Hypoxia and differentiation in human neuroblastoma cells

Abstract

The childhood tumour neuroblastoma is derived from immature cells of the sympathetic nervous system, which have become arrested at different maturation stages. Neuroblastoma is a malignancy with a high degree of heterogeneity, and there is a correlation between a poor differentiation status and a more aggressive phenotype. The MYCN gene is amplified in approximately 25% of neuroblastoma tumours and is correlated to an aggressive tumour phenotype. A role of MYCN in keeping these tumours at an immature stage has been suggested. However, we show that over-expression of MYCN in non-amplified neuroblastoma cells do not restrain their capacity to differentiate. Furthermore, in a panel of 28 neuroblastoma tumours and 27 cell lines, we do not see a correlation between the degree of MYCN expression and the expression of neuronal or neuroendocrine marker genes. Previously hypoxia and/or nutrient deprivation has been suggested to induce a neuroendocrine lineage shift in neuroblastoma tumours. However, when growing neuroblastoma cell lines at hypoxia and/or low glucose conditions, we observe a down-regulation of both neuronal and chromaffin marker genes. Instead, genes normally expressed in early neural crest cells are induced. We therefore propose that hypoxia and/or glucose deficiency induce a dedifferentiation of neuroblastoma cells, thereby rendering them a more aggressive phenotype. In addition, when growing neuroblastoma cells at hypoxia and/or without glucose, we find that hypoxia protects from glucose-deprivation induced cell death. This further adds to the malignant potential of hypoxic neuroblastoma cells. We have further investigated the hypoxia inducible transcription factors HIF-1a and HIF-2a in neuroblastoma cells and find that they have separate patterns of activation over time and in response to different oxygen levels