Advertisement

Journal of Neuro-Oncology

, Volume 84, Issue 1, pp 9–19 | Cite as

PAX6 increases glioma cell susceptibility to detachment and oxidative stress

  • Jason Y. Chang
  • Yuanjie Hu
  • Eric Siegel
  • Latarsha Stanley
  • Yi-Hong Zhou
Lab Investigation–human/animal tissue

Abstract

Purpose

Glioblastoma multiforme (GBM) is an incurable malignant glioma which is very resistant to radiation and alkylating agent-based chemotherapy. Necrosis is a hallmark for GBM and the layer surrounding necrotic area is packed with cells which are now believed to be those migrating out of the necrotic area. Oxidative stress is a condition that GBM cells encounter in the necrotic zone, which is one of the stressful conditions that GBM cells need to resist in order to survive. Our previous studies revealed that low PAX6 expression is a favorable molecular trait for survival acquired by GBM cells because PAX6 could suppress cell invasion and tumorigenicity. Since detachment of cells from GBM is an early event for cell migration and subsequent invasion, we examined whether PAX6 is involved in cell survival after detachment.

Experimental Design

PAX6 over-expression was achieved in glioma cells transiently (by adenoviral-mediated transient over-expression) or stably (by the establishment of stable cell lines after transfection). The effect of PAX6 over-expression on the survival and growth of glioma cells after detachment from the culture was determined.

Result

Our data revealed that GBM cells (with their low PAX6 levels) survived the detachment procedure well. However, PAX6 over-expression attenuated GBM cell recovery of growth after detachment-induced stress. Importantly, intracellular reactive oxygen species (ROS) levels increased following cell detachment and that PAX6 over-expressing cells retained higher level of ROS than control cells. This may be partially responsible for the impaird growth rate after cell detachment. Addition of anti-oxidant improved the cell viability of PAX6 over-expressing cells, but did not restore their ability to proliferate.

Conclusion

To survive, GBM cells must resist oxidative stress in the necrotic zone as well as the intracellular ROS generated during detachment. Since PAX6 over-expression in low PAX6-expressing glioma cells attenuated cell survival and growth after detachment, these results suggest that a reduced PAX6 expression may be a molecular trait that gives glioma cells a real selection advantage over other cell types to survive in stressful conditions, thus resulting in expansion of their population.

Keywords

PAX6 Glioma cell Detachment Oxidative stress 

Abbreviations

CS

Calf serum

GBM

Glioblastoma multiforme

FU

Fluorescence units

H2DCF-DA

2′,7′-dichlorodihydrofluorescein diacetate

HBSS

Hank’s Balanced Salt Solution

JC-1

5,5′, 6,6′-tetrachloro-1,1′, 3,3′-tetraethylbenzimidazolocarbocyanine iodide

MTT

(3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide)

ROS

Reactive oxygen species

SF

Serum free

Notes

Acknowledgments

This work was supported by the Arkansas Cancer Research Center Tobacco Settlement Fund, American Cancer Society IRG to YH Zhou, Arkansas BRIN program to L Stanley and YH Zhou, and by the Research to Prevent Blindness to JY Chang.

References

  1. 1.
    Kleihues P, Louis DN, Scheithauer BW et al (2002) The WHO classification of tumors of the nervous system. J Neuropathol Exp Neurol 61:215–25; discussion 226–19Google Scholar
  2. 2.
    Maher EA, Furnari FB, Bachoo RM et al (2001) Malignant glioma: genetics and biology of a grave matter. Genes Dev 15:1311–1333PubMedCrossRefGoogle Scholar
  3. 3.
    Godard S, Getz G, Delorenzi M et al (2003) Classification of human astrocytic gliomas on the basis of gene expression: a correlated group of genes with angiogenic activity emerges as a strong predictor of subtypes. Cancer Res 63:6613–6625PubMedGoogle Scholar
  4. 4.
    Gaspar LE, Fisher BJ, Macdonald DR et al (1992) Supratentorial malignant glioma: patterns of recurrence and implications for external beam local treatment. Int J Radiat Oncol Biol Phys 24:55–57PubMedGoogle Scholar
  5. 5.
    Pope WB, Sayre J, Perlina A et al (2005) MR imaging correlates of survival in patients with high-grade gliomas. AJNR Am J Neuroradiol 26:2466–2474PubMedGoogle Scholar
  6. 6.
    Brat DJ, Castellano-Sanchez AA, Hunter SB et al (2004) Pseudopalisades in glioblastoma are hypoxic, express extracellular matrix proteases, and are formed by an actively migrating cell population. Cancer Res 64:920–927PubMedCrossRefGoogle Scholar
  7. 7.
    Iida T, Furuta A, Kawashima M et al (2001) Accumulation of 8-oxo-2′-deoxyguanosine and increased expression of hMTH1 protein in brain tumors. Neuro-Oncol 3:73–81PubMedCrossRefGoogle Scholar
  8. 8.
    Kalaria RN, Premkumar DR, Pax AB et al (1996) Production and increased detection of amyloid beta protein and amyloidogenic fragments in brain microvessels, meningeal vessels and choroid plexus in Alzheimer’s disease. Brain Res Mol Brain Res 35:58–68PubMedCrossRefGoogle Scholar
  9. 9.
    Ziche M, Morbidelli L (2000) Nitric oxide and angiogenesis. J Neuro-Oncol 50:139–148CrossRefGoogle Scholar
  10. 10.
    Brown NS, Bicknell R (2001) Hypoxia and oxidative stress in breast cancer. Oxidative stress: its effects on the growth, metastatic potential and response to therapy of breast cancer. Breast Cancer Res 3: 323–327PubMedCrossRefGoogle Scholar
  11. 11.
    Schwartzbaum JA, Cornwell DG (2000) Oxidant stress and glioblastoma multiforme risk: serum antioxidants, gamma-glutamyl transpeptidase, and ferritin. Nutr Cancer 38:40–49PubMedCrossRefGoogle Scholar
  12. 12.
    Sposto R, Ertel IJ, Jenkin RD et al (1989) The effectiveness of chemotherapy for treatment of high grade astrocytoma in children: results of a randomized trial. A report from the Childrens Cancer Study Group. J Neurooncol 7:165–167PubMedCrossRefGoogle Scholar
  13. 13.
    Fine HA, Dear KB, Loeffler JS et al (1993) Meta-analysis of radiation therapy with and without adjuvant chemotherapy for malignant gliomas in adults. Cancer 71:2585–2597PubMedCrossRefGoogle Scholar
  14. 14.
    Stewart LA (2002) Chemotherapy in adult high-grade glioma: a systematic review and meta-analysis of individual patient data from 12 randomised trials. Lancet 359:1011–1018PubMedCrossRefGoogle Scholar
  15. 15.
    Zhou YH, Tan F, Hess KR et al (2003) The expression of PAX6, PTEN, vascular endothelial growth factor, and epidermal growth factor receptor in gliomas: relationship to tumor grade and survival. Clin Cancer Res 9:3369–3375PubMedGoogle Scholar
  16. 16.
    Zhou YH, Wu X, Tan F et al (2005) PAX6 suppresses growth of human glioblastoma cells. J Neuro-Oncol 71:223–229CrossRefGoogle Scholar
  17. 17.
    Ke LD, Fueyo J, Chen X et al (1998) A novel approach to glioma gene therapy: down-regulation of the vascular endothelial growth factor in glioma cells using ribozymes. Int J Oncol 12:1391–1396PubMedGoogle Scholar
  18. 18.
    Garg TK, Chang JY (2004) 15-deoxy-delta 12, 14-Prostaglandin J2 prevents reactive oxygen species generation and mitochondrial membrane depolarization induced by oxidative stress. BMC Pharmacol 4:6PubMedCrossRefGoogle Scholar
  19. 19.
    Garg T, Chang J (2003) Oxidative stress causes ERK phosphorylation and cell death in cultured retinal pigment epithelium: prevention of cell death by AG126 and 15-deoxy-delta 12, 14-PGJ2. BMC Ophthalmol 3:5PubMedCrossRefGoogle Scholar
  20. 20.
    Mayes DA, Hu Y, Teng Y et al (2006) PAX6 Suppresses the invasiveness of glioblastoma cells and the expression of the matrix metalloproteinase 2 gene. Cancer Res 66:9809–9817PubMedCrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media, LLC 2007

Authors and Affiliations

  • Jason Y. Chang
    • 1
    • 2
  • Yuanjie Hu
    • 3
  • Eric Siegel
    • 4
  • Latarsha Stanley
    • 5
  • Yi-Hong Zhou
    • 3
  1. 1.Department of Neurobiology and Developmental SciencesUniversity of Arkansas for Medical SciencesLittle RockUSA
  2. 2.Department of OphthalmologyUniversity of Arkansas for Medical SciencesLittle RockUSA
  3. 3.Department of Neurological SurgeryUniversity of California, IrvineIrvineUSA
  4. 4.Department of BiostatisticsUniversity of Arkansas for Medical SciencesLittle RockUSA
  5. 5.Arkansas Biomedical Research Infrastructure NetworkUniversity of Arkansas for Medical SciencesLittle RockUSA

Personalised recommendations