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Journal of Cellular Biochemistry

Nucleus pulposus cells express HIF-1α under normoxic culture conditions: A metabolic adaptation to the intervertebral disc microenvironment

Authors

  • Makarand V. Risbud,

    Corresponding author
    1. Department of Orthopaedic Surgery and Graduate Program in Tissue Engineering and Regenerative Medicine, Thomas Jefferson University, Philadelphia, Pennsylvania, 19107
    • Department of Orthopaedic Surgery, 1015 Walnut Street, Suite 501 Curtis Bldg., Thomas Jefferson University, Philadelphia, PA 19107.
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  • Asha Guttapalli,

    1. Department of Orthopaedic Surgery and Graduate Program in Tissue Engineering and Regenerative Medicine, Thomas Jefferson University, Philadelphia, Pennsylvania, 19107
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  • David G. Stokes,

    1. Division of Rheumatology, Thomas Jefferson University, Philadelphia 19107, Pennsylvania
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  • David Hawkins,

    1. Division of Rheumatology, Thomas Jefferson University, Philadelphia 19107, Pennsylvania
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  • Keith G. Danielson,

    1. Department of Orthopaedic Surgery and Graduate Program in Tissue Engineering and Regenerative Medicine, Thomas Jefferson University, Philadelphia, Pennsylvania, 19107
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  • Thomas P. Schaer,

    1. Comparative Orthopaedics Research Laboratory, University of Pennsylvania Veterinary School of Medicine, New Bolton Center, Kennett Square, Philadelphia, Pennsylvania, 19348
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  • Todd J. Albert,

    1. Department of Orthopaedic Surgery and Graduate Program in Tissue Engineering and Regenerative Medicine, Thomas Jefferson University, Philadelphia, Pennsylvania, 19107
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  • Irving M. Shapiro

    1. Department of Orthopaedic Surgery and Graduate Program in Tissue Engineering and Regenerative Medicine, Thomas Jefferson University, Philadelphia, Pennsylvania, 19107
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Abstract

Nucleus pulposus (NP) cells of the intervertebral disc reside in an environment that has a limited vascular supply and generate energy through anaerobic glycolysis. The goal of the present study was to examine the expression and regulation of HIF-1α, a transcription factor that regulates oxidative metabolism in nucleus pulposus cells. Nucleus pulposus cells were isolated from rat, human, and sheep disc and maintained at either 21% or 2% oxygen for various time periods. Cells were also treated with desferrioxamine (Dfx), a compound that mimics the effects of hypoxia (Hx). Expression and function of HIF-1α were assessed by immunofluorescence microscopy, Western blot analysis, gel shift assays, and luciferase reporter assays. In normoxia (Nx), rat, sheep, and human nucleus pulposus cells consistently expressed the HIF-1α subunit. Unlike other skeletal cells, when maintained under low oxygen tension, the nucleus pulposus cells exhibited a minimal induction in HIF-1α protein levels. Electromobility shift assays confirmed the functional binding of normoxic HIF-1α protein to its putative DNA binding motif. A dual luciferase reporter assay showed increased HIF-1α transcriptional activity under hypoxia compared to normoxic level, although this induction was small when compared to HeLa and other cell types. These results indicate that normoxic stabilization of HIF-1α is a metabolic adaptation of nucleus pulposus cells to a unique oxygen-limited microenvironment. The study confirmed that HIF-1α can be used as a phenotypic marker of nucleus pulposus cells. J. Cell. Biochem. 98: 152–159, 2006. © 2006 Wiley-Liss, Inc.

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