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

Sawada, Michio; Nakashima, Shigeru; Banno, Yoshiko; Yamakawa, Haruki; Takenaka, Katsunobu; Shinoda, Jun; Nishimura, Yasuaki; Sakai, Noboru; Nozawa, Yoshinori

doi:10.1038/sj.onc.1203699

Cited by 88 publications

(56 citation statements)

References 60 publications

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“…On the other hand, the in¯uence of proapoptotic Bax on ceramide-mediated cell death has only been indirectly studied in C6 rat glioma cells. It was shown that overexpression of Bax in these cells enhanced etoposide-induced apoptosis without a ecting ceramide formation after etoposide treatment (Sawada et al, 2000b). From their results the authors concluded that Bax promotes apoptosis regardless of ceramide formation.…”

Section: Introductionmentioning

confidence: 65%

Ceramide induces mitochondrial activation and apoptosis via a Bax-dependent pathway in human carcinoma cells

et al. 2002

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The intracellular pathways leading to mitochondrial activation and subsequent cell death in the ceramidemediated stress response have been intensively studied in recent years. Experimental evidence has been provided that ceramide-induced apoptosis is inhibited by overexpression of antiapoptotic proteins of the Bcl-2 family. However, the direct e ect of proapoptotic gene products, e.g. Bax, on ceramide-induced death signalling has not yet been studied in detail. In the present work, we show by measurement of mitochondrial permeability transition, cytochrome c release, activation of caspase-3 and DNA fragmentation that ceramide-induced apoptosis is marginal in Bax-negative DU 145 cells. Reconstitution of Bax by generation of DU 145 cells stably expressing this proapoptotic factor, clearly enhanced ceramideinduced apoptosis at all levels of the mitochondrial signalling cascade. Using the broad-range caspase inhibitor zVAD-fmk and zDEVD-fmk, an inhibitor of caspase-3-like activities, we demonstrate that the ceramide-induced mitochondrial activation in Bax-transfected DU 145 cells is caspase-independent. On the other hand, apoptotic events located downstream of the mitochondria, e.g. DNA fragmentation, were shown to be caspase-dependent. This in¯uence of Bax on ceramide-induced apoptosis was con®rmed in another cellular system: whereas Bax-positive HCT116 wild type cells were very sensitive towards induction of cell death by C 2 -ceramide, sensitivity of Bax knock-out HCT116 cells was signi®cantly reduced. Thus, we conclude that Bax is a key activator of ceramide-mediated death pathways.

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Section: Introductionmentioning

confidence: 65%

Ceramide induces mitochondrial activation and apoptosis via a Bax-dependent pathway in human carcinoma cells

et al. 2002

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“…Interestingly, this effect of BEX2 is similar to the protective function of Bcl-2 overexpression against the ceramide-mediated mitochondrial apoptosis and cytochrome c release. 31,32 Moreover, BEX2 partially protected the breast cancer cells against STS-mediated mitochondrial apoptosis (Fig. 3c), which indicates that the anti-apoptotic function of BEX2 is not limited to the ceramide response.…”

Section: Discussionmentioning

confidence: 93%

BEX2 regulates mitochondrial apoptosis and G1 cell cycle in breast cancer

Naderi¹,

Liu²,

Bennett³

2010

Intl Journal of Cancer

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We have recently demonstrated that BEX2 is differentially expressed in primary breast tumors and BEX2 expression is required for the Nerve Growth factor inhibition of ceramide-induced apoptosis in breast cancer. In this study we investigate the functional role of BEX2 in the survival and growth of breast cancer cells. We demonstrate that BEX2 downregulation induces mitochondrial apoptosis and sensitizes breast cancer cells to the pro-apoptotic effects of ceramide, doxorubicin and staurosporine. In addition, BEX2 overexpression protects the breast cancer cells against mitochondrial apoptosis. We show that this effect of BEX2 is mediated through the modulation of Bcl-2 protein family, which involves the positive regulation of anti-apoptotic member Bcl-2 and the negative regulation of pro-apoptotic members BAD, BAK1 and PUMA. Moreover, our data suggests that BEX2 expression is required for the normal cell cycle progression during G1 in breast cancer cells through the regulation of cyclin D1 and p21. To further support the significance of BEX2 in the pathogenesis of breast cancer we demonstrate that BEX2 overexpression is associated with a higher activation of the Bcl-2/NF-jB pathway in primary breast tumors. Furthermore, we show that BEX2 downregulation results in a higher expression and activity of protein phosphatase 2A. The modulation of protein phosphatase 2A, which is also known to mediate the cellular response to ceramide, provides a possible mechanism to explain the BEX2-mediated cellular effects. This study demonstrates that BEX2 has a significant role in the regulation of mitochondrial apoptosis and G1 cell cycle in breast cancer.

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“…The activities of both N-and A-SMases were determined using a mixed micelle assay system. [6][7][8] For measuring N-SMase activity, the membrane fractions (20 mg protein) were mixed with [choline-methyl- 14 C]SM (40 000 c.p.m. in 1 nmol of bovine brain SM in 0.25% Triton X-100 solubilized by sonication) in 0.1 M Tris/HCl buffer (pH 7.4) containing 6 mM MgCl 2 , and the reaction mixture was incubated for 30 min at 371C.…”

Section: Smase Assaymentioning

confidence: 99%

“…5 We previously reported the molecular mechanism by which etoposide, an anticancer drug, induces ceramide generation in human glioma cells. [6][7][8] However, the role of ceramide in radiation-induced apoptosis in gliomas remains to be clarified.…”

Section: Introductionmentioning

confidence: 99%

p53-Independent ceramide formation in human glioma cells during γ-radiation-induced apoptosis

Hara

Nakashima

Kiyono³

et al. 2004

Cell Death Differ

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Although the p53 tumor-suppressor gene product plays a critical role in apoptotic cell death induced by DNA-damaging chemotherapeutic agents, human glioma cells with functional p53 were more resistant to c-radiation than those with mutant p53. U-87 MG cells with wild-type p53 were resistant to c-radiation. U87-W E6 cells that lost functional p53, by the expression of type 16 human papillomavirus E6 oncoprotein, became susceptible to radiation-induced apoptosis. The formation of ceramide by acid sphingomyelinase (A-SMase), but not by neutral sphingomyelinase, was associated with p53-independent apoptosis. SR33557 (2-isopropyl-1-(4-[3-Nmethyl-N-(3,4-dimethoxybphenethyl)amino]propyloxy)benzene-sulfonyl) indolizine, an inhibitor of A-SMase, suppressed radiation-induced apoptotic cell death. In contrast, radiationinduced A-SMase activation was blocked in glioma cells with endogenous functional p53. The expression of acid ceramidase was induced by c-radiation, and was more evident in cells with functional p53. N-oleoylethanolamine, which is known to inhibit ceramidase activity, unexpectedly downregulated acid ceramidase and accelerated radiation-induced apoptosis in U87-W E6 cells. Moreover, cells with functional p53 could be sensitized to c-radiation by N-oleoylethanolamine, which suppressed radiation-induced acid ceramidase expression and then enhanced ceramide formation. Sensitization to cradiation was also observed in U87-MG cells depleted of functional p53 by retroviral expression of small interfering RNA. These results indicate that ceramide may function as a mediator of p53-independent apoptosis in human glioma cells in response to c-radiation, and suggest that p53-dependent expression of acid ceramidase and blockage of A-SMase activation play pivotal roles in protection from c-radiation of cells with endogenous functional p53.

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

Cited by 88 publications

References 60 publications

Ceramide induces mitochondrial activation and apoptosis via a Bax-dependent pathway in human carcinoma cells

Ceramide induces mitochondrial activation and apoptosis via a Bax-dependent pathway in human carcinoma cells

BEX2 regulates mitochondrial apoptosis and G1 cell cycle in breast cancer

p53-Independent ceramide formation in human glioma cells during γ-radiation-induced apoptosis

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