Involvement of nuclear factorkB in the regulation of cyclooxygenase-2 expression by interleukin-1 in rheumatoid synoviocytes
| Autor: | Leslie J. Crofford, Timothy Hla, Bing Tan, Conor J. McCarthy |
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| Rok vydání: | 1997 |
| Předmět: |
Electrophoresis
medicine.medical_specialty Time Factors Blotting Western Immunology Biology Gene Expression Regulation Enzymologic Arthritis Rheumatoid Rheumatology Internal medicine Sense (molecular biology) Gene expression medicine Humans Immunology and Allergy Pharmacology (medical) Electrophoretic mobility shift assay RNA Messenger Promoter Regions Genetic Enhancer Transcription factor Messenger RNA Synovial Membrane NF-kappa B Transcription Factor RelA RNA Oligonucleotides Antisense Molecular biology Blot Endocrinology Prostaglandin-Endoperoxide Synthases Interleukin-1 |
| Zdroj: | Arthritis & Rheumatism. 40:226-236 |
| ISSN: | 1529-0131 0004-3591 |
| Popis: | To evaluate involvement of the transcription factor nuclear factor kappa B (NF-kappa B) in the increased expression of cyclooxygenase-2 (COX-2) stimulated by interleukin-1 beta (IL-1 beta) in primary rheumatoid synoviocytes.We treated early-passage rheumatoid synoviocytes with IL-1 beta and examined the time course of NF-kappa B translocation to the nucleus by Western blot analysis, as well as NF-kappa B binding to the COX-2 promoter/enhancer by electrophoretic mobility shift assay. We correlated the time course of NF-kappa B binding with expression of COX-2 messenger RNA (mRNA) and protein. Synoviocytes were then treated with either sense or antisense phosphorothioate-modified oligonucleotides derived from the transcription start site of the human NF-kappa B p65 RNA. We analyzed NF-kappa B binding to the COX-2 promoter and COX-2 protein levels after these treatments.IL-1 beta rapidly stimulated the translocation of the p65, p50, and c-rel NF-kappa B subunits from the cytoplasm to the nucleus. Electrophoretic mobility shift assay demonstrated binding to 2 NF-kappa B sites within the COX-2 promoter/enhancer, with a time course identical to that of nuclear localization of NF-kappa B. Supershift analysis revealed that binding activity was due primarily to the p65-p50 heterodimer and the p50 homodimer. With appropriate lag time after NF-kappa B binding, COX-2 mRNA and protein were increased. Pretreatment of RA synoviocytes with NF-kappa B p65 antisense oligonucleotides resulted in decreased binding to the COX-2 promoter and decreased COX-2 protein expression.These data demonstrate that signaling via the NF-kappa B pathway is involved in regulation of COX-2 expression induced by IL-1 beta in RA synoviocytes. |
| Databáze: | OpenAIRE |
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