Cited2 modulates hypoxia‐inducible factor–dependent expression of vascular endothelial growth factor in nucleus pulposus cells of the rat intervertebral disc

Abstract

Objective. To determine whether nucleus pulposus cells of the intervertebral disc express hypoxiainducible factor 2␣ (HIF-2␣), and to assess the role of HIF-1 and HIF-2 in controlling cited2 and vascular endothelial growth factor (VEGF) expression.Methods. Rat cells were cultured under normoxic (21% O 2 ) or hypoxic (2% O 2 ) conditions, and expression and promoter activity of HIF-2 target genes were evaluated. Gain-or loss-of-function experiments were performed to investigate the contribution of HIF isoforms to cited2 activity as well as the role of cited2 in regulating VEGF expression.Results. We found that HIF-2␣ protein was expressed in vivo and that protein and messenger RNA expression were similar under both normoxic and hypoxic conditions. However, there was a significant increase in HIF-2␣ transactivation under hypoxic conditions. With respect to functional activity, unlike the case in most other tissues, HIF-2 failed to increase the transcriptional activities of superoxide dismutase 2 and frataxin, 2 common target genes involved in radical dismutation. However, under hypoxic conditions, HIF-2 preferentially regulated the expression and promoter activity of cited2, a p300 binding protein. When HIF-2␣ or HIF-1␣ was suppressed, cited2 promoter activity was inhibited. Finally, we showed that forced expression or suppression of cited2 resulted in corresponding changes in expression of VEGF, a common target gene for HIF-1 and HIF-2 in the nucleus pulposus cells.Conclusion. Results of this study indicate that in nucleus pulposus cells, HIF-2 and HIF-1 modulate their own transcriptional activity through cited2. We suggest that the 2 arms of the regulatory circuit serve to maintain survival activities and inhibit angiogenesis in the healthy disc.

Hypoxia-inducible Factor (HIF)-1α and CCN2 Form a Regulatory Circuit in Hypoxic Nucleus Pulposus Cells

“…Furthermore, our study shows that in contrast to HIF-1␣, CCN2 is a positive regulator of HIF-2␣ expression. This is relevant, as we have previously reported that in nucleus pulposus, HIF-2␣ controls the expression of cited2, a p300 interacting protein (29). By competing with HIF-␣ for p300 binding, cited2 suppresses transcriptional activity of HIF (29).…”

“…This is relevant, as we have previously reported that in nucleus pulposus, HIF-2␣ controls the expression of cited2, a p300 interacting protein (29). By competing with HIF-␣ for p300 binding, cited2 suppresses transcriptional activity of HIF (29). Thus, it is possible that through induction of the HIF-2␣-cited2 circuit, CCN2 may decrease HIF-1␣ activity and target gene expression.…”

Hypoxia promotes noncanonical autophagy in nucleus pulposus cells independent of MTOR and HIF1A signaling

“…[2][3][4][5] NP cells adapt to this hypoxic niche by modulating a key transcription factor, HIF1A (hypoxia inducible factor 1 a subunit), to promote cell survival, matrix synthesis, and to regulate glycolytic metabolism. [6][7][8][9][10][11][12][13] A recent study using NP-specific conditional Hif1a knockout mice also shows that HIF1A is required for postnatal NP cell survival in vivo. 14 Autophagy is a highly conserved cellular process where organelles and cytosolic proteins are encapsulated within target of rapamycin [serine/threonine kinase]) resulting in increased autophagy.…”

Responses and adaptations of intervertebral disc cells to microenvironmental stress: a possible central role of autophagy in the adaptive mechanism

“…However, only HIF1b was found in rat AF cells in vivo (93). In a hypoxic microenvironment, the a-subunits of HIF are stable and dimerize with the other subunits to bind hypoxia response elements (HREs) and promote the expression of protective mRNAs and proteins, such as vascular endothelial growth factor (VEGF), cited2 and galectin-3 to inhibit apoptosis (94)(95)(96). With regard to the ECM, HIF-1 was reported to promote the synthesis of aggrecan, which is the major proteoglycan expressed in discs (97).…”