Apoptosis induction preceded by mitochondrial depolarization in multiple myeloma cell line U266 by 2-aminophenoxazine-3-one

Ken Shirato, Kazuhiko Imaizumi, Keisuke Miyazawa, Akira Takasaki, Junichiro Mizuguchi, Xiao Fang Che, Shinichi Akiyama, Akio Tomoda

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    Abstract

    The aim of the present study was to investigate the mechanism of apoptosis in human multiple myeloma cell line, U266, caused by 2-aminophenoxazine-3-one (Phx-3). Flow-cytometrical and morphological analyses showed that Phx-3 increased the population of annexin V-positive cells including early stage apoptotic cells and late stage apoptotic cells and induced DNA fragmentation or apoptotic body formation in U266 cells, indicating that Phx-3 induced the apoptosis of U266 cells. Activity of caspase-3 was extensively increased in U266 cells treated with Phx-3 time-dependently within 24 h, but this Phx-3-stimulated activity of the enzyme in the cells was completely cancelled by the addition of N-benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (z-VAD-fmk), a pan-caspase inhibitor. The addition of z-VAD-fmk almost blocked the apoptotic effect of Phx-3 against U266 cells, indicating that Phx-3-induced apoptosis of U266 cells was dependent on a caspase signaling pathway. Moreover, the apoptosis of U266 cells occurred after the induction of cell cycle arrest of the cells in the S and G2/M phase, the loss of mitochondrial membrane potential, and activation of caspase-3 reached maximum, which were caused by Phx-3 within 24 h. These results support the views that the apoptosis of U266 cells caused by Phx-3 may be preceded by the cell cycle arrest, depolarization of mitochondria and activation of caspase-3. These results support the view that Phx-3 may be utilized in future as chemotherapeutic agent against multiple myeloma which is extremely refractory to chemotherapy.

    Original languageEnglish
    Pages (from-to)62-67
    Number of pages6
    JournalBiological and Pharmaceutical Bulletin
    Volume31
    Issue number1
    DOIs
    Publication statusPublished - 2008 Jan

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    Keywords

    • 2-aminophenoxazine-3-one
    • Apoptosis
    • Caspase-3
    • Mitochondrial membrane potential
    • Multiple myeloma

    ASJC Scopus subject areas

    • Molecular Medicine
    • Pharmacology, Toxicology and Pharmaceutics(all)

    Cite this

    Shirato, K., Imaizumi, K., Miyazawa, K., Takasaki, A., Mizuguchi, J., Che, X. F., Akiyama, S., & Tomoda, A. (2008). Apoptosis induction preceded by mitochondrial depolarization in multiple myeloma cell line U266 by 2-aminophenoxazine-3-one. Biological and Pharmaceutical Bulletin, 31(1), 62-67. https://doi.org/10.1248/bpb.31.62