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Influence of Bax or Bcl-2 overexpression on the ceramide-dependent apoptotic pathway in glioma cells

Oncogene volume 19, pages 3508–3520 (2000)Cite this article

A Corrigendum to this article was published on 30 November 2006

This article has been updated

Abstract

Ceramide has recently been regarded as a potential mediator of apoptosis. In the present study, the effects of Bcl-2 and Bax on the ceramide-mediated apoptotic pathways were examined in glioma cells overexpressing Bcl-2 or Bax. Etoposide, cisplatin and tumor necrosis factor-α induced apoptosis of C6 rat glioma cells which was associated with ceramide formation due to activation of neutral sphingomyelinase, followed by release of mitochondrial cytochrome c into the cytosol and activation of caspases-9 and -3. The growth of C6 cells stably overexpressing either Bcl-2 or Bax was almost equal to that of the vector-transfected cells. Bax overexpression enhanced etoposide-induced apoptosis through acceleration of cytochrome c release and caspases activation. However, Bax had no effect on ceramide formation. Similar findings were obtained in C6 cells and U87-MG human glioblastoma cells which were transiently overexpressed with Bax. In contrast, Bcl-2 overexpression resulted in a retardation of the apoptotic process via prevention of cytochrome c release and caspases activation, and ceramide formation was also blocked when Bcl-2 was highly overexpressed in glioma cells. In addition, transient overexpression of Bcl-xL also exerted inhibitory effects on ceramide formation and apoptotic cell death induced by etoposide. These results indicate that Bax promotes apoptosis regardless of ceramide formation and that Bcl-2 or Bcl-xL prevents ceramide formation by repressing neutral sphingomyelinase as well as ceramide-induced cytochrome c release.

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Change history

Abbreviations

TNF:

tumor necrosis factor

SM:

sphingomyelin

SMase:

sphingomyelinase

ROS:

reactive oxygen species

FBS:

fetal bovine serum

DMEM:

Dulbecco's modified Eagle's medium

HPTLC:

high performance thin-layer chromatography

PBS:

phosphate-buffered saline

DMSO:

dimethyl sulfoxide

AMC:

7-amino-4-methylcoumarin

SDS–PAGE:

sodium dodecylsulfate polyacrylamide gel electrophoresis

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Acknowledgements

We are grateful to Dr KM Rich for the C6/Bax and C6/Neo cells, and Dr Y Tsujimoto for the mammalian expression vectors containing human bcl-2, human bcl-xL, and mouse bax. This work was supported in part by Grants-in-Aid for Scientific Research on Priority Areas (10212204), (B) (11557104) and (C) (10670136, 11671364) from The Ministry of Education, Science, Sports and Culture of Japan, Special Coordination Funds for Promoting Science and Technology from The Science and Technology Agency of Japan, and The Naito Foundation.

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Authors and Affiliations

  1. Department of Neurosurgery, Gifu University School of Medicine, Tsukasamachi-40, 500-8705, Gifu, Japan

    Motoshi Sawada, Haruki Yamakawa, Katsunobu Takenaka, Jun Shinoda, Yasuaki Nishimura & Noboru Sakai

  2. Department of Biochemistry, Gifu University School of Medicine, Tsukasamachi-40, 500-8705, Gifu, Japan

    Shigeru Nakashima & Yoshiko Banno

  3. Gifu International Institute of Biotechnology, Mitake-cho, 505-0116, Kani-gun, Gifu, Japan

    Yoshinori Nozawa

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Sawada, M., Nakashima, S., Banno, Y. et al. Influence of Bax or Bcl-2 overexpression on the ceramide-dependent apoptotic pathway in glioma cells. Oncogene 19, 3508–3520 (2000). https://doi.org/10.1038/sj.onc.1203699

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