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Identifying altered gene expression in neuroblastoma cells preceding apoptosis

Abstract

Purpose

Concomitant differentiation and partial inhibition of proteasome trigger cell death in a neuroblastoma cell line (NBP2). Neither induction of differentiation nor partial inhibition of proteasome alone affects the viability of NBP2 cells. We wanted to identify genes whose expression alters under concomitant conditions and may account for cell death.

Methods

We used gel electrophoresis to analyze total genomic DNA for the detection of DNA fragmentation. Affymetrix Murine Genome U74A version 2 microarray was used to screen for ∼6,000 functionally characterized genes and ∼6,000 expressed sequence tags (ESTs). Real time PCR (RT-PCR) was performed to provide an accurate assessment of changes in gene expression.

Results

Concomitant differentiation and partial inhibition of proteasome trigger apoptosis, characterized by genomic DNA fragmentation in NBP2 cells. We found that the expression of 41 genes changed 2.5-fold or more primarily under concomitant conditions midway through apoptosis. Based on real time PCR, the expression of galectin-3, glycosylated 96, a leucine zipper protein (LRG-21), and endothelial cell activated protein C receptor (EPCR) increased between 50–500-fold, whereas the expression of Polo serine/threonine kinase, N-myc, and Histone H2A.1 decreased ranging from 8 to 37 fold. Altered expression of galectin-3, EPCR, and LRG-21 was detected as early as 2–8 h post simultaneous conditions.

Conclusion

We identified new genes that might be involved in apoptotic events in neuroblastoma cells.

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Acknowledgments

We thank Dr. Kelly Ambler and Dr. Andrea Koenig for a critical reading of this manuscript. We also thank Karen Helm, Mike Ashton, and Phillip Eckels for their help on flow cytometry at UCHSC and Denver Health Medical Center. This work was supported by a grant from NIH/NIA (R01AG18285).

Author information

Correspondence to Piruz Nahreini.

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Real Time PCR Analysis of Fold Changes in Gene Expression (DOC 34.5 kb)

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Nahreini, P., Yan, X., Andreatta, C.P. et al. Identifying altered gene expression in neuroblastoma cells preceding apoptosis. J Cancer Res Clin Oncol 134, 411–419 (2008). https://doi.org/10.1007/s00432-007-0303-0

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Keywords

  • Neuroblastoma
  • Apoptosis
  • Affymetrix microarray
  • Proteasome inhibition
  • Differentiation
  • Mitochondria
  • Neurodegeneration