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Glucocorticoids inhibit the induction of nitric oxide synthase II by down-regulating cytokine-induced activity of transcription factor nuclear factor-kappa B.
Mol Pharmacol 1996; 49(1):15-21MP

Abstract

Incubation of human A549/8 cells with human interleukin-1 beta (50 units/ml), interferon-gamma (100 units/ml), and tumor necrosis factor-alpha (10 ng/ml) (cytomix) resulted in a marked expression of the mRNA of the inducible nitric oxide synthase (NOS II). This induction was prevented by cycloheximide. Dexamethasone markedly reduced cytokine-induced NOS II mRNA concentrations; this reduction was prevented by RU 38486 (mifepristone). Pyrrolidine dithiocarbamate, an inhibitor of nuclear factor-kappa B (NF-kappa B) activation, also significantly decreased cytomix-induced NOS II mRNA levels. When A549/8 cells were transfected with a construct containing 1570-bp 5'-flanking sequence of the murine NOS II gene cloned before a reporter gene, the murine NOS II promoter was induced up to 20-fold with cytomix but not with bacterial lipopolysaccharide. Dexamethasone as well as pyrrolidine dithiocarbamate inhibited this induction. In electrophoretic mobility shift assays, nuclear protein extracts from cytomix-induced, but not from unstimulated cells, significantly slowed the migration of an oligonucleotide containing the NF-kappa B-binding site. This band shift was markedly reduced by dexamethasone. On the other hand, cytomix-induced nuclear protein content of NF-kappa B p65 and NF-kappa B p50 was not reduced by dexamethasone (as analyzed by Western blot). Dexamethasone also did not reduce cytomix-induced expression of NF-kappa B p65 mRNA or enhance the expression of NF-kappa B inhibitor mRNA. The human and murine NOS II promoters also contain consensus sequences for activating protein-1 (AP-1) binding. However, AP-1 binding activity of nuclear extracts of A549/8 cells was not enhanced by cytomix or inhibited by dexamethasone. These data suggest that the activated glucocorticoid receptor prevents (by a protein/protein interaction) the binding of transcription factor NF-kappa B, but not AP-1, to the NOS II promoter, thereby inhibiting the induction of NOS II transcription.

Authors+Show Affiliations

Department of Pharmacology, Johannes Gutenberg University, Mainz, Germany.No affiliation info availableNo affiliation info availableNo affiliation info available

Pub Type(s)

Journal Article
Research Support, Non-U.S. Gov't

Language

eng

PubMed ID

8569701

Citation

Kleinert, H, et al. "Glucocorticoids Inhibit the Induction of Nitric Oxide Synthase II By Down-regulating Cytokine-induced Activity of Transcription Factor Nuclear Factor-kappa B." Molecular Pharmacology, vol. 49, no. 1, 1996, pp. 15-21.
Kleinert H, Euchenhofer C, Ihrig-Biedert I, et al. Glucocorticoids inhibit the induction of nitric oxide synthase II by down-regulating cytokine-induced activity of transcription factor nuclear factor-kappa B. Mol Pharmacol. 1996;49(1):15-21.
Kleinert, H., Euchenhofer, C., Ihrig-Biedert, I., & Förstermann, U. (1996). Glucocorticoids inhibit the induction of nitric oxide synthase II by down-regulating cytokine-induced activity of transcription factor nuclear factor-kappa B. Molecular Pharmacology, 49(1), pp. 15-21.
Kleinert H, et al. Glucocorticoids Inhibit the Induction of Nitric Oxide Synthase II By Down-regulating Cytokine-induced Activity of Transcription Factor Nuclear Factor-kappa B. Mol Pharmacol. 1996;49(1):15-21. PubMed PMID: 8569701.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Glucocorticoids inhibit the induction of nitric oxide synthase II by down-regulating cytokine-induced activity of transcription factor nuclear factor-kappa B. AU - Kleinert,H, AU - Euchenhofer,C, AU - Ihrig-Biedert,I, AU - Förstermann,U, PY - 1996/1/1/pubmed PY - 1996/1/1/medline PY - 1996/1/1/entrez SP - 15 EP - 21 JF - Molecular pharmacology JO - Mol. Pharmacol. VL - 49 IS - 1 N2 - Incubation of human A549/8 cells with human interleukin-1 beta (50 units/ml), interferon-gamma (100 units/ml), and tumor necrosis factor-alpha (10 ng/ml) (cytomix) resulted in a marked expression of the mRNA of the inducible nitric oxide synthase (NOS II). This induction was prevented by cycloheximide. Dexamethasone markedly reduced cytokine-induced NOS II mRNA concentrations; this reduction was prevented by RU 38486 (mifepristone). Pyrrolidine dithiocarbamate, an inhibitor of nuclear factor-kappa B (NF-kappa B) activation, also significantly decreased cytomix-induced NOS II mRNA levels. When A549/8 cells were transfected with a construct containing 1570-bp 5'-flanking sequence of the murine NOS II gene cloned before a reporter gene, the murine NOS II promoter was induced up to 20-fold with cytomix but not with bacterial lipopolysaccharide. Dexamethasone as well as pyrrolidine dithiocarbamate inhibited this induction. In electrophoretic mobility shift assays, nuclear protein extracts from cytomix-induced, but not from unstimulated cells, significantly slowed the migration of an oligonucleotide containing the NF-kappa B-binding site. This band shift was markedly reduced by dexamethasone. On the other hand, cytomix-induced nuclear protein content of NF-kappa B p65 and NF-kappa B p50 was not reduced by dexamethasone (as analyzed by Western blot). Dexamethasone also did not reduce cytomix-induced expression of NF-kappa B p65 mRNA or enhance the expression of NF-kappa B inhibitor mRNA. The human and murine NOS II promoters also contain consensus sequences for activating protein-1 (AP-1) binding. However, AP-1 binding activity of nuclear extracts of A549/8 cells was not enhanced by cytomix or inhibited by dexamethasone. These data suggest that the activated glucocorticoid receptor prevents (by a protein/protein interaction) the binding of transcription factor NF-kappa B, but not AP-1, to the NOS II promoter, thereby inhibiting the induction of NOS II transcription. SN - 0026-895X UR - https://www.unboundmedicine.com/medline/citation/8569701/Glucocorticoids_inhibit_the_induction_of_nitric_oxide_synthase_II_by_down_regulating_cytokine_induced_activity_of_transcription_factor_nuclear_factor_kappa_B_ L2 - http://molpharm.aspetjournals.org/cgi/pmidlookup?view=long&pmid=8569701 DB - PRIME DP - Unbound Medicine ER -