Lipopolysaccharide-induced expression of cyclooxygenase-2 in mouse macrophages is inhibited by chloromethylketones and a direct inhibitor of NF-kappa B translocation.
In macrophages, cyclooxygenase-2 (COX-2) is induced by cytokines, mitogens, or endotoxin. The present study investigates whether inhibitors of the nuclear transcription factor NF-kappa B affect lipopolysaccharide (LPS)-mediated expression of COX-2 mRNA, protein, and activity in the macrophage cell line J774.1A. The activation of COX-2 was assessed by measuring the accumulation of prostaglandin (PG) E2 by radioimmunoassay. Expression of COX-2 mRNA and protein was detected by Northern and Western blot analysis, respectively. In the absence of LPS, mouse macrophages did not express COX-2 and generated low amounts of prostaglandin (PG) E2. Treatment of J774.1A with LPS (0.1-30 micrograms/ml) caused expression of COX-2 protein and activity. Induction of COX-2 activity along with the induction of COX-2 mRNA and protein by LPS was attenuated by the serine protease inhibitors N-alpha-tosyl-L-phenylalanine chloromethyl ketone (TPCK) and N-alpha-tosyl-L-lysine chloromethyl ketone (TLCK). A cell permeable peptide and a direct inhibitor of NF-kappa B translocation, SN50, attenuated the accumulation of PGE2 in cell supernatant in a concentration-dependent manner. Our results show that induction of COX-2 by LPS in macrophages involves activation of NF-kappa B and point to a possible therapeutic use of protease inhibitors in inflammatory processes.
Abate
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A
Department of Pharmacology and Toxicology, School of Pharmacy, Martin Luther University, Halle (Saale), Germany.
Oberle
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Schröder
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H
D000818:Animals; D015152:Blotting, Northern; D015153:Blotting, Western; D002460:Cell Line; D002470:Cell Survival; D051546:Cyclooxygenase 2; D015232:Dinoprostone / Q000502:physiology; D004305:Dose-Response Relationship, Drug; D007527:Isoenzymes / Q000096:biosynthesis*; D008070:Lipopolysaccharides / Q000037:antagonists & inhibitors*; D008264:Macrophages / Q000502:physiology*; D051379:Mice; D016328:NF-kappa B / Q000378:metabolism*; D010455:Peptides / Q000494:pharmacology; D011451:Prostaglandin-Endoperoxide Synthases / Q000096:biosynthesis*; D013347:Subcellular Fractions / Q000737:chemistry; D013997:Time Factors; D014107:Tosyllysine Chloromethyl Ketone / Q000494:pharmacology*; D014108:Tosylphenylalanyl Chloromethyl Ketone / Q000494:pharmacology*