Lysophosphatidylcholine-induced modulation of Ca2+-activated K+channels contributes to ROS-dependent proliferation of cultured human endothelial cells

Proliferation of endothelial cells plays a crucial role in the process of atherosclerotic plaque destabilization. The major component of oxidized low-density lipoprotein lysophosphatidylcholine (LPC) has been shown to promote endothelial proliferation by increasing the production of reactive oxygen species (ROS). Since K+ channels are known to control the cell cycle, we investigated the role of Ca2+-activated K+ channels (BKCa) in the regulation of LPC-induced endothelial proliferation and ROS generation. A significant increase of cell growth induced by LPC (20 mumol/l; cell counts (CCs): +87%, thymidin incorporation: +89%; n = 12, P < 0.01) was observed, which was inhibited by the BKCa inhibitor iberiotoxin (IBX; 100 nmol/l), by the NAD(P)H-oxidase inhibitor diphenyleneiodonium (5 mumol/l) and by transfection with antisense (AS) oligonucleotides against NAD(P)H oxidase, whereas N-G-monomethyl-L-arginine (L-NMMA) further increased LPC-induced cell growth. Using the patch-clamp technique a significant increase of BKCa open-state probability (control: 0.004 +/- 0.002; LPC: 0.104 +/- 0.035; n = 21, P < 0.05) by LPC was observed. Using dichlorofluorescein fluorescence microscopy a significant increase of ROS induced by LPC was reported, that was blocked by IBX and Ca2+ antagonists. Intracellular Ca2+ measurements revealed a capacitative Ca2+ influx caused by LPC. Bioactivity of nitric oxide (NO) was measured using a [H-3]-cGMP radioimmunoassay. LPC significantly decreased acetylcholine-induced NO synthesis. LPC significantly increased cGMP levels in endothelial cells transfected with AS, which was blocked by IBX. In conclusion, our results demonstrate that LPC activates BKCa thereby increasing ROS production which induces endothelial proliferation. In addition LPC-induced BKCa-activation contributes to increased cGMP levels, if ROS production is prevented by AS. (C) 2004 Elsevier Ltd. All rights reserved.