EXTENDED REPORT

Oestrogens inhibit interleukin 1ß-mediated nitric oxide synthase expression in articular chondrocytes through nuclear factor-B impairment

Pascal Richette^1,2, Marie-France Dumontier¹, Khadija Tahiri¹, Magdalena Widerak¹, Antoine Torre¹, Mourad Benallaloua¹, François Rannou¹, Marie-Therese Corvol¹, Jean-François Savouret¹

¹ INSERM-UMR-747, Universite Paris Descartes, UFR Biomedicale, Paris, France
² AP-HP, Groupe Hospitalier Lariboisiere, Federation de Rhumatologie, Paris, F-75010, France; Universite Paris Diderot, Faculte de Medecine Xavier Bichat, Paris, France

Correspondence to:
Dr J-F Savouret
INSERM UMR-747, Universite Paris Descartes, UFR Biomedicale, 45 rue des Saints Peres, 75006Paris, France; jean-francois.savouret{at}univ-paris5.fr

Objectives: To investigate the presence and functionality of oestrogen receptor (ER) in interleukin (IL)1ß-treated rabbit articular chondrocytes in culture, and to determine the mechanisms of 17ß oestradiol (E2) effects on IL1ß-induced inducible nitric oxide synthase (iNOS) expression.

Methods: The presence and functionality of ER were investigated by immunocytochemistry and transient expression of an E2-responsive reporter construct. iNOS expression and production were determined by transient expression of a chimeric iNOS promoter–luciferase construct and protein immunoblotting. Nitric oxide (NO) production was determined by the Griess reaction. DNA-binding activities of nuclear factor-B (NF-B) and activated protein 1 were determined by electrophoretic mobility shift assay (EMSA)–ELISA assays. Nuclear translocation of p65 was studied by immunocytochemistry.

Results: ER was identified in the nucleus of chondrocytes. ER efficiently transactivated a transiently expressed E2-responsive construct. On IL1ß treatment, ER partially diffused from its nuclear localisation into the cytoplasm and its transactivation ability was impaired. Nevertheless, E2, tamoxifen and raloxifene efficiently inhibited IL1ß-induced NO production (–34%, –31% and –36%, respectively). E2 decreased IL1ß-induced iNOS protein expression (–40%). Transient expression of an iNOS promoter construct strongly suggested that iNOS expression was inhibited at the transcriptional level, and EMSA-ELISA assays showed that E2 reduced (–60%) the IL1ß-induced p65 DNA-binding capacity. Finally, the p65 nuclear translocation induced by IL1ß was also strongly decreased by E2.

Conclusions: Our data support a reciprocal antagonism between oestrogens and IL1ß, ultimately resulting in the decrease of cytokine-dependent NO production through transcriptional inhibition of iNOS expression. This effect was associated with selective inhibition of p65 DNA binding and nuclear translocation.

Abbreviations: Ap-1, actovated protein 1; ER, oestrogen receptor ; ERE, oestrogen responsive element; ERT, oestrogen replacement therapy; iNOS, inducible nitric oxide synthase; NF-B, nuclear factor B; SERM, selective oestrogen receptor modulators

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