Annona muricata leaves induced apoptosis in A549 cells through mitochondrial-mediated pathway and involvement of NF-κB
© Moghadamtousi et al.; licensee BioMed Central Ltd. 2014
Received: 19 June 2014
Accepted: 5 August 2014
Published: 15 August 2014
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.
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.
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.
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.
KeywordsAnnona muricata Lung cancer Apoptosis Caspase Mitochondria NF-κB
Lung cancer as one of the critical causes of cancer death throughout the world has the prevalent complication of apoptosis resistance against different anticancer agents . Due to the typical asymptomatic progression of lung cancer at an early stage, it is normally diagnosed at an advanced stage (56%). In spite of all the development in chemoradiation and surgical techniques, the 5-year survival rate for patients with advanced stage disease is still 3.6% . In addition, numerous lung cancer survivors suffer from lung dysfunction, particularly patients with the lung surgical history . Thereby, continued research into the development of safe and efficient new anticancer agents against lung cancer cells is urgently necessary for further improvements in cancer therapy.
Apoptosis is a critical physiological process responsible for the homeostatic mechanism and maintenance of cell populations in tissues . Due to the close correlation between the mechanism of apoptosis and the effect of anticancer agents, extensive research has been done on this mode of cell death . The accumulation of reactive oxygen species (ROS) in cancer cells is a critical factor for the induction of apoptosis by natural products [6, 7], since it will result in oxidative DNA damage following by a collapse in mitochondrial membrane potential (MMP) and leakage of cytochrome c, which lead to the activation of the caspase cascade . Furthermore, the perturbation in the expression level of Bax and Blc-2 proteins is an important factor to determine the susceptibility of tumor cells to anticancer agents . Previous anticancer studies also proved that constitutive activation of the ubiquitous transcription factor of NF-κB (nuclear factor-kappa B) is involved in governing the promoting tumor progression of solid and hemopoietic malignancies [10, 11]. Therefore, anticancer agents with the ability to suppress the NF-κB translocation are effectively induce the apoptosis in cancer cells.
Annona muricata L. known as gravel, guanabana and soursop is a member of Custard-Apple plants in the Annonaceae family due to a custard-like texture of its fruit. It is a small deciduous tree with a height of 5–8 m and roundish canopy . This popular fruit tree has been widely cultivated in many tropical countries and traditionally used for an array of diseases and ailments . Previous studies demonstrated a significant cytotoxicity for A. muricata leaves against various cancer cell lines without affecting the normal cells [14, 15]. Due to this tremendous antiproliferative effect, A. muricata was described as “the cancer killer” . Ethanolic extract of A. muricata leaves was suggested to have apoptosis-inducing potential against myelogenous leukemic K562 cells, although the detailed mechanism of action has not been explained . Amongst constituents isolated from A. muricata leaves, namely annonaceous acetogenins, alkaloids and essential oils, annonaceous acetogenins are strongly implied to be responsible for the promising anticancer effect . The principle objective of this study was to examine how A. muricata leaves affecting A549 lung cancer cells, and to investigate the possible mechanism of action involved in this effect.
Plant material and extraction procedures
The plant species (Annona muricata) collected from Ipoh, Malaysia, was authenticated by Dr. Yong Kien Thai, an ethnobotanist from the department of Biological Sciences, University of Malaya. The voucher specimen number for this plant is KLU47978. The air-dried leaves of A. muricata (1 kg) were cut into fine pieces using a mill grinder and soaked in n-hexane (1500 mL, three times) in conical flasks for four days at room temperature (25–27°C). The n-hexane extract was filtered and the residues were sequentially re-extracted with ethyl acetate (1500 ml, three times) and methanol (1500 ml, three times) using the same method. The resultant filtrate was concentrated to dryness by a Buchi R110 Rotavapor (Buchi Labortechnik AG, Flawil, Switzerland) at 40°C and stored at – 30°C until use. The isolated extracts were dissolved in dimethyl sulfoxide (DMSO) for further experiments.
Cell culture and MTT assay
MCF-7 (human breast cancer cells), MDA-MB-231 (human breast cancer cells), A549 (human lung cancer cells), HepG2 (human hepatoma cells) and WRL-68 (human hepatic cells) cell lines were obtained from American Type Cell Collection (ATCC, Manassas, VA, USA). Cells were cultured in RPMI-1640 medium (Sigma, St. Louis, MO, USA) supplemented with 10% FBS (PAA, Pasching, Austria), 100 U/mL penicillin (PAA) and 50 μg/mL amphotericin B (PAA) at 37°C with 5% CO2. The negative control for all the assays was represented by the untreated medium containing vehicle DMSO (0.1%).
The cytotoxicity of the extracts was determined using the MTT assay as originally described by Mossman . Briefly, cells were treated with different concentrations (1.56, 3.12, 6.25, 12.5, 25, 50 and 100 μg/mL) of three isolated extracts (hexane, ethyl acetate and methanol) and curcumin (positive control) in 96-well plates and incubated for 72 h. After the incubation time, MTT dye (20 μL, 5 mg/mL, Sigma) was added to the cells for 4 h followed by incubation with DMSO for 10 min. The colorimetric assay was measured at the absorbance of 570 nm using a microplate reader (Asys UVM340, Eugendorf, Austria). The antiproliferative potential of the extracts was expressed as IC50 values. As an ethyl acetate extract of the leaves (AMEAE) demonstrated the lowest IC50 value against lung cancer A549 cells, we used only AMEAE to continue this study against A549 cells.
LDH release assay
To confirm the cytotoxicity of AMEAE, we carried out lactate dehydrogenase (LDH) release assay using Pierce™ LDH Cytotoxicity Assay Kit (Thermo Scientific™, Pittsburgh, PA, USA). In brief, A549 cells were treated with AMEAE at different concentrations for 48 h. The supernatant of treated A549 cells was transferred into 96-well plate to assess the LDH activity. Triton X-100 (2%) served as a positive control was used to completely lyse the cells and release the maximum LDH. Next, the LDH reaction solution (100 μl) was added to the cells for 30 min. The red color intensity presenting the LDH activity was measured by the absorbance at 490 using a Tecan Infinite®200 Pro (Tecan, Männedorf, Switzerland) microplate reader. The level of released LDH from treated cells was expressed as a percentage of positive control.
Acridine orange/propidium iodide (AO/PI) double staining assay
Morphological changes induced by AMEAE in A549 cells were analyzed using Acridine orange/Propidium iodide (AO/PI) double staining assay. Briefly, A549 cells were seeded in 60 mm2 culture dishes followed by treatment with AMEAE (10 μg/mL) for 24, 48 and 72 h. After the incubation time, extract-untreated and treated A549 cells were harvested and washed with PBS. Then, the pellets were stained with 10 μg/mL of AO/PI (1 mg/mL). The stained cells were then observed under a BX51 UV-fluorescent microscope (Olympus, Tokyo, Japan) within 30 min.
Induction of the early and late apoptosis by AMEAE was further studied via Annexin-V/PI staining assay. Briefly, A549 cells (1 × 106) were plated in 60 mm2 culture dishes and treated with vehicle DMSO and AMEAE (10 μg/mL) for 24, 48 and 72 h. After harvest of adherent and suspension cells and washing them twice with PBS, they were re-suspended in Annexin-V binding buffer (BD Biosciences, San Jose, CA, USA) and stained with Annexin-V-FITC (BD) and PI (Sigma) according to the manufacturer’s instructions. The fluorescence intensity of A549 cells was then analyzed by flow cytometry (BD FACSCanto™ II, San Jose, CA, USA) through quadrant statistics for necrotic and apoptotic cell populations. PI was used for detection of the late apoptosis and necrosis while Annexin-V was consumed for the detection of the early and late apoptosis.
Cell cycle assay
A flow cytometry analysis was carried out to determine the cell cycle distribution in treated A549 cells with AMEAE. In brief, A549 cells (5 × 104 cells/mL) were treated with AMEAE (10 μg/mL) for 24, 48 and 72 h. After fixation with cold ethanol, cells were washed with PBS and stained with PI (50 μl, 10 mg/mL) for 1 h at 37°C. In addition, RNase A (10 mg/mL) was also used to limit the ability of the PI to bind only to DNA molecules. The stained cells were analyzed for DNA content using flow cytometer (BD FACSCanto™ II).
Reactive oxygen species (ROS) assay
The effect of AMEAE on the ROS formation in A549 cells was determined by ROS assay. Briefly, treated lung cancer cells with AMEAE at different concentrations in 96-well plates were incubated for 24 h. After the incubation time, the treated cells were stained with dihydroethidium (DEH) at 2.5 μg/mL and Hoechst 33342 (500 nM) dyes for 30 min. Then, cells were fixed with paraformaldehyde (3.5%) for 15 min and washed with PBS twice. The Cellomics ArrayScan HCS reader was used to measure the ROS generation in treated A549 cells.
To further determine the role of ROS generation in AMEAE-induced antiproliferative effect, A549 cells were treated with antioxidants prior to treatment with AMEAE and the cell viability was measured after 24 h. In brief, A549 cells in the exponential phase of growth were supplemented with antioxidants superoxide dismutase (SOD, 300 U/mL) and catalase (400 U/mL) for 1 h prior to AMEAE (20 μg/mL) treatment for 24 h. After incubation time, the cell viability analysis was carried out using a microplate reader (Asys UVM340, Eugendorf, Austria).
Multiple cytotoxicity assay
To simultaneously determine the crucial apoptotic events in A549 cells after treatment with AMEAE, we used Cellomics Multiparameter Cytotoxicity 3 Kit (Thermo Scientific™, Pittsburgh, PA, USA). Briefly, lung cancer A549 cells were seeded into 96-well plates for 24 h. The cells were treated with AMEAE at different concentrations prior to staining the cells with cell permeability and mitochondrial membrane potential (MMP) dyes. Then, cells were fixed and blocked with 1X blocking buffer according to the manufacture’s protocol. Next, primary cytochrome c antibody and secondary DyLight 649 conjugated goat antimouse IgG were added for 1 h. Nuclei of treated cells were also stained with Hoechst 33342 dye. Stained A549 cells in 96-well plates were analyzed using ArrayScan high content screening (HCS) system.
Bioluminescent assays for caspase-8, -9 and -3/7 activities
A dose-dependent study on the caspase-8, -9 and -3/7 activation was carried out using Caspase-Glo® 3/7, 8 and 9 kit (Promega, Madison, WI, USA). In brief, a total of 5 × 103 A549 cells were seeded per well in a white 96-well microplate and incubated with different concentrations of AMEAE for 24 h. Then, caspase-Glo reagent (100 μl) was added to the cells for 30 min. The induced activation of caspases was measured using a Tecan Infinite®200 Pro (Tecan, Männedorf, Switzerland) microplate reader.
To determine the protein expression of cleaved caspase-3 and -9, western blot analysis was carried out as previously described in detail . In brief, A549 cells treated with vehicle DMSO or AMEAE at different concentrations were washed with PBS and lysed in ice-cold Radio Immuno Precipitation Assay (RIPA) buffer. Cell extracts (80 μg protein) were subjected to sodium dodecyl sulfate–polyacrylamide gel electrophoresis (SDS–PAGE), transferred to nitrocellulose membrane, probed with anti-β-actin, anti-cleaved caspase-9 and anti-cleaved caspase-3 (Cell Signaling Technology, Danvers, MA, USA). HRP-conjugated secondary antibodies were used followed by the detection of protein expression using the ECL plus chemiluminescence kit (Amersham Biosciences, Piscataway, NJ, USA).
Quantitative PCR analysis
The expression of the Bax and Bcl-2 in treated A549 cells was analyzed by quantitative PCR analysis. After treatment of A549 cells with the AMEAE extract at different concentrations for 24 h, Zymo Research Quick-RNA™ MiniPrep kit (Zymo Research, Freiburg, Germany) was used to isolate total RNAs according to the manufacture’s protocol. Then, High Capacity RNA-to-cDNA™ kit (Applied Biosystems, Foster City, CA, USA) was used to synthesize complementary DNAs. Quantitative PCR was carried out with TaqMan® Gene Expression Assays and TaqMan® Fast Advanced Master Mix using the Applied Biosystems StepOnePlus™ system. GAPDH was used to normalize all data. The IDs for TaqMan® Gene Expression Assays used in this study are GAPDH: Hs02758991_g1, Bcl-2: Hs00608023_m1 and Bax: Hs00180269_m1.
Measurement of NF-κB activity
The Cellomics ArrayScan HCS system was used to analyze the suppressive effect of AMEAE on the nuclear translocation of NF-κB induced by TNF-α. The experiment was carried out using Cellomics nucleus factor-κB (NF-κB) activation kit (Thermo Scientific) as previously described . In brief, A549 cells (1.0 × 104 cells/well) were treated with AMEAE at different concentrations in a 96-well plate for 3 h. The treated A549 cells were stimulated by TNF-α (1 ng/mL) for 30 min. Then, cells were fixed and stained according to the manufacturer’s protocol and analyzed using Array Scan HCS Reader and Cytoplasm to Nucleus Translocation Bioapplication software.
Data are presented as mean ± SEM of three individual experiments. Statistical analysis was performed with a one-way ANOVA analysis using the Prism statistical software package (GraphPad Software, USA). Differences were considered as being significant at *p < 0.05.
Results and discussion
AMEAE inhibited the proliferation of cancer cells
IC 50 values of A. muricata leaves extracts on five different cell lines after 72 h treatment
49.92 ± 2.23
38.72 ± 0.99
21.05 ± 0.42
77.92 ± 2.23
89.53 ± 3.93
6.39 ± 0.43
11.36 ± 0.67
5.09 ± 0.41
9.3 ± 0.91
47.10 ± 1.23
85.58 ± 3.55
> 100 *
7.65 ± 0.55
9.34 ± 0.76
11.32 ± 1.54
17.66 ± 1.21
54.24 ± 2.21
Induction of LDH release by AMEAE
Quantification of apoptosis using fluorescence microscope and AO/PI double-staining
Induction of early and late apoptosis using flow cytometry analysis
AMEAE-induced G1 cell cycle arrest
ROS generation induced by AMEAE
Mitochondria-initiated events induced by AMEAE
During apoptosis, mitochondrial membrane potential is frequently disrupted due to the formation of permeability transition pores or the insertions of proapoptotic proteins, such as, Bid or Bax in the mitochondrial membrane . A variety of non-receptor mediated stimuli, including free radicals, radiation, hypoxia and toxin can trigger the intrinsic signaling pathway . The stimuli produce intracellular signals that cause loss in MMP and the opening of the mitochondrial permeability transition pore. These changes in the inner mitochondrial membrane lead to the release of various proteins from the intermembrane space into the cytosol . The main group of released proteins consists of serine protease HtrA2/Omi, cytochrome c and Smac/DIABLO. Cytochrome c forms apoptosome through binding to pro-caspase-9 as well as Apaf-1. Apoptosome activates caspase-9 following with activation of caspase-3 [36, 37].
AMEAE induced caspase-8, -9 and -3/7 activation
AMEAE induced upregulation of Bax and downregulation of Bcl-2 at the gene expression level
NF-κB translocation suppressed by AMEAE
In conclusion, the anticancer potential of ethyl acetate extract of A. muricata leaves was supported by the evidence provided in the present study, including lactate dehydrogenase leakage, reactive oxygen species generation, loss in mitochondrial membrane potential, increase in the level of cytochrome c, upregulation of Bax, downregulation of Bcl-2 and activation of initiator and executioner caspases. The antiproliferative effect of AMEAE was accompanied by cell cycle arrest at G1 phase and suppression of NF-κB translocation. The results confirmed the involvement of intrinsic pathways in induced apoptosis.
A. muricata leaves ethyl acetate extract
High content screening
Nuclear factor-kappa B
Mitochondrial membrane potential
Reactive oxygen species (ROS)
Bcl-2 associated X protein
B-cell lymphoma protein 2
This research was supported by the High Impact Research Chancellery Grant UM.C/625/1/HIR/175 from the University of Malaya, University of Malaya Research Grant (RP001-2012C) and Postgraduate Research Fund (PG118-2013A).
- Ramezanpour M, da Silva KB, Sanderson BJ: Venom present in sea anemone (heteractis magnifica) induces apoptosis in non-small-cell lung cancer A549 cells through activation of mitochondria-mediated pathway. Biotechnol Lett. 2012, 36 (3): 1-7.Google Scholar
- Siegel R, DeSantis C, Virgo K, Stein K, Mariotto A, Smith T, Cooper D, Gansler T, Lerro C, Fedewa S, Lin C, Leach C, Cannady RS, Cho H, Scoopa S, Hachey M, Kirch R, Jemal A, Ward E: Cancer treatment and survivorship statistics, 2012. CA-Cancer J Clin. 2012, 62 (4): 220-241. 10.3322/caac.21149.View ArticlePubMedGoogle Scholar
- Mitsudomi T, Morita S, Yatabe Y, Negoro S, Okamoto I, Tsurutani J, Seto T, Satouchi M, Tada H, Hirashima T, Asami K, Katakami N, Takada M, Yoshioka H, Shibata K, Kudoh S, Shimizu E, Saito H, Toyooka S, Nakagawa K, Fukuoka M: Gefitinib versus cisplatin plus docetaxel in patients with non-small-cell lung cancer harbouring mutations of the epidermal growth factor receptor (WJTOG3405): an open label, randomised phase 3 trial. Lancet Oncol. 2010, 11 (2): 121-128. 10.1016/S1470-2045(09)70364-X.View ArticlePubMedGoogle Scholar
- Kim SS, Cho HJ, Kang JY, Kang HK, Yoo TK: Inhibition of androgen receptor expression with small interfering RNA enhances cancer cell apoptosis by suppressing survival factors in androgen insensitive, late stage LNCaP cells. Sci World J. 2013, 2013: doi:10.1155/2013/519397Google Scholar
- Ocker M, Höpfner M: Apoptosis-modulating drugs for improved cancer therapy. Eur Surg Res. 2012, 48 (3): 111-120. 10.1159/000336875.View ArticlePubMedGoogle Scholar
- Schumacker PT: Reactive oxygen species in cancer cells: live by the sword, die by the sword. Cancer Cell. 2006, 10 (3): 175-176. 10.1016/j.ccr.2006.08.015.View ArticlePubMedGoogle Scholar
- Moghadamtousi SZ, Goh BH, Chan CK, Shabab T, Kadir HA: Biological activities and phytochemicals of swietenia macrophylla king. Molecules. 2013, 18 (9): 10465-10483. 10.3390/molecules180910465.View ArticlePubMedGoogle Scholar
- Simon H-U, Haj-Yehia A, Levi-Schaffer F: Role of reactive oxygen species (ROS) in apoptosis induction. Apoptosis. 2000, 5 (5): 415-418. 10.1023/A:1009616228304.View ArticlePubMedGoogle Scholar
- Martinou J-C, Youle RJ: Mitochondria in apoptosis: Bcl-2 family members and mitochondrial dynamics. Dev Cell. 2011, 21 (1): 92-101. 10.1016/j.devcel.2011.06.017.View ArticlePubMedPubMed CentralGoogle Scholar
- Yang F, Jove V, Chang S, Hedvat M, Liu L, Buettner R, Tian Y, Scuto A, Wen W, Yip MLR, Meter TV, Yen Y, Jove R: Bortezomib induces apoptosis and growth suppression in human medulloblastoma cells, associated with inhibition of AKT and NF-kB signaling, and synergizes with an ERK inhibitor. Cancer Biol Ther. 2012, 13 (6): 349-357. 10.4161/cbt.19239.View ArticlePubMedPubMed CentralGoogle Scholar
- Van Waes C: Nuclear factor-κB in development, prevention, and therapy of cancer. Clin Cancer Res. 2007, 13 (4): 1076-1082. 10.1158/1078-0432.CCR-06-2221.View ArticlePubMedGoogle Scholar
- Adewole SO, Caxton-Martins EA: Morphological changes and hypoglycemic effects of annona muricata Linn.(annonaceae) leaf aqueous extract on pancreatic β-cells of streptozotocin-treated diabetic rats. Afr J Biomed Res. 2006, 9 (3): 173-187.Google Scholar
- Adewole SO, Ojewole JA: Protective effects of annona muricata Linn. (annonaceae) leaf aqueous extract on serum lipid profiles and oxidative stress in hepatocytes of streptozotocin-treated diabetic rats. Afr J Tra Compl Alter Med. 2009, 6 (1): 30-Google Scholar
- George VC, Kumar DN, Rajkumar V, Suresh P, Ashok R: Quantitative assessment of the relative antineoplastic potential of the n-butanolic leaf extract of annona muricata Linn. In normal and immortalized human cell lines. Asian Pac J Cancer P. 2012, 13 (2): 699-704. 10.7314/APJCP.2012.13.2.699.View ArticleGoogle Scholar
- Mishra S, Ahmad S, Kumar N, Sharma BK: Annona muricata (the cancer killer): a review. Glob J Pharma Res. 2013, 2 (1): 1613-1618.Google Scholar
- Ezirim AU, Okachi VI, James AB, Adebeshi OA, Ogunnowo S, Odeghe OB: Induction of apoptosis in myelogenous leukemic K562 cells by ethanolic leaf extract of annona muricata. Indian J Drugs Dis. 2013, 2 (3): 142-151.Google Scholar
- Zeng L, Wu F-E, Oberlies NH, McLaughlin JL, Sastrodihadjo S: Five new monotetrahydrofuran ring acetogenins from the leaves of annona muricata. J Nat Prod. 1996, 59 (11): 1035-1042. 10.1021/np960447e.View ArticlePubMedGoogle Scholar
- Mosmann T: Rapid colorimetric assay for cellular growth and survival: application to proliferation and cytotoxicity assays. J Immunol Methods. 1983, 65 (1): 55-63.View ArticlePubMedGoogle Scholar
- Liew SY, Looi CY, Paydar M, Cheah FK, Leong KH, Wong WF, Mustafa MR, Litaudon M, Awang K: Subditine, a new monoterpenoid indole alkaloid from bark of nauclea subdita (korth.) steud. Induces apoptosis in human prostate cancer cells. PLoS One. 2014, 9 (2): e87286-10.1371/journal.pone.0087286.View ArticlePubMedPubMed CentralGoogle Scholar
- Looi CY, Moharram B, Paydar M, Wong YL, Leong KH, Mohamad K, Arya A, Wong WF, Mustafa MR: Induction of apoptosis in melanoma A375 cells by a chloroform fraction of centratherum anthelminticum (L.) seeds involves NF-kappaB, p53 and Bcl-2-controlled mitochondrial signaling pathways. BMC Complem Altern M. 2013, 13 (1): 166-10.1186/1472-6882-13-166.View ArticleGoogle Scholar
- Chan FK-M, Moriwaki K, De Rosa MJ: Detection of Necrosis by Release of Lactate Dehydrogenase Activity. Immune Homeostasis. Edited by: Snow AL, Lenardo MJ. 2013, New York: Humana Press, 65-70.View ArticleGoogle Scholar
- Häcker G: The morphology of apoptosis. Cell Tissue Res. 2000, 301 (1): 5-17. 10.1007/s004410000193.View ArticlePubMedGoogle Scholar
- Ovadje P, Chatterjee S, Griffin C, Tran C, Hamm C, Pandey S: Selective induction of apoptosis through activation of caspase-8 in human leukemia cells (Jurkat) by dandelion root extract. J Ethnopharmacol. 2011, 133 (1): 86-91. 10.1016/j.jep.2010.09.005.View ArticlePubMedGoogle Scholar
- Rajagopalan V, Hannun YA: Sphingolipid Metabolism and Signaling as a Target for Cancer Treatment. Cell Death Signaling in Cancer Biology and Treatment. Edited by: Johnson DE. 2013, New York: Springer press, 205-229.View ArticleGoogle Scholar
- Ding X, Zhu F-S, Li M, Gao S-G: Induction of apoptosis in human hepatoma SMMC-7721 cells by solamargine from solanum nigrum L. J Ethnopharmacol. 2012, 139 (2): 599-604. 10.1016/j.jep.2011.11.058.View ArticlePubMedGoogle Scholar
- Elmore S: Apoptosis: a review of programmed cell death. Toxicol Pathol. 2007, 35 (4): 495-516. 10.1080/01926230701320337.View ArticlePubMedPubMed CentralGoogle Scholar
- Arur S, Uche UE, Rezaul K, Fong M, Scranton V, Cowan AE, Mohler W, Han DK: Annexin I is an endogenous ligand that mediates apoptotic cell engulfment. Dev Cell. 2003, 4 (4): 587-598. 10.1016/S1534-5807(03)00090-X.View ArticlePubMedGoogle Scholar
- Park M-T, Lee S-J: Cell cycle and cancer. J Biochem Mol Biol. 2003, 36 (1): 60-65. 10.5483/BMBRep.2003.36.1.060.View ArticlePubMedGoogle Scholar
- Pucci B, Kasten M, Giordano A: Cell cycle and apoptosis. Neoplasia. 2000, 2 (4): 291-299. 10.1038/sj.neo.7900101.View ArticlePubMedPubMed CentralGoogle Scholar
- Zorofchian Moghadamtousi S, Karimian H, Khanabdali R, Razavi M, Firoozinia M, Zandi K, Kadir HA: Anticancer and antitumor potential of fucoidan and fucoxanthin. Two main metabolites isolated from brown algae. Sci World J. 2014, 2014: doi:10.1155/2014/768323Google Scholar
- Li T, Kon N, Jiang L, Tan M, Ludwig T, Zhao Y, Baer R, Gu W: Tumor suppression in the absence of p53-mediated cell-cycle arrest, apoptosis, and senescence. Cell. 2012, 149 (6): 1269-1283. 10.1016/j.cell.2012.04.026.View ArticlePubMedPubMed CentralGoogle Scholar
- Adams DJ, Boskovic ZV, Theriault JR, Wang AJ, Stern AM, Wagner BK, Shamji AF, Schreiber SL: High-throughput screening identifies small-molecule enhancers of reactive oxygen species that are nontoxic or cause genotype-selective cell death. ACS Chem Biol. 2013, 8 (5): 923-929. 10.1021/cb300653v.View ArticlePubMedPubMed CentralGoogle Scholar
- Circu ML, Aw TY: Reactive oxygen species, cellular redox systems, and apoptosis. Free Radic Biol Med. 2010, 48 (6): 749-762. 10.1016/j.freeradbiomed.2009.12.022.View ArticlePubMedPubMed CentralGoogle Scholar
- Kushnareva Y, Andreyev AY, Kuwana T, Newmeyer DD: Bax activation initiates the assembly of a multimeric catalyst that facilitates Bax pore formation in mitochondrial outer membranes. PLoS Biol. 2012, 10 (9): e1001394-10.1371/journal.pbio.1001394.View ArticlePubMedPubMed CentralGoogle Scholar
- Saelens X, Festjens N, Walle LV, Grup MV, Loo GV, Vandenabeele P: Toxic proteins released from mitochondria in cell death. Oncogene. 2012, 23 (16): 2861-2874. 2012View ArticleGoogle Scholar
- Hill MM, Adrain C, Duriez PJ, Creagh EM, Martin SJ: Analysis of the composition, assembly kinetics and activity of native Apaf-1 apoptosomes. EMBO J. 2004, 23 (10): 2134-2145. 10.1038/sj.emboj.7600210.View ArticlePubMedPubMed CentralGoogle Scholar
- Mondragón L, Orzáez M, Gortat A, Sancho M, Messeguer A, Vicent MJ, Perez-Paya E: Molecules That Bind a Central Protein Component of the Apoptosome, Apaf-1, and Modulate Its Activity. Apoptosome. Edited by: Cecconi F, D'Amelio M. 2010, Netherlands: Springer press, 75-94.View ArticleGoogle Scholar
- Callus B, Vaux D: Caspase inhibitors: viral, cellular and chemical. Cell Death Differ. 2006, 14 (1): 73-78.View ArticlePubMedGoogle Scholar
- Delgado ME, Olsson M, Lincoln FA, Zhivotovsky B, Rehm M: Determining the contributions of caspase-2, caspase-8 and effector caspases to intracellular VDVADase activities during apoptosis initiation and execution. BBA-Mol Cell Res. 2013, 1833 (10): 2279-2292.Google Scholar
- Cory S, Adams JM: The Bcl2 family: regulators of the cellular life-or-death switch. Nat Rev Cancer. 2002, 2 (9): 647-656. 10.1038/nrc883.View ArticlePubMedGoogle Scholar
- Zamzami N, Kroemer G: The mitochondrion in apoptosis: how Pandora’s box opens. Nat Rev Mol Cell Bio. 2001, 2 (1): 67-71. 10.1038/35048073.View ArticleGoogle Scholar
- Bhardwaj A, Sethi G, Vadhan-Raj S, Bueso-Ramos C, Takada Y, Gaur Y, Nair AS, Shishodia S, Aggarwal BB: Resveratrol inhibits proliferation, induces apoptosis, and overcomes chemoresistance through down-regulation of STAT3 and nuclear factor-κB–regulated antiapoptotic and cell survival gene products in human multiple myeloma cells. Blood. 2007, 109 (6): 2293-2302. 10.1182/blood-2006-02-003988.View ArticlePubMedGoogle Scholar
- Baldwin AS: Control of oncogenesis and cancer therapy resistance by the transcription factor NF-κB. J Clin Invest. 2001, 107 (3): 241-246. 10.1172/JCI11991.View ArticlePubMedPubMed CentralGoogle Scholar
- The pre-publication history for this paper can be accessed here:http://www.biomedcentral.com/1472-6882/14/299/prepub
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