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Targeting 14-3-3 protein, difopein induces apoptosis of human glioma cells and suppresses tumor growth in mice

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Abstract

14-3-3 protein has emerged as critical regulators of diverse cellular responses. Previous studies found that strong 14-3-3 protein expression was observed and associated with tumor genesis and progression in glioma. Here, we further elucidated the role of 14-3-3 protein in apoptosis of human glioma U251 and U87 cells by global inhibition of 14-3-3 functions with a general 14-3-3 antagonist, difopein. In vitro, morphological observation and DNA laddering assay showed that difopein-treated glioma cells displayed outstanding apoptosis characteristics, such as nuclear fragmentation, appearance of membrane-enclosed apoptotic bodies and DNA laddering fragment. Moreover, flow cytometric detection of phosphatidylserine externalization indicated that difopein-induced apoptosis occurred in a time-dependent manner. Interestingly, inhibiting 14-3-3 with small interfere RNA also induce apoptosis of human glioma U251 cells. Furthermore, RT-PCR and western blot assay further substantiated that difopein had strong effects to induce glioma cell apoptosis through down-regulating Bcl-2, up-regulating Bax and activating caspase-9 and caspase-3. In vivo, retroviral vector was constructed and retroviral-mediated transfer of difopein to glioma was implanted in nude mice. Difopein effectively hindered proliferation and triggered apoptosis of tumor cells implanted into nude mice. This work not only reveals a critical role of 14-3-3 in apoptosis suppression in glioma cells, but also identifies and validates 14-3-3 as a potential molecular target for anticancer therapeutic development.

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Acknowledgments

Supported by the National Natural Science Foundation of China (39970752) and Scientific and Technological Project of ShaanXi Province (2008K09-09). We gratefully acknowledge Michael Brenner (university of Alabama in Birmingham, USA) for providing pGfa2-LacZ plasmid, Wei Zhang (Biochemistry department in our university) for providing retroviral vector pLncx, and Haian Fu (Department of Pharmacology in Emory University, USA) for providing pSCM138 plasmid. We thank Jielai Xia’s (statistics department in our university) help in statistical analysis. We thank for Xiaoyan Chen (immune department in our university) assistance with flow cytometry, Angang Yang (immune department in our university) and Haian Fu for their comments on the manuscript. We thank Juan Li, Guo Geng, Wei Lin, Wei Zhang, LuHua Zhang, Yuhai Zhang and Jinxiang Chen for their helps in the process of major revision. And we also thank Junli Huo and Wanjuan Yang for material preparing for this research.

Conflict of interest statement

The authors declare that they have no conflict of interest.

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Authors and Affiliations

  1. Institute of Neurosurgery, Xijing Hospital, Fourth Military Medical University, Changle Western Road 17#, 710032, Xi’an, Shaanxi Province, People’s Republic of China

    Weidong Cao, Xiaoliang Yang, Jie Zhou, Xiang Zhang & Zhou Fei

  2. Pharmacology Department, Fourth Military Medical University, Changle Western Road 17#, 710032, Xi’an, Shaanxi Province, People’s Republic of China

    Zenghui Teng

  3. Department of Neurosurgery, Central Hospital of Xi’an City, Fifth Western Road 161#, 710003, Xi’an, Shaanxi Province, People’s Republic of China

    Lei Cao

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  1. Weidong Cao
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  2. Xiaoliang Yang
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  3. Jie Zhou
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  4. Zenghui Teng
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  5. Lei Cao
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  6. Xiang Zhang
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  7. Zhou Fei
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Corresponding author

Correspondence to Xiang Zhang.

Additional information

The authors Weidong Cao and Xiaoliang Yang contributed equally to this work.

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Cao, W., Yang, X., Zhou, J. et al. Targeting 14-3-3 protein, difopein induces apoptosis of human glioma cells and suppresses tumor growth in mice. Apoptosis 15, 230–241 (2010). https://doi.org/10.1007/s10495-009-0437-4

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  • DOI: https://doi.org/10.1007/s10495-009-0437-4

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