Induction of cyclooxygenase-2 by heat shock protein 60 in macrophages and endothelial cells.
The 60-kDa heat shock protein (HSP60), an endogenous ligand for the toll-like 4 receptor, is generated in response to inflammation, tissue injury, and/or stress and stimulates macrophages to produce cytotoxic and proinflammatory mediators including nitric oxide, tumor necrosis factor (TNF)-alpha, interleukin (IL)-6, and IL-12. In the present studies we report that HSP60 is an effective inducer of cyclooxygenase-2 (COX-2) in macrophages, as well as endothelial cells. In both cell types, the synthesis of COX-2 was coordinate with induction of nitric oxide synthase (NOS)-2 and with nitric oxide production. With the use of promoter constructs in transient transfection assays, optimal expression of COX-2 in macrophages was found to require nuclear factor (NF)-kappaB, the cAMP-response element (CRE), and NF-IL-6, but not the E-box. Mobility shift assays revealed that HSP60 induced NF-kappaB and CRE binding activity, while CCAAT/enhancer binding protein (C/EBP), which binds to NF-IL-6, was constitutively active in the cells. Both c-Jun and CRE binding protein (CREB) bound to the CRE, while C/EBP-beta bound to NF-IL-6. These data indicate that NF-kappaB, C/EBP-beta, c-Jun, and CREB are important in HSP60-induced expression of COX-2. The c-Jun-NH(2)-terminal kinase (JNK), p44/42 mitogen-activated protein (MAP) kinase [extracellular signal-regulated kinase 1/2 (ERK1/2)], and p38 MAP kinase were rapidly activated by HSP60 in the macrophages. PD-98059, an inhibitor of phosphorylation of ERK1/2, caused a marked inhibition of HSP60-induced COX-2 and NOS-2 expression. Unexpectedly, SB-203580, a p38 kinase antagonist, was found to block HSP60-induced expression of COX-2, but not NOS-2. These data indicate that both ERK1/2 kinase and p38 kinase play a role in regulating HSP60-induced expression of COX-2.
Billack
Blase
B
Department of Pharmacology and Toxicology, Rutgers University, Piscataway, NJ 08854, USA.
Heck
Diane E
DE
Mariano
Thomas M
TM
Gardner
Carol R
CR
Sur
Runa
R
Laskin
Debra L
DL
Laskin
Jeffrey D
JD
D000818:Animals; D022762:CCAAT-Enhancer-Binding Proteins / Q000378:metabolism; D002478:Cells, Cultured; D018834:Chaperonin 60 / Q000494:pharmacology*; D017362:Cyclic AMP Response Element-Binding Protein / Q000378:metabolism; D051546:Cyclooxygenase 2; D004727:Endothelium / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D004790:Enzyme Induction / Q000187:drug effects*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation / Q000187:drug effects*; D017930:Genes, Reporter; D007371:Interferon-gamma / Q000494:pharmacology; D007527:Isoenzymes / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism*; D008070:Lipopolysaccharides / Q000494:pharmacology; D008264:Macrophages / Q000166:cytology / Q000187:drug effects / Q000201:enzymology*; D051379:Mice; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D016297:Mutagenesis, Site-Directed; D009569:Nitric Oxide / Q000096:biosynthesis; D019001:Nitric Oxide Synthase / Q000235:genetics / Q000378:metabolism; D052247:Nitric Oxide Synthase Type II; D011401:Promoter Regions, Genetic; D011451:Prostaglandin-Endoperoxide Synthases / Q000235:genetics / Q000378:metabolism*; D012333:RNA, Messenger / Q000096:biosynthesis; D051381:Rats; D012045:Regulatory Sequences, Nucleic Acid / Q000502:physiology; D014157:Transcription Factors / Q000378:metabolism; D014162:Transfection
D022762:CCAAT-Enhancer-Binding Proteins; D018834:Chaperonin 60; D017362:Cyclic AMP Response Element-Binding Protein; D004791:Enzyme Inhibitors; D007527:Isoenzymes; D008070:Lipopolysaccharides; D012333:RNA, Messenger; D014157:Transcription Factors; D009569:Nitric Oxide; D007371:Interferon-gamma; D019001:Nitric Oxide Synthase; D052247:Nitric Oxide Synthase Type II; C496317:Nos2 protein, mouse; C496318:Nos2 protein, rat; D051546:Cyclooxygenase 2; D011451:Prostaglandin-Endoperoxide Synthases; D020928:Mitogen-Activated Protein Kinases