Dataset Information

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

ABSTRACT: 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 ?-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 ?g/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 ?g/mL) and high concentration (50 ?g/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.

SUBMITTER: Foo JB

PROVIDER: S-EPMC4096536 | biostudies-literature | 2014

REPOSITORIES: biostudies-literature

ACCESS DATA

Publications

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

Foo Jhi Biau JB Yazan Latifah Saiful LS Tor Yin Sim YS Armania Nurdin N Ismail Norsharina N Imam Mustapha Umar MU Yeap Swee Keong SK Cheah Yoke Kqueen YK Abdullah Rasedee R Ismail Maznah M

BMC complementary and alternative medicine 20140619

<h4>Background</h4>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.<h4>Methods</h4>Dillenia suffruticosa root was extracted by sequential solvent extraction. The anti-proliferative activity of DCM-DS was determined by using MTT ...[more]

PMID: 24947113

Similar Datasets

Project description:BackgroundIndigofera suffruticosa Mill. is used as a folk medicine for treating patients with leukemia, however very little is known regarding the molecular mechanism of its anti-leukemic activity and the chemical profile of the active extract. The present study aimed to reveal the molecular effect of I. suffruticosa aerial parts extract (ISAE) on leukemia cells and its chemical constituents.MethodsCytotoxicity of ISAE were determined by resazurin viability assay, multitox - Glo multiplex cytotoxicity assay, and Annexin V staining assay. Cell cycle profiles were revealed by propidium iodide staining assay. The effects of ISAE on G2/M arrest signaling and DNA damage were evaluated by Western blot assay and phospho-H2A.X staining assay. The chemical profile of ISAE were determined by tandem mass spectroscopy and molecular networking approach.ResultsWe showed that the acute lymphoblastic leukemia cell line Jurkat cell was more responsive to ISAE treatment than other leukemia cell lines. In contrast, ISAE did not induce cytotoxic effects in normal fibroblast cells. Cell cycle analysis revealed that ISAE triggered G2/M arrest in Jurkat cells in dose- and time-dependent manners. Elevation of annexin V-stained cells and caspase 3/7 activity suggested ISAE-induced apoptosis. Furthermore, ISAE alone could increase the phosphorylation of CDK1 at Y15 and activate the ATR/CHK1/Wee1/CDC25C signaling pathway. However, the addition of caffeine, a widely used ATR inhibitor to ISAE, reduced the phosphorylation of ATR, CHK1, and CDK1, as well as G2/M arrest in Jurkat cells. Moreover, increased phospho-H2A.X stained cells indicated the involvement of DNA damage in the anti-leukemic effect of ISAE. Finally, qualitative analysis using UPLC-tandem mass spectroscopy and molecular networking revealed that tryptanthrin was the most abundant organoheterocyclic metabolite in ISAE. At equivalent concentrations to ISAE, tryptanthrin induced G2/M arrest of Jurkat cells, which can be prevented by caffeine.ConclusionsISAE causes G2/M arrest via activating ATR/CHK1/CDK1 pathway and tryptanthrin is one of the active components of ISAE. Our findings provide subtle support to the traditional use of I. suffruitcosa in leukemia management in folk medicine.

Project description:CD95 is best known for its ability to induce apoptosis via a well-characterized pathway involving caspase-mediated proteolytic events. However, in apoptosis-resistant cell lines of diverse cancer types stimulation of CD95 primarily has pro-tumorigenic effects that affect many of the hallmarks of cancer. For instance, in colon cancer cells with a mutant KRAS gene CD95 primarily promotes invasion and metastasis. In the current study, we further investigated the context dependency of the consequences of CD95 activation in colon cancer. We used a series of patient-derived three-dimensional colon cancer cultures and studied their response to stimulation with CD95 ligand (CD95L). CD95L had a strong inhibitory effect on the clone-forming capacity of five out of nine cultures. In line with previous work, these cultures all had a wild-type KRAS gene and expressed high levels of CD95. Furthermore, the most sensitive cultures were characterized by microsatellite instability (MSI) and deficient mismatch repair. The reduced clonogenic growth of MSI-type colonospheres resulting from chronic CD95 stimulation was only partly due to apoptosis as many tumor cells survived treatment, yet were unable to regenerate clones. CD95 stimulation caused an irreversible cell cycle arrest, which was associated with cytokine secretion, similar to the senescence-associated secretory phenotype (SASP), and expression of senescence-associated ?-galactosidase. In human colon cancer cohorts, CD95 expression was strongly correlated with the recently identified consensus molecular subtype 1 (CMS1), which mainly consists of MSI-high tumors, and with two independent SASP signatures. Mechanistically, CD95-induced senescence was caused by chronic DNA damage via caspase-activated DNAse resulting in p53 activation and p21 expression, with a minor contribution of the SASP. We conclude that induction of senescence is a hitherto unrecognized consequence of high CD95 expression, which appears to be most relevant for CMS1.

Dataset Information

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

Publications

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

Similar Datasets

OmicsDI is part of the ELIXIR infrastructure

Tweets