Thrombin induces NO release from cultured rat microglia via protein kinase C, mitogen-activated protein kinase, and NF-kappa B.
Microglia, brain resident macrophages, become activated in brains injured due to trauma, ischemia, or neurodegenerative diseases. In this study, we found that thrombin treatment of microglia induced NO release/inducible nitric-oxide synthase expression, a prominent marker of activation. The effect of thrombin on NO release increased dose-dependently within the range of 5-20 units/ml. In immunoblot analyses, inducible nitric-oxide synthase expression was detected within 9 h after thrombin treatment. This effect of thrombin was significantly reduced by protein kinase C inhibitors, such as Go6976, bisindolylmaleimide, and Ro31-8220. Within 15 min, thrombin activated three subtypes of mitogen-activated protein kinases: extracellular signal-regulated kinase, p38, and c-Jun N-terminal kinase/stress-activated protein kinase. Inhibition of the extracellular signal-regulated kinase pathway and p38 reduced the NO release of thrombin-treated microglia. Thrombin also activated nuclear factor kappaB (NF-kappaB) within 5 min, and N-acetyl cysteine, an inhibitor of NF-kappaB, reduced NO release. However, thrombin receptor agonist peptide (an agonist of protease activated receptor-1 (PAR-1)), could not mimic the effect of thrombin, and cathepsin G, a PAR-1 inhibitor, did not reduce the effect of thrombin. These results suggest that thrombin can activate microglia via protein kinase C, mitogen-activated protein kinases, and NF-kappaB but that this occurs independently of PAR-1.
Ryu
J
J
Department of Pharmacology, Ajou University School of Medicine, Suwon, 442-721, Korea.
Pyo
H
H
Jou
I
I
Joe
E
E
D000111:Acetylcysteine / Q000494:pharmacology; D000818:Animals; D002227:Carbazoles / Q000494:pharmacology; D002478:Cells, Cultured; D002540:Cerebral Cortex / Q000166:cytology; D004790:Enzyme Induction; D005419:Flavonoids / Q000494:pharmacology; D007093:Imidazoles / Q000494:pharmacology; D007211:Indoles / Q000494:pharmacology; D008301:Maleimides / Q000494:pharmacology; D017628:Microglia / Q000166:cytology / Q000187:drug effects*; D020928:Mitogen-Activated Protein Kinases / Q000037:antagonists & inhibitors / Q000378:metabolism*; D016328:NF-kappa B / Q000378:metabolism*; D009569:Nitric Oxide / Q000378:metabolism*; D019001:Nitric Oxide Synthase / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D011493:Protein Kinase C / Q000037:antagonists & inhibitors / Q000378:metabolism*; D011506:Proteins / Q000494:pharmacology; D011725:Pyridines / Q000494:pharmacology; D051381:Rats; D017207:Rats, Sprague-Dawley; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin / Q000819:agonists / Q000378:metabolism; D015398:Signal Transduction; D013917:Thrombin / Q000494:pharmacology*
D002227:Carbazoles; D005419:Flavonoids; D007093:Imidazoles; D007211:Indoles; D008301:Maleimides; D016328:NF-kappa B; D011506:Proteins; D011725:Pyridines; D044463:Receptor, PAR-1; D018179:Receptors, Thrombin; C081021:Go 6976; D009569:Nitric Oxide; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496318:Nos2 protein, rat; D011493:Protein Kinase C; D020928:Mitogen-Activated Protein Kinases; D013917:Thrombin; C088060:bisindolylmaleimide; C093642:SB 203580; C093973:2-(2-amino-3-methoxyphenyl)-4H-1-benzopyran-4-one; C064758:Ro 31-8220; D000111:Acetylcysteine