Induction of cell cycle arrest and apoptosis in caspase-3 deficient MCF-7 cells by Dillenia suffruticosa root extract via multiple signalling pathways

BMC Complementary and Alternative Medicine

Maznah Ismail²  Rasedee Abdullah¹  Yoke Kqueen Cheah⁴  Swee Keong Yeap³  Mustapha Umar Imam²  Norsharina Ismail²  Nurdin Armania⁴  Yin Sim Tor²  Latifah Saiful Yazan⁴  Jhi Biau Foo² 

[1] Department of Pathology and Microbiology, Faculty of Veterinary Medicine, Universiti Putra Malaysia, 43400 Serdang, Malaysia;Laboratory of Molecular Biomedicine, Institute of Bioscience, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia;Laboratory of Vaccines & Immunotherapeutics, Institute of Bioscience, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia;Department of Biomedical Science, Faculty of Medicine and Health Sciences, Universiti Putra Malaysia, 43400 UPM Serdang, Selangor, Malaysia

关键词: Oxidative stress;    Apoptosis;    Cell cycle;    Dichloromethane extract;    Dillenia suffruticosa;   

Others  :  1220091
DOI  :  10.1186/1472-6882-14-197

 received in 2013-11-28, accepted in 2014-06-13,  发布年份 2014

【 摘 要 】

Background

Dillenia suffruticosa root dichloromethane extract (DCM-DS) has been reported to exhibit strong cytotoxicity towards breast cancer cells. The present study was designed to investigate the cell cycle profile, mode of cell death and signalling pathways of DCM-DS-treated human caspase-3 deficient MCF-7 breast cancer cells.

Methods

Dillenia suffruticosa root was extracted by sequential solvent extraction. The anti-proliferative activity of DCM-DS was determined by using MTT assay. The mode of cell death was evaluated by using inverted light microscope and Annexin-V/PI-flow cytometry analysis. Cell cycle analysis and measurement of intracellular reactive oxygen species (ROS) were performed by using flow cytometry. MCF-7 cells were co-treated with antioxidants α-tocopherol and ascorbic acid to evaluate whether the cell death was mainly due to oxidative stress. GeXP-based multiplex system was employed to investigate the expression of apoptotic, growth and survival genes in MCF-7 cells. Western blot analysis was performed to confirm the expression of the genes.

Results

DCM-DS was cytotoxic to the MCF-7 cells in a time-and dose-dependent manner. The IC₅₀ values of DCM-DS at 24, 48 and 72 hours were 20.3 ± 2.8, 17.8 ± 1.5 and 15.5 ± 0.5 μg/mL, respectively. Cell cycle analysis revealed that DCM-DS induced G₀/G₁ and G₂/M phase cell cycle arrest in MCF-7 cells at low concentration (12.5 and 25 μg/mL) and high concentration (50 μg/mL), respectively. Although Annexin-V/PI-flow cytometry analysis has confirmed that DCM-DS induced apoptosis in MCF-7 cells, the distinct characteristics of apoptosis such as membrane blebbing, chromatin condensation, nuclear fragmentation and formation of apoptotic bodies were not observed under microscope. DCM-DS induced formation of ROS in MCF-7 cells. Nevertheless, co-treatment with antioxidants did not attenuate the cell death at low concentration of DCM-DS. The pro-apoptotic gene JNK was up-regulated whereby anti-apoptotic genes AKT1 and ERK1/2 were down-regulated in a dose-dependent manner. Western blot analysis has confirmed that DCM-DS significantly up-regulated the expression of pro-apoptotic JNK1, pJNK and down-regulated anti-apoptotic AKT1, ERK1 in MCF-7 cells.

Conclusion

DCM-DS induced cell cycle arrest and apoptosis in MCF-7 cells via multiple signalling pathways. It shows the potential of DCM-DS to be developed to target the cancer cells with mutant caspase-3.

【 授权许可】

   
2014 Foo et al.; licensee BioMed Central Ltd.

【 图 表 】

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