Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Brkic, L.
Acyl chain-dependent effect of lysophosphatidylcholine on cyclooxygenase (COX)-2 expression in endothelial cells
[ Dissertation ] Medical University of Graz; 2013. pp. 77
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- Autor*innen der Med Uni Graz:
- Brkic Lada
- Betreuer*innen:
- Frank Sasa
- Marsche Gunther
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- Abstract:
- Endothelial lipase (EL) is a serum phospholipase produced mainly by vascular endothelial cells. EL generates substantial amounts of saturated palmitoyl lysophosphatidyl-choline (16:0 LPC) and unsaturated oleoyl (18:1 LPC), linoleyl (18:2 LPC) and arachidonoyl lysophosphatidyl-choline (20:4 LPC) by cleaving its major substrate high-density lipoprotein (HDL) phosphatidylcholine. Because EL is active on the surface of vascular endothelial cells and its expression is upregulated by inflammation, EL-derived LPCs generated in close proximity of vascular endothelium might be important modulators of endothelial function. While 16:0 LPC is an established activator of various signaling cascades and modulator of expression of various genes including cyclooxygenase (COX)-2, almost nothing is known about signaling and gene-expression properties of unsaturated LPCs. Therefore, the aim of the present study was to examine the signaling properties of unsaturated LPCs in comparison to 16:0 LPC with an emphasis on intracellular Ca2+ homeostasis, activation of MAP kinases and nuclear transcription factors as well as modulation of COX-2 expression. Quantitative PCR experiment showed that 16:0, 18:1, 20:4, but not 18:2 LPC raised COX-2 mRNA expression in human umbilical vein endothelial cell-derived cell line EA.hy 926, with profoundly different potencies and kinetics. Similarly, Western blot analysis revealed profound differences in the capacity and kinetics of the tested LPCs to upregulate COX-2 protein. Interestingly, COX-e protein was increased by 18:2 but not with 18:1 LPC. Experiments performed in the presence of pharmacological inhibitors of various signalling pathways revealed the involvement of intracellular Ca2+ and p38 MAPK in LPC-elicited COX-2 expression. All tested LPCs were capable of increasing intracellular Ca2+ concentration, however with different potencies as found by fluorescence spectrometry in Fura-2/AM loaded EA.hy 926 cells. The LPC-elicited increase in [Ca2+]i was dependent on phospholipase C (PLC) and inositol-triphosphate receptor (IP3R). Furthermore 16:0. 18:1 and 20:4 LPC induced p38 MAPK activation with markedly different kinetics, as found by monitoring p38 MAPK phosphorylation by Western blot. The involvement and markedly different relative contribution of selected transcription factors, including cyclic AMP responsive element binding protein (CREB), c-Jun and nuclear factor kappa B (NF-¿B), in LPC-induced COX-2 upregulation was found upon knock-down of those transcription factors by siRNA approach. Collectively, the obtained data strongly argue that the tested LPCs exhibit remarkably different, acyl chain-related potencies and kinetics of COX-2 induction in human endothelial cells, dependent on intracellular Ca2+ and p38 MAPK, as well as nuclear factors NF-¿B, c-Jun and CREB. Considering their high plasma levels together with their simultaneous action on endothelial COX-2 in vivo, the tested LPCs emerged as potent and important regulators of vascular (patho)biology.