Proinflammatory gene induction by platelet-activating factor mediated via its cognate nuclear receptor.
It has been postulated that intracellular binding sites for platelet-activating factor (PAF) contribute to proinflammatory responses to PAF. Isolated nuclei from porcine cerebral microvascular endothelial cells (PCECs) produced PAF-molecular species in response to H(2)O(2). Using FACS analysis, we demonstrated the expression of PAF receptors on cell and nuclear surfaces of PCECs. Confocal microscopy studies performed on PCECs, Chinese hamster ovary cells stably overexpressing PAF receptors, and isolated nuclei from PCECs also showed a robust nuclear distribution of PAF receptors. Presence of PAF receptors at the cell nucleus was further revealed in brain endothelial cells by radioligand binding experiments, immunoblotting, and in situ in brain by immunoelectron microscopy. Stimulation of nuclei with methylcarbamate-PAF evoked a decrease in cAMP production and a pertussis toxin-sensitive rise in nuclear calcium, unlike observations in plasma membrane, which exhibited a pertussis toxin-insensitive elevation in inositol phosphates. Moreover, on isolated nuclei methylcarbamate-PAF evoked the expression of proinflammatory genes inducible nitric oxide synthase and cyclooxygenase-2 (COX-2) and was associated with augmented extracellular signal-regulated kinase 1/2 phosphorylation and NF-kappaB binding to the DNA consensus sequence. COX-2 expression was prevented by mitogen-activated protein kinase kinase/extracellular signal-regulated kinase 1/2 and NF-kappaB inhibitors. This study describes for the first time the nucleus as a putative organelle capable of generating PAF and expresses its receptor, which upon stimulation induces the expression of the proinflammatory gene COX-2.
Marrache
A Marilise
AM
Department of Pharmacology and Therapeutics, McGill University, Montreal, Quebec, Canada.
Gobeil
Fernand
F
Bernier
Sylvie G
SG
Stankova
Jana
J
Rola-Pleszczynski
Marek
M
Choufani
Sanaa
S
Bkaily
Ghassan
G
Bourdeau
Annie
A
Sirois
Martin G
MG
Vazquez-Tello
Alejandro
A
Fan
Li
L
Joyal
Jean-Sébastien
JS
Filep
Janos G
JG
Varma
Daya R
DR
Ribeiro-Da-Silva
Alfredo
A
Chemtob
Sylvain
S
D000818:Animals; D001665:Binding Sites / Q000235:genetics; D016466:CHO Cells; D002467:Cell Nucleus / Q000378:metabolism / Q000648:ultrastructure; D002478:Cells, Cultured; D016384:Consensus Sequence; D006224:Cricetinae; D051546:Cyclooxygenase 2; D004247:DNA / Q000235:genetics / Q000378:metabolism; D004730:Endothelium, Vascular / Q000378:metabolism / Q000648:ultrastructure; D015971:Gene Expression Regulation, Enzymologic; D018836:Inflammation Mediators / Q000378:metabolism*; D007527:Isoenzymes / Q000235:genetics; D008854:Microscopy, Electron; D019001:Nitric Oxide Synthase / Q000235:genetics; D052247:Nitric Oxide Synthase Type II; D010972:Platelet Activating Factor / Q000378:metabolism*; D010980:Platelet Membrane Glycoproteins / Q000378:metabolism*; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D011956:Receptors, Cell Surface / Q000378:metabolism*; D043562:Receptors, G-Protein-Coupled; D015398:Signal Transduction; D013552:Swine; D015533:Transcriptional Activation
D018836:Inflammation Mediators; D007527:Isoenzymes; D010972:Platelet Activating Factor; D010980:Platelet Membrane Glycoproteins; D012333:RNA, Messenger; D011956:Receptors, Cell Surface; D043562:Receptors, G-Protein-Coupled; C039351:platelet activating factor receptor; D004247:DNA; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases