; Sphingosine kinase-1; PKC; MAPK; ECL; BSA; DMEM; SDS-PAGE; Histamine; PDGF; SK; GAPDH; PKC isoenzyme
• ISSN: 1388-1981; 0006-3002; 1879-2618; 1878-2434
• DOI: 10.1016/j.bbalip.2006.02.007

Sphingosine 1-phosphate (S1P) is a potent mitogenic signal generated from sphingosine by the action of sphingosine kinases (SKs). In this study, we show that in the human arterial endothelial cell line EA.hy 926 histamine induces a time-dependent upregulation of the SK-1 mRNA and protein expression which is followed by increased SK-1 activity. A similar upregulation of SK-1 is also observed with the direct protein kinase C activator 12- O-tetradecanoylphorbol-13-acetate (TPA). In contrast, SK-2 activity is not affected by neither histamine nor TPA. The increased SK-1 protein expression is due to stimulated de novo synthesis since cycloheximide inhibited the delayed SK-1 protein upregulation. Moreover, the increased SK-1 mRNA expression results from an increased promoter activation by histamine and TPA. In mechanistic terms, the transcriptional upregulation of SK-1 is dependent on PKC and the extracellular signal-regulated protein kinase (ERK) cascade since staurosporine and the MEK inhibitor U0126 abolish the TPA-induced SK-1 induction. Furthermore, the histamine effect is abolished by the H 1-receptor antagonist diphenhydramine, but not by the H 2-receptor antagonist cimetidine. Parallel to the induction of SK-1, histamine and TPA stimulate an increased migration of endothelial cells, which is prevented by depletion of the SK-1 by small interfering RNA (siRNA). To appoint this specific cell response to a specific PKC isoenzyme, siRNA of PKC-α, -δ, and -ε were used to selectively downregulate the respective isoforms. Interestingly, only depletion of PKC-α leads to a complete loss of TPA- and histamine-triggered SK-1 induction and cell migration. In summary, these data show that PKC-α activation in endothelial cells by histamine-activated H 1-receptors, or by direct PKC activators leads to a sustained upregulation of the SK-1 protein expression and activity which, in turn, is critically involved in the mechanism of endothelial cell migration.