gms | German Medical Science

63. Jahrestagung der Deutschen Gesellschaft für Neurochirurgie (DGNC)
Joint Meeting mit der Japanischen Gesellschaft für Neurochirurgie (JNS)

Deutsche Gesellschaft für Neurochirurgie (DGNC) e. V.

13. - 16. Juni 2012, Leipzig

Resistance to hypoxia-induced, BNIP3-mediated cell death contributes to the increase of CD133+ cell population in human glioblastoma-derived cultures in lowered oxygen tension

Meeting Abstract

  • U. Kahlert - Laboratory of Molecular Neurosurgery, Dept. of Stereotactic and Functional Neurosurgery, University Medical Center, Freiburg, Germany
  • D. Maciaczyk - Laboratory of Molecular Neurosurgery, Dept. of Stereotactic and Functional Neurosurgery, University Medical Center, Freiburg, Germany
  • M. Döbrössy - Laboratory of Molecular Neurosurgery, Dept. of Stereotactic and Functional Neurosurgery, University Medical Center, Freiburg, Germany
  • C.H. Herold-Mende - Division of Neurosurgical Research, Dept. of Neurosurgery, Heidelberg, Germany
  • G. Nikkhah - Laboratory of Molecular Neurosurgery, Dept. of Stereotactic and Functional Neurosurgery, University Medical Center, Freiburg, Germany
  • J. Maciaczyk - Dept. of General Neurosurgery, University Medical Center, Freiburg, Germany

Deutsche Gesellschaft für Neurochirurgie. Japanische Gesellschaft für Neurochirurgie. 63. Jahrestagung der Deutschen Gesellschaft für Neurochirurgie (DGNC), Joint Meeting mit der Japanischen Gesellschaft für Neurochirurgie (JNS). Leipzig, 13.-16.06.2012. Düsseldorf: German Medical Science GMS Publishing House; 2012. DocFR.07.03

DOI: 10.3205/12dgnc216, URN: urn:nbn:de:0183-12dgnc2160

Veröffentlicht: 4. Juni 2012

© 2012 Kahlert et al.
Dieser Artikel ist ein Open Access-Artikel und steht unter den Creative Commons Lizenzbedingungen (http://creativecommons.org/licenses/by-nc-nd/3.0/deed.de). Er darf vervielfältigt, verbreitet und öffentlich zugänglich gemacht werden, vorausgesetzt dass Autor und Quelle genannt werden.


Gliederung

Text

Objective: Brain tumor stem-like cells (BTSC) are responsible for tumor initiation, propagation and resistance to standard therapies. Previous reports showed hypoxia-induced increase of BTSC expressing CD133 together with over-expression of other stem cell-related genes and more pronounced clonogenic capacity of the cultures. In this study, we investigated the mechanisms responsible for hypoxia-dependent induction of CD133+ subpopulation in glioblastoma-derived cell cultures.

Methods: Glioblastoma-derived cell lines were propagated under free floating culture conditions and subjected to either normoxic (21%) or hypoxic (3%) environmental oxygen tensions. Growth kinetic was assessed with CellTiter 96® Cell Proliferation Assay. CD133 expression was investigated with flow cytometry. Additionally, quantitative rtPCR-based expression analysis of stem cell-related and pro/anti-apoptotic genes of bulk cultures as well as FACS-sorted CD133± subpopulations followed by Western blot protein analysis was performed. To modify the epigenetic regulation of gene expression hypoxic cultures were treated with 10 μM of de-methylation agent 5’-Azacitidine.

Results: Cell cultures exposed to hypoxia showed several fold increase of CD133+ BTSC population. Interestingly, growth rate of hypoxic cultures was significantly lower than their normoxic counterparts. Both increase of CD133+ subpopulation and deceleration of the growth kinetic were reversible following transfer to normoxic conditions. Exposure to hypoxia induced a BNIP3-dependant apoptosis preferentially in CD133- cells as shown by gene expression (up to 28 fold increase of BNIP3 mRNA levels) and protein analysis. In contrast, CD133+ cells showed a down-regulation of BNIP3 expression, owing to the methylation of its promoter and overexpression of anti-apoptotic genes Bcl-2 and Survivin (up to 8 and 6 fold respectively) that led to their better survival under hypoxia. Application of AZA resulted in dramatic increase of BNIP3 expression levels in CD133+ cells under hypoxia and complete abrogation of hypoxia-induced increase of CD133+ subpopulation.

Conclusions: Resistance of CD133+ BTSC against hypoxia-induced BNIP3-mediated apoptosis is dependent on the selective silencing of BNIP3 promoter and contributes to their better survival in lowered oxygen tension. This could be abrogated by AZA application indicating the possible efficacy of novel therapy focused on eradication of BTSC by modifications of epigenetic regulation of gene expression.