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Daphnoretin-induced apoptosis in HeLa cells: a possible mitochondria-dependent pathway

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Abstract

Daphnoretin is a bicoumarin compound isolated from a natural product, Wikstroemia indica, which has been used to treat many diseases. It has strong antiviral and anti-tumor activities. Taking the anti-tumor activity of daphnoretin as a starting point, the present study aimed to test the pro-apoptotic effect of daphnoretin and its underlying mechanism in HeLa cells. The inhibitory effects of daphnoretin on viability and proliferation of HeLa cells were determined by the MTT assay. Daphnoretin-induced apoptotic morphological changes were analyzed by mitochondrial membrane potential and Hoechst staining. The number and stage of apoptotic HeLa cells were determined by flow cytometry. Gene expression was determined by reverse-transcription polymerase chain reaction. Protein expression was determined by western blot. The caspase activity of HeLa cells was detected by a caspase-3 and caspase-9 colorimetric assay kit. We found that daphnoretin significantly inhibited HeLa cells’ viability by the MTT assay and flow cytometry. The nuclei of the apoptotic cells exhibited strong, blue fluorescence in Hoechst staining. Bax mRNA and protein levels were increased while bcl-2 mRNA levels were decreased after daphnoretin treatment. Daphnoretin also activated both caspase-3 and caspase-9. These findings suggest that daphnoretin promotes apoptosis of HeLa cells in a mitochondria-mediated way. Daphnoretin therefore has potential to be a promising drug to treat uterine cervix cancer.

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Acknowledgments

We would like to thank Dr. Stefanie MAERZ for shepherding the manuscript. The current study was mainly supported by Shanghai Science Foundation for Youths (Project Number: 2009Y016) and Clinical Pharmacy Fund of Shanghai Pharmaceutical Association (Project Number: 2010-YY-01-15).

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Authors and Affiliations

  1. Department of Pharmacology, School of Pharmacy, Fudan University, 826 Zhang Heng Road, Pudong New Area, Shanghai, 201203, China

    Jun-Tao Kan, Ze-Yu Cheng, Yi-Zhun Zhu & Wei Guo

  2. Institute of Biomedical Sciences, Fudan University, 826 Zhang Heng Road, Pudong New Area, Shanghai, 201203, China

    Jun-Tao Kan & Yi-Zhun Zhu

  3. Obstetrics and Gynecology Hospital, Fudan University, Shanghai, 200011, China

    Zhen-Yu Yang & Xian-Li Wang

Authors
  1. Zhen-Yu Yang
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  2. Jun-Tao Kan
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  3. Ze-Yu Cheng
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  4. Xian-Li Wang
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  5. Yi-Zhun Zhu
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  6. Wei Guo
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Corresponding author

Correspondence to Wei Guo.

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Zhen-Yu Yang, Jun-Tao Kan contributed equally to this work and are co-first authors for this paper.

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Yang, ZY., Kan, JT., Cheng, ZY. et al. Daphnoretin-induced apoptosis in HeLa cells: a possible mitochondria-dependent pathway. Cytotechnology 66, 51–61 (2014). https://doi.org/10.1007/s10616-013-9536-8

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  • DOI: https://doi.org/10.1007/s10616-013-9536-8

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