Oleoyl-Lysophosphatidylcholine Limits Endothelial Nitric Oxide Bioavailability by Induction of Reactive Oxygen Species

Abstract

Previously we reported modulation of endothelial prostacyclin and interleukin-8 production, cyclooxygenase-2 expression and vasorelaxation by oleoyl- lysophosphatidylcholine (LPC 18:1). In the present study, we examined the impact of this LPC on nitric oxide (NO) bioavailability in vascular endothelial EA.hy926 cells. Basal NO formation in these cells was decreased by LPC 18:1. This was accompanied with a partial disruption of the active endothelial nitric oxide synthase (eNOS)- dimer, leading to eNOS uncoupling and increased formation of reactive oxygen species (ROS). The LPC 18:1-induced ROS formation was attenuated by the superoxide scavenger Tiron, as well as by the pharmacological inhibitors of eNOS, NADPH oxidases, flavin-containing enzymes and superoxide dismutase (SOD). Intracellular ROS-formation was most prominent in mitochondria, less pronounced in cytosol and undetectable in endoplasmic reticulum. Importantly, Tiron completely prevented the LPC 18:1-induced decrease in NO bioavailability in EA.hy926 cells. The importance of the discovered findings for more in vivo like situations was analyzed by organ bath experiments in mouse aortic rings. LPC 18:1 attenuated the acetylcholine-induced, endothelium dependent vasorelaxation and massively decreased NO bioavailability. We conclude that LPC 18:1 induces eNOS uncoupling and unspecific superoxide production. This results in NO scavenging by ROS, a limited endothelial NO bioavailability and impaired vascular function.

Lysophosphatidylcholine induces cytotoxicity/apoptosis and IL-8 production of human endothelial cells: Related mechanisms

“…The above event is related to the induction of ROS over-production by LPC. Recently oleoyl-LPC has been shown to cause a decrease of NO production, and eNOS uncoupling, but increase of ROS production in EAHY endothelial cells [ 13 ]. Peng et al (2010) also found the inhibition of NO, stimulation of ROS by LPC to stimulate caspase 3-dependent apoptotic response in endothelial cells [ 10 ].…”

MicroRNA-142-3p improves vascular relaxation in uremia

“…Wire myography was done as previously described [20]. Briefly, mouse aortic rings 2 mm in length were isolated and positioned in small wire myograph chambers (Danish MyoTechnology, Aarhus, Denmark) containing physiological salt solution (PSS) (114 mM NaCl, 4.7 mM KCl, 0.8 mM KH 2 PO 4 , 1.2 mM MgCl 2 , 2.5 mM CaCl 2 , 25 mM NaHCO 3 and 11 mM d -glucose pH 7.4).…”

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“…EA.hy 926 cells were plated (250,000 cells/well) onto 6 well plates. After 24 h, confluent cells were treated with 100 μg/mL EV-HDL or EL-HDL in DMEM without FCS for 5 or 16 h. Intracellular conversion of L- [ 3 H]arginine into L-[ 3 H]citrulline was measured as previously described [ 17 ]. Briefly, after incubation with EV-HDL or EL-HDL, cells were washed and incubated at 37 °C with 50 mmol/L Tris buffer, pH 7.4, containing 100 mmol/L NaCl, 5 mmol/L KCl, 1 mmol/L MgCl 2 , 3 mmol/L CaCl 2 , 5% (vol/vol) FCS, and L-[2,3- 3 H]arginine (~106 dpm) in the absence or presence of 0.5 μmol/L calcium iono- phore A23187.…”

“…Next, the rings were contracted with increasing concentrations of norepinephrine (NE) (1 nmol/L – 0.3 μmol/L) (Sigma-Aldrich) to produce 80% of the maximum contraction achieved by modified PSS followed by endothelium-dependent relaxation to cumulatively increasing concentrations of acetylcholine chloride (1 nmol/L – 0.3 μmol/L) (Sigma- Aldrich). After washout and equilibration, the rings were constricted with one dose of NE in the presence or absence of L-NNA (300 μmol/L) to 80% of the maximal constriction achieved by modified PSS [ 17 ]. Thereafter, 100 μg/mL EV-HDL protein or EL-HDL protein were added to the rings in the absence or presence of L-NNA (300 μmol/L).…”

“…After relaxation achieved by switching from modified PSS to PSS, the rings were exposed to the required NE concentrations for 10% of the maximal KCl-induced constriction, followed by addition of L-NNA (300 μmol/L). The ratio of constriction observed after and before addition of L-NNA is indicative of NO availability, whereby a higher ratio indicates a higher NO availability [ 17 ].…”