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2-oxoglutarate downregulates expression of vascular endothelial growth factor and erythropoietin through decreasing hypoxia-inducible factor-1α and inhibits angiogenesis

Authors

  • Ken Matsumoto,

    1. Department of Clinical and Experimental Hematology, Doctoral Program in Advanced Biomedical Applications, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan
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  • Shigehiko Imagawa,

    Corresponding author
    1. Doctoral Program of Sports Medicine, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan
    • Doctoral Program of Sports Medicine, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki 305-8577, Japan.
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  • Naoshi Obara,

    1. Department of Clinical and Experimental Hematology, Doctoral Program in Advanced Biomedical Applications, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan
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  • Norio Suzuki,

    1. Center for Tsukuba Advanced Research Alliance, University of Tsukuba, Tsukuba, Ibaraki, Japan
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  • Satoru Takahashi,

    1. Department of Anatomy and Embryology, Biomolecular and Integrated Medical Sciences, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan
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  • Toshiro Nagasawa,

    1. Department of Clinical and Experimental Hematology, Doctoral Program in Advanced Biomedical Applications, Graduate School of Comprehensive Human Sciences, University of Tsukuba, Tsukuba, Ibaraki, Japan
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  • Masayuki Yamamoto

    1. Center for Tsukuba Advanced Research Alliance, University of Tsukuba, Tsukuba, Ibaraki, Japan
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Abstract

In oxygenated cells, hypoxia-inducible factor-1 (HIF-1) α subunits are rapidly degraded by a mechanism that involves ubiquitination by the von Hippel–Lindau tumor suppressor E3 ligase complex using 2-oxoglutarate as a substrate. We examined the effect of 2-oxoglutarate on the production of erythropoietin and vascular endothelial growth factor (VEGF). The expression of erythropoietin and VEGF protein were dose-dependently downregulated in Hep3B cells by the addition of 2-oxoglutarate. The promoter activity of VEGF-luciferase was dose-dependently downregulated by the addition of 2-oxoglutarate. Gel mobility shift assays revealed that the addition of 2-oxoglutarate dose-dependently inhibited HIF-1 binding activity, but did not affect GATA binding activity. Western blot analysis revealed that 2-oxoglutarate dose-dependently inhibited the HIF-1α protein level in Hep3B cells in hypoxic conditions. However, MG132 (the proteasome inhibitor) rescued the inhibition of HIF-1α protein expression by 2-oxoglutarate. Furthermore, under hypoxic conditions, 2-oxoglutarate dose-dependently inhibited tube formation in in vitro angiogenesis assays. These results indicate that 2-oxoglutarate treatment may be useful for the inhibition of angiogenesis. J. Cell. Physiol. 209: 333–340, 2006. © 2006 Wiley-Liss, Inc.

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