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

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.