Induction of heme oxygenase-1 attenuates lipopolysaccharide-induced cyclooxygenase-2 expression in mouse brain endothelial cells.
BACKGROUND
Prostaglandin E2 (PGE2), an arachidonic acid metabolite converted by cyclooxygenase-2 (COX-2), plays important roles in the regulation of endothelial functions in response to bacterial infection. The enzymatic activity of COX-2 can be down-regulated by heme oxygenase-1 (HO-1) induction. However, the mechanisms underlying HO-1 modulating COX-2 protein expression are not known.


OBJECTIVE
The aim of the present study was to investigate whether the up-regulation of HO-1 regulates COX-2 expression induced by lipopolysaccharide (LPS), an endotoxin produced by Gram negative bacteria, in mouse brain endothelial cells (bEnd.3)


METHODS
Cultured bEnd.3 cells were used to investigate LPS-induced COX-2 expression and PGE2 production. Cobalt protoporphyrin IX (CoPP, an HO-1 inducer), infection with a recombinant adenovirus carried with HO-1 gene (Adv-HO-1), or zinc protoporphyrin (ZnPP, an HO-1 inhibitor) was used to stimulate HO-1 induction or inhibit HO-1 activity. The expressions of COX-2 and HO-1 were evaluated by western blotting. PGE2 levels were detected by an enzyme-linked immunoassay. Hemoglobin (a chelator of carbon monoxide, CO, one of metabolites of HO-1) and CO-RM2 (a CO releasing molecule) were used to investigate the mechanisms of HO-1 regulating COX-2 expression.


RESULTS
We found that LPS-induced COX-2 expression and PGE2 production were mediated through NF-κB (p65) via activation of Toll-like receptor 4 (TLR4). LPS-induced COX-2 expression was inhibited by HO-1 induction by pretreatment with CoPP or infection with Adv-HO-1. This inhibitory effect of HO-1 was reversed by pretreatment with either ZnPP or hemoglobin. Pretreatment with CO-RM2 also inhibited TLR4/MyD88 complex formation, NF-κB (p65) activation, COX-2 expression, and PGE2 production induced by LPS.


CONCLUSIONS
We show here a novel inhibition of HO-1 on LPS-induced COX-2/PGE2 production in bEnd.3. Our results reinforce the emerging role of cerebral endothelium-derived HO-1 as a protector against cerebral vascular inflammation triggered by bacterial infection.
Department of Physiology and Pharmacology, Chang Gung University, Kwei-San, Tao-Yuan, Taiwan.
Ruey-Horng
RH
Shih
Chuen-Mao
CM
Yang
D000818:Animals; D002478:Cells, Cultured; D002560:Cerebrovascular Circulation / Q000187:drug effects* / Q000502:physiology; D051546:Cyclooxygenase 2 / Q000235:genetics / Q000378:metabolism*; D015232:Dinoprostone / Q000378:metabolism; D042783:Endothelial Cells / Q000166:cytology / Q000187:drug effects* / Q000201:enzymology*; D004730:Endothelium, Vascular / Q000166:cytology* / Q000187:drug effects*; D051547:Heme Oxygenase-1 / Q000235:genetics / Q000378:metabolism*; D006801:Humans; D008070:Lipopolysaccharides / Q000494:pharmacology*; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism; D015854:Up-Regulation
D008070:Lipopolysaccharides; D016328:NF-kappa B; D051547:Heme Oxygenase-1; D051546:Cyclooxygenase 2; D015232:Dinoprostone