stilbenes and Adenocarcinoma-of-Lung

Although it is well documented that endoplasmic reticulum (ER) stress and mitochondrial dysfunction are associated with apoptosis, little is known about whether they are involved in the apoptotic cell death induced by resveratrol and arsenic trioxide (ATO) combination. In this study, we identified a series of sensitization effects of resveratrol on human lung adenocarcinoma A549 cells to ATO treatment, with the combination index (CI) of resveratrol and ATO less than 1. Then, we demonstrated that ER stress was contributed to this synergistic effect, which was manifested by increased the expression levels of ER stress hallmarks, including 78-kDa glucose-regulated protein (GRP 78), caspase 12 and C/EBP-homologous protein (CHOP), In addition, mitochondrial dysfunction was observed after exposure of A549 cells to resveratrol or/and ATO, which was displayed by some alterations of mitochondria-related events, such as loss of mitochondrial membrane potential, cytochrome c release and changes of Bax and Bcl-2 expressions. Our results further demonstrated that resveratrol and ATO-induced ER stress and mitochondrial dysfunction were mediated by reactive oxygen species (ROS), showing that pre-treatment of N-acetyl-l-cysteine, a potent ROS scavenger, restored the ER stress and mitochondrial dysfunction in cells co-treated with resveratrol and ATO, thereby leading to the reduction of the apoptosis. Collectively, these results clearly suggest that ROS-mediated ER stress and mitochondrial dysfunction were involved in the apoptosis induced by resveratrol and ATO in A549 cells, which provides a novel insight into the molecular mechanisms of resveratrol-mediated ATO-sensitization.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Antineoplastic Agents; Apoptosis; Arsenic Trioxide; Arsenicals; Cell Line, Tumor; Drug Synergism; Endoplasmic Reticulum Stress; Humans; Lung; Lung Neoplasms; Mitochondria; Oxides; Reactive Oxygen Species; Resveratrol; Stilbenes

To explore the synergistically effects and mechanisms of resveratrol (RES) enhanced the oxidative stress and apoptotic cell death induced by As2O3 (arsenic trioxide).. According to the result of MTT assay, human lung adenocarcinoma A549 cells were divided into four treatment groups as follow: control group, single RES or As2O3 treated group and the group treated with RES and As2O3. Then the differences of cell viability, colony formation, level of ROS, GSH content, mitochondrial membrane potential and apoptosis rate were compared with single or combined treatment. In addition, pre-treatment with L-buthionine sulfoximine (L-BSO), the inhibitory of GSH synthesis, was used to identify the role of GSH in synergistically apoptosis induced by RES and As2O3.. The detected results demonstrated that RES could effectively inhibited the growth of A549 cells when its concentration above 20 μmol/L, and inhibition effects was concentration dependent manner. The rate of colony formation, GSH content, mitochondrial membrane potential and apoptosis rate in combination group were significantly lower than that of single RES or As2O3 treatment group (P < 0.05), whereas, RES markedly increased the level of ROS, the expression of cytochrome c and caspase 3 induced by As2O3. When pre-treatment with BSO before RES and As2O3 combination incubation, beside the apoptosis rate was increased from 30.0% to 77.7%, the GSH content was sharply depleted while ROS massively accumulated in intracellular.. RES could significantly intensified the effects of As2O3 in inhibiting the proliferation, depleting GSH content, ROS accumulation, mitochondrial membrane potential decline and cytochrome c releasing, thus leading to cells apoptosis via Cas-3 activation in a mitochondria-dependent pathway.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Anticarcinogenic Agents; Antineoplastic Agents; Antineoplastic Combined Chemotherapy Protocols; Apoptosis; Arsenic Trioxide; Arsenicals; Buthionine Sulfoximine; Caspase 3; Drug Synergism; Glutathione; Humans; Lung Neoplasms; Membrane Potential, Mitochondrial; Mitochondria; Oxidation-Reduction; Oxides; Resveratrol; Stilbenes; Tumor Cells, Cultured

Resveratrol has been shown to be a potential chemopreventive and anticancer agent, inducing apoptosis in a variety of cancer cells. The present study was performed to evaluate the effect of resveratrol on A549 human lung adenocarcinoma epithelial cells. 3-(4,5-Dimethylthiazol-2-Yl)-2,5-diphenyltetrazolium bromide evaluation demonstrated that the exposure of cells to increasing concentrations of resveratrol (0-175 µM) for 24 h resulted in a decrease in cell viability (IC50 85.5 µM). Annexin V/propidium iodide double stain verified apoptosis in A549 cells, while negligible cell cytotoxity (≥ 0.5 %) was observed in all untreated incubations. Using colorimetric assay kits, induction of caspase-3, but not of caspase-8, activity was detected in response to resveratrol (> 130 µM). Confirmatory evidence of this finding was provided by Western blotting, indicating expression of cleaved caspase-3 levels in a concentration-dependent manner with a minimum resveratrol concentration of 65 µM required for activation of this protease, while that of caspase-8 remained unaffected. The apoptotic process was associated with reactive oxygen species production in a concentration-dependent manner, evidenced by microscopic examination and fluorescence-activated cell sorting analysis using the 2',7'-dichlorofluorescein diacetate assay. In the presence of the mitochondrial electron transport chain inhibitor rotenone, reactive oxygen species production and the concomitant apoptotic cell population were significantly reduced. This finding suggested that the resveratrol-induced apoptosis was mediated via a mitochondrial pathway alignment in human A549 cells. Although effective levels were observed at high concentrations, the outcome may well differ under in vivo conditions. Finally, experiments reaffirmed the chemical instability of trans-resveratrol, suggesting the need for protection of the solutions from extended exposure to light.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Anti-Inflammatory Agents, Non-Steroidal; Apoptosis; Caspase 3; Caspase 8; Cell Line, Tumor; Cell Survival; Epithelial Cells; Fluoresceins; Humans; Lung Neoplasms; Mitochondria; Reactive Oxygen Species; Resveratrol; Stilbenes; Tetrazolium Salts; Thiazoles

Lung adenocarcinoma is the most common subtype of non-small cell lung cancer and the leading cause of cancer death worldwide. In this study, we investigated the effect of resveratrol (Res) on lung adenocarcinoma A549 cells and its potential mechanism. We found after Res treatment, the interspace of A549 cells decreased and granular material increased in the cell nucleus. These changes were remarkably correlated with the increased concentration of Res. Res induces apoptosis in A549 cells and inhibits cell proliferation in a dose-dependent manner. We further showed that after Res treatment, expression of p53, Bax, and cleaved caspase-3 protein was dramatically up-regulated, while expression of Bcl-2 and the ratio of Bcl-2/Bax were down-regulated. Our study demonstrates that Res inhibits proliferation and induces apoptosis of A549 cells through regulation of p53, Bax, Bcl-2, and cleaved caspase-3 expression.

Topics: A549 Cells; Adenocarcinoma; Adenocarcinoma of Lung; Antineoplastic Agents, Phytogenic; Apoptosis; bcl-2-Associated X Protein; Blotting, Western; Caspase 3; Cell Proliferation; Humans; Lung Neoplasms; Proto-Oncogene Proteins c-bcl-2; Resveratrol; Stilbenes; Tumor Suppressor Protein p53

Although pterostilbene (PTE) has been shown to have potent antitumor activities against various cancer types, the molecular mechanisms of these activities remain unclear. In this study, we investigated the antitumor activity of PTE against human lung adenocarcinoma in vitro and in vivo and explored the role of the Notch1 signaling pathway in this process. PTE treatment resulted in a dose- and time-dependent decrease in the viability of A549 cells. Additionally, PTE exhibited strong antitumor activity, as evidenced not only by a reduced mitochondrial membrane potential (MMP) and a decreased intracellular glutathione content but also by increases in the apoptotic index and the level of reactive oxygen species (ROS). Furthermore, PTE treatment induced the activation of the Notch1 Intracellular Domain (NICD) protein and activated Hes1. DAPT (a gamma secretase inhibitor) and Notch1 siRNA prevented the induction of NICD and Hes1 activation by PTE treatment and sensitized the cells to PTE treatment. The down-regulation of Notch signaling also prevented the activation of pro-survival pathways (most notably the PI3K/Akt pathway) after PTE treatment. In summary, lung adenocarcinoma cells may enhance Notch1 activation as a protective mechanism in response to PTE treatment. Combining a gamma secretase inhibitor with PTE treatment may represent a novel approach for treating lung adenocarcinoma by inhibiting the survival pathways of cancer cells.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Animals; Antineoplastic Agents; Apoptosis; Cell Line, Tumor; Cell Shape; Cell Survival; Dipeptides; Drug Resistance, Neoplasm; Drug Synergism; Glutathione; Humans; Lung Neoplasms; Male; Membrane Potential, Mitochondrial; Mice; Mice, Nude; Proto-Oncogene Proteins c-akt; Reactive Oxygen Species; Receptor, Notch1; Signal Transduction; Stilbenes; Tumor Burden; Xenograft Model Antitumor Assays

The potential benefits of resveratrol as an anticancer (proapoptosis) and antioxidant (pro-survival) compound have been studied extensively. However, the role of resveratrol in modulation of the toxicity induced by sodium arsenite (NaAsO₂) is still unclear. In the present study, we examined the effects of resveratrol on NaAsO₂-induced cytotoxicity, DNA and chromosomal damage, cell cycle progression, apoptosis and oxidative stress in human lung adenocarcinoma epithelial (A549) cell line at concentrations from 1 to 20 μM after 24h exposure. Our results revealed that at 1 and 5 μM, resveratrol was found to exert benefit effects, promoting cell viability and proliferation over 24h NaAsO₂ exposure, whereas, resveratrol was showed to inhibit cell survival under the same condition at 20 μM. Corresponding to the opposing effect of resveratrol at low vs. high concentrations, DNA and chromosomal damage, cell apoptotic rate and level of oxidative stress were also alleviated by lower concentrations (1, 5 μM) of resveratrol, but exacerbated by higher concentration (20 μM) resveratrol. Our study implicates that resveratrol is the most beneficial to cells at 1 and 5 μM and caution should be taken in applying resveratrol as an anticancer therapeutic agent or nutraceutical supplement due to its concentration dependent effect.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Antineoplastic Agents, Phytogenic; Antioxidants; Apoptosis; Arsenites; Cell Line, Tumor; Cell Proliferation; Cell Survival; Chromosome Breakage; Comet Assay; DNA Damage; Glutathione; Humans; Lung Neoplasms; Micronuclei, Chromosome-Defective; Mutagens; Neoplastic Stem Cells; Osmolar Concentration; Oxidation-Reduction; Oxidative Stress; Resveratrol; Sodium Compounds; Stilbenes

Based on our recent findings that resveratrol, a natural plant polyphenol found in red grape skins as well as other food products, induces apoptosis via a caspase-independent intrinsic pathway in human lung adenocarcinoma cells, this study is designed to explore whether SB203580, a p38 inhibitor, potentiates the resveratrol-induced apoptosis of human lung adenocarcinoma (A549) cells. We found that pretreatment with SB203580 enhanced the resveratrol-induced apoptosis by accelerating the intrinsic apoptotic pathway including Bax activation, loss of mitochondrial membrane potential, and activation of both caspase-9 and -3. Although treatment with resveratrol alone did not induce caspase-8 activation, cotreatment with both SB203580 and resveratrol not only enhanced FasL cleavage but also activated caspase-8, indicating that the extrinsic apoptotic pathway may be involved in the synergistic effect. Collectively, we for the first time demonstrate that SB203580 synergistically enhances the resveratrol-induced apoptosis by accelerating Bax-mediated intrinsic pathway and initiating extrinsic pathway, suggesting a possible alternative therapeutic strategy for human lung cancer.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Antineoplastic Agents, Phytogenic; Apoptosis; Caspase 3; Caspase 8; Caspase 9; Cell Line, Tumor; Cell Survival; Drug Synergism; Enzyme Inhibitors; Gene Expression Regulation, Neoplastic; Humans; Imidazoles; Lung Neoplasms; Membrane Potential, Mitochondrial; p38 Mitogen-Activated Protein Kinases; Pyridines; Resveratrol; Stilbenes

Resveratrol (RV), a natural plant polyphenol widely present in foods such as grapes, wine, and peanuts, has an ability to inhibit various stages of carcinogenesis in vitro and in vivo. In this report, we explored the roles of intrinsic and extrinsic apoptotic pathways during RV-induced apoptosis in human lung adenocarcinoma (ASTC-a-1) cells. After exposure of cells to different concentrations of RV, we found that RV induced concentration-dependent apoptosis. Fluorometric substrates assay and western blotting (WB) analysis showed that caspase-8 was not activated, which was further verified by monitoring the cleavage of Bid to tBid using fluorescence resonance energy transfer (FRET) microscopy imaging inside single living cells, indicating that extrinsic apoptotic pathway was not involved in RV-induced apoptosis. In addition, inhibition of caspases-3 or -9 but not caspase-8 using the specific inhibitors of caspases modestly but significantly attenuated RV-induced apoptosis. Moreover, flow cytometry (FCM) analysis showed that RV treatment induced time-dependent loss of mitochondrial membrane potential (∆ψ(m)), in combination with the activation of caspases-3 and -9; we therefore concluded that RV-induced apoptosis involved the intrinsic apoptotic pathway. It is noteworthy that RV treatment induced translocation of AIF from mitochondria to nucleus in a time dependent manner, and that knockdown of AIF remarkably attenuated RV-induced apoptosis. Collectively, our findings demonstrate that RV induces caspase-8-independent apoptosis via AIF and to a lesser extent caspase-9-dependent mitochondrial pathway in ASTC-a-1 cells.

Topics: Adenocarcinoma; Adenocarcinoma of Lung; Apoptosis; Apoptosis Inducing Factor; Caspase 3; Caspase 8; Caspase 9; Cell Line, Tumor; Cell Nucleus; Cell Survival; Enzyme Activation; Humans; Lung Neoplasms; Membrane Potential, Mitochondrial; Mitochondria; Protein Transport; Resveratrol; RNA Interference; Stilbenes