Annona muricata leaves induced apoptosis in A549 cells through mitochondrial-mediated pathway and involvement of NF-κB

BMC Complementary and Alternative Medicine

Hamed Karimian³  Elham Rouhollahi¹  Mohammadjavad Paydar¹  Habsah Abdul Kadir²  Soheil Zorofchian Moghadamtousi² 

[1] Department of Pharmacology, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia;Biomolecular Research Group, Biochemistry Program, Institute of Biological Sciences, Faculty of Science, University of Malaya, Kuala Lumpur, Malaysia;Department of Pharmacy, Faculty of Medicine, University of Malaya, Kuala Lumpur, Malaysia

关键词: NF-κB;    Mitochondria;    Caspase;    Apoptosis;    Lung cancer;    Annona muricata;   

Others  :  1086973
DOI  :  10.1186/1472-6882-14-299

 received in 2014-06-19, accepted in 2014-08-05,  发布年份 2014

【 摘 要 】

Background

Annona muricata leaves have been reported to have antiproliferative effects against various cancer cell lines. However, the detailed mechanism has yet to be defined. The current study was designed to evaluate the molecular mechanisms of A. muricata leaves ethyl acetate extract (AMEAE) against lung cancer A549 cells.

Methods

The effect of AMEAE on cell proliferation of different cell lines was analyzed by MTT assay. High content screening (HCS) was applied to investigate the suppression of NF-κB translocation, cell membrane permeability, mitochondrial membrane potential (MMP) and cytochrome c translocation from mitochondria to cytosol. Reactive oxygen species (ROS) formation, lactate dehydrogenase (LDH) release and activation of caspase-3/7, -8 and -9 were measured while treatment. The western blot analysis also carried out to determine the protein expression of cleaved caspase-3 and -9. Flow cytometry analysis was used to determine the cell cycle distribution and phosphatidylserine externalization. Quantitative PCR analysis was performed to measure the gene expression of Bax and Bcl-2 proteins.

Results

Cell viability analysis revealed the selective cytotoxic effect of AMEAE towards lung cancer cells, A549, with an IC₅₀ value of 5.09 ± 0.41 μg/mL after 72 h of treatment. Significant LDH leakage and phosphatidylserine externalization were observed in AMEAE treated cells by fluorescence analysis. Treatment of A549 cells with AMEAE significantly elevated ROS formation, followed by attenuation of MMP via upregulation of Bax and downregulation of Bcl-2, accompanied by cytochrome c release to the cytosol. The incubation of A549 cells with superoxide dismutase and catalase significantly attenuated the cytotoxicity caused by AMEAE, indicating that intracellular ROS plays a pivotal role in cell death. The released cytochrome c triggered the activation of caspase-9 followed by caspase-3. In addition, AMEAE-induced apoptosis was accompanied by cell cycle arrest at G₀/G₁ phase. Moreover, AMEAE suppressed the induced translocation of NF-κB from cytoplasm to nucleus.

Conclusions

Our data showed for the first time that the ethyl acetate extract of Annona muricata inhibited the proliferation of A549 cells, leading to cell cycle arrest and programmed cell death through activation of the mitochondrial-mediated signaling pathway with the involvement of the NF-kB signalling pathway.

【 授权许可】

   
2014 Moghadamtousi et al.; licensee BioMed Central Ltd.

【 图 表 】

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