Hypoxia sensitizes human malignant glioma cells towards CD95L‐induced cell death

Abstract

Death ligands such as CD95 ligand (CD95L) have limited activity against glioma cells under normoxic conditions. Hypoxia is a critical aspect of the microenvironment of gliomas in vivo. We investigated the effect of co-exposure to acute hypoxia and CD95 ligand in three human malignant glioma cell lines with different susceptibility to CD95L under normoxic conditions. Hypoxia sensitized all three cell lines towards CD95L-induced cell death. Co-exposure resulted in apoptotic changes in the early phase, with gradual conversion to secondary necrosis with increasing length of hypoxia. The mitochondrial injury induced by hypoxia was enhanced by co-treatment, and caspase cleavage became prominent. Inhibition of the epidermal growth factor receptor (EGFR), although sensitizing glioma cells to CD95L under normoxia, protects glioma cells from hypoxia by reducing energy consumption. However, the opposing effects of EGFR signalling on death induced by CD95L or hypoxia were neutralized by co-exposure to hypoxia and CD95L. Furthermore, inhibition of protein synthesis by cycloheximide also reduced glucose consumption and conferred protection from hypoxia, but did not modulate CD95L-induced cell death under hypoxic conditions. These results suggest that death ligands may be useful to target hypoxic tumour cells resistant to conventional therapies or to complement strategies aiming at the induction of tumour hypoxia.

Synthesis of cytochrome c oxidase 2: a p53-dependent metabolic regulator that promotes respiratory function and protects glioma and colon cancer cells from hypoxia-induced cell death

“…Profound hypoxia (0.1% O 2 ) was induced by incubating cells in Gas Pak pouches for anaerobic culture (Becton-Dickinson, Heidelberg, Germany) (Steinbach et al, 2003). Moderate hypoxia (1% O 2 ) was induced in a Labotect incubator (Goettingen, Germany).…”

“…Thereafter, cells were washed with cold phosphate-buffered saline and lysed in lysis buffer (50 mM Tris-HCl (pH 8), 120 mM NaCl, 5 mM EDTA, 0.5% NP-40) containing protease inhibitors (Roche, Mannheim, Germany). Cellular lysates were prepared as described (Steinbach et al, 2003) and subjected to sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Membranes were probed with antibodies to human p53 (sc-263; Santa Cruz, San Diego, CA, USA), murine p53 (p53-Ab-4; Merck, Darmstadt, Germany), GAPDH (MAB374; Chemicon, Nuernberg, Germany), AMPKa (2531; Cell Signaling, Frankfurt am Main, Germany), phospho-AMPKa (2531; Cell Signaling), acetyl coA carboxylase (3662; Cell Signaling), phospho-acetyl coA carboxylase (3661; Cell Signaling), actin (SC1616; Santa Cruz) or phospho-4E binding protein 1 (9459; Cell Signaling).…”

“…The ATP concentration was determined by luciferase assay with the CLS II kit (Boehringer, Mannheim, Germany) (Steinbach et al, 2003).…”

Mutant and Misfolded Human Growth Hormone is Rapidly Degraded Through the Proteasomal Degradation Pathway in a Cellular Model for Isolated Growth Hormone Deficiency Type II

“…Cells synchronically grown in cell culture flasks were prepared for electron microscopy as described earlier (13). Specimens were analysed and documented with an EM 10A electron microscope (Zeiss, Oberkochen, Germany).…”

Epidermal growth factor receptor and mammalian target of rapamycin as therapeutic targets in malignant glioma: current clinical status and perspectives

“…However, under the conditions of the tumor microenvironment, inhibition of the EGFR-PI3K/Akt-mTOR cascade may also have negative consequences, because it may affect the sensitivity towards starvation. We have recently reported that, in malignant glioma cells, inhibition of EGFR, Akt and mTOR profoundly reduces energy consumption and thereby provides protection against hypoxic stress [26][27][28]. Given the increasingly evident role of hypoxia as a key selective pressure for solid tumors and the prominent occurrence of hypoxia in gliomas, spontaneously or potentially induced by antiangiogenic therapies [29,30], this mechanism may limit the efficacy of anti-EGFR and anti-mTOR therapies.…”