Analysis of inflammatory gene induction by oxidized phospholipids in vivo by quantitative real-time RT-PCR in comparison with effects of LPS.
Oxidized phospholipids are thought to play a role in the development of atherosclerosis and other chronic inflammatory processes. In this study, we analyzed the expression of inflammatory genes induced by oxidized L-alpha-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholin (OxPAPC) in vitro and in vivo using quantitative real-time reverse transcriptase-polymerase chain reaction (RT-PCR). Cultured human umbilical vein endothelial cells (HUVEC) and monocyte-like U937 cells were treated with OxPAPC or lipopolysaccharide (LPS) for 3 h. For in vivo studies, OxPAPC or LPS was injected intravenously into female C57Bl/6J mice and different tissues were isolated after 3 h. We found that both OxPAPC and LPS induced expression of early growth response factor 1 (EGR-1) and monocyte chemoattractant protein 1 (MCP-1) in HUVEC and of JE, the mouse homologue of MCP-1, in liver and heart. Interestingly, OxPAPC but not LPS increased expression of heme oxygenase 1 (HO-1) in U937 cells, HUVEC, aorta, heart, liver, and isolated blood cells. In contrast, E-selectin was selectively induced by LPS, but not by OxPAPC. Finally, OxPAPC-induced expression of HO-1 was blocked by a platelet-activating factor (PAF) receptor antagonist. We conclude that oxidized phospholipids are biologically active in vivo and exert a specific response inducing a pattern of genes that is different from that induced by LPS. In addition, we demonstrate that the quantitative real-time RT-PCR technology is a proper tool to investigate differential inflammatory gene induction in vivo.
Kadl
Alexandra
A
Department of Vascular Biology and Thrombosis Research, University of Vienna, Schwarzspanierstrasse 17, 1090 Vienna, Austria.
Huber
Joakim
J
Gruber
Florian
F
Bochkov
Valery N
VN
Binder
Bernd R
BR
Leitinger
Norbert
N
D000465:Algorithms; D000818:Animals; D001324:Autoantigens / Q000096:biosynthesis; D018932:Chemokine CCL2; D046508:Culture Techniques; D018076:DNA, Complementary / Q000096:biosynthesis / Q000235:genetics; D004268:DNA-Binding Proteins / Q000096:biosynthesis; D051766:Early Growth Response Protein 1; D005260:Female; D005786:Gene Expression Regulation / Q000187:drug effects*; D017874:Immediate-Early Proteins; D007249:Inflammation / Q000235:genetics* / Q000473:pathology*; D007275:Injections, Intravenous; D008070:Lipopolysaccharides / Q000494:pharmacology*; D008099:Liver / Q000187:drug effects / Q000378:metabolism; D051379:Mice; D008810:Mice, Inbred C57BL; D010084:Oxidation-Reduction; D010713:Phosphatidylcholines / Q000494:pharmacology*; D010743:Phospholipids / Q000008:administration & dosage / Q000494:pharmacology*; D010980:Platelet Membrane Glycoproteins / Q000037:antagonists & inhibitors; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D011956:Receptors, Cell Surface / Q000037:antagonists & inhibitors; D043562:Receptors, G-Protein-Coupled; D020133:Reverse Transcriptase Polymerase Chain Reaction; D014157:Transcription Factors / Q000096:biosynthesis; D015533:Transcriptional Activation; D015854:Up-Regulation / Q000187:drug effects
D001324:Autoantigens; C491768:Ccl2 protein, mouse; D018932:Chemokine CCL2; D018076:DNA, Complementary; D004268:DNA-Binding Proteins; D051766:Early Growth Response Protein 1; C486109:Egr1 protein, mouse; D017874:Immediate-Early Proteins; D008070:Lipopolysaccharides; D010713:Phosphatidylcholines; D010743:Phospholipids; D010980:Platelet Membrane Glycoproteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; D014157:Transcription Factors; C472349:oxidized-L-alpha-1-palmitoyl-2-arachidonoyl-sn-glycero-3-phosphorylcholine; C039351:platelet activating factor receptor