Liver x receptor agonists inhibit cytokine-induced osteopontin expression in macrophages through interference with activator protein-1 signaling pathways.

Daisuke Ogawa, Jeffrey F. Stone, Yasunori Takata, Florian Blaschke, Van H. Chu, Dwight A. Towler, Ronald E. Law, Willa A. Hsueh, Dennis Bruemmer

Research output: Contribution to journalArticle

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Abstract

Osteopontin (OPN) is a proinflammatory cytokine and adhesion molecule implicated in the chemoattraction of monocytes and in cell-mediated immunity. We have recently reported that genetic OPN-deficiency attenuates the development of atherosclerosis in apoE-/- mice identifying OPN as potential target for pharmacological intervention in atherosclerosis. Synthetic agonists for the Liver X Receptor (LXR), members of the nuclear hormone receptor superfamily, prevent the development of atherosclerosis by regulating cholesterol homeostasis and suppressing inflammatory gene expression in macrophages. We demonstrate here that LXR ligands inhibit cytokine-induced OPN expression in macrophages. Two synthetic LXR ligands, T0901317 and GW3965, inhibited TNF-alpha, IL-1beta, INF-gamma and lipopolysaccharide induced OPN mRNA and protein expression in RAW 264.7 macrophages. Transient transfection experiments revealed that LXR ligands suppress cytokine-induced OPN promoter activity. Deletion analysis, heterologous promoter assays, and site-directed mutagenesis identified an activator protein-1 (AP-1) consensus site at -76 relative to the initiation site that supports OPN transcription in macrophages and mediates the effects of LXR ligands to inhibit OPN transcription. Electrophoretic mobility shift and chromatin immunoprecipitation assays indicated that LXR agonists inhibit cytokine-induced c-Fos and phospho-c-Jun binding to this AP-1 site. Cytokine-induced c-Fos and phospho-c-Jun protein expression was inhibited by LXR ligands and overexpression of c-Fos and c-Jun reversed the inhibitory effect of LXR ligands on OPN promoter activity in transactivation assays. Finally, treatment of C57BL/6J mice with LXR ligands inhibited OPN expression in peritoneal macrophages indicating that the observed effects of LXR ligands to inhibit OPN expression are applicable in vivo. These observations identify the regulation of macrophage OPN expression as a mechanism whereby LXR ligands may impact macrophage inflammatory responses and atherosclerosis. The full text of this article is available online at http://circres.ahajournals.org.

Original languageEnglish
JournalCirculation Research
Volume96
Issue number7
Publication statusPublished - Apr 15 2005
Externally publishedYes

Fingerprint

Osteopontin
Transcription Factor AP-1
Macrophages
Cytokines
Liver
Ligands
Atherosclerosis
Liver X Receptors
Proto-Oncogene Proteins c-jun
Chromatin Immunoprecipitation
Peritoneal Macrophages
Apolipoproteins E
Cytoplasmic and Nuclear Receptors
Site-Directed Mutagenesis
Inbred C57BL Mouse
Cellular Immunity
Transcriptional Activation
Transfection
Lipopolysaccharides
Monocytes

ASJC Scopus subject areas

  • Physiology
  • Cardiology and Cardiovascular Medicine

Cite this

Liver x receptor agonists inhibit cytokine-induced osteopontin expression in macrophages through interference with activator protein-1 signaling pathways. / Ogawa, Daisuke; Stone, Jeffrey F.; Takata, Yasunori; Blaschke, Florian; Chu, Van H.; Towler, Dwight A.; Law, Ronald E.; Hsueh, Willa A.; Bruemmer, Dennis.

In: Circulation Research, Vol. 96, No. 7, 15.04.2005.

Research output: Contribution to journalArticle

Ogawa, D, Stone, JF, Takata, Y, Blaschke, F, Chu, VH, Towler, DA, Law, RE, Hsueh, WA & Bruemmer, D 2005, 'Liver x receptor agonists inhibit cytokine-induced osteopontin expression in macrophages through interference with activator protein-1 signaling pathways.', Circulation Research, vol. 96, no. 7.
Ogawa, Daisuke ; Stone, Jeffrey F. ; Takata, Yasunori ; Blaschke, Florian ; Chu, Van H. ; Towler, Dwight A. ; Law, Ronald E. ; Hsueh, Willa A. ; Bruemmer, Dennis. / Liver x receptor agonists inhibit cytokine-induced osteopontin expression in macrophages through interference with activator protein-1 signaling pathways. In: Circulation Research. 2005 ; Vol. 96, No. 7.
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abstract = "Osteopontin (OPN) is a proinflammatory cytokine and adhesion molecule implicated in the chemoattraction of monocytes and in cell-mediated immunity. We have recently reported that genetic OPN-deficiency attenuates the development of atherosclerosis in apoE-/- mice identifying OPN as potential target for pharmacological intervention in atherosclerosis. Synthetic agonists for the Liver X Receptor (LXR), members of the nuclear hormone receptor superfamily, prevent the development of atherosclerosis by regulating cholesterol homeostasis and suppressing inflammatory gene expression in macrophages. We demonstrate here that LXR ligands inhibit cytokine-induced OPN expression in macrophages. Two synthetic LXR ligands, T0901317 and GW3965, inhibited TNF-alpha, IL-1beta, INF-gamma and lipopolysaccharide induced OPN mRNA and protein expression in RAW 264.7 macrophages. Transient transfection experiments revealed that LXR ligands suppress cytokine-induced OPN promoter activity. Deletion analysis, heterologous promoter assays, and site-directed mutagenesis identified an activator protein-1 (AP-1) consensus site at -76 relative to the initiation site that supports OPN transcription in macrophages and mediates the effects of LXR ligands to inhibit OPN transcription. Electrophoretic mobility shift and chromatin immunoprecipitation assays indicated that LXR agonists inhibit cytokine-induced c-Fos and phospho-c-Jun binding to this AP-1 site. Cytokine-induced c-Fos and phospho-c-Jun protein expression was inhibited by LXR ligands and overexpression of c-Fos and c-Jun reversed the inhibitory effect of LXR ligands on OPN promoter activity in transactivation assays. Finally, treatment of C57BL/6J mice with LXR ligands inhibited OPN expression in peritoneal macrophages indicating that the observed effects of LXR ligands to inhibit OPN expression are applicable in vivo. These observations identify the regulation of macrophage OPN expression as a mechanism whereby LXR ligands may impact macrophage inflammatory responses and atherosclerosis. The full text of this article is available online at http://circres.ahajournals.org.",
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AU - Stone, Jeffrey F.

AU - Takata, Yasunori

AU - Blaschke, Florian

AU - Chu, Van H.

AU - Towler, Dwight A.

AU - Law, Ronald E.

AU - Hsueh, Willa A.

AU - Bruemmer, Dennis

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