Yonsei Med J.  2006 Aug;47(4):551-557. 10.3349/ymj.2006.47.4.551.

Hydrogen Peroxide Upregulates TNF-Related Apoptosis-Inducing Ligand (TRAIL) Expression in Human Astroglial Cells, and Augments Apoptosis of T Cells

• Kwon D
• Choi IH

• KMID: 1779506
• DOI: http://doi.org/10.3349/ymj.2006.47.4.551

Abstract

The brain is particularly vulnerable to oxygen free radicals, and these radicals have been implicated in the pathology of several neurological disorders. In this study, the modulation of TNF-related apoptosis-inducing ligand (TRAIL) expression by oxidative stress was shown in LN215 cells, an astroglioma cell line. Hydrogen peroxide (H2O2) treatment increased TRAIL expression in LN215 cells and H2O2-induced TRAIL augmented apoptosis in Peer cells, a cell line sensitive to TRAIL- mediated cell death. Our findings suggest that the upregulation of TRAIL in astroglial cells may abrogate immune cell effector functions.

Keyword

Astroglial cells; hydrogen peroxide; TRAIL; cyclosporin A; apoptosis; oxidative stress; neuroimmunology

MeSH Terms

*Up-Regulation
TNF-Related Apoptosis-Inducing Ligand/*biosynthesis
T-Lymphocytes/*metabolism
Ribonucleases/metabolism
Oxidative Stress
Immunosuppressive Agents/pharmacology
Hydrogen Peroxide/*pharmacology
Humans
*Gene Expression Regulation, Neoplastic
Cyclosporine/pharmacology
Cell Line, Tumor
Astrocytes/*metabolism
*Apoptosis
Anoxia
Allergy and Immunology

Figure

• 

  Fig. 1 Gene expression of TRAIL in human astroglial cells after treatment with hydrogen peroxide (H2O2) in a dose or time-dependent manner. (A) LN215 cells were treated with varying doses of hydrogen peroxide (0 to 800 µM) for 4 hrs. (B) LN215 cells were incubated with hydrogen peroxide 800 µM for various times (0 to 24 hrs). Total RNA was measured for TRAIL mRNA by RT-PCR (A) or RNase protection assay (B). Cells not treated with H2O2 were used as control (CNTL). The data are representative of three experiments.

• 

  Fig. 2 Expression of TRAIL in human astroglial cells after treatment with H2O2 in a dose or time-dependent manner. (A) LN215 cells were treated with varying doses of hydrogen peroxide (0 to 800 µM) for 18 hrs. (C) LN215 cells were incubated with hydrogen peroxide 800 µM for various times (0 to 24 hrs). For western blot, 20 µg of total protein was loaded in 12% gel and anti-TRAIL monoclonal Ab was used as a primary Ab. (B) FACS analysis of TRAIL expression on LN215 were treated with 800 µM of hydrogen peroxide for 18 hrs. The data are representative of three independent experiments.

• 

  Fig. 3 Inhibition of TRAIL expression by cyclosporin A (CsA) in human astroglial cells after treatment with H2O2. (A) LN215 cells were pre-incubated with CsA (0.01 to 10 µM) for 30 min, were treated with hydrogen peroxide (800 µM) for 4 hrs and then gene expression of TRAIL were measured by RT-PCR. Cells not treated with H2O2 were used as a control. β-actin was used as the internal control. The fold induction was calculated as follows: (TRAIL/β-actin intensity in the study group)/(TRAIL/β-actin intensity in the control group). The fold induction value represents mean ± SD of triplicate experiments. (B) LN215 cells were pre-incubated with CsA (1 µM) for 30 min, were treated with hydrogen peroxide (800 µM) for 18 hrs and then expression of TRAIL were measured by western blot. The data are representative of three independent experiments.

• 

  Fig. 4 Induction of apoptosis in Peer cells by human astroglial cells treated with H2O2. LN215 cells were incubated with 800 µM of H2O2 for 18 hrs in serum free medium. LN215 cells were washed to remove H2O2 and then co-cultured with Peer cells for 24 hrs. Effector cells (LN215 cells) and target cells (Peer cells) were co-cultured at a ratio of 4 : 1. Cell death was measured by staining with annexin V. Control LN215 cells were not treated with H2O2, but maintained in serum free condition for 18 hrs. LN215 were pre-incubated for 30 min with CsA (1 µg/mL), and incubated with H2O2. The fold induction value represents a mean ± SD of triplicate experiments.

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