Nitric oxide inhibits endothelial IL-1[beta]-induced ICAM-1 gene expression at the transcriptional level decreasing Sp1 and AP-1 activity.
BACKGROUND
Nitric oxide (NO) has frequently been shown to inhibit leukocyte adherence to activated endothelium thus displaying anti-adhesive and immunosuppressive activities. A molecular mechanism contributing to this effect is described.


METHODS
Primary murine aortic endothelial cells were activated with interleukin (IL)-1beta to express intercellular adhesion molecule-1 (ICAM-1) mRNA in the presence or absence of the physiological spontaneous NO-donor S-nitrosocysteine. Subsequently, semiquantitative RT-PCR and gel shift assays with nuclear extracts were performed to analyse the effects of NO on ICAM-1 mRNA expression and on the activity of transcription factors involved in ICAM-1 transcription. In addition, luciferase reporter gene activity of cytokine-activated cells transiently transfected with an ICAM-1 promoter-luciferase construct and cultured in the presence of the slow-releasing NO-donor DETA/NO was determined.


RESULTS
NO at subtoxic concentrations decreases IL-1beta-induced endothelial ICAM-1 mRNA expression. This inhibition occurs at the transcriptional level, as NO affects IL-1b-induced ICAM-1 promoter activity in transiently transfected cells. Using gel-shift assays and double-stranded oligonucleotide consensus sequences of the known transcription factor binding sites of the ICAM-1 promoter, Sp1 and AP-1 were identified as transcriptional activators of IL-1beta-driven ICAM-1 expression. The DNA binding of both of these transcription factors to specific binding sites of the ICAM-1 promoter was decreased in MAEC exposed to NO.


CONCLUSIONS
Our studies indicate that the anti-adhesive effect of NO concentrations equivalent to high-output NO synthesis is mediated, at least in part, by inhibition of ICAM-1 expression via a concerted action of NO on the redox-sensitive transcriptional activators Sp1 and AP-1. This molecular mechanism may contribute to the anti-inflammatory actions of NO synthesized by the inducible NO synthase.
Berendji-Grün
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Research Group Immunobiology, Medical Department, Heinrich-Heine-University of Düsseldorf, Germany.
Kolb-Bachofen
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Kröncke
K D
KD
D000818:Animals; D002478:Cells, Cultured; D003545:Cysteine / Q000031:analogs & derivatives* / Q000494:pharmacology; D004730:Endothelium, Vascular / Q000187:drug effects / Q000502:physiology*; D005786:Gene Expression Regulation / Q000187:drug effects / Q000502:physiology; D006801:Humans; D018799:Intercellular Adhesion Molecule-1 / Q000096:biosynthesis* / Q000235:genetics; D007375:Interleukin-1 / Q000494:pharmacology* / Q000502:physiology; D051379:Mice; D009569:Nitric Oxide / Q000502:physiology*; D020030:Nitric Oxide Donors / Q000494:pharmacology*; D012333:RNA, Messenger / Q000096:biosynthesis / Q000235:genetics; D026403:S-Nitrosothiols / Q000494:pharmacology; D016329:Sp1 Transcription Factor / Q000502:physiology*; D018808:Transcription Factor AP-1 / Q000502:physiology*; D014158:Transcription, Genetic
D007375:Interleukin-1; D020030:Nitric Oxide Donors; D012333:RNA, Messenger; D026403:S-Nitrosothiols; D016329:Sp1 Transcription Factor; D018808:Transcription Factor AP-1; D018799:Intercellular Adhesion Molecule-1; D009569:Nitric Oxide; C021315:S-nitrosocysteine; D003545:Cysteine