Sample records for a549 cell apoptosis

  1. [Study on thaspine in inducing apoptosis of A549 cell].

    PubMed

    Zhang, Yan-min; He, Lang-chong

    2007-04-01

    To investigate the effect of thaspine on the cellular proliferation, apoptosis and cell cycle in A549 cell line. A549 cell was cultured with different concentrations of thaspine. Cellular proliferation was detected with MTT, apoptosis and cell cycle were checked with Flow Cytometer, and change of microstructure was observed by transmission electron microscope. Thaspine could inhibit the proliferation and induce apoptosis of A549 cell in a time-dose dependent manner. Cell cycle was significantly stopped at the S phase by thaspine with FCM technology. Under electronic microscope, the morphology of A549 cell showed nuclear karyopycnosis, chromatin agglutination and typical apoptotic body when the cell was treated with thaspine. Thaspine has the effects of anti-tumor and inducing apoptosis.

  2. Artesunate induces AIF-dependent apoptosis in A549 cells

    NASA Astrophysics Data System (ADS)

    Zhou, Chen-juan; Chen, Tong-Sheng

    2012-03-01

    Artesunate (ART), a semi-synthetic derivative of the sesquiterpene artemisinin extracted from the Chinese herb Artemisia annua, exerts a broad spectrum of clinical activity against human cancers. It has been shown that ART induces cancer cells death through apoptosis pathway. This study investigated whether ART treatment induced reactive oxygen species (ROS)-dependent cell death in the apoptosis fashion in human lung adenocarconoma A549 cell line and the proapoptotic protein apoptosis inducing factor (AIF) is involved in ART-induced apoptosis. Cells treated with ART exhibited typical apoptotic morphology as chromatin condensation, margination and shrunken nucleus. ART treatment also induced a loss of mitochondrial membrane potential and AIF release from mitochondria. Silencing AIF can remarkable attenuated ART-induced apoptosis. Collectively, ART induces apoptosis by caspase-independent intrinsic pathway in A549 cells.

  3. [Apoptosis inducing effect of Hechanpian on human lung adenocarcinoma A549 cells].

    PubMed

    Xiong, Shao-Quan; Zhou, Dai-Han; Lin, Li-Zhu

    2010-06-01

    To study the apoptosis inducing effects of Hechanpian (HCP) on human lung adenocarcinoma A549 cells. HCP containing rat serum was prepared and applied on A549 cells. The cell growth inhibition rate was tested by MTT assay; the effect of HCP on cell apoptosis was observed with Propidium iodide (PI) staining and flow cytometry analysis; the mRNA expression of epidermal growth factor receptor (EGFR) was detected through RT-PCR. The growth of A549 cells was obviously inhibited after being treated by HCP containing serum, and the cells presented an apoptotic change. The cell apoptosis rate after treated by serum containing 10% and 20% HCP was 20.5% and 33.2%, respectively, significantly higher than that in the control (6.1% in cells didn't treated with HCP, P < 0.05). Compared with control, EGFR mRNA expression in HCP treated cells was significantly lower (P < 0.05). HCP has apoptosis inducing effect on A549 cell, and its molecular mechanism is probably correlated with the inhibition of EGFR gene transcription.

  4. Ghrelin ameliorates the human alveolar epithelial A549 cell apoptosis induced by lipopolysaccharide

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Huang, Chunrong; Zheng, Haichong; He, Wanmei

    Ghrelin is a gastric acyl-peptide that plays an inhibitory role in cell apoptosis. Herein we investigate the protective effects of ghrelin in LPS-induced apoptosis of human alveolar epithelial A549 cells, along with the possible molecular mechanisms. LPS exposure impaired cell viability and increased apoptosis of A549 cells significantly in concentration- and time-dependent manners embodied in increased Bax and cleaved caspase-3 production, coupled with decreased Bcl-2 levels. Simultaneously, LPS remarkably decreased the expression of phosphatidylinositol 3 kinase/protein kinase B (PI3K/Akt) and extracellular signal-regulated kinas (ERK) in A549 cells. However, ghrelin'pretreatment ameliorated LPS-caused alterations in the ratio of Bax/Bcl-2 and cleaved caspase-3 expression, whereas activatedmore » the PI3K/Akt and ERK signaling. These results demonstrate that ghrelin lightens LPS-induced apoptosis of human alveolar epithelial cells partly through activating the PI3K/Akt and ERK pathway and thereby might benefit alleviating septic ALI. -- Graphical abstract: Ghrelin ameliorates the human alveolar epithelial A549 cells apoptosis induced by lipopolysaccharide partly through activating the PI3K/Akt and ERK pathway. Display Omitted -- Highlights: •It has been observed that LPS insult significantly increased apoptosis in A549 cells. •Both Akt and ERK signaling are critical adapter molecules to mediate the ghrelin-mediated proliferative effect. •Ghrelin may have a therapeutic effect in the prevention of LPS-induced apoptosis.« less

  5. Induction of ER Stress-Mediated Apoptosis by α-Lipoic Acid in A549 Cell Lines

    PubMed Central

    Kim, Jong In; Lee, Chang Min; Park, Eok-Sung; Kim, Ki Nyun; Kim, Hyung Chul; Lee, Hae Young

    2012-01-01

    Background α-Lipoic acid (α-LA) has been studied as an anticancer agent as well as a therapeutic agent for diabetes and obesity. We performed this study to evaluate the anticancer effects and mechanisms of α-LA in a lung cancer cell line, A549. Materials and Methods α-LA-induced apoptosis of A549 cells was detected by fluorescence-activated cell sorting analysis and a DNA fragmentation assay. Expression of apoptosis-related genes was analyzed by western blot and reverse transcription-polymerase chain reaction analyses. Results α-LA induced apoptosis and DNA fragmentation in A549 cells in a dose- and time-dependent manner. α-LA increased caspase activity and the degradation of poly (ADP-ribose) polymerase. It induced expression of endoplasmic reticulum (ER) stress-related genes, such as glucose-regulated protein 78, C/EBP-homologous protein, and the short form of X-box binding protein-1, and decreased expression of the anti-apoptotic protein, X-linked inhibitor of apoptosis protein. Reactive oxygen species (ROS) production was induced by α-LA, and the antioxidant N-acetyl-L-cysteine decreased the α-LA-induced increase in expression of apoptosis and ER stress-related proteins. Conclusion α-LA induced ER stress-mediated apoptosis in A549 cells via ROS. α-LA may therefore be clinically useful for treating lung cancer. PMID:22363901

  6. Seleno-short-chain chitosan induces apoptosis in human non-small-cell lung cancer A549 cells through ROS-mediated mitochondrial pathway.

    PubMed

    Zhao, Yana; Zhang, Shaojing; Wang, Pengfei; Fu, Shengnan; Wu, Di; Liu, Anjun

    2017-12-01

    Seleno-short-chain chitosan (SSCC) is a synthesized chitosan derivative. In this study, antitumor activity and underlying mechanism of SSCC on human non-small-cell lung cancer A549 cells were investigated in vitro. The MTT assay showed that SSCC could inhibit cell viability in a dose- and time-dependent manner, and 200 μg/ml SSCC exhibited significantly toxic effects on A549 cells. The cell cycle assay showed that SSCC triggered S phase cell cycle arrest in a dose- and time-dependent manner, which was related to a downregulation of S phase associated cyclin A. The DAPI staining and Annexin V-FITC/PI double staining identified that the SSCC could induce A549 cells apoptosis. Further studies found that SSCC led to the generation of reactive oxygen species (ROS) and the disruption of mitochondrial membrane potential (MMP) by DCFH-DA and Rhodamin 123 staining, respectively. Meanwhile, free radical scavengers N-acetyl-L-cysteine (NAC) pretreatment confirmed that SSCC-induced A549 cells apoptosis was associated with ROS generation. Furthermore, real-time PCR and western blot assay showed that SSCC up-regulated Bax and down-regulated Bcl-2, subsequently incited the release of cytochrome c from mitochondria to cytoplasm, activated the increase of cleaved-caspase 3 and finally induced A549 cells apoptosis in vitro. In general, the present study demonstrated that SSCC induced A549 cells apoptosis via ROS-mediated mitochondrial apoptosis pathway.

  7. Curcumin promotes apoptosis in A549/DDP multidrug-resistant human lung adenocarcinoma cells through an miRNA signaling pathway

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zhang, Jian, E-mail: zhangjian197011@yahoo.com; Zhang, Tao; Ti, Xinyu

    2010-08-13

    Research highlights: {yields} Curcumin had anti-cancer effects on A549/DDP multidrug-resistant human lung adenocarcinoma cells {yields} Curcumin promotes apoptosis in A549/DDP cells through a miRNA signaling pathway {yields} Curcumin induces A549/DDP cell apoptosis by downregulating miR-186* {yields} miR-186* may serve as a potential gene therapy target for refractory lung cancer that is sensitive to curcumin -- Abstract: Curcumin extracted from the rhizomes of Curcuma longa L. has been shown to have inhibitory effects on cancers through its anti-proliferative and pro-apoptotic activities. Emerging evidence demonstrates that curcumin can overcome drug resistance to classical chemotherapies. Thus, the mechanisms underlying the anti-tumor activities ofmore » curcumin require further study. In our study, we first demonstrated that curcumin had anti-cancer effects on A549/DDP multidrug-resistant human lung adenocarcinoma cells. Further studies showed that curcumin altered miRNA expression; in particular, significantly downregulated the expression of miR-186* in A549/DDP. In addition, transfection of cells with a miR-186* inhibitor promoted A549/DDP apoptosis, and overexpression of miR-186* significantly inhibited curcumin-induced apoptosis in A549/DDP cells. These observations suggest that miR-186* may serve as a potential gene therapy target for refractory lung cancer that is sensitive to curcumin.« less

  8. β-elemene reverses the drug resistance of lung cancer A549/DDP cells via the mitochondrial apoptosis pathway.

    PubMed

    Yao, Cheng-Cai; Tu, Yuan-Rong; Jiang, Jie; Ye, Sheng-Fang; Du, Hao-Xin; Zhang, Yi

    2014-05-01

    β-elemene (β-ELE) is a new anticancer drug extracted from Curcuma zedoaria Roscoe and has been widely used to treat malignant tumors. Recent studies have demonstrated that β-ELE reverses the drug resistance of tumor cells. To explore the possible mechanisms of action of β-ELE, we investigated its effects on cisplatin-resistant human lung adenocarcinoma A549/DDP cells. The effects of β-ELE on the growth of A549/DDP cells in vitro were determined by MTT assay. Apoptosis was assessed by fluorescence microscopy with Hoechst 33258 staining and flow cytometry with Annexin V-FITC/PI double staining. Mitochondrial membrane potential was assessed using JC-1 fluorescence probe and laser confocal scanning microscopy, and intracellular reactive oxygen species levels were measured by 2',7'-dichlorofluorescein-diacetate staining and flow cytometry. Cytosolic glutathione content was determined using GSH kits. The expression of cytochrome c, caspase-3, procaspase-3 and the Bcl-2 family proteins was assessed by western blotting. The results demonstrated that β-ELE inhibited the proliferation of A549/DDP cells in a time- and dose-dependent manner. Furthermore, β-ELE enhanced the sensitivity of A549/DDP cells to cisplatin and reversed the drug resistance of A549/DDP cells. Consistent with a role in activating apoptosis, β-ELE decreased mitochondrial membrane potential, increased intracellular reactive oxygen species concentration and decreased the cytoplasmic glutathione levels in a time- and dose-dependent manner. The combination of β-ELE and cisplatin enhanced the protein expression of cytochrome c, caspase-3 and Bad, and reduced protein levels of Bcl-2 and procaspase-3 in the A549/DDP lung cancer cells. These results define a pathway of procaspase‑3-β-ELE function that involves decreased mitochondrial membrane potential, leading to apoptosis triggered by the release of cytochrome c into the cytoplasm and the modulation of apoptosis-related genes. The reversal of drug

  9. Novel synthetic chalcones induce apoptosis in the A549 non-small cell lung cancer cells harboring a KRAS mutation.

    PubMed

    Wang, Yiqiang; Hedblom, Andreas; Koerner, Steffi K; Li, Mailin; Jernigan, Finith E; Wegiel, Barbara; Sun, Lijun

    2016-12-01

    A series of novel chalcones were synthesized by the Claisen-Schmidt condensation reaction of tetralones and 5-/6-indolecarboxaldehydes. Treatment of human lung cancer cell line harboring KRAS mutation (A549) with the chalcones induced dose-dependent apoptosis. Cell cycle analyses and Western blotting suggested the critical role of the chalcones in interrupting G2/M transition of cell cycle. SAR study demonstrated that substituent on the indole N atom significantly affects the anticancer activity of the chalcones, with methyl and ethyl providing the more active compounds (EC 50 : 110-200nM), Compound 1g was found to be >4-fold more active in the A549 cells (EC 50 : 110nM) than in prostate (PC3) or pancreatic cancer (CLR2119, PAN02) cells. Furthermore, compound 1l selectively induced apoptosis of lung cancer cells A549 (EC 50 : 0.55μM) but did not show measurable toxicity in the normal lung bronchial epithelial cells (hBEC) at doses as high as 10μM, indicating specificity towards cancer cells. Copyright © 2016 Elsevier Ltd. All rights reserved.

  10. Group B Streptococcus serotypes III and V induce apoptosis and necrosis of human epithelial A549 cells.

    PubMed

    Da Costa, Andréia Ferreira Eduardo; Pereira, Camila Serva; Santos, Gabriela Da Silva; Carvalho, Técia Maria Ulisses; Hirata, Raphael; De Mattos-Guaraldi, Ana Luiza; Rosa, Ana Cláudia De Paula; Nagao, Prescilla Emy

    2011-05-01

    Although group B Streptococcus (GBS) has been classically described as an exclusively extracellular pathogen, growing evidence suggests that it may be internalized by epithelial cells. However, the fates of intracellular GBS and of infected respiratory epithelial cells remain unclear. Little is known about the bacterial components involved in these processes. The present study investigated the bacterial internalization by A549 cells and the apoptosis/necrosis of the infected human epithelial cells. The morphological changes in A549 cells observed from 2 h post-infection with GBS included vacuolization and the formation of apoptotic bodies. Flow cytometry revealed that 81.2% of apoptotic A549 cells were infected with GBS serotype III 90356-liquor. Moreover, a double-staining assay using propidium iodide (PI)/Annexin V (AV) gave information about the numbers of viable (PI-/AV-) (18.27%) vs. early apoptotic (PI-/AV+) (73.83%) and late apoptotic cells (PI+/AV+) (7.37%) during infection of A549 cells with GBS III 90356-liquor. In addition, 37% necrotic cells were observed in A549 cells infected with GBS serotype V 90186-blood. In conclusion, GBS serotypes III and V induce apoptosis of epithelial cells in the early stages of GBS infection, resulting in tissue destruction, bacterial spreading and, in consequence, invasive disease or systemic infection.

  11. Induction of apoptosis in non-small cell lung carcinoma A549 cells by PGD₂ metabolite, 15d-PGJ₂.

    PubMed

    Wang, Jun-Jie; Mak, Oi-Tong

    2011-11-01

    PGD2 (prostaglandin D2) is a mediator in various pathophysiological processes, including inflammation and tumorigenesis. PGD2 can be converted into active metabolites and is known to activate two distinct receptors, DP (PGD2 receptor) and CRTH2/DP2 (chemoattractant receptor-homologous molecule expressed on Th2 cells). In the past, PGD2 was thought to be involved principally in the process of inflammation. However, in recent years, several studies have shown that PGD2 has anti-proliferative ability against tumorigenesis and can induce cellular apoptosis via activation of the caspase-dependent pathway in human colorectal cancer cells, leukaemia cells and eosinophils. In the lung, where PGD2 is highly released when sensitized mast cells are challenged with allergen, the mechanism of PGD2-induced apoptosis is unclear. In the present study, A549 cells, a type of NSCLC (non-small cell lung carcinoma), were treated with PGD2 under various conditions, including while blocking DP and CRTH2/DP2 with the selective antagonists BWA868C and ramatroban respectively. We report here that PGD2 induces A549 cell death through the intrinsic apoptotic pathway, although the process does not appear to involve either DP or CRTH2/DP2. Similar results were also found with H2199 cells, another type of NSCLC. We found that PGD2 metabolites induce apoptosis effectively and that 15d-PGJ2 (15-deoxy-Δ12,14-prostaglandin J2) is a likely candidate for the principal apoptotic inducer in PGD2-induced apoptosis in NSCLC A549 cells.

  12. [HDAC1 expression and effect of TSA on proliferation and apoptosis of A549 cells].

    PubMed

    Huang, Hong; Zhang, Zhen-Xiang; Xu, Yong-Jian; Shao, Jing-Fang

    2003-09-01

    Histone deacetylase (HDAC) shows a high expression in many cancer cells and the inhibitor of HDAC1, trichostatin A (TSA), can inhibit the growth of cancer cells. Hypoxia is a common feature of malignant tumors. This paper was designed to investigate the expression of HDAC1 of A549 cell strains in hypoxia condition and the effect of TSA on their proliferation and apoptosis. The authors designed 1 normoxia group (control group) and 5 hypoxia groups (test groups): hypoxia 6h group (A), TSA + hypoxia 6h (B), hypoxia 12h group (C), hypoxia 24h group (D), TSA + hypoxia 24h (E), hypoxia 48h group (F). The expression of HDAC1 in A549 cells was examined using Western blot analysis. Proliferation, the apoptotic rates of A549 cells and the effect of TSA on them were determined using MTT method, immunohistochemistry, TUNEL method, and flow cytometry. The expression of mRNA of HDAC1 and the effect of TSA on it were determined using reverse transcription-polymerase chain reaction (RT-PCR). The A values expressed by HDAC1 in A549 cell strains were 138+/-11 in the control group, 78+/-4, 86+/-5, 124+/-3, and 120+/-9 in test groups A, C, D, and F, respectively. The A values of HDAC1mRNA versus the A values of beta-Atin mRNA were 0.68+/-0.03 in the control group, 0.46+/-0.03, 0.45+/-0.02, 0.70+/-0.03, and 0.33+/-0.02 in test groups A, C, D, and F, respectively. The A values of the expression of PCNA in A549 cell strains were 0.13+/-0.03 in the control group, 0.10+/-0.02, 0.11+/-0.02, 0.16+/-0.02, and 0.11+/-0.03 in test groups A, B, D, and E, respectively. The A values of MTT in A549 cell strains were 0.50+/-0.06 in the control group, 0.41+/-0.04, 0.45+/-0.03, 0.59+/-0.02, and 0.45+/-0.03 in test groups A, B, D, and E, respectively. The A values of positive cells of apoptosis in A549 cell strains were 0.16+/-0.04 in the control group, 0.18+/-0.02, 0.18+/-0.05, 0.20+/-0.05, and 0.23+/-0.05 in test groups A, B, D, and E, respectively. The apoptotic rates in A549 cells were 1.11% in the

  13. Mechanisms underlying regulation of cell cycle and apoptosis by hnRNP B1 in human lung adenocarcinoma A549 cells.

    PubMed

    Han, Juan; Tang, Feng-ming; Pu, Dan; Xu, Dan; Wang, Tao; Li, Weimin

    2014-01-01

    Overexpression of heterogeneous nuclear ribonucleoprotein B1 (hnRNP B1), a nuclear RNA binding protein, has been reported to occur in early-stage lung cancer and in premalignant lesions. DNA-dependent protein kinase (DNA-PK) is known to be involved in the repair of double-strand DNA breaks. Reduced capacity to repair DNA has been associated with the risk of lung cancer. We investigated a link between hnRNP B1 and DNA-PK and their effects on proliferation, cell cycle, and apoptosis in the human lung adenocarcinoma cell line A549. We found that hnRNP B1 and DNA-PK interact with each other in a complex fashion. Reducing hnRNP B1 expression in A549 cells with the use of RNAi led to upregulation of p53 activity through upregulation of DNA-PK activity but without inducing p53 expression. Further, suppression of hnRNP B1 in A549 cells slowed cell proliferation, promoted apoptosis, and induced cell cycle arrest at the G1 stage. The presence of NU7026 reduced the arrest of cells at the G1 stage and reduced the apoptosis rate while promoting cell growth. Taken together, our results demonstrate that by regulating DNA-PK activity, hnRNP B1 can affect p53-mediated cell cycle progression and apoptosis, resulting in greater cell survival and subsequent proliferation.

  14. Effect of Withaferin A on A549 cellular proliferation and apoptosis in non-small cell lung cancer.

    PubMed

    Cai, Yong; Sheng, Zhao-Ying; Chen, Yun; Bai, Chong

    2014-01-01

    To explore the effect of Withaferin A on A549 cellular proliferation and apoptosis in non-small cell lung cancer (NSCLC). NSCNC cell line A549 was selected to explore the effect of Withaferin A on A549 cellular proliferation, apoptosis and the PI3K/Akt signal pathway capable of regulating tumor biological behavior by assessment of cellular proliferation, cellular apoptotic rates and cellular cycling as well as by immuno-blotting. Withaferin A could inhibit A549 cellular proliferation and the control rate was dosage-dependent (P<0.05), which also increased time-dependently with the same dosage of Withaferin A (P<0.05). The apoptotic indexes in A549 cells treated with 0, 2.5, 5.0, 10.0 and 20.0 μmol·L-1 Withaferin A for 48 h were significantly different (P<0.05). In addition, the apoptotic rates of each group in both early and advanced stages were higher than those in 0 μmol·L-1 (P<0.05), which were evidently higher after 48 h than those after 24 h (P<0.05). A549 cells treated by Withaferin A for 48 h were markedly lower in Bcl-2 level and obviously higher in Bax and cleaved caspase-3 levels than those treated by 0 μmol·L-1 Withaferin A (P<0.05), and there were significant differences among 5, 10 and 20 μmol·L-1 Withaferin A (P<0.05). The ratios of A549 cells treated by Withaferin A for 48 h in G0/G1 stage were higher than those in 0 μmol·L-1 , while those in S and G2/M stages were obviously lower than those in G2/M stage, and there were significant differences in 5.0, 10.0 and 20.0 μmol·L-1 Withaferin A (P<0.05). Additionally, p-Akt/Akt values were in reverse association with dosage, and the differences were significant (P<0.05). Withaferin A can inhibit the proliferation and apoptosis of A549 cells by suppressing activation of the PI3K/Akt pathways.

  15. Osthole induces G2/M arrest and apoptosis in lung cancer A549 cells by modulating PI3K/Akt pathway

    PubMed Central

    2011-01-01

    Background To explore the effects of Osthole on the proliferation, cell cycle and apoptosis of human lung cancer A549 cells. Methods Human lung cancer A549 cells were treated with Osthole at different concentrations. Cell proliferation was measured using the MTT assay. Cell cycle was evaluated using DNA flow cytometry analysis. Induction of apoptosis was determined by flow cytometry and fluorescent microscopy. The expressions of Cyclin B1, p-Cdc2, Bcl-2, Bax, t-Akt and p-Akt were evaluated by Western blotting. Results Osthole inhibited the growth of human lung cancer A549 cells by inducing G2/M arrest and apoptosis. Western blotting demonstrated that Osthole down-regulated the expressions of Cyclin B1, p-Cdc2 and Bcl-2 and up-regulated the expressions of Bax in A549 cells. Inhibition of PI3K/Akt signaling pathway was also observed after treating A549 cells with Osthole. Conclusions Our findings suggest that Osthole may have a therapeutic application in the treatment of human lung cancer. PMID:21447176

  16. [Astaxanthin inhibits proliferation and promotes apoptosis of A549 lung cancer cells via blocking JAK1/STAT3 pathway].

    PubMed

    Wu, Chuntao; Zhang, Jinji; Liu, Tienan; Jiao, Guimei; Li, Changzai; Hu, Baoshan

    2016-06-01

    Objective To investigate the anti-tumor effects of astaxanthin on A549 lung cancer cells and the related mechanisms. Methods A549 cells were cultured with various concentrations of astaxanthin (20, 40, 60, 80, 100 μmol/L), and DMSO at the same concentrations served as vehicle controls. The viability of A549 cells was detected by CCK-8 assay; cell cycle and apoptosis were observed by flow cytometry; and the expressions of B-cell lymphoma-2 (Bcl-2), Bcl-2 associated X protein (Bax), signal transducers and activators of transcription 3 (STAT3), and Janus kinase 1 (JAK1) were evaluated by Western blotting. Results CCK-8 assay showed that astaxanthin decreased the proliferation of A549 cells in a dose-dependent manner. Flow cytometry showed that astaxanthin increased the number of cells in the G0/G1 phase and induced apoptosis in A549 cells. Western blotting showed that astaxanthin up-regulated the expression of Bax and down-regulated the expressions of Bcl-2, STAT3 and JAK1. Conclusion Astaxanthin functions as a potent inhibitor of A549 lung cancer cell growth by targeting JAK1/STAT3 signaling pathway.

  17. G4-Tetra DNA Duplex Induce Lung Cancer Cell Apoptosis in A549 Cells

    NASA Astrophysics Data System (ADS)

    Xu, Xiaobo; Zhao, YiZhuo; Lu, Hu; Fu, Cuiping; Li, Xiao; Jiang, Liyan; Li, Shanqun

    2016-10-01

    The specific DNA is typically impermeable to the plasma membrane due to its natural characters, but DNA tetra structures (DTNs) can be readily uptake by cells in the absence of transfection agents, providing a new strategy to deliver DNA drugs. In this research, the delivery efficiency of tetrahedral DNA nanostructures was measured on adenocarcinomic human alveolar basal epithelial (A549) cells via delivering AS1411 (G4). The DNA tetra-AS1411 complex was rapidly and abundantly uptake by A549 cells, and the induced apoptosis was enhanced. Furthermore, biodistribution in mouse proved the rapid clearance from non-targeted organs in vivo. This study improved the understanding of potential function in DNA-based drug delivery and proved that DTNs-AS1411 could be potentially useful for the treatment of lung cancer.

  18. Phloretin induces apoptosis of non-small cell lung carcinoma A549 cells via JNK1/2 and p38 MAPK pathways.

    PubMed

    Min, Jie; Huang, Kenan; Tang, Hua; Ding, Xinyu; Qi, Chen; Qin, Xiong; Xu, Zhifei

    2015-12-01

    Phloretin (Ph) existing in apples, pears and various vegetables is known to have antitumor activities in several cancer cell lines. However, little is known about its effect on human lung cancer cells. The aim of the present study was to see whether Ph could induce apoptosis of non-small cell lung cancer (NSCLC) cells, and explore the possible underlying mechanism of action. We found that Ph markedly induced cell apoptosis of NSCLC cell line A549, and inhibited the migration of A549 cells in a dose-dependent manner. The expression level of BAX, cleaved caspase-3 and -9, and degraded form of PARP was increased and Bcl-2 was decreased after Ph treatment. In addition, the phosphorylation of P38 MAPK, ERK1/2 and JNK1/2 was increased in a dose‑dependent manner in parallel with Ph treatment. Inhibition of P38 MAPK and JNK1/2 by specific inhibitors significantly abolished the Ph-induced activation of the caspase-3 and -9. In vivo tumor-suppression assay further indicated that Ph (20 mg/kg) displayed a more significant inhibitory effect on A549 xenografts in tumor growth. All these findings indicate that Ph is able to inhibit NSCLC A549 cell growth by inducing apoptosis through P38 MAPK and JNK1/2 pathways, and therefore may prove to be an adjuvant to the treatment of NSCLC.

  19. Formoxanthone C, isolated from Cratoxylum formosum ssp. pruniflorum, reverses anticancer drug resistance by inducing both apoptosis and autophagy in human A549 lung cancer cells.

    PubMed

    Kaewpiboon, Chutima; Boonnak, Nawong; Kaowinn, Sirichat; Chung, Young-Hwa

    2018-02-15

    Multidrug resistance (MDR) cancer toward cancer chemotherapy is one of the obstacles in cancer therapy. Therefore, it is of interested to use formoxanthone C (1,3,5,6-tetraoxygenated xanthone; XanX), a natural compound, which showed cytotoxicity against MDR human A549 lung cancer (A549RT-eto). The treatment with XanX induced not only apoptosis- in A549RT-eto cells, but also autophagy-cell death. Inhibition of apoptosis did not block XanX-induced autophagy in A549RT-eto cells. Furthermore, suppression of autophagy by beclin-1 small interfering RNAs (siRNAs) did not interrupt XanX-induced apoptosis, indicating that XanX can separately induce apoptosis and autophagy. Of interest, XanX treatment reduced levels of histone deacetylase 4 (HDAC4) protein overexpressed in A549RT-etocells. The co-treatment with XanX and HDAC4 siRNA accelerated both autophagy and apoptosis more than that by XanX treatment alone, suggesting survival of HDAC4 in A549RT-eto cells. XanX reverses etoposide resistance in A549RT-eto cells by induction of both autophagy and apoptosis, and confers cytotoxicity through down-regulation of HDAC4. Copyright © 2017. Published by Elsevier Ltd.

  20. Phloretin induces apoptosis of non-small cell lung carcinoma A549 cells via JNK1/2 and p38 MAPK pathways

    PubMed Central

    MIN, JIE; LI, XU; HUANG, KENAN; TANG, HUA; DING, XINYU; QI, CHEN; QIN, XIONG; XU, ZHIFEI

    2015-01-01

    Phloretin (Ph) existing in apples, pears and various vegetables is known to have antitumor activities in several cancer cell lines. However, little is known about its effect on human lung cancer cells. The aim of the present study was to see whether Ph could induce apoptosis of non-small cell lung cancer (NSCLC) cells, and explore the possible underlying mechanism of action. We found that Ph markedly induced cell apoptosis of NSCLC cell line A549, and inhibited the migration of A549 cells in a dose-dependent manner. The expression level of BAX, cleaved caspase-3 and -9, and degraded form of PARP was increased and Bcl-2 was decreased after Ph treatment. In addition, the phosphorylation of P38 MAPK, ERK1/2 and JNK1/2 was increased in a dose-dependent manner in parallel with Ph treatment. Inhibition of P38 MAPK and JNK1/2 by specific inhibitors significantly abolished the Ph-induced activation of the caspase-3 and -9. In vivo tumor-suppression assay further indicated that Ph (20 mg/kg) displayed a more significant inhibitory effect on A549 xenografts in tumor growth. All these findings indicate that Ph is able to inhibit NSCLC A549 cell growth by inducing apoptosis through P38 MAPK and JNK1/2 pathways, and therefore may prove to be an adjuvant to the treatment of NSCLC. PMID:26503828

  1. Ionizing Radiation Potentiates Dihydroartemisinin-Induced Apoptosis of A549 Cells via a Caspase-8-Dependent Pathway

    PubMed Central

    Chen, Tongsheng; Chen, Min; Chen, Jingqin

    2013-01-01

    This report is designed to explore the molecular mechanism by which dihydroartemisinin (DHA) and ionizing radiation (IR) induce apoptosis in human lung adenocarcinoma A549 cells. DHA treatment induced a concentration- and time-dependent reactive oxygen species (ROS)-mediated cell death with typical apoptotic characteristics such as breakdown of mitochondrial membrane potential (Δψm), caspases activation, DNA fragmentation and phosphatidylserine (PS) externalization. Inhibition of caspase-8 or -9 significantly blocked DHA-induced decrease of cell viability and activation of caspase-3, suggesting the dominant roles of caspase-8 and -9 in DHA-induced apoptosis. Silencing of proapoptotic protein Bax but not Bak significantly inhibited DHA-induced apoptosis in which Bax but not Bak was activated. In contrast to DHA treatment, low-dose (2 or 4 Gy) IR induced a long-playing generation of ROS. Interestingly, IR treatment for 24 h induced G2/M cell cycle arrest that disappeared at 36 h after treatment. More importantly, IR synergistically potentiated DHA-induced generation of ROS, activation of caspase-8 and -3, irreparable G2/M arrest and apoptosis, but did not enhance DHA-induced loss of Δψm and activation of caspase-9. Taken together, our results strongly demonstrate the remarkable synergistic efficacy of combination treatment with DHA and low-dose IR for A549 cells in which IR potentiates DHA-induced apoptosis largely by enhancing the caspase-8-mediated extrinsic pathway. PMID:23536891

  2. Oxidative stress mediated apoptosis induced by nickel ferrite nanoparticles in cultured A549 cells.

    PubMed

    Ahamed, Maqusood; Akhtar, Mohd Javed; Siddiqui, Maqsood A; Ahmad, Javed; Musarrat, Javed; Al-Khedhairy, Abdulaziz A; AlSalhi, Mohamad S; Alrokayan, Salman A

    2011-05-10

    Due to the interesting magnetic and electrical properties with good chemical and thermal stabilities, nickel ferrite nanoparticles are being utilized in many applications including magnetic resonance imaging, drug delivery and hyperthermia. Recent studies have shown that nickel ferrite nanoparticles produce cytotoxicity in mammalian cells. However, there is very limited information concerning the toxicity of nickel ferrite nanoparticles at the cellular and molecular level. The aim of this study was to investigate the cytotoxicity, oxidative stress and apoptosis induction by well-characterized nickel ferrite nanoparticles (size 26 nm) in human lung epithelial (A549) cells. Nickel ferrite nanoparticles induced dose-dependent cytotoxicity in A549 cells demonstrated by MTT, NRU and LDH assays. Nickel ferrite nanoparticles were also found to induce oxidative stress evidenced by generation of reactive oxygen species (ROS) and depletion of antioxidant glutathione (GSH). Further, co-treatment with the antioxidant L-ascorbic acid mitigated the ROS generation and GSH depletion due to nickel ferrite nanoparticles suggesting the potential mechanism of oxidative stress. Quantitative real-time PCR analysis demonstrated that following the exposure of A549 cells to nickel ferrite nanoparticles, the level of mRNA expressions of cell cycle checkpoint protein p53 and apoptotic proteins (bax, caspase-3 and caspase-9) were significantly up-regulated, whereas the expression of anti-apoptotic proteins (survivin and bcl-2) were down-regulated. Moreover, activities of caspase-3 and caspase-9 enzymes were also significantly higher in nickel ferrite nanoparticles exposed cells. To the best of our knowledge this is the first report showing that nickel ferrite nanoparticles induced apoptosis in A549 cells through ROS generation and oxidative stress via p53, survivin, bax/bcl-2 and caspase pathways. Copyright © 2011 Elsevier Ireland Ltd. All rights reserved.

  3. A polysaccharide fraction of adlay seed (Coixlachryma-jobi L.) induces apoptosis in human non-small cell lung cancer A549 cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Lu, Xiangyi; Liu, Wei; Wu, Junhua

    Highlights: Black-Right-Pointing-Pointer A polysaccharide from adlay seed, its molecular mass, optical rotation and sugars was determined. Black-Right-Pointing-Pointer We demonstrated that a polysaccharide from adlay can induce apoptosis in cancer cells. Black-Right-Pointing-Pointer The polysaccharide inhibited the metabolism and proliferation of NSCLC A549 cells. Black-Right-Pointing-Pointer The polysaccharide may trigger apoptosis via the mitochondria-dependent pathway. -- Abstract: Different seed extracts from Coix lachryma-jobi (adlay seed) have been used for the treatment of various cancers in China, and clinical data support the use of these extracts for cancer therapy; however, their underlying molecular mechanisms have not been well defined. A polysaccharide fraction, designated asmore » CP-1, was extracted from the C.lachryma-jobi L. var. using the ethanol subsiding method. CP-1 induced apoptosis in A549 cells in a dose-dependent manner, as determined by MTT assay. Apoptotic bodies were observed in the cells by scanning electronic microscopy. Apoptosis and DNA accumulation during S-phase of the cell cycle were determined by annexin V-FITC and PI staining, respectively, and measured by flow cytometry. CP-1 also extended the comet tail length on single cell gel electrophoresis, and disrupted the mitochondrial membrane potential. Further analysis by western blotting showed that the expression of caspase-3 and caspase-9 proteins was increased. Taken together, our results demonstrate that CP-1 is capable of inhibiting A549 cell proliferation and inducing apoptosis via a mechanism primarily involving the activation of the intrinsic mitochondrial pathway. The assay data suggest that in addition to its nutritional properties, CP-1 is a very promising candidate polysaccharide for the development of anti-cancer medicines.« less

  4. Molecular mechanisms underlying mangiferin-induced apoptosis and cell cycle arrest in A549 human lung carcinoma cells.

    PubMed

    Shi, Wei; Deng, Jiagang; Tong, Rongsheng; Yang, Yong; He, Xia; Lv, Jianzhen; Wang, Hailian; Deng, Shaoping; Qi, Ping; Zhang, Dingding; Wang, Yi

    2016-04-01

    Mangiferin, which is a C‑glucosylxanthone (1,3,6,7-tetrahydroxyxanthone-C2-β-D-glucoside) purified from plant sources, has recently gained attention due to its various biological activities. The present study aimed to determine the apoptotic effects of mangiferin on A549 human lung adenocarcinoma cells. In vitro studies demonstrated that mangiferin exerted growth‑inhibitory and apoptosis-inducing effects against A549 cells. In addition, mangiferin exhibited anti-tumor properties in A549 xenograft mice in vivo. Mangiferin triggered G2/M phase cell cycle arrest via downregulating the cyclin-dependent kinase 1-cyclin B1 signaling pathway, and induced apoptotic cell death by inhibiting the protein kinase C-nuclear factor-κB pathway. In addition, mangiferin was able to enhance the antiproliferative effects of cisplatin on A549 cells, thus indicating the potential for a combined therapy. Notably, mangiferin exerted anticancer effects in vivo, where it was able to markedly decrease the volume and weight of subcutaneous tumor mass, and expand the lifespan of xenograft mice. The present study clarified the molecular mechanisms underlying mangiferin-induced antitumor activities, and suggested that mangiferin may be considered a potential antineoplastic drug for the future treatment of cancer.

  5. Venom present in sea anemone (Heteractis magnifica) induces apoptosis in non-small-cell lung cancer A549 cells through activation of mitochondria-mediated pathway.

    PubMed

    Ramezanpour, Mahnaz; da Silva, Karen Burke; Sanderson, Barbara J S

    2014-03-01

    Lung cancer is a major cause of cancer deaths throughout the world and the complexity of apoptosis resistance in lung cancer is apparent. Venom from Heteractis magnifica caused dose-dependent decreases in survival of the human non-small-cell lung cancer cell line, as determined by the MTT and Crystal Violet assays. The H. magnifica venom induced cell cycle arrest and induced apoptosis of A549 cells, as confirmed by annexin V/propidium iodide staining. The venom-induced apoptosis in A549 cells was characterized by cleavage of caspase-3 and a reduction in the mitochondrial membrane potential. Interestingly, crude extracts from H. magnifica had less effect on the survival of non-cancer cell lines. In the non-cancer cells, the mechanism via which cell death occurred was through necrosis not apoptosis. These findings are important for future work using H. magnifica venom for pharmaceutical development to treat human lung cancer.

  6. Molecular mechanisms underlying mangiferin-induced apoptosis and cell cycle arrest in A549 human lung carcinoma cells

    PubMed Central

    SHI, WEI; DENG, JIAGANG; TONG, RONGSHENG; YANG, YONG; HE, XIA; LV, JIANZHEN; WANG, HAILIAN; DENG, SHAOPING; QI, PING; ZHANG, DINGDING; WANG, YI

    2016-01-01

    Mangiferin, which is a C-glucosylxanthone (1,3,6,7-tetrahydroxyxanthone-C2-β-D-glucoside) purified from plant sources, has recently gained attention due to its various biological activities. The present study aimed to determine the apoptotic effects of mangiferin on A549 human lung adenocarcinoma cells. In vitro studies demonstrated that mangiferin exerted growth-inhibitory and apoptosis-inducing effects against A549 cells. In addition, mangiferin exhibited anti-tumor properties in A549 xenograft mice in vivo. Mangiferin triggered G2/M phase cell cycle arrest via down-regulating the cyclin-dependent kinase 1-cyclin B1 signaling pathway, and induced apoptotic cell death by inhibiting the protein kinase C-nuclear factor-κB pathway. In addition, mangiferin was able to enhance the antiproliferative effects of cisplatin on A549 cells, thus indicating the potential for a combined therapy. Notably, mangiferin exerted anticancer effects in vivo, where it was able to markedly decrease the volume and weight of subcutaneous tumor mass, and expand the lifespan of xenograft mice. The present study clarified the molecular mechanisms underlying mangiferin-induced antitumor activities, and suggested that mangiferin may be considered a potential antineoplastic drug for the future treatment of cancer. PMID:26935347

  7. Apoptosis of human lung adenocarcinoma A549 cells induced by prodigiosin analogue obtained from an entomopathogenic bacterium Serratia marcescens.

    PubMed

    Zhou, Wei; Jin, Zhi-Xiong; Wan, Yong-Ji

    2010-12-01

    An entomopathogenic bacterial strain SCQ1 was isolated from silkworm (Bombyx mori) and identified as Serratia marcescens via 16S rRNA gene analysis. This strain produces a red pigment that causes acute septicemia of silkworm. The red pigment of strain SCQ1 was identified as prodigiosin analogue (PGA) with various reported biological activities. In this study, we found that low concentration of PGA showed significant anticancer activity in human lung adenocarcinoma A549 cells, but has little effect in human bone marrow stem cells, in vitro. By exposure to different concentrations of PGA for 24 h, morphological changes and the MTT assay showed that A549 cell line was very sensitive to PGA, with IC(50) value about 2.2 mg/L. Early stage of apoptosis was detected by flow cytometry while A549 cells were treated with PGA for 4 and 12 h, respectively. The proportion of dead cells was increased with treatment time or the concentrations of PGA, but it was inversely proportional to that of apoptotic cells. These results indicate that PGA obtained from strain SCQ1 induces apoptosis in A549 cells, but the molecular mechanisms of cell death are complicated, and the S. marcescens strain SCQ1 may serve as a source of the anticancer compound, PGA.

  8. Oleiferoside W from the roots of Camellia oleifera C. Abel, inducing cell cycle arrest and apoptosis in A549 cells.

    PubMed

    Wu, Jiang-Ping; Kang, Nai-Xin; Zhang, Mi-Ya; Gao, Hong-Wei; Li, Xiao-Ran; Liu, Yan-Li; Xu, Qiong-Ming; Yang, Shi-Lin

    2017-07-06

    Camellia oleifera C. Abel has been widely cultivated in China, and a group of bioactive constituents such as triterpeniod saponin have been isolated from C. oleifera C. Abel. In the current study, a new triterpeniod saponin was isolated from the EtOH extract of the roots of C. oleifera C. Abel, named as oleiferoside W, and the cytotoxic properties of oleiferoside W were evaluated in non-small cell lung cancer A549 cells. At the same time the inducing apoptosis, the depolarization of mitochondrial membrane potential (Δψ), the up-regulation of related pro-apoptotic proteins, such as cleaved-PARP, cleaved-caspase-3, and the down-regulation of anti-apoptotic marker Bcl-2/Bax were measured on oleiferoside W. Furthermore, the function, inducing the generation of reactive oxygen species (ROS) and apoptosis, of oleiferoside W could be reversed by N-acetylcysteine (NAC). In conclusion, our findings showed that oleiferoside W induced apoptosis involving mitochondrial pathway and increasing intracellular ROS production in the A549 cells, suggesting that oleiferoside W may have the possibility to be a useful anticancer agent for therapy in lung cancer.

  9. Effects of karanjin on cell cycle arrest and apoptosis in human A549, HepG2 and HL-60 cancer cells.

    PubMed

    Guo, Jian-Ru; Chen, Qian-Qian; Lam, Christopher Wai-Kei; Zhang, Wei

    2015-07-26

    We have investigated the potential anticancer effects of karanjin, a principal furanoflavonol constituent of the Chinese medicine Fordia cauliflora, using cytotoxic assay, cell cycle arrest, and induction of apoptosis in three human cancer cell lines (A549, HepG2 and HL-60 cells). MTT cytotoxic assay showed that karanjin could inhibit the proliferation and viability of all three cancer cells. The induction of cell cycle arrest was observed via a PI (propidium iodide)/RNase Staining Buffer detection kit and analyzed by flow cytometry: karanjin could dose-dependently induce cell cycle arrest at G2/M phase in the three cell lines. Cell apoptosis was assessed by Annexin V-FITC/PI staining: all three cancer cells treated with karanjin exhibited significantly increased apoptotic rates, especially in the percentage of late apoptosis cells. Karanjin can induce cancer cell death through cell cycle arrest and enhance apoptosis. This compound may be effective clinically for cancer pharmacotherapy.

  10. Shikonin Induces Apoptosis, Necrosis, and Premature Senescence of Human A549 Lung Cancer Cells through Upregulation of p53 Expression

    PubMed Central

    Yeh, Yueh-Chiao; Liu, Tsun-Jui; Lai, Hui-Chin

    2015-01-01

    Shikonin, a natural naphthoquinone pigment isolated from Lithospermum erythrorhizon, has been reported to suppress growth of various cancer cells. This study was aimed to investigate whether this chemical could also inhibit cell growth of lung cancer cells and, if so, works via what molecular mechanism. To fulfill this, A549 lung cancer cells were treated with shikonin and then subjected to microscopic, biochemical, flow cytometric, and molecular analyses. Compared with the controls, shikonin significantly induced cell apoptosis and reduced proliferation in a dose-dependent manner. Specially, lower concentrations of shikonin (1–2.5 μg/mL) cause viability reduction; apoptosis and cellular senescence induction is associated with upregulated expressions of cell cycle- and apoptotic signaling-regulatory proteins, while higher concentrations (5–10 μg/mL) precipitate both apoptosis and necrosis. Treatment of cells with pifithrin-α, a specific inhibitor of p53, suppressed shikonin-induced apoptosis and premature senescence, suggesting the role of p53 in mediating the actions of shikonin on regulation of lung cancer cell proliferation. These results indicate the potential and dose-related cytotoxic actions of shikonin on A549 lung cancer cells via p53-mediated cell fate pathways and raise shikonin a promising adjuvant chemotherapeutic agent for treatment of lung cancer in clinical practice. PMID:25737737

  11. β-Sitosterol targets Trx/Trx1 reductase to induce apoptosis in A549 cells via ROS mediated mitochondrial dysregulation and p53 activation.

    PubMed

    Rajavel, Tamilselvam; Packiyaraj, Pandian; Suryanarayanan, Venkatesan; Singh, Sanjeev Kumar; Ruckmani, Kandasamy; Pandima Devi, Kasi

    2018-02-01

    β-Sitosterol (BS), a major bioactive constituent present in plants and vegetables has shown potent anticancer effect against many human cancer cells, but the underlying mechanism remain elusive on NSCLC cancers. We found that BS significantly inhibited the growth of A549 cells without harming normal human lung and PBMC cells. Further, BS treatment triggered apoptosis via ROS mediated mitochondrial dysregulation as evidenced by caspase-3 & 9 activation, Annexin-V/PI positive cells, PARP inactivation, loss of MMP, Bcl-2-Bax ratio alteration and cytochrome c release. Moreover, generation of ROS species and subsequent DNA stand break were found upon BS treatment which was reversed by addition of ROS scavenger (NAC). Indeed BS treatment increased p53 expression and its phosphorylation at Ser15, while silencing the p53 expression by pifithrin-α, BS induced apoptosis was reduced in A549 cells. Furthermore, BS induced apoptosis was also observed in NCI-H460 cells (p53 wild) but not in the NCI-H23 cells (p53 mutant). Down-regulation of Trx/Trx1 reductase contributed to the BS induced ROS accumulation and mitochondrial mediated apoptotic cell death in A549 and NCI-H460 cells. Taken together, our findings provide evidence for the novel anti-cancer mechanism of BS which could be developed as a promising chemotherapeutic drug against NSCLC cancers.

  12. A flavonoid isolated from Streptomyces sp. (ERINLG-4) induces apoptosis in human lung cancer A549 cells through p53 and cytochrome c release caspase dependant pathway.

    PubMed

    Balachandran, C; Sangeetha, B; Duraipandiyan, V; Raj, M Karunai; Ignacimuthu, S; Al-Dhabi, N A; Balakrishna, K; Parthasarathy, K; Arulmozhi, N M; Arasu, M Valan

    2014-12-05

    The aim of this study was to investigate the anticancer activity of a flavonoid type of compound isolated from soil derived filamentous bacterium Streptomyces sp. (ERINLG-4) and to explore the molecular mechanisms of action. Cytotoxic properties of ethyl acetate extract was carried out against A549 lung cancer cell line using the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT) assay. Cytotoxic properties of isolated compound were investigated in A549 lung cancer cell line, COLO320DM cancer cell line and Vero cells. The compound showed potent cytotoxic properties against A549 lung cancer cell line and moderate cytotoxic properties against COLO320DM cancer cell line. Isolated compound showed no toxicity up to 2000 μg/mL in Vero cells. So we have chosen the A549 lung cancer cell line for further anticancer studies. Intracellular visualization was done by using a laser scanning confocal microscope. Apoptosis was measured using DNA fragmentation technique. Treatment of the A549 cancer cells with isolated compound significantly reduced cell proliferation, increased formation of fragmented DNA and apoptotic body. Activation of caspase-9 and caspase-3 indicated that compound may be inducing intrinsic and extrinsic apoptosis pathways. Bcl-2, p53, pro-caspases, caspase-3, caspase-9 and cytochrome c release were detected by western blotting analysis after compound treatment (123 and 164 μM). The activities of pro-caspases-3, caspase-9 cleaved to caspase-3 and caspase-9 gradually increased after the addition of isolated compound. But Bcl-2 protein was down regulated after treatment with isolated compound. Molecular docking studies showed that the compound bound stably to the active sites of caspase-3 and caspase-9. These results strongly suggest that the isolated compound induces apoptosis in A549 cancer cells via caspase activation through cytochrome c release from mitochondria. The present results might provide helpful suggestions for the design of

  13. Cancer Associated Fibroblast-Derived Hepatocyte Growth Factor Inhibits the Paclitaxel-Induced Apoptosis of Lung Cancer A549 Cells by Up-Regulating the PI3K/Akt and GRP78 Signaling on a Microfluidic Platform

    PubMed Central

    Xu, Zhiyun; He, Tianrui; Li, Encheng; Guo, Zhe; Liu, Fen; Jiang, Chunmeng; Wang, Qi

    2015-01-01

    Tumor stroma and growth factors provide a survival environment to tumor cells and can modulate their chemoresistance by dysregulating several signal pathways. In this study, we fabricated a three-dimensional (3D) microfluidic chip using polydimethylsiloxane (PDMS) to investigate the impact of hepatocyte growth factor (HGF) from cancer-associated fibroblasts (CAF) on the Met/PI3K/AKT activation, glucose regulatory protein (GRP78) expression and the paclitaxel-induced A549 cell apoptosis. With a concentration gradient generator, the assembled chip was able to reconstruct a tumor microenvironment in vitro. We found high levels of HGF in the supernatants of CAF and the CAF matrix from the supernatants of activated HFL1 fibroblasts or HGF enhanced the levels of Met, PI3K and AKT phosphorylation and GRP78 expression in A549 cells cultured in a 3D cell chamber, which was abrogated by anti-HGF. Inhibition of Met attenuated the CAF matrix-enhanced PI3K/AKT phosphorylation and GRP78 expression while inhibition of PI3K reduced GRP78 expression, but not Met phosphorylation in A549 cells. Inhibition of GRP78 failed to modulate the CAF matrix-enhanced Met/PI3K/AKT phosphorylation in A549 cells. Furthermore, inhibition of PI3K or GRP78 enhanced spontaneous and paclitaxel-induced A549 cell apoptosis. Moreover, treatment with the CAF matrix inhibited spontaneous and medium or high dose of paclitaxel-induced A549 cell apoptosis. Inhibition of PI3K or GRP78 attenuated the CAF matrix-mediated inhibition on paclitaxel-induced A549 cell apoptosis. Our data indicated that HGF in the CAF matrix activated the Met/PI3K/AKT and up-regulated GRP78 expression, promoting chemoresistance to paclitaxel-mediated apoptosis in A549 cells. Our findings suggest that the microfluidic system may represent an ideal platform for signaling research and drug screening. PMID:26115510

  14. Phytol shows anti-angiogenic activity and induces apoptosis in A549 cells by depolarizing the mitochondrial membrane potential.

    PubMed

    Sakthivel, Ravi; Malar, Dicson Sheeja; Devi, Kasi Pandima

    2018-06-13

    In the present study, the antiproliferative activity of phytol and its mechanism of action against human lung adenocarcinoma cell line A549 were studied in detail. Results showed that phytol exhibited potent antiproliferative activity against A549 cells in a dose and time-dependent manner with an IC 50 value of 70.81 ± 0.32 μM and 60.7 ± 0.47 μM at 24 and 48 h, respectively. Phytol showed no adverse toxic effect in normal human lung cells (L-132), but mild toxic effect was observed when treated with maximum dose (67 and 84 μM). No membrane-damaging effect was evidenced by PI staining and SEM analysis. The results of mitochondrial membrane potential analysis, cell cycle analysis, FT-IR and Western blotting analysis clearly demonstrated the molecular mechanism of phytol as induction of apoptosis in A549 cells, as evidenced by formation of shrinked cell morphology with membrane blebbing, depolarization of mitochondrial membrane potential, increased cell population in the sub-G0 phase, band variation in the DNA and lipid region, downregulation of Bcl-2, upregulation of Bax and the activation of caspase-9 and -3. In addition, phytol inhibited the CAM vascular growth as evidenced by CAM assay, which positively suggests that phytol has anti-angiogenic potential. Taken together, these findings clearly demonstrate the mode of action by which phytol induces cell death in A549 lung adenocarcinoma cells. Copyright © 2018 Elsevier Masson SAS. All rights reserved.

  15. Anticancer activity of polysaccharide from Glehnia littoralis on human lung cancer cell line A549.

    PubMed

    Wu, Jun; Gao, Weiping; Song, Zhuoyue; Xiong, Qingping; Xu, Yingtao; Han, Yun; Yuan, Jun; Zhang, Rong; Cheng, Yunbo; Fang, Jiansong; Li, Weirong; Wang, Qi

    2018-01-01

    The purpose of this study was to investigate the anticancer activity of polysaccharide (PGL) from Glehnia littoralis on human lung cancer cell line A549. Based on MTT assay, the results suggested that PGL could significantly reduce A549 cells proliferation in a time- and dose-dependent manner. In addition, PGL displayed an inhibitory activity for the A549 cells migration in Transwell migration assay. The results from both flow cytometry analysis and Hochst 3342 staining of apoptotic cells indicated that PGL could promote apoptosis, and induce cycle arrest of A549 cells. Moreover, immunofluorescence assay elucidated PGL could also down-regulate expression of proliferating cell nuclear antigen (PCNA). Overall, these results showed that PGL exerts a strong anticancer action through inhibiting the A549 cells migration, proliferation and inducing cell apoptosis. It could be a new source of natural anticancer agent against lung cancer with potential value in supplements and medicine. Copyright © 2017 Elsevier B.V. All rights reserved.

  16. SB203580 enhances the RV-induced loss of mitochondrial membrane potential and apoptosis in A549 cells

    NASA Astrophysics Data System (ADS)

    Li, Hai-yang; Zhuang, Cai-ping; Wang, Xiao-ping; Chen, Tong-sheng

    2012-03-01

    Resveratrol (RV), a naturally occurring phytoalexin, is known to possess a wide spectrum of chemopreventive and chemotherapeutic effects in various stages of human tumors. p38, a member of the mitogen-activated protein kinase (MAPK) superfamily, is always activated by some extracellular stimulus to regulate many cellular signal transduction pathways, such as apoptosis, proliferation, and inflammation and so on. In this report, we assessed the effect of SB203580, a specific inhibitor of p38 MAPK signaling pathway, on the RV-induced apoptosis in human lung adenocarcinoma (A549) cells. CCK-8 assay showed that pretreatment with SB203580 significantly enhanced the cytotoxicity of RV, which was further verified by analyzing the phosphatidylserine externalization using flow cytometry. In order to further confirm whether SB203580 accelerated apoptosis via the intrinsic apoptosis pathway, we analyzed the dysfunction of mitochondrial membrane potential (Δψm) of cells stained with rhodamine 123 by using flow cytometry after treatment with RV in the absence and presence of SB203580. Our data for the first time reported that p38 inhibitor SB203580 enhanced the RV-induced apoptosis via a mitochondrial pathway.

  17. Paraquat induces extrinsic pathway of apoptosis in A549 cells by induction of DR5 and repression of anti-apoptotic proteins, DDX3 and GSK3 expression.

    PubMed

    Hathaichoti, Sasiphen; Visitnonthachai, Daranee; Ngamsiri, Pronrumpa; Niyomchan, Apichaya; Tsogtbayar, Oyu; Wisessaowapak, Churaibhon; Watcharasit, Piyajit; Satayavivad, Jutamaad

    2017-08-01

    Paraquat (PQ) is a bipyridyl derivative herbicide known to cause lung toxicity partly through induction of apoptosis. Here we demonstrated that PQ caused apoptosis in A549 cells. PQ increased cleavage of caspase-8 and Bid, indicating caspase-8 activation and truncated Bid, the two key mediators of extrinsic apoptosis. Additionally, PQ treatment caused an increase in DR5 (death receptor-5) and caspase-8 interaction, indicating formation of DISC (death-inducing signaling complex). These results indicate that PQ induces apoptosis through extrinsic pathway in A549 cells. Moreover, PQ drastically increased DR5 expression and membrane localization. Furthermore, PQ caused prominent concentration dependent reductions of DDX3 (the DEAD box protein-3) and GSK3 (glycogen synthase kinase-3) which can associate with DR5 and prevent DISC formation. Additionally, PQ decreased DR5-DDX3 interaction, suggesting a reduction of DDX3/GSK3 anti-apoptotic complex. Inhibition of GSK3, which is known to promote extrinsic apoptosis by its pharmacological inhibitor, BIO accentuated PQ-induced apoptosis. Moreover, GSK3 inhibition caused a further decrease in PQ-reduced DR5-DDX3 interaction. Taken together, these results suggest that PQ may induce extrinsic pathway of apoptosis in A549 cells through upregulation of DR5 and repression of anti-apoptotic proteins, DDX3/GSK3 leading to reduction of anti-apoptotic complex. Copyright © 2017 Elsevier Ltd. All rights reserved.

  18. A novel polysaccharide from Sargassum integerrimum induces apoptosis in A549 cells and prevents angiogensis in vitro and in vivo.

    PubMed

    Liu, Ge; Kuang, Shan; Wu, Shimei; Jin, Weihua; Sun, Chaomin

    2016-05-24

    Many polysaccharides isolated from plants have exhibited promising antitumor activities. The aim of this study is to investigate the antitumor activity of the novel polysaccharide named SPS from Sargassum integerrimum, elucidate the underlying anticancer mechanism in a human lung cancer cell line A549, and evaluate its anti-angiogenic activity both in vitro and in vivo. The results show that SPS significantly reduces A549 cells viability in a dose- and time-dependent manner via MTT method. Flow cytometry analysis indicates that SPS could induce cell apoptosis, the loss of mitochondrial membrane potential (MMP), generation of reactive oxygen species (ROS) and G2/M phase cell cycle arrest of A549 cells. Up-regulation of the expressions of P53 and Bax, down-regulation of the expression of Bcl-2, and activation of cleaved caspase-3, caspase-9 and PARP are also detected by western blotting after the treatment of SPS. In addition, SPS inhibits the proliferation, migration and cord formation of human umbilical vein endothelial cells (HUVECs) in vitro, and prevents the vascular development of zebrafish embryos in vivo. Altogether, our data prove the anticancer and anti-angiogenesis properties of SPS, and provide further insights into the potential pharmacological application of SPS as antitumor and anti-angiogenic agent against lung cancer.

  19. Anti-tumor activity and mechanism of apoptosis of A549 induced by ruthenium complex.

    PubMed

    Sun, Dongdong; Mou, Zhipeng; Li, Nuan; Zhang, Weiwei; Wang, Yazhe; Yang, Endong; Wang, Weiyun

    2016-12-01

    Two new ruthenium (II) polypyridyl complexes [Ru(MeIm) 4 (pip)] 2+ (1) and [Ru(MeIm) 4 (4-npip)] 2+ (2) were synthesized under the guidance of computational studies (DFT). Their binding property to human telomeric G-quadruplex studied by UV-Vis absorption spectroscopy, the fluorescent resonance energy transfer (FRET) melting assay and circular dichroism (CD) spectroscopy for validating the theoretical prediction. Both of them were evaluated for their potential anti-proliferative activity against four human tumor cell lines. Complex 2 shows growth inhibition against all the cell lines tested, especially the human lung tumor cell (A549). The RTCA analysis not only validated the inhibition activity but also showed the ability of reducing A549 cells' migration. DNA-flow cytometric analysis, mitochondrial membrane potential (ΔΨm) and the scavenger measurements of reactive oxygen species (ROS) analysis carried out to investigate the mechanism of cell growth inhibition and apoptosis-inducing effect of complex 2. The results demonstrated that complex 2 induces tumor cells apoptosis by acting on both mitochondrial homeostasis destruction and death receptor signaling pathways. And those suggested that complex 2 could be a candidate for further evaluation as a chemotherapeutic agent against human tumor.

  20. Chlorogenic acid regulates apoptosis and stem cell marker-related gene expression in A549 human lung cancer cells.

    PubMed

    Yamagata, Kazuo; Izawa, Yuri; Onodera, Daiki; Tagami, Motoki

    2018-04-01

    Previous studies indicated that chlorogenic acid, a compound present in many fruits and vegetables, has anti-cancer activities. We report that chlorogenic acid regulates the expression of apoptosis-related genes and self-renewal-related stem cell markers in cancer cells. The lung cancer cell line A549 was cultured with or without chlorogenic acid. The presence of chlorogenic acid decreased cell proliferation as measured by MTT activity. Polymerase chain reaction (PCR) showed that treatment of cells with chlorogenic acid reduced the expression of BCL2 but increased that of both BAX and CASP3. Chlorogenic acid enhanced annexin V expression as measured using fluorescently labeled annexin V. Chlorogenic acid also induced p38 MAPK and JNK gene expression. Meanwhile, several agents, including SB203580 (p38 MAP kinase inhibitor), N-acetylcysteine (antioxidant inhibitor), dipyridamole (phosphodiesterase inhibitor), and apocynin (NADPH-oxidase inhibitor) blocked chlorogenic acid-induced BAX gene expression. Chlorogenic acid reduced gene expression levels of stem cell-associated markers NANOG, POU5F1, and SOX2. Together these results indicate that chlorogenic acid affects the expression of apoptosis-related genes that are part of oxidative stress and p38 MAP-dependent pathways, as well as genes encoding stem cell markers. In conclusion, chlorogenic acid may contribute to the polyphenolic anti-cancer effect associated with consumption of vegetables and fruits.

  1. A novel small molecule, Rosline, inhibits growth and induces caspase-dependent apoptosis in human lung cancer cells A549 through a reactive oxygen species-dependent mechanism.

    PubMed

    Zhao, Ting; Feng, Yang; Jin, Wenling; Pan, Hui; Li, Haizhou; Zhao, Yang

    2016-06-01

    Chemical screening using synthetic small molecule libraries has provided a huge amount of novel active molecules. It generates lead compound for drug development and brings focus on molecules for mechanistic investigations on many otherwise intangible biological processes. In this study, using non-small cell lung cancer cell A549 to screen against a structurally novel and diverse synthetic small molecule library of 2,400 compounds, we identified a molecule named rosline that has strong anti-proliferation activity on A549 cells with a 50% cell growth inhibitory concentration (IC50 ) of 2.87 ± 0.39 µM. We showed that rosline treatment increased the number of Annexin V-positive staining cell, as well as G2/M arrest in their cell cycle progression. Further, we have demonstrated that rosline induces a decrease of mitochondrial membrane potential (Δφm ) and an increase of caspases 3/7 and 9 activities in A549 cells, although having no effect on the activity of caspase 8. Moreover, we found that rosline could induce the production of reactive oxygen species (ROS) and inhibit the phosphorylation of signaling molecule Akt in A549 cells. Alternatively, an antioxidant N-acetyl-L-cysteine (NAC) significantly attenuated rosline's effects on the mitochondrial membrane potential, caspases 3/7 and 9 activities, cell viabilities and the phosphorylation of Akt. Our results demonstrated that ROS played an important role in the apoptosis of A549 cells induced by rosline. © 2016 International Federation for Cell Biology.

  2. MiR-21 suppresses the anticancer activities of curcumin by targeting PTEN gene in human non-small cell lung cancer A549 cells.

    PubMed

    Zhang, W; Bai, W; Zhang, W

    2014-08-01

    Curcumin, a natural phytochemical, exhibits potent anticancer activities. Here, we sought to determine the molecular mechanisms underlying the cytotoxic effects of curcumin against human non-small cell lung cancer (NSCLC) cells. MTT assay and annexin-V/PI staining were used to analyze the effects of curcumin on the proliferation and apoptosis of A549 cells. The expression of microRNA-21 in curcumin-treated A549 cells was measured by quantitative real-time polymerase chain reaction assay. The protein level of phosphatase and tensin homolog (PTEN), a putative target of microRNA-21, was determined by Western blot analysis. Transfection of A549 cells with microRNA-21 mimic or PTEN small interfering RNA was performed to modulate the expression of microRNA-21 and PTEN under the treatment of curcumin. Curcumin at 20-40 μM inhibited cell proliferation and induced apoptosis in A549 cells. Curcumin treatment produced a dose-dependent and significant (P < 0.05) suppression of microRNA-21 expression, compared to untreated A549 cells. Moreover, the protein level of PTEN, a putative target of microRNA-21, was significantly elevated in curcumin-treated A549 cells, as determined by Western blot analysis. Transfection of A549 cells with microRNA-21 mimic or PTEN small interfering RNA significantly (P < 0.05) reversed the growth suppression and apoptosis induction by curcumin, compared to corresponding controls. Our data suggest a novel molecular mechanism in which inhibition of microRNA-21 and upregulation of PTEN mediate the anticancer activities of curcumin in NSCLC cells. Suppression of microRNA-21 may thus have therapeutic benefits against this malignancy.

  3. [The mechanism of docetaxel-induced apoptosis in human lung cancer cells].

    PubMed

    Li, Y; Shi, T; Zhao, W

    2000-05-01

    To study the mechanism of docetaxel-induced apoptosis. Morphological study, DNA gel electrophoresis, flow cytometry and fluorescin labeled Annexin V to detect apoptosis, RT-PCR to detect the gene related with apoptosis. Human lung cancer A549 cells treated with docetaxel induced cell cycle arrest at G2M phase, leading to apoptosis. The morphology of A549 showed nuclear chromatine condensation and fragmentation. Typical ladder pattern of DNA fragmentation was observed. Sub-G1 peak was found by flow cytometry. Transcription of Fas gene was enhanced, while no change in c-myc and bcl-2 genes. Annexin labeling results revealed the co-existence of cell apoptosis and necrosis in docetaxel-treated A549 cells. Docetaxel induces apoptosis and necrosis of human lung cancer. The induction of apoptosis may be related to expression of Fas.

  4. The production of reactive oxygen species and the mitochondrial membrane potential are modulated during onion oil-induced cell cycle arrest and apoptosis in A549 cells.

    PubMed

    Wu, Xin-jiang; Stahl, Thorsten; Hu, Ying; Kassie, Fekadu; Mersch-Sundermann, Volker

    2006-03-01

    Protective effects of Allium vegetables against cancers have been shown extensively in experimental animals and epidemiologic studies. We investigated cell proliferation and the induction of apoptosis by onion oil extracted from Allium cepa, a widely consumed Allium vegetable, in human lung cancer A549 cells. GC/MS analysis suggested that propyl sulfides but not allyl sulfides are major sulfur-containing constituents of onion oil. Onion oil at 12.5 mg/L significantly induced apoptosis (13% increase of apoptotic cells) as indicated by sub-G1 DNA content. It also caused cell cycle arrest at the G2/M phase; 25 mg/L onion oil increased the percentage of G2/M cells almost 6-fold compared with the dimethyl sulfoxide control. The action of onion oil may occur via a reactive oxygen species-dependent pathway because cell cycle arrest and apoptosis were blocked by the antioxidants N-acetylcysteine and exogenous glutathione. Marked collapse of the mitochondrial membrane potential suggested that dysfunction of the mitochondria may be involved in the oxidative burst and apoptosis induced by onion oil. Expression of phospho-cdc2 and phospho-cyclin B1 were downregulated by onion oil, perhaps accounting for the G2/M arrest. Overall, these results suggest that onion oil may exert chemopreventive action by inducing cell cycle arrest and apoptosis in tumor cells.

  5. mTOR inhibition of cardamonin on antiproliferation of A549 cells is involved in a FKBP12 independent fashion.

    PubMed

    Tang, Ying; Fang, Qi; Shi, Daohua; Niu, Peiguang; Chen, Yaoyao; Deng, Jie

    2014-03-18

    Cardamonin has previously demonstrated that it had an antiproliferative effect on vascular smooth muscle cells by inhibiting the activity of mammalian target of rapamycin (mTOR). The antiproliferative effect and the possible mechanism of combining with mTOR of cardamonin were investigated on A549 cells. Cell proliferation, cell cycle and apoptosis were measured by methyl thiazolyl tetrazolium (MTT) and flow cytometry, respectively. mTOR and 12 kDa FK506 binding protein (FKBP12) were transfected into A549 cells by Lipofectamine. Western blots were used to examine the mTOR expressions and its activities, and the expressions of 70 kDa ribosomal S6 kinase (p70S6K), FKBP12 and Interleukin-2 (IL-2), respectively. Treated with cardamonin, the proliferation of A549 cells was inhibited. Meanwhile, cell cycle was blocked and DNA synthesis was decreased whereas cell apoptosis was promoted, and the activation of mTOR and p70S6K was decreased by cardamonin. Transfected with mTOR or FKBP12, proliferation of A549 cells was increased. Rapamycin had a similar degree of effect on antiproliferation of both transfected cells. However, the antiproliferative effect of cardamonin on mTOR transfected cells was stronger than that on FKBP12 transfected cells. Both rapamycin and cardamonin decreased the phosphorylation of mTOR and p70S6K in two kinds of transfected cells. Cardamonin had no effect on the expression of FKBP12 and IL-2, whereas the expressions were decreased by rapamycin. Cardamonin inhibited proliferation and induced apoptosis of A549 cells via mTOR. It might directly interact with mTOR independently of binding with FKBP12. Copyright © 2014 Elsevier Inc. All rights reserved.

  6. Chlorella vulgaris Induces Apoptosis of Human Non-Small Cell Lung Carcinoma (NSCLC) Cells.

    PubMed

    Zhang, Zhi-Dong; Liang, Kai; Li, Kun; Wang, Guo-Quan; Zhang, Ke-Wei; Cai, Lei; Zhai, Shui-Ting; Chou, Kuo-Chen

    2017-01-01

    Chlorella vulgaris (C. vulgaris), a unicellular green microalga, has been widely used as a food supplement and reported to have antioxidant and anticancer properties. The current study was designed to assess the cytotoxic, apoptotic, and DNA-damaging effects of C. vulgaris growth factor (CGF), hot water C. vulgaris extracts, inlung tumor A549 and NCI-H460 cell lines. A549 cells, NCI-H460 cells, and normal human fibroblasts were treated with CGF at various concentrations (0-300 μg/ml) for 24 hr. The comet assay and γH2AX assay showed DNA damage in A549 and NCI-H460 cells upon CGF exposure. Evaluation of apoptosis by the TUNEL assay and DNA fragmentation analysis by agarose gel electrophoresis showed that CGF induced apoptosis in A549 and NCI-H460 cells. Chlorella vulgaris hot water extract induced apoptosis and DNA damage in human lung carcinoma cells. CGF can thus be considered a potential cytotoxic or genotoxic drug for treatment of lung carcinoma. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  7. Effects of TGF-β signaling blockade on human A549 lung adenocarcinoma cell lines.

    PubMed

    Xu, Cheng-Cheng; Wu, Lei-Ming; Sun, Wei; Zhang, Ni; Chen, Wen-Shu; Fu, Xiang-Ning

    2011-01-01

    Transforming growth factor β (TGF-β) is overexpressed in a wide variety of cancer types including lung adenocarcinoma (LAC), and the TGF-β signaling pathway plays an important role in tumor development. To determine whether blockade of the TGF-β signaling pathway can inhibit the malignant biological behavior of LAC, RNA interference (RNAi) technology was used to silence the expression of TGF-β receptor, type II (TGFβRII) in the LAC cell line, A549, and its effects on cell proliferation, invasion and metastasis were examined. Three specific small interfering RNAs (siRNAs) designed for targeting human TGFβRII were transfected into A549 cells. The expression of TGFβRII was detected by Western blot analysis. Cell proliferation was measured by MTT and clonogenic assays. Cell apoptosis was assessed by flow cytometry. The invasion and metastasis of A549 cells were investigated using the wound healing and Matrigel invasion assays. The expression of PI3K, phosphorylated Smad2, Smad4, Akt, Erk1/2, P38 and MMPs was detected by Western blot analysis. The TGFβRII siRNA significantly reduced the expression of TGFβRII in A549 cells. The knockdown of TGFβRII in A549 cells resulted in the suppression of cell proliferation, invasion and metastasis and induced cell apoptosis. In addition to the Smad-dependent pathway, independent pathways including the Erk MAPK, PI3K/Akt and p38 MAPK pathways, as well as the expression of MMPs and VEGF, were inhibited. In conclusion, TGF-β signaling is required for LAC progression. Therefore, the blockade of this signaling pathway by the down-regulation of TGFβRII using SiRNA may provide a potential gene therapy for LAC.

  8. MG132 as a proteasome inhibitor induces cell growth inhibition and cell death in A549 lung cancer cells via influencing reactive oxygen species and GSH level.

    PubMed

    Han, Yong Hwan; Park, Woo Hyun

    2010-07-01

    Carbobenzoxy-Leu-Leu-leucinal (MG132) as a proteasome inhibitor has been shown to induce apoptotic cell death through formation of reactive oxygen species (ROS). In the present study, we evaluated the effects of MG132 on the growth of A549 lung cancer cells in relation to cell growth, ROS and glutathione (GSH) levels. Treatment with MG132 inhibited the growth of A549 cells with an IC(50) of approximately 20 microM at 24 hours. DNA flow cytometric analysis indicated that 0.5 approximately 30 microM MG132 induced a G1 phase arrest of the cell cycle in A549 cells. Treatment with 10 or 30 microM MG132 also induced apoptosis, as evidenced by sub-G1 cells and annexin V staining cells. This was accompanied by the loss of mitochondrial membrane potential (MMP; Delta psi m). The intracellular ROS levels including O(2) (*-) were strongly increased in 10 or 30 microM MG132-treated A549 cells but were down-regulated in 0.1, 0.5 or 1 microM MG132-treated cells. Furthermore, 10 or 30 microM MG132 increased mitochondrial O(2) (*- ) level but 0.1, 0.5 or 1 microM MG132 decreased that. In addition, 10 or 30 microM MG132 induced GSH depletion in A549 cells. In conclusion, MG132 inhibited the growth of human A549 cells via inducing the cell cycle arrest as well as triggering apoptosis, which was in part correlated with the changes of ROS and GSH levels. Our present data provide important information on the anti-growth mechanisms of MG132 in A549 lung cancer cells in relation to ROS and GSH.

  9. Apatinib resensitizes cisplatin-resistant non-small cell lung carcinoma A549 cell through reversing multidrug resistance and suppressing ERK signaling pathway.

    PubMed

    Liu, Z-L; Jin, B-J; Cheng, C-G; Zhang, F-X; Wang, S-W; Wang, Y; Wu, B

    2017-12-01

    To observe the reversal effect of apatinib on the resistance to cisplatin (DDP) of A549/cisplatin (A549/DDP) cells and its relevant mechanism. A549/DDP cells were treated with the control method, apatinib alone, DDP alone and DDP combined with apatinib. The cell proliferation was detected by the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay and the cell clone formation assay. The cell apoptosis was detected by Hoechst 33258 staining and annexin V and propidium iodide (PI) double labeling. The changes in apoptotic proteins, multidrug resistance protein 1 (MDR1) and extracellular signal-regulated kinase (ERK) signaling pathway proteins in each group after treatment were detected by Western blotting. MTT assay results showed that compared with A549 cells, A549/DDP cells had obvious resistance to DDP. MTT assay and cell clone formation assay revealed that the tumor inhibition rate of the sub-lethal dose of apatinib (10 μM) combined with DDP was higher than that of DDP alone. The apoptosis detection results indicated that the proportion of apoptotic cells in the apatinib (10 μM) combined with DDP group was significantly increased. Western blotting results revealed that compared with that in parental A549 cells, the expression level of MDR1 in A549/DDP cells was significantly increased, and the ERK signaling pathway was activated. In the apatinib combined with DDP group, the levels of cleaved caspase-3, cleaved caspase-9 and B-cell lymphoma-2 (Bcl-2)-associated X (BAX) proteins were significantly upregulated, while the level of Bcl-2 proteins was downregulated. Apatinib could inhibit the expression of MDR1 and the activity of the ERK signaling pathway in a dose-dependent manner. Apatinib can restore the sensitivity of A549/DDP cells to DDP by down-regulating the expression level of MDR1 and inhibiting the activity of the ERK signaling pathway.

  10. TRAIL overexpression co-regulated by Egr1 and HRE enhances radiosensitivity of hypoxic A549 cells depending on its apoptosis inducing role.

    PubMed

    Yang, Yan-Ming; Fang, Fang; Li, Xin; Yu, Lei; Wang, Zhi-Cheng

    2017-01-01

    Ionizing radiation can upregulate the expression levels of TRAIL and enhance tumor cell apoptosis. While Early growth response 1 (Egr1) gene promoter has radiation inducible characteristics, the expression for exogenous gene controlled by Egr1 promoter could be enhanced by ionizing radiation, but its efficiency is limited by tissue hypoxia. Hypoxia response elements (HREs) are important hypoxic response regulatory sequences and sensitivity enhancers. Therefore, we chose TRAIL as the gene radiotherapy to observe whether it is regulated by Egr1 and HER and its effects on A549 cells and its mechanism. The pcDNA3.1-Egr1-TRAIL (pc-E-hsT) and pcDNA3.1-HRE/Egr1-TRAIL (pc-H/E-hsT) plasmids containing Egr1-hsTRAIL and HRE/Egr1-hsTRAIL were transfected into A549 cells, the cells were treated by hypoxia and radiation. The TRAIL mRNA in the cells and protein concentration in the culture supernatants were measured by RT-PCR and ELISA, respectively. Mean lethal dose D0 value was evaluated with colony forming assay. The cell apoptotic rates were analyzed by FCM and TUNEL assay. Expression of DR4, DR5 and cleaved caspase-3 proteins were analyzed by western blotting. It showed that TRAIL mRNA expression and TRAIL concentration all significantly increased under hypoxia and/or radiation. D0 value of pc-H/E‑hsT transfected cells under hypoxia was lowest, indicating more high radiosensitivity. Hypoxia could not cause the pc-E-hsT transfected cell apoptotic rate increase, but there were promoting effects in pc-H/E-hsT transfected cells. DR4 had not obvious change in pc-E-hsT and pc-H/E-hsT transfected cells under normoxic and hypoxic condition, otherwise, DR5 and cleaved caspase-3 increased mostly in pc-H/E-hsT transfected cells under hypoxic condition. TRAIL overexpression was co-regulated by Egr1 and HRE. TRAIL might promote hypoxic A549 cell radiosensitivity and induce apoptosis depending on DR5 to caspase-3 pathways.

  11. Effect of taxol from Pestalotiopsis mangiferae on A549 cells-In vitro study.

    PubMed

    Kathiravan, Govindarajan; Sureban, Sripathi M

    2009-12-01

    Pestalotiopsis mangiferae Coelomycete fungi were used to examine the production of taxol. The taxol isolated from this fungus is biologically active against cancer cell lines were investigated for its antiproliferative activity in human Non Small Cell Lung Cancer A549 cells. The results showed that the methylene chloride extraction of Pestalotiopsis mangiferae inhibited the proliferation of A 549 cells as measured by MTT and Trypan blue assay. Flow cytometric analysis showed that methylene chloride extraction of Pestalotiopsis mangiferae blocked cell cycle progression in G0/G1 phase. In addition fungal taxol induced A549 cell apoptosis as determined by propidium iodide staining. Further the percentage of LDH release was increased at increasing concentrations which is a measure of cell death. The levels of sialic acid levels and DNA, RNA and protein levels were decreased after treatment with methylene chloride extraction of Pestalotiopsis mangiferae. We suggests that methylene chloride extraction of Pestalotiopsis mangiferae might be considered for future therapeutic application with further studies against lung cancer.

  12. Dehydrobruceine B enhances the cisplatin-induced cytotoxicity through regulation of the mitochondrial apoptotic pathway in lung cancer A549 cells.

    PubMed

    Huang, Zhuqing; Yang, Guotao; Shen, Tao; Wang, Xiaoning; Li, Haizhen; Ren, Dongmei

    2017-05-01

    Dehydrobruceine B (DHB) is a quassinoid isolated from Brucea javanica. We have shown previously that DHB induced apoptosis on two kinds of lung cancer cell lines, A549 and NCI-H292. In the present study, we investigated the interactions of DHB and cisplatin (CDDP) on apoptotic-related cancer cell death. Synergistic effects on cell proliferation and apoptosis were observed when A549 cells were treated with DHB plus CDDP. DHB combined CDDP exposure increased depolarization of mitochondrial membrane potential (MMP) and release of cytochrome c from mitochondria into the cytoplasm. The combination treatment also enhanced protein expression of Bax, reduced the protein levels of Bcl-xL and Bcl-2, and increased the cleavage of caspase-3, caspase-9 and poly (ADP-ribose) polymerase (PARP). These results indicated that DHB sensitized A549 cells to cisplatin by regulating the mitochondrial apoptotic pathway. High constitutive expression of Nrf2 was found in A549 cells, which enhance the resistance of cancer cells to chemotherapeutic agents including cisplatin. DHB reduced the protein levels of Nrf2 and its target genes, which may contribute to the increase of intracellular ROS level, consequently, induced mitochondria apoptosis. These results generated a rationale for further investigation of DHB combined with CDDP as a potential therapeutic strategy in lung cancer. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  13. Chrysophanol-induced cell death (necrosis) in human lung cancer A549 cells is mediated through increasing reactive oxygen species and decreasing the level of mitochondrial membrane potential.

    PubMed

    Ni, Chien-Hang; Yu, Chun-Shu; Lu, Hsu-Feng; Yang, Jai-Sing; Huang, Hui-Ying; Chen, Po-Yuan; Wu, Shin-Hwar; Ip, Siu-Wan; Chiang, Su-Yin; Lin, Jaung-Geng; Chung, Jing-Gung

    2014-05-01

    Chrysophanol (1,8-dihydroxy-3-methylanthraquinone) is one of the anthraquinone compounds, and it has been shown to induce cell death in different types of cancer cells. The effects of chrysophanol on human lung cancer cell death have not been well studied. The purpose of this study is to examine chrysophanol-induced cytotoxic effects and also to investigate such influences that involved apoptosis or necrosis in A549 human lung cancer cells in vitro. Our results indicated that chrysophanol decreased the viable A549 cells in a dose- and time-dependent manner. Chrysophanol also promoted the release of reactive oxygen species (ROS) and Ca(2+) and decreased the levels of mitochondria membrane potential (ΔΨm ) and adenosine triphosphate in A549 cells. Furthermore, chrysophanol triggered DNA damage by using Comet assay and DAPI staining. Importantly, chrysophanol only stimulated the cytocheome c release, but it did not activate other apoptosis-associated protein levels including caspase-3, caspase-8, Apaf-1, and AIF. In conclusion, human lung cancer A549 cells treated with chrysophanol exhibited a cellular pattern associated with necrotic cell death and not apoptosis in vitro. © 2012 Wiley Periodicals, Inc. Environ Toxicol 29: 740-749, 2014. Copyright © 2012 Wiley Periodicals, Inc., a Wiley company.

  14. Effect of taxol from Pestalotiopsis mangiferae on A549 cells-In vitro study

    PubMed Central

    Kathiravan, Govindarajan; Sureban, Sripathi M.

    2009-01-01

    Pestalotiopsis mangiferae Coelomycete fungi were used to examine the production of taxol. The taxol isolated from this fungus is biologically active against cancer cell lines were investigated for its antiproliferative activity in human Non Small Cell Lung Cancer A549 cells. The results showed that the methylene chloride extraction of Pestalotiopsis mangiferae inhibited the proliferation of A 549 cells as measured by MTT and Trypan blue assay. Flow cytometric analysis showed that methylene chloride extraction of Pestalotiopsis mangiferae blocked cell cycle progression in G0/G1 phase. In addition fungal taxol induced A549 cell apoptosis as determined by propidium iodide staining. Further the percentage of LDH release was increased at increasing concentrations which is a measure of cell death. The levels of sialic acid levels and DNA, RNA and protein levels were decreased after treatment with methylene chloride extraction of Pestalotiopsis mangiferae. We suggests that methylene chloride extraction of Pestalotiopsis mangiferae might be considered for future therapeutic application with further studies against lung cancer. PMID:25206246

  15. Depletion of hepatoma-derived growth factor-related protein-3 induces apoptotic sensitization of radioresistant A549 cells via reactive oxygen species-dependent p53 activation

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Yun, Hong Shik; Hong, Eun-Hee; Department of Chemistry, College of Natural Sciences, Hanyang University, Seoul 133-791

    2013-09-27

    Highlights: •HRP-3 is a radiation- and anticancer drug-responsive protein in A549 cells. •Depletion of HRP-3 induces apoptosis of radio- and chemoresistant A549 cells. •Depletion of HRP-3 promotes ROS generation via inhibition of the Nrf2/HO-1 pathway. •Depletion of HRP-3 enhances ROS-dependent p53 activation and PUMA expression. -- Abstract: Biomarkers based on functional signaling have the potential to provide greater insight into the pathogenesis of cancer and may offer additional targets for anticancer therapeutics. Here, we identified hepatoma-derived growth factor-related protein-3 (HRP-3) as a radioresistance-related gene and characterized the molecular mechanism by which its encoded protein regulates the radio- and chemoresistant phenotypemore » of lung cancer-derived A549 cells. Knockdown of HRP-3 promoted apoptosis of A549 cells and potentiated the apoptosis-inducing action of radio- and chemotherapy. This increase in apoptosis was associated with a substantial generation of reactive oxygen species (ROS) that was attributable to inhibition of the Nrf2/HO-1 antioxidant pathway and resulted in enhanced ROS-dependent p53 activation and p53-dependent expression of PUMA (p53 upregulated modulator of apoptosis). Therefore, the HRP-3/Nrf2/HO-1/ROS/p53/PUMA cascade is an essential feature of the A549 cell phenotype and a potential radiotherapy target, extending the range of targets in multimodal therapies against lung cancer.« less

  16. Perfluorocarbon reduces cell damage from blast injury by inhibiting signal paths of NF-κB, MAPK and Bcl-2/Bax signaling pathway in A549 cells

    PubMed Central

    Li, Huaidong; Li, Chunsun; Yang, Zhen; Li, Yanqin; She, Danyang; Cao, Lu; Wang, Wenjie; Liu, Changlin; Chen, Liangan

    2017-01-01

    Background and objective Blast lung injury is a common type of blast injury and has very high mortality. Therefore, research to identify medical therapies for blast injury is important. Perfluorocarbon (PFC) is used to improve gas exchange in diseased lungs and has anti-inflammatory functions in vitro and in vivo. The aim of this study was to determine whether PFC reduces damage to A549 cells caused by blast injury and to elucidate its possible mechanisms of action. Study design and methods A549 alveolar epithelial cells exposed to blast waves were treated with and without PFC. Morphological changes and apoptosis of A549 cells were recorded. PCR and enzyme-linked immunosorbent assay (ELISA) were used to measure the mRNA or protein levels of IL-1β, IL-6 and TNF-α. Malondialdehyde (MDA) levels and superoxide dismutase (SOD) activity levels were detected. Western blot was used to quantify the expression of NF-κB, Bax, Bcl-2, cleaved caspase-3 and MAPK cell signaling proteins. Results A549 cells exposed to blast wave shrank, with less cell-cell contact. The morphological change of A549 cells exposed to blast waves were alleviated by PFC. PFC significantly inhibited the apoptosis of A549 cells exposed to blast waves. IL-1β, IL-6 and TNF-α cytokine and mRNA expression levels were significantly inhibited by PFC. PFC significantly increased MDA levels and decreased SOD activity levels. Further studies indicated that NF-κB, Bax, caspase-3, phospho-p38, phosphor-ERK and phosphor-JNK proteins were also suppressed by PFC. The quantity of Bcl-2 protein was increased by PFC. Conclusion Our research showed that PFC reduced A549 cell damage caused by blast injury. The potential mechanism may be associated with the following signaling pathways: 1) the signaling pathways of NF-κB and MAPK, which inhibit inflammation and reactive oxygen species (ROS); and 2) the signaling pathways of Bcl-2/Bax and caspase-3, which inhibit apoptosis. PMID:28323898

  17. Novel taspine derivative 12k inhibits cell growth and induces apoptosis in lung cell carcinoma.

    PubMed

    Dai, Bingling; Wang, Wenjie; Liu, Rui; Wang, Hongying; Zhang, Yanmin

    2015-03-01

    Taspine is an active compound in anticancer agent development. 12k was synthesized with taspine as lead compound bearing biphenyl scaffold and showed potent anticancer activity. Here, we investigated the effect of taspine derivative 12k on A549 lung cells. We showed that 12k not only decreased significantly A549 cell viability, A549 cell colony formation but also impaired A549 cell migration. Moreover, 12k treatment blocked cell cycle progression by increasing cell number in S phase to 42.80% for 6 μmol/L vs. 28.86% for control while decreasing cell number in G1 phase. Accordingly, this was associated with an increase protein expression of cyclin E and a decrease protein expression of cyclin D1, cyclin B1 and its associated CDK1 (cdc2). Meanwhile, we found that 12k induced A549 cell apoptosis, which was closely associated with the effect of the Bcl-2 family. Increase of Bad, Bak and Bax expression levels, decrease of Bcl-2 and Mcl-1 expression levels were observed. SiRNA knockdown of c-myc in A549 cells significantly attenuated tumor inhibition effects of 12k. In conclusion, our results demonstrate that 12k has an inhibitory effect on growth of A549 cell by inducing cell cycle arrest and apoptosis. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  18. Induction of cell death by pyropheophorbide-α methyl ester-mediated photodynamic therapy in lung cancer A549 cells.

    PubMed

    Tu, Ping-Hua; Huang, Wen-Jun; Wu, Zhan-Ling; Peng, Qing-Zhen; Xie, Zhi-Bin; Bao, Ji; Zhong, Ming-Hua

    2017-03-01

    Pyropheophorbide-α methyl ester (MPPa) was a promising photosensitizer with stable chemical structure, strong absorption, higher tissue selectivity and longer activation wavelengths. The present study investigated the effect of MPPa-mediated photodynamic treatment on lung cancer A549 cells as well as the underlying mechanisms. Cell Counting Kit-8 was employed for cell viability assessment. Reactive oxygen species levels were determined by fluorescence microscopy and flow cytometry. Cell morphology was evaluated by Hoechst staining and transmission electron microscopy. Mitochondrial membrane potential, cellular apoptosis and cell cycle distribution were evaluated flow-cytometrically. The protein levels of apoptotic effectors were examined by Western blot. We found that the photocytotoxicity of MPPa showed both drug- and light- dose dependent characteristics in A549 cells. Additionally, MPPa-PDT caused cell apoptosis by reducing mitochondrial membrane potential, increasing reactive oxygen species (ROS) production, inducing caspase-9/caspase-3 signaling activation as well as cell cycle arrest at G 0 /G 1 phase. These results suggested that MPPa-PDT mainly kills cells by apoptotic mechanisms, with overt curative effects, indicating that MPPa should be considered a potent photosensitizer for lung carcinoma treatment. © 2017 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

  19. Effects of exogenous IL-37 on the biological characteristics of human lung adenocarcinoma A549 cells and the chemotaxis of regulatory T cells.

    PubMed

    Chen, Yu-Hua; Zhou, Bi-Yun; Wu, Guo-Cai; Liao, De-Quan; Li, Jing; Liang, Si-Si; Wu, Xian-Jin; Xu, Jun-Fa; Chen, Yong-Hua; Di, Xiao-Qing; Lin, Qiong-Yan

    2018-02-14

    This study aims to investigate the effects of exogenous interleukin (IL)-37 on the biological characteristics of human lung adenocarcinoma A549 cells and the chemotaxis of regulatory T (Treg) cells. After isolating the CD4+ CD25+ Treg cells from the peripheral blood, flow cytometry was used to detect the purity of the Treg cells. A549 cells were divided into blank (no transfection), empty plasmid (transfection with pIRES2-EGFP empty plasmid) or IL-37 group (transfection with pIRES2-EGFP-IL-37 plasmid). RT-PCR was used to detect mRNA expression of IL-37 and ELISA to determine IL-37 and MMP-9 expressions. Western blotting was applied to detect the protein expressions of PCNA, Ki-67, Cyclin D1, CDK4, cleaved caspase-3 and cleaved caspase-9. MTT assay, flow cytometry, scratch test and transwell assay were performed to detect cell proliferation, cycle, apoptosis, migration and invasion. Effect of exogenous IL-37 on the chemotaxis of Treg cells was measured through transwell assay. Xenograft models in nude mice were eastablished to detect the impact of IL-37 on A549 cells. The IL-37 group had a higher IL-37 expression, cell apoptosis in the early stage and percentage of cells in the G0/G1 phase than the blank and empty plasmid groups. The IL-37 group had a lower MMP-9 expression, optical density (OD), percentage of cells in the S and G2/M phases, migration, invasion and chemotaxis of CD4+CD25+ Foxp3+ Treg cells. The xenograft volume and weight of nude mice in the IL-37 group were lower than those in the blank and empty plasmid groups. Compared with the blank and empty plasmid groups, the IL-37 group had significantly reduced expression of PCNA, Ki-67, Cyclin D1 and CDK4 but elevated expression of cleaved caspase-3 and cleaved caspase-9. Therefore, exogenous IL-37 inhibits the proliferation, migration and invasion of human lung adenocarcinoma A549 cells as well as the chemotaxis of Treg cells while promoting the apoptosis of A549 cells.

  20. Hinokitiol Inhibits Migration of A549 Lung Cancer Cells via Suppression of MMPs and Induction of Antioxidant Enzymes and Apoptosis

    PubMed Central

    Jayakumar, Thanasekaran; Liu, Chao-Hong; Wu, Guan-Yi; Lee, Tzu-Yin; Manubolu, Manjunath; Hsieh, Cheng-Ying; Yang, Chih-Hao; Sheu, Joen-Rong

    2018-01-01

    Hinokitiol, a natural monoterpenoid from the heartwood of Calocedrus formosana, has been reported to have anticancer effects against various cancer cell lines. However, the detailed molecular mechanisms and the inhibiting roles of hinokitiol on adenocarcinoma A549 cells remain to be fully elucidated. Thus, the current study was designed to evaluate the effect of hinokitiol on the migration of human lung adenocarcinoma A549 cells in vitro. The data demonstrates that hinokitiol does not effectively inhibit the viability of A549 cells at up to a 10 µM concentration. When treated with non-toxic doses (1–5 µM) of hinokitiol, the cell migration is markedly suppressed at 5 µM. Hinokitiol significantly reduced p53 expression, followed by attenuation of Bax in A549 cells. A dose-dependent inhibition of activated caspase-9 and -3 was observed in the presence of hinokitiol. An observed increase in protein expression of matrix metalloproteinases (MMPs) -2/-9 in A549 cells was significantly inhibited by hinokitiol. Remarkably, when A549 cells were subjected to hinokitiol (1–5 µM), there was an increase in the activities of antioxidant enzymes catalase (CAT) and superoxide dismutase (SOD) from the reduction in cells. In addition, the incubation of A549 cells with hinokitiol significantly activated the cytochrome c expression, which may be triggered by activation of caspase-9 followed by caspase-3. These observations indicate that hinokitiol inhibited the migration of lung cancer A549 cells through several mechanisms, including the activation of caspases-9 and -3, induction of p53/Bax and antioxidant CAT and SOD, and reduction of MMP-2 and -9 activities. It also induces cytochrome c expression. These findings demonstrate a new therapeutic potential for hinokitiol in lung cancer chemoprevention. PMID:29565268

  1. Hinokitiol Inhibits Migration of A549 Lung Cancer Cells via Suppression of MMPs and Induction of Antioxidant Enzymes and Apoptosis.

    PubMed

    Jayakumar, Thanasekaran; Liu, Chao-Hong; Wu, Guan-Yi; Lee, Tzu-Yin; Manubolu, Manjunath; Hsieh, Cheng-Ying; Yang, Chih-Hao; Sheu, Joen-Rong

    2018-03-22

    Hinokitiol, a natural monoterpenoid from the heartwood of Calocedrus formosana , has been reported to have anticancer effects against various cancer cell lines. However, the detailed molecular mechanisms and the inhibiting roles of hinokitiol on adenocarcinoma A549 cells remain to be fully elucidated. Thus, the current study was designed to evaluate the effect of hinokitiol on the migration of human lung adenocarcinoma A549 cells in vitro. The data demonstrates that hinokitiol does not effectively inhibit the viability of A549 cells at up to a 10 µM concentration. When treated with non-toxic doses (1-5 µM) of hinokitiol, the cell migration is markedly suppressed at 5 µM. Hinokitiol significantly reduced p53 expression, followed by attenuation of Bax in A549 cells. A dose-dependent inhibition of activated caspase-9 and -3 was observed in the presence of hinokitiol. An observed increase in protein expression of matrix metalloproteinases (MMPs) -2/-9 in A549 cells was significantly inhibited by hinokitiol. Remarkably, when A549 cells were subjected to hinokitiol (1-5 µM), there was an increase in the activities of antioxidant enzymes catalase (CAT) and superoxide dismutase (SOD) from the reduction in cells. In addition, the incubation of A549 cells with hinokitiol significantly activated the cytochrome c expression, which may be triggered by activation of caspase-9 followed by caspase-3. These observations indicate that hinokitiol inhibited the migration of lung cancer A549 cells through several mechanisms, including the activation of caspases-9 and -3, induction of p53/Bax and antioxidant CAT and SOD, and reduction of MMP-2 and -9 activities. It also induces cytochrome c expression. These findings demonstrate a new therapeutic potential for hinokitiol in lung cancer chemoprevention.

  2. Enhancement of recombinant myricetin on the radiosensitivity of lung cancer A549 and H1299 cells

    PubMed Central

    2014-01-01

    Objective Myricetin, a common dietary flavonoid is widely distributed in fruits and vegetables, and is used as a health food supplement based on its immune function, anti-oxidation, anti-tumor, and anti-inflammatory properties. The aim of this study was to investigate the effects of myricetin on combination with radiotherapy enhance radiosensitivity of lung cancer A549 and H1299 cells. Methods A549 cells and H1299 cells were exposed to X-ray with or without myricetin treatment. Colony formation assays, CCK-8 assay, flow cytometry and Caspase-3 level detection were used to evaluate the radiosensitization activity of myricetin on cell proliferation and apoptosis in vitro. Nude mouse tumor xenograft model was built to assessed radiosensitization effect of myricetin in vivo. Results Compared with the exposed group without myricetin treatment, the groups treated with myricetin showed significantly suppressed cell surviving fraction and proliferation, increased the cell apoptosis and increased Caspase-3 protein expression after X-ray exposure in vitro. And in vivo assay, growth speed of tumor xenografts was significantly decreased in irradiated mice treated with myricetin. Conclusions The study demonstrated both in vitro and in vivo evidence that combination of myricetin with radiotherapy can enhance tumor radiosensitivity of pulmonary carcinoma A549 and H1299 cells, and myricetin could be a potential radiosensitizer for lung cancer therapy. Virtual slides The virtual slide(s) for this article can be found here: http://www.diagnosticpathology.diagnomx.eu/vs/5791518001210633 PMID:24650056

  3. In vitro effects of nicotine on the non-small-cell lung cancer line A549.

    PubMed

    Gao, Tao; Zhou, Xue-Liang; Liu, Sheng; Rao, Chang-Xiu; Shi, Wen; Liu, Ji-Chun

    2016-04-01

    To investigate in vitro effects of nicotine on the non-small-cell lung cancer line A549. The case-control study was conducted at the First Affiliated Hospital of Nanchang University from 1st January to 30th June, 2014 and comprised A549 cells which were treated with a series of concentrations of nicotine (0.01 µM, 0.1 µM, 1 µM and 10 µM) for 24 hours. Control cells were incubated under the same conditions without the addition of nicotine. Cell growth was detected by monotetrazolium salt [3-(4, 5-dimethyl-2-thiazolyl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay. Cell apoptosis was detected by Haematoxylin and Eosin staining, immunofluorescence analysis of Filamentous actin and electron microscope observation. Nicotine had no significant effect on A549 cell growth at the dose of 0.01µM (p>0.05), but had significant growth inhibitory effects at the doses of 0.1µM, 1µM and 10µM (p< 0.05 each). A significant decrease in cell numbers was observed on staining (p< 0.05). Significant changes in the size and shape of cells and concomitant changes in cytoskeletons and organelles were observed by immunofluorescence and electron microscope observation (p< 0.05). The growth inhibitory effects of nicotine on A549 cells were found to be dose-dependent.

  4. Plasmodium circumsporozoite protein suppresses the growth of A549 cells via inhibiting nuclear transcription factor κB.

    PubMed

    Deng, Xu-Feng; Zhou, Dong; Liu, Quan-Xing; Zheng, Hong; Ding, Yan; Xu, Wen-Yue; Min, Jia-Xin; Dai, Ji-Gang

    2018-05-01

    Blocking the activation of nuclear factor κB (NF-κB) is a promising strategy for the treatment of non-small cell lung cancer. The circumsporozoite protein (CSP), a key component of the sporozoite stage of the malaria parasite, was previously reported to block NF-κB activation in hepatocytes. Therefore, in the present study, the effect of CSP on the growth of the human lung cancer cell line, A549, was investigated. It was demonstrated that transfection with a recombinant plasmid expressing CSP was able to inhibit the proliferation of A549 cells in a dose-dependent manner and induce the apoptosis of A549 cells. A NF-κB gene reporter assay indicated that CSP and its nuclear localization signal (NLS) motif were able to equally suppress the activation of NF-κB following stimulation with human recombinant tumor necrosis factor (TNF)-α in A549 cells. Furthermore, western blot analysis indicated that NLS did not affect the phosphorylation and degradation of IκB, but was able to markedly inhibit the nuclear translocation of NF-κB in TNF-α stimulated A549 cells. Therefore, the data suggest that CSP may be investigated as a potential novel NF-κB inhibitor for the treatment of lung cancer.

  5. Inhibition of lung cancer cells A549 and H460 by curcuminoid extracts and nanoemulsions prepared from Curcuma longa Linnaeus.

    PubMed

    Chang, Hong-Bin; Chen, Bing-Huei

    2015-01-01

    The objectives of this study were to explore the inhibition mechanism of lung cancer cells A549 and H460 by curcuminoid extracts and nanoemulsions prepared from Curcuma longa Linnaeus. In addition, human bronchus epithelial cell line BEAS-2B (normal cell) was selected for comparison. A high-performance liquid chromatography (HPLC) method was developed to separate and quantify the various curcuminoids in C. longa extract, including curcumin (1,714.5 μg/mL), demethoxycurcumin (1,147.4 μg/mL), and bisdemethoxycurcumin (190.2 μg/mL). A high-stability nanoemulsion composed of Tween 80, water, and curcuminoid extract was prepared, with mean particle size being 12.6 nm. The cell cycle was retarded at G2/M for both the curcuminoid extract and nanoemulsion treatments; however, the inhibition pathway may be different. H460 cells were more susceptible to apoptosis than A549 cells for both curcuminoid extract and nanoemulsion treatments. Growth of BEAS-2B remained unaffected for both the curcuminoid extract and nanoemulsion treatments, with a concentration range from 1 to 4 μg/mL. Also, the activities of caspase-3, caspase-8, and caspase-9 followed a dose-dependent increase for both A549 and H460 cells for both the treatments, accompanied by a dose-dependent increase in cytochrome C expression and a dose-dependent decrease in CDK1 expression. Interestingly, a dose-dependent increase in cyclin B expression was shown for A549 cells for both the treatments, while a reversed trend was found for H460 cells. Both mitochondria and death receptor pathways may be responsible for apoptosis of both A549 and H460 cells.

  6. Hyaluronic acid-fabricated nanogold delivery of the inhibitor of apoptosis protein-2 siRNAs inhibits benzo[a]pyrene-induced oncogenic properties of lung cancer A549 cells

    NASA Astrophysics Data System (ADS)

    Lin, Chung-Ming; Kao, Wei-Chien; Yeh, Chun-An; Chen, Hui-Jye; Lin, Shinn-Zong; Hsieh, Hsien-Hsu; Sun, Wei-Shen; Chang, Chih-Hsuan; Hung, Huey-Shan

    2015-03-01

    Benzo[a]pyrene (BaP), a component of cooking oil fumes (COF), promotes lung cancer cell proliferation and survival via the induction of inhibitor of apoptosis protein-2 (IAP-2) proteins. Thus knockdown of IAP-2 would be a promising way to battle against lung cancer caused by COF. Functionalized gold nanoparticle (AuNP) is an effective delivery system for bio-active materials. Here, biocompatible hyaluronic acid (HA) was fabricated into nanoparticles to increase the target specificity by binding to CD44-over-expressed cancer cells. IAP-2-specific small-interfering RNA (siRNAs) or fluorescein isothiocyanate (FITC) were then incorporated into AuNP-HA. Conjugation of IAP-2 siRNA into AuNPs-HA was verified by the UV-vis spectrometer and Fourier transform infrared spectrometer. Further studies showed that AuNP-HA/FITC were effectively taken up by A549 cells through CD44-mediated endocytosis. Incubation of BaP-challenged cells with AuNP-HA-IAP-2 siRNAs silenced the expression of IAP-2, decreased cell proliferation and triggered pronounced cell apoptosis by the decrease in Bcl-2 protein and the increase in Bax protein as well as the active form of caspases-3. The BaP-elicited cell migration and enzymatic activity of the secreted matrix metalloproteinase-2 were also substantially suppressed by treatment with AuNP-HA-IAP-2 siRNAs. These results indicated that IAP-2 siRNAs can be efficiently delivered into A549 cells by functionalized AuNP-HA to repress the IAP-2 expression and BaP-induced oncogenic events, suggesting the potential therapeutic application of IAP-2 siRNA or other siRNA-conjugated AuNP-HA composites to COF-induced lung cancer and other gene-caused diseases in the future.

  7. Calotropin from Asclepias curasavica induces cell cycle arrest and apoptosis in cisplatin-resistant lung cancer cells.

    PubMed

    Mo, En-Pan; Zhang, Rong-Rong; Xu, Jun; Zhang, Huan; Wang, Xiao-Xiong; Tan, Qiu-Tong; Liu, Fang-Lan; Jiang, Ren-Wang; Cai, Shao-Hui

    2016-09-16

    Calotropin (M11), an active compound isolated from Asclepias curasavica L., was found to exert strong inhibitory and pro-apoptotic activity specifically against cisplatin-induced resistant non-small cell lung cancer (NSCLC) cells (A549/CDDP). Molecular mechanism study revealed that M11 induced cell cycle arrest at the G2/M phase through down-regulating cyclins, CDK1, CDK2 and up-regulating p53 and p21. Furthermore, M11 accelerated apoptosis through the mitochondrial apoptotic pathway which was accompanied by increase Bax/Bcl-2 ratio, decrease in mitochondrial membrane potential, increase in reactive oxygen species production, activations of caspases 3 and 9 as well as cleavage of poly ADP-ribose polymerase (PARP). The activation and phosphorylation of JNK was also found to be involved in M11-induced apoptosis, and SP610025 (specific JNK inhibitor) partially prevented apoptosis induced by M11. In contrast, all of the effects that M11 induce cell cycle arrest and apoptosis in A549/CDDP cells were not significant in A549 cells. Drugs with higher sensitivity against resistant tumor cells than the parent cells are rather rare. Results of this study supported the potential application of M11 on the non-small lung cancer (NSCLC) with cisplatin resistance. Copyright © 2016 Elsevier Inc. All rights reserved.

  8. The Chromone Alkaloid, Rohitukine, Affords Anti-Cancer Activity via Modulating Apoptosis Pathways in A549 Cell Line and Yeast Mitogen Activated Protein Kinase (MAPK) Pathway

    PubMed Central

    Safia; Kamil, Mohd; Jadiya, Pooja; Sheikh, Saba; Haque, Ejazul; Nazir, Aamir; Lakshmi, Vijai; Mir, Snober S.

    2015-01-01

    The field of cancer research and treatment has made significant progress, yet we are far from having completely safe, efficient and specific therapies that target cancer cells and spare the healthy tissues. Natural compounds may reduce the problems related to cancer treatment. Currently, many plant products are being used to treat cancer. In this study, Rohitukine, a natural occurring chromone alkaloid extracted from Dysoxylum binectariferum, was investigated for cytotoxic properties against budding yeast as well as against lung cancer (A549) cells. We endeavored to specifically study Rohitukine in S. cerevisiae in the context of MAPK pathways as yeast probably represents the experimental model where the organization and regulation of MAPK pathways are best understood. MAPK are evolutionarily conserved protein kinases that transfer extracellular signals to the machinery controlling essential cellular processes like growth, migration, differentiation, cell division and apoptosis. We aimed at carrying out hypothesis driven studies towards targeting the important network of cellular communication, a critical process that gets awry in cancer. Employing mutant strains of genetic model system Saccharomyces cerevisiae. S. cerevisiae encodes five MAPKs involved in control of distinct cellular responses such as growth, differentiation, migration and apoptosis. Our study involves gene knockouts of Slt2 and Hog1 which are functional homologs of human ERK5 and mammalian p38 MAPK, respectively. We performed cytotoxicity assay to evaluate the effect of Rohitukine on cell viability and also determined the effects of drug on generation of reactive oxygen species, induction of apoptosis and expression of Slt2 and Hog1 gene at mRNA level in the presence of drug. The results of this study show a differential effect in the activity of drug between the WT, Slt2 and Hog1 gene deletion strain indicating involvement of MAPK pathway. Further, we investigated Rohitukine induced cytotoxic

  9. Inhibition of lung cancer cells A549 and H460 by curcuminoid extracts and nanoemulsions prepared from Curcuma longa Linnaeus

    PubMed Central

    Chang, Hong-Bin; Chen, Bing-Huei

    2015-01-01

    The objectives of this study were to explore the inhibition mechanism of lung cancer cells A549 and H460 by curcuminoid extracts and nanoemulsions prepared from Curcuma longa Linnaeus. In addition, human bronchus epithelial cell line BEAS-2B (normal cell) was selected for comparison. A high-performance liquid chromatography (HPLC) method was developed to separate and quantify the various curcuminoids in C. longa extract, including curcumin (1,714.5 μg/mL), demethoxycurcumin (1,147.4 μg/mL), and bisdemethoxycurcumin (190.2 μg/mL). A high-stability nanoemulsion composed of Tween 80, water, and curcuminoid extract was prepared, with mean particle size being 12.6 nm. The cell cycle was retarded at G2/M for both the curcuminoid extract and nanoemulsion treatments; however, the inhibition pathway may be different. H460 cells were more susceptible to apoptosis than A549 cells for both curcuminoid extract and nanoemulsion treatments. Growth of BEAS-2B remained unaffected for both the curcuminoid extract and nanoemulsion treatments, with a concentration range from 1 to 4 μg/mL. Also, the activities of caspase-3, caspase-8, and caspase-9 followed a dose-dependent increase for both A549 and H460 cells for both the treatments, accompanied by a dose-dependent increase in cytochrome C expression and a dose-dependent decrease in CDK1 expression. Interestingly, a dose-dependent increase in cyclin B expression was shown for A549 cells for both the treatments, while a reversed trend was found for H460 cells. Both mitochondria and death receptor pathways may be responsible for apoptosis of both A549 and H460 cells. PMID:26345201

  10. Lipocalin 2, a new GADD153 target gene, as an apoptosis inducer of endoplasmic reticulum stress in lung cancer cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Hsin, I-Lun; Hsiao, Yueh-Chieh; Institute of Medicine, Chung Shan Medical University, Taichung 40201, Taiwan

    2012-09-15

    Endoplasmic reticulum (ER) stress is activated under severe cellular conditions. GADD153, a member of the C/EBP family, is an unfolded protein response (UPR) responsive transcription factor. Increased levels of lipocalin 2, an acute phase protein, have been found in several epithelial cancers. The aim of this study is to investigate the function of lipocalin 2 in lung cancer cells under ER stress. Treatment with thapsigargin, an ER stress activator, led to increases in cytotoxicity, ER stress, apoptosis, and lipocalin 2 expression in A549 cells. GADD153 silencing decreased lipocalin 2 expression in A549 cells. On chromatin immunoprecipitation assay, ER stress increasedmore » GADD153 DNA binding to lipocalin 2 promoter. Furthermore, silencing of lipocalin 2 mitigated ER stress-mediated apoptosis in A549 cells. Our findings demonstrated that lipocalin 2 is a new GADD153 target gene that mediates ER stress-induced apoptosis. Highlights: ► We demonstrate that Lipocalin 2 is a new GADD153 target gene. ► Lipocalin 2 mediates ER stress-induced apoptosis. ► ER stress-induced lipocalin 2 expression is calcium-independent in A549 cells. ► Lipocalin 2 dose not play a major role in ER stress-induced autophagy.« less

  11. Genistein decreases A549 cell viability via inhibition of the PI3K/AKT/HIF‑1α/VEGF and NF‑κB/COX‑2 signaling pathways.

    PubMed

    Zhang, Juan; Su, Hongzheng; Li, Qingfeng; Li, Jing; Zhao, Qianfeng

    2017-04-01

    Genistein is an important chemopreventive agent against atherosclerosis and cancer. However, whether genistein is effective in the treatment of lung cancer, and its underlying mechanism, remains to be determined. The present study demonstrated that genistein treatment of A549 lung cancer cells decreased viability in a dose‑ and time‑dependent manner, and induced apoptosis. Additionally, A549 cells exhibited significantly increased reactive oxygen species formation and cytochrome‑c leakage, and activated caspase‑3, B‑cell lymphoma 2‑associated X protein and apoptosis inducing factor expression levels, which are involved in the mitochondrial apoptosis pathway. Furthermore, the phosphatidylinositol‑4,5‑biphosphate 3‑kinase (PI3K)/protein kinase B (AKT)/hypoxia‑inducible factor‑1α (HIF‑1α) and nuclear factor‑κB (NF‑κB)/cyclooxygenase‑2 (COX‑2) signaling pathways were significantly downregulated by genistein treatment. In conclusion, reduced proliferation and increased apoptosis in A549 lung cancer cells was associated with inhibition of the PI3K/AKT/HIF‑1α/ and NF‑κB/COX‑2 signaling pathways, which implicates genistein as a potential chemotherapeutic agent for the treatment of lung cancer.

  12. The antitumor effect of tanshinone IIA on anti-proliferation and decreasing VEGF/VEGFR2 expression on the human non-small cell lung cancer A549 cell line.

    PubMed

    Xie, Jun; Liu, Jiahui; Liu, Heng; Liang, Shihui; Lin, Meigui; Gu, Yueyu; Liu, Taoli; Wang, Dongmei; Ge, Hui; Mo, Sui-Lin

    2015-11-01

    The effects of tanshinone IIA on the proliferation of the human non-small cell lung cancer cell line A549 and its possible mechanism on the VEGF/VEGFR signal pathway were investigated. The exploration of the interaction between tanshinone IIA and its target proteins provides a feasible platform for studying the anticancer mechanism of active components of herbs. The CCK-8 assay was used to evaluate the proliferative activity of A549 cells treated with tanshinone IIA (2.5-80 μmol/L) for 24, 48 and 72 h, respectively. Flow cytometry was used for the detection of cell apoptosis and cell cycle perturbation. VEGF and VEGFR2 expression were studied by Western blotting. The binding mode of tanshinone IIA within the crystal structure of the VEGFR2 protein was evaluated with molecular docking analysis by use of the CDOCKER algorithm in Discovery Studio 2.1. The CCK-8 results showed that tanshinone IIA can significantly inhibit A549 cell proliferation in a dose- and time-dependent manner. Flow cytometry results showed that the apoptosis rate of tested group was higher than the vehicle control, and tanshinone IIA-treated cells accumulated at the S phase, which was higher than the vehicle control. Furthermore, the expression of VEGF and VEGFR2 was decreased in Western blot. Finally, molecular docking analysis revealed that tanshinone IIA could be stably docked into the kinase domain of VEGFR2 protein with its unique modes to form H-bonds with Cys917 and π-π stacking interactions with Val848. In conclusion, tanshinone IIA may suppress A549 proliferation, induce apoptosis and cell cycle arrest at the S phase. This drug may suppress angiogenesis by targeting the protein kinase domains of VEGF/VEGFR2.

  13. The antitumor effect of tanshinone IIA on anti-proliferation and decreasing VEGF/VEGFR2 expression on the human non-small cell lung cancer A549 cell line

    PubMed Central

    Xie, Jun; Liu, Jiahui; Liu, Heng; Liang, Shihui; Lin, Meigui; Gu, Yueyu; Liu, Taoli; Wang, Dongmei; Ge, Hui; Mo, Sui-lin

    2015-01-01

    The effects of tanshinone IIA on the proliferation of the human non-small cell lung cancer cell line A549 and its possible mechanism on the VEGF/VEGFR signal pathway were investigated. The exploration of the interaction between tanshinone IIA and its target proteins provides a feasible platform for studying the anticancer mechanism of active components of herbs. The CCK-8 assay was used to evaluate the proliferative activity of A549 cells treated with tanshinone IIA (2.5−80 μmol/L) for 24, 48 and 72 h, respectively. Flow cytometry was used for the detection of cell apoptosis and cell cycle perturbation. VEGF and VEGFR2 expression were studied by Western blotting. The binding mode of tanshinone IIA within the crystal structure of the VEGFR2 protein was evaluated with molecular docking analysis by use of the CDOCKER algorithm in Discovery Studio 2.1. The CCK-8 results showed that tanshinone IIA can significantly inhibit A549 cell proliferation in a dose- and time-dependent manner. Flow cytometry results showed that the apoptosis rate of tested group was higher than the vehicle control, and tanshinone IIA-treated cells accumulated at the S phase, which was higher than the vehicle control. Furthermore, the expression of VEGF and VEGFR2 was decreased in Western blot. Finally, molecular docking analysis revealed that tanshinone IIA could be stably docked into the kinase domain of VEGFR2 protein with its unique modes to form H-bonds with Cys917 and π–π stacking interactions with Val848. In conclusion, tanshinone IIA may suppress A549 proliferation, induce apoptosis and cell cycle arrest at the S phase. This drug may suppress angiogenesis by targeting the protein kinase domains of VEGF/VEGFR2. PMID:26713270

  14. Cytotoxicity and gene expression profiling of polyhexamethylene guanidine hydrochloride in human alveolar A549 cells.

    PubMed

    Jung, Ha-Na; Zerin, Tamanna; Podder, Biswajit; Song, Ho-Yeon; Kim, Yong-Sik

    2014-06-01

    In Korea, lung disease of children and pregnant women associated with humidifier disinfectant use has become a major concern. A common sterilizer is polyhexamethylene guanidine (PHMG), a member of the guanidine family of antiseptics. This study was done to elucidate the putative cytotoxic effect of PHMG and the PHMG-mediated altered gene expression in human alveolar epithelial A549 cells in vitro. Cell viability analyses revealed the potent cytotoxicity of PHMG, with cell death evident at as low as 5 μg/mL. Death was dose- and time-dependent, and was associated with formation of intracellular reactive oxygen species, and apoptosis significantly, at even 2 μg/mL concentration. The gene expression profile in A549 cells following 24 h exposure to 5 μg/mL of PHMG was investigated using DNA microarray analysis. Changes in gene expression relevant to the progression of cell death included induction of genes related to apoptosis, autophagy, fibrosis, and cell cycle. However, the expressions of genes encoding antioxidant and detoxifying enzymes were down-regulated or not affected. The altered expression of selected genes was confirmed by quantitative reverse transcription-polymerase chain reaction and Western blot analyses. The collective data suggest that PHMG confers cellular toxicity through the generation of intracellular reactive oxygen species and alteration of gene expression. Copyright © 2014 Elsevier Ltd. All rights reserved.

  15. Methyl methanesulfonate induces necroptosis in human lung adenoma A549 cells through the PIG-3-reactive oxygen species pathway.

    PubMed

    Jiang, Ying; Shan, Shigang; Chi, Linfeng; Zhang, Guanglin; Gao, Xiangjing; Li, Hongjuan; Zhu, Xinqiang; Yang, Jun

    2016-03-01

    Methyl methanesulfonate (MMS) is an alkylating agent that can induce cell death through apoptosis and necroptosis. The molecular mechanisms underlying MMS-induced apoptosis have been studied extensively; however, little is known about the mechanism for MMS-induced necroptosis. Therefore, we first established MMS-induced necroptosis model using human lung carcinoma A549 cells. It was found that, within a 24-h period, although MMS at concentrations of 50, 100, 200, 400, and 800 μM can induce DNA damage, only at higher concentrations (400 and 800 μM) MMS treatment lead to necroptosis in A549 cells, as it could be inhibited by the specific necroptotic inhibitor necrostatin-1, but not the specific apoptotic inhibitor carbobenzoxy-valyl-alanyl-aspartyl-[O-methyl]-fluoromethylketone (Z-VAD-fmk). MMS-induced necroptosis was further confirmed by the induction of the necroptosis biomarkers including the depletion of cellular NADH and ATP and leakage of LDH. This necroptotic cell death was also concurrent with the increased expression of p53, p53-induced gene 3 (PIG-3), high mobility group box-1 protein (HMGB1), and receptor interaction protein kinase (RIP) but not the apoptosis-associated caspase-3 and caspase-9 proteins. Elevated reactive oxygen species (ROS) level was also involved in this process as the specific ROS inhibitor (4-amino-2,4-pyrrolidine-dicarboxylic acid (APDC)) can inhibit the necroptotic cell death. Interestingly, knockdown of PIG-3 expression by small interfering RNA (siRNA) treatment can inhibit the generation of ROS. Taken together, these results suggest that MMS can induce necroptosis in A549 cells, probably through the PIG-3-ROS pathway.

  16. A methoxyflavanone derivative from the Asian medicinal herb (Perilla frutescens) induces p53-mediated G2/M cell cycle arrest and apoptosis in A549 human lung adenocarcinoma.

    PubMed

    Abd El-Hafeez, Amer Ali; Fujimura, Takashi; Kamei, Rikiya; Hirakawa, Noriko; Baba, Kenji; Ono, Kazuhisa; Kawamoto, Seiji

    2017-07-14

    Perilla frutescens is an Asian dietary herb consumed as an essential seasoning in Japanese cuisine as well as used for a Chinese medicine. Here, we report that a newly found methoxyflavanone derivative from P. frutescens (Perilla-derived methoxyflavanone, PDMF; 8-hydroxy-5,7-dimethoxyflavanone) shows carcinostatic activity on human lung adenocarcinoma, A549. We found that treatment with PDMF significantly inhibited cell proliferation and decreased viability through induction of G 2 /M cell cycle arrest and apoptosis. The PDMF stimulation induces phosphorylation of tumor suppressor p53 on Ser15, and increases its protein amount in conjunction with up-regulation of downstream cyclin-dependent kinase inhibitor p21 Cip1/Waf1 and proapoptotic caspases, caspase-9 and caspase-3. We also found that small interfering RNA knockdown of p53 completely abolished the PDMF-induced G 2 /M cell cycle arrest, and substantially abrogated its proapoptotic potency. These results suggest that PDMF represents a useful tumor-preventive phytochemical that triggers p53-driven G 2 /M cell cycle arrest and apoptosis.

  17. Crocidolite asbestos causes an induction of p53 and apoptosis in cultured A-549 lung carcinoma cells.

    PubMed

    Pääkkö, P; Rämet, M; Vähäkangas, K; Korpela, N; Soini, Y; Turunen, S; Jaworska, M; Gillissen, A

    1998-01-01

    A number of genotoxic chemicals and agents, such as benzo(a)pyrene and ultraviolet light, are able to induce nuclear accumulation of p53 protein. Usually, this response is transient and a consequence of stabilization of the wild-type p53 protein. After withdrawal of the exposure, the amount of p53 protein returns to a normal level within hours or a few days. We have studied the p53 response to the exposure of crocidolite asbestos in A-549 lung carcinoma cells using three different methods, i.e., p53 immunohistochemistry, Western blotting and metabolic labelling followed by p53 immunoprecipitation. With these techniques we demonstrate a dose-dependent p53 nuclear response to crocidolite exposure. The half-life of p53 protein in A-549 lung carcinoma cells cultured in serum-free media increased from 30 up to 80 min, and the protein reacted with a wild-type specific antibody suggesting that it was in a wild-type conformation. In situ 3'-end labelling of A-549 cells demonstrated a dose-dependent increase in apoptotic activity. Our data support the idea that increased apoptotic activity, induced by crocidolite, is mediated by p53.

  18. A novel herbal formula induces cell cycle arrest and apoptosis in association with suppressing the PI3K/AKT pathway in human lung cancer A549 cells.

    PubMed

    Xiong, Fei; Jiang, Miao; Huang, Zhenzhou; Chen, Meijuan; Chen, Kejun; Zhou, Jing; Yin, Lian; Tang, Yuping; Wang, Mingyan; Ye, Lihong; Zhan, Zhen; Duan, Jinao; Fu, Haian; Zhang, Xu

    2014-03-01

    In recent years, the incidence of lung cancer, as well as the mortality rate from this disease, has increased. Moreover, because of acquired drug resistance and adverse side effects, the effectiveness of current therapeutics used for the treatment of lung cancer has decreased significantly. Chinese medicine has been shown to have significant antitumor effects and is increasingly being used for the treatment of cancer. However, as the mechanisms of action for many Chinese medicines are undefined, the application of Chinese medicine for the treatment of cancer is limited. The formula tested has been used clinically by the China National Traditional Chinese Medicine Master, Professor Zhonging Zhou for treatment of cancer. In this article, we examine the efficacy of Ke formula in the treatment of non-small cell lung cancer and elucidate its mechanism of action. A Balb/c nude mouse xenograft model using A549 cells was previously established. The mice were randomly divided into normal, mock, Ke, cisplatin (DDP), and co-formulated (Ke + DDP) groups. After 15 days of drug administration, the animals were sacrificed, body weight and tumor volume were recorded, and the tumor-inhibiting rate was calculated. A cancer pathway finder polymerase chain reaction array was used to monitor the expression of 88 genes in tumor tissue samples. The potential antiproliferation mechanism was also investigated by Western blot analysis. Ke formula minimized chemotherapy-related weight loss in tumor-bearing mice without exhibiting distinct toxicity. Ke formula also inhibited tumor growth, which was associated with the downregulation of genes in the PI3K/AKT, MAPK, and WNT/β-catenin pathways. The results from Western blot analyses further indicated that Ke blocked the cell cycle progression at the G1/S phase and induced apoptosis mainly via the PI3K/AKT pathway. Ke formula inhibits tumor growth in an A549 xenograft mouse model with no obvious side effects. Moreover, Ke exhibits synergistic

  19. Effects of SASH1 on lung cancer cell proliferation, apoptosis, and invasion in vitro.

    PubMed

    Chen, En-guo; Chen, Yanfan; Dong, Liang-liang; Zhang, Ji-song

    2012-10-01

    The purposes of this study were to investigate the effects of the SASH1 gene on the growth, proliferation, apoptosis, invasiveness, and metastatic potential of lung cancer cells and explore the potential use of SASH1 for the treatment of human lung cancer. The SASH1 gene was cloned into the pcDNA3.1 eukaryotic expression vector, and SASH1 shRNA were designed and constructed. The resulting constructs were transfected into A549 human lung cancer cells, and the changes in the relevant biological characteristics of the cells overexpressing SASH1 and cells with downregulated expression of SASH1 were analyzed using the MTT assay, transwell invasion assay, and flow cytometry. The effects of the SASH1 gene on the expression of cyclin D1, Bcl-2, and MMP-2/9 were also concurrently examined. In the A549 cells from the pcDNA3.1-SASH1 transfected group, cell viability, proliferation, and migration were significantly reduced compared to the control cells (p = 0.039, p = 0.013), and a cell cycle arrest in G1 was observed. The A549 cells transfected with the SASH1 shRNA demonstrated significantly higher cell viabilities, proliferation, and migration compared to the control cells (p = 0.012, p = 0.045). Additionally, the percentage of A549 cells undergoing apoptosis was significantly higher in the pcDNA3.1-SASH1 transfected cells and significantly lower in the SASH1 shRNA transfected cells compared to the control cells (p = 0.010, p = 0.000). The cyclin D1, Bcl-2, and MMP-9/2 protein expression levels were significantly lower in the pcDNA3.1-SASH1-transfected cells and were significantly higher in the SASH1 shRNA-transfected cells than that in the control cells. The SASH1 gene may inhibit A549 cell growth and proliferation as well as promote cellular apoptosis. The overexpression of the SASH1 gene may also be related to the decreased migration of A549 human lung cancer cells.

  20. Umbelliprenin is cytotoxic against QU-DB large cell lung cancer cell line but anti-proliferative against A549 adenocarcinoma cells

    PubMed Central

    2012-01-01

    Background Umbelliprenin is a natural compound, belonging to the class of sesquiterpene coumarins. Recently, umbelliprenin has attracted the researchers' attention for its antitumor activities against skin tumors. Its effect on lung cancer is largely unknown. The aim of our study was to investigate the effects of this natural compound, which is expected to have low adverse effects, on lung cancer. Methods The QU-DB large cell and A549 adenocarcinoma lung cancer cell lines were treated with umbelliprenin. IC50 values were estimated using methyl thiazolely diphenyl-tetrazolium bromide (MTT) assay, in which a decrease in MTT reduction can occur as a result of cell death or cell proliferation inhibition. To quantify the rate of cell death at IC50 values, flow cytometry using Annexin V-FITC (for apoptotic cells), and propidium iodide (for necrotic cells) dyes were employed. Results Data from three independent MTT experiments in triplicate revealed that IC50 values for QU-DB and A549 were 47 ± 5.3 μM and 52 ± 1.97 μM, respectively. Annexin V/PI staining demonstrated that umbelliprenin treatment at IC50 induced 50% cell death in QU-DB cells, but produced no significant death in A549 cells until increasing the umbelliprenin concentration to IC80. The pattern of cell death was predominantly apoptosis in both cell lines. When peripheral blood mononuclear cells were treated with 50 μM and less concentrations of umbelliprenin, no suppressive effect was observed. Conclusions We found cytotoxic/anti-proliferative effects of umbelliprenin against two different types of lung cancer cell lines. PMID:23351548

  1. Integrated High Throughput Analysis Identifies GSK3 as a Crucial Determinant of p53-Mediated Apoptosis in Lung Cancer Cells.

    PubMed

    Zhang, Yu; Zhu, Chenyang; Sun, Bangyao; Lv, Jiawei; Liu, Zhonghua; Liu, Shengwang; Li, Hai

    2017-01-01

    p53 dysfunction is frequently observed in lung cancer. Although restoring the tumour suppressor function of p53 is recently approved as a putative strategy for combating cancers, the lack of understanding of the molecular mechanism underlying p53-mediated lung cancer suppression has limited the application of p53-based therapies in lung cancer. Using RNA sequencing, we determined the transcriptional profile of human non-small cell lung carcinoma A549 cells after treatment with two p53-activating chemical compounds, nutlin and RITA, which could induce A549 cell cycle arrest and apoptosis, respectively. Bioinformatics analysis of genome-wide gene expression data showed that distinct transcription profiles were induced by nutlin and RITA and 66 pathways were differentially regulated by these two compounds. However, only two of these pathways, 'Adherens junction' and 'Axon guidance', were found to be synthetic lethal with p53 re-activation, as determined via integrated analysis of genome-wide gene expression profile and short hairpin RNA (shRNA) screening. Further functional protein association analysis of significantly regulated genes associated with these two synthetic lethal pathways indicated that GSK3 played a key role in p53-mediated A549 cell apoptosis, and then gene function study was performed, which revealed that GSK3 inhibition promoted p53-mediated A549 cell apoptosis in a p53 post-translational activity-dependent manner. Our findings provide us with new insights regarding the mechanism by which p53 mediates A549 apoptosis and may cast light on the development of more efficient p53-based strategies for treating lung cancer. © 201 The Author(s). Published by S. Karger AG, Basel.

  2. Blockage of glycolysis by targeting PFKFB3 alleviates sepsis-related acute lung injury via suppressing inflammation and apoptosis of alveolar epithelial cells.

    PubMed

    Gong, Yuanqi; Lan, Haibing; Yu, Zhihong; Wang, Meng; Wang, Shu; Chen, Yu; Rao, Haiwei; Li, Jingying; Sheng, Zhiyong; Shao, Jianghua

    2017-09-16

    Sepsis-related acute lung injury (ALI) is characterized by excessive lung inflammation and apoptosis of alveolar epithelial cells resulting in acute hypoxemic respiratory failure. Recent studies indicated that anaerobic glycolysis play an important role in sepsis. However, whether inhibition of aerobic glycolysis exhibits beneficial effect on sepsis-induced ALI is not known. In vivo, a cecal ligation and puncture (CLP)-induced ALI mouse model was set up and mice treated with glycolytic inhibitor 3PO after CLP. The mice treated with the 3PO ameliorated the survival rate, histopathological changes, lung inflammation, lactate increased and lung apoptosis of mice with CLP-induced sepsis. In vitro, the exposure of human alveolar epithelial A549 cells to lipopolysaccharide (LPS) resulted in cell apoptosis, inflammatory cytokine production, enhanced glycolytic flux and reactive oxygen species (ROS) increased. While these changes were attenuated by 3PO treatment. Sequentially, treatment of A549 cells with lactate caused cell apoptosis and enhancement of ROS. Pretreatment with N-acetylcysteine (NAC) significantly lowered LPS and lactate-induced the generation of ROS and cell apoptosis in A549 cells. Therefore, these results indicate that anaerobic glycolysis may be an important contributor in cell apoptosis of sepsis-related ALI. Moreover, LPS specifically induces apoptotic insults to A549 cell through lactate-mediated enhancement of ROS. Copyright © 2017 Elsevier Inc. All rights reserved.

  3. White Tea Extract Induces Apoptosis in Nonsmall Cell Lung Cancer Cells– The Role of PPAR-γ and 15-Lipoxygenases

    PubMed Central

    Mao, Jenny T.; Nie, Wen-Xian; Tsu, I-Hsien; Jin, Yu-Sheng; Rao, Jian yu; Lu, Qing-Yi; Zhang, Zuo-Feng; Go, Vay Liang W.; Serio, Kenneth J.

    2010-01-01

    Purpose Emerging preclinical data suggests that tea possess anticarcinogenic and antimutagenic properties. We therefore hypothesize that white tea extract (WTE) is capable of favorably modulating apoptosis, a mechanism associated with lung tumorigenesis. Experimental Design We examined the effects of physiologically relevant doses of WTE on the induction of apoptosis in the nonsmall cell lung cancer (NSCLC) cell lines, A549 (adenocarcinoma) and H520 (squamous cell carcinoma) cells. We further characterized the molecular mechanisms responsible for the WTE-induced apoptosis, including the induction of PPAR-γ and the 15-lipoxygenase (15-LOX) signaling pathway. Results We found that WTE was effective in inducing apoptosis in both A549 and H520 cells, and inhibition of PPAR-γ with GW 9662 partially reversed the WTE-induced apoptosis. We further demonstrate that WTE increased PPAR-γ activation and mRNA expression, concomitantly increased 15-HETE release, and up-regulated 15-LOX-1 and 2 mRNA expression by A549 cells. Inhibition of 15-LOX with NGDA, as well as caffeic acid, abrogated the WTE-induced PPAR-γ activation and up-regulation of PPAR-γ mRNA expression in A549 cells. WTE also induced cyclin-dependent kinase inhibitor 1A (CDKN1A) mRNA expression and activated caspase 3. Inhibition of caspase 3 abrogated the WTE-induced apoptosis. Conclusions Our findings indicate that WTE is capable of inducing apoptosis in NSCLC cell lines. The induction of apoptosis appears to be mediated, in part, through the up-regulation of the PPAR-γ and 15-LOX signaling pathways, with enhanced activation of caspase 3. Our findings support the future investigation of WTE as an antineoplastic and chemopreventive agent for lung cancer. PMID:20668019

  4. MLKL-PITPα signaling-mediated necroptosis contributes to cisplatin-triggered cell death in lung cancer A549 cells.

    PubMed

    Jing, Lin; Song, Fei; Liu, Zhenyu; Li, Jianghua; Wu, Bo; Fu, Zhiguang; Jiang, Jianli; Chen, Zhinan

    2018-02-01

    Necroptosis has been reported to be involved in cisplatin-induced cell death, but the mechanisms underlying the occurrence of necroptosis are not fully elucidated. In this study, we show that apart from apoptosis, cisplatin induces necroptosis in A549 cells. The alleviation of cell death by two necroptosis inhibitors-necrostatin-1 (Nec-1) and necrosulfonamide (NSA), and the phosphorylation of mixed lineage kinase domain-like protein (MLKL) at serine 358, suggest the involvement of receptor-interacting protein kinase 1 (RIPK1)-RIPK3-MLKL signaling in cisplatin-treated A549 cells. Additionally, the initiation of cisplatin-induced necroptosis relies on autocrine tumor necrosis factor alpha (TNF-α). Furthermore, we present the first evidence that phosphatidylinositol transfer protein alpha (PITPα) is involved in MLKL-mediated necroptosis by interacting with the N terminal MLKL on its sixth helix and the preceding loop, which facilitates MLKL oligomerization and plasma membrane translocation in necroptosis. Silencing of PITPα expression interferes with MLKL function and reduces cell death. Our data elucidate that cisplatin-treated lung cancer cells undergo a new type of programmed cell death called necroptosis and shed new light on how MLKL translocates to the plasma membrane. Copyright © 2017 Elsevier B.V. All rights reserved.

  5. Cadmium induces oxidative stress and apoptosis in lung epithelial cells.

    PubMed

    Kiran Kumar, K M; Naveen Kumar, M; Patil, Rajeshwari H; Nagesh, Rashmi; Hegde, Shubha M; Kavya, K; Babu, R L; Ramesh, Govindarajan T; Sharma, S Chidananda

    2016-11-01

    Cadmium (Cd) is one of the well-known highly toxic environmental and industrial pollutants. Cd first accumulates in the nucleus and later interacts with zinc finger proteins of antiapoptotic genes and inhibit the binding of transcriptional factors and transcription. However, the role of Cd in oxidative stress and apoptosis is less understood. Hence, the present study was undertaken to unveil the mechanism of action. A549 cells were treated with or without Cd and cell viability was measured by MTT assay. Treatment of cells with Cd shows reduced viability in a dose-dependent manner with IC 50 of 45 μM concentration. Cd significantly induces the reactive oxygen species (ROS), lipid peroxidation followed by membrane damage with the leakage of lactate dehydrogenase (LDH). Cells with continuous exposure of Cd deplete the antioxidant super oxide dismutase (SOD) and glutathione peroxidase (GSH-Px) enzymes. Further, analysis of the expression of genes involved in apoptosis show that both the extrinsic and intrinsic apoptotic pathways were involved. Death receptor marker tumor necrosis factor-α (TNF-α), executor caspase-8 and pro-apoptotic gene (Bax) were induced, while antiapoptotic gene (Bcl-2) was decreased in Cd-treated cells. Fluorescence-activated cell sorting (FACS) analysis further confirms the induction of apoptosis in Cd-treated A549 cells.

  6. Cytotoxic, Antiproliferative and Apoptotic Effects of Perillyl Alcohol and Its Biotransformation Metabolite on A549 and HepG2 Cancer Cell Lines.

    PubMed

    Oturanel, Ceren E; Kıran, İsmail; Özşen, Özge; Çiftçi, Gülşen A; Atlı, Özlem

    2017-01-01

    A monoterpene, perillyl alcohol, has attracted attention in medicinal chemistry since it exhibited chemo-preventive and therapeutic properties against a variety of cancers. In the present work, it was aimed to obtain derivatives of perillyl alcohol through microbial biotransformation and investigate their anticancer activities against A549 and HepG2 cancer cell lines. Biotransformation studies were carried out in a α-medium for 7 days at 25oC. XTT assay was performed to investigate the anticancer activities of perillyl alcohol and its biotransformation metabolite, dehydroperillic acid, against A549 and HepG2 cell lines and their selectivity using healthy cell line, NIH/3T3. Cell proliferation ELISA, BRDU (colorimetric) assay was used for measurement of proliferation in replicative cells in which DNA synthesis occurs. Flow cytometric analyses were also carried out for measuring apoptotic cell percentages, caspase 3 activation and mitochondrial membrane potential. Biotransformation of perillyl alcohol with Fusarium culmorum yielded dehydroperillic acid in a yield of 20.4 %. In in vitro anticancer studies, perillyl alcohol was found to exert cytotoxicity against HepG2 cell line with an IC50 value of 409.2 μg/mL. However, this effect was not found to be selective because of its higher IC50 (250 μg/mL) value against NIH/3T3 cell line. On the other hand, dehydroperillic acid was found to be effective and also selective against A549 cell line with an IC50 value of 125 μg/mL and a selectivity index (SI) value of 400. Apoptosis inducing effects of dehydroperillic acid was better in A549 cell line. Dehydroperillic acid may be a good candidate for therapy of lung adenocarcinoma and may show this anticancer activity by inducing apoptosis. Copyright© Bentham Science Publishers; For any queries, please email at epub@benthamscience.org.

  7. Bu-Zhong-Yi-Qi Decoction, the Water Extract of Chinese Traditional Herbal Medicine, Enhances Cisplatin Cytotoxicity in A549/DDP Cells through Induction of Apoptosis and Autophagy

    PubMed Central

    Xiong, Ying

    2017-01-01

    Cisplatin is one of the most active cytotoxic agents for non-small cell lung cancer (NSCLC) treatment. However, the development of cisplatin resistance is common. Bu-Zhong-Yi-Qi decoction (BZYQD), a Chinese traditional herbal medicine, is widely used for the enhancement of antitumor effect in other medications. In this study, we evaluated the effect and drug-resistance reversal mechanism of BZYQD combined with cisplatin on cisplatin-resistant A549/DDP cells. Our results showed that BZYQD exhibited direct cytotoxic and chemosensitizing effects. Cotreatment with BZYQD and cisplatin induced intrinsic apoptotic pathways which were measured by condensed nuclear chromatin, Annexin V/PI apoptosis assay, and apoptosis related proteins expression. In addition, cotreatment with BZYQD and cisplatin also activated autophagy, as indicated by an increase in LC3 puncta, classical autophagosomes and/or autolysosomes, and an accumulation of LC3-II and ATG7 protein. Finally, cotreatment with BZYQD and cisplatin resulted in the generation of ROS and scavenging ROS by NAC almost completely suppressing cell death. These results suggest that cotreatment with BZYQD and cisplatin might reverse cisplatin resistance by inducing ROS accumulation, which activates apoptosis and autophagy by oxidative stress. The combination of BZYQD and cisplatin may represent a novel approach in treatment for NSCLC and thus offer a new target for chemotherapy. PMID:28154825

  8. Xylitol induces cell death in lung cancer A549 cells by autophagy.

    PubMed

    Park, Eunjoo; Park, Mi Hee; Na, Hee Sam; Chung, Jin

    2015-05-01

    Xylitol is a widely used anti-caries agent that has anti-inflammatory effects. We have evaluated the potential of xylitol in cancer treatment. It's effects on cell proliferation and cytotoxicity were measured by MTT assay and LDH assay. Cell morphology and autophagy were examined by immunostaining and immunoblotting. Xylitol inhibited cell proliferation in a dose-dependent manner in these cancer cells: A549, Caki, NCI-H23, HCT-15, HL-60, K562, and SK MEL-2. The IC50 of xylitol in human gingival fibroblast cells was higher than in cancer cells, indicating that it is more specific for cancer cells. Moreover, xylitol induced autophagy in A549 cells that was inhibited by 3-methyladenine, an autophagy inhibitor. These results indicate that xylitol has potential in therapy against lung cancer by inhibiting cell proliferation and inducing autophagy of A549 cells.

  9. Antimetastatic Effects of Phyllanthus on Human Lung (A549) and Breast (MCF-7) Cancer Cell Lines

    PubMed Central

    Lee, Sau Har; Jaganath, Indu Bala; Wang, Seok Mui; Sekaran, Shamala Devi

    2011-01-01

    Background Current chemotherapeutic drugs kill cancer cells mainly by inducing apoptosis. However, they become ineffective once cancer cell has the ability to metastasize, hence the poor prognosis and high mortality rate. Therefore, the purpose of this study was to evaluate the antimetastatic potential of Phyllanthus (P. niruri, P. urinaria, P. watsonii, and P. amarus) on lung and breast carcinoma cells. Methodology/Principal Findings Cytotoxicity of Phyllanthus plant extracts were first screened using the MTS reduction assay. They were shown to inhibit MCF-7 (breast carcinoma) and A549 (lung carcinoma) cells growth with IC50 values ranging from 50–180 µg/ml and 65–470 µg/ml for methanolic and aqueous extracts respectively. In comparison, they have lower toxicity on normal cells with the cell viability percentage remaining above 50% when treated up to 1000 µg/ml for both extracts. After determining the non-toxic effective dose, several antimetastasis assays were carried out and Phyllanthus extracts were shown to effectively reduce invasion, migration, and adhesion of both MCF-7 and A549 cells in a dose-dependent manner, at concentrations ranging from 20–200 µg/ml for methanolic extracts and 50–500 µg/ml for aqueous extracts. This was followed by an evaluation of the possible modes of cell death that occurred along with the antimetastatic activity. Phyllanthus was shown to be capable of inducing apoptosis in conjunction with its antimetastastic action, with more than three fold increase of caspases-3 and -7, the presence of DNA-fragmentation and TUNEL-positive cells. The ability of Phyllanthus to exert antimetastatic activities is mostly associated to the presence of polyphenol compounds in its extracts. Conclusions/Significance The presence of polyphenol compounds in the Phyllanthus plant is critically important in the inhibition of the invasion, migration, and adhesion of cancer cells, along with the involvement of apoptosis induction. Hence

  10. Houttuynia cordata Thunb extract modulates G0/G1 arrest and Fas/CD95-mediated death receptor apoptotic cell death in human lung cancer A549 cells

    PubMed Central

    2013-01-01

    Background Houttuynia cordata Thunb (HCT) is commonly used in Taiwan and other Asian countries as an anti-inflammatory, antibacterial and antiviral herbal medicine. In this study, we investigated the anti-human lung cancer activity and growth inhibition mechanisms of HCT in human lung cancer A549 cells. Results In order to investigate effects of HCT on A549 cells, MTT assay was used to evaluate cell viability. Flow cytometry was employed for cell cycle analysis, DAPI staining, and the Comet assay was used for DNA fragmentation and DNA condensation. Western blot analysis was used to analyze cell cycle and apoptotic related protein levels. HCT induced morphological changes including cell shrinkage and rounding. HCT increased the G0/G1 and Sub-G1 cell (apoptosis) populations and HCT increased DNA fragmentation and DNA condensation as revealed by DAPI staining and the Comet assay. HCT induced activation of caspase-8 and caspase-3. Fas/CD95 protein levels were increased in HCT-treated A549 cells. The G0/G1 phase and apoptotic related protein levels of cyclin D1, cyclin A, CDK 4 and CDK 2 were decreased, and p27, caspase-8 and caspase-3 were increased in A549 cells after HCT treatment. Conclusions The results demonstrated that HCT-induced G0/G1 phase arrest and Fas/CD95-dependent apoptotic cell death in A549 cells PMID:23506616

  11. A novel synthetic compound exerts effective anti-tumour activity in vivo via the inhibition of tubulin polymerisation in A549 cells.

    PubMed

    Yan, Jun; Pang, Yanqing; Sheng, Jianfeng; Wang, Yali; Chen, Jie; Hu, Jinhui; Huang, Ling; Li, Xingshu

    2015-09-01

    Microtubules are critical elements that are involved in a wide range of cellular processes, and thus, they have become an attractive target for many anticancer drugs. A novel synthesised compound, 12P, was identified as new microtubule inhibitor. This compound inhibits tubulin polymerisation through binding to the colchicine-binding site of tubulin. 12P exhibits excellent anti-proliferative activities against a panel of human cancer cell lines, with IC₅₀ values range from 9 to 55nM. Interestingly, compound 12P also displayed equally potent cytotoxicity against several drug-resistant cell lines, and it showed high selectivity for active human umbilical vein endothelial cells (HUVECs). Further flow cytometric analysis showed that 12P induces G₂/M phase arrest and apoptosis in A549 cells. Cellular studies have revealed that the induction of apoptosis by 12P was associated with a collapse of mitochondrial membrane potential (MMP), accumulation of reactive oxygen species (ROS), alterations in the expression of some cell cycle-related proteins (e.g. Cyclin B1, Cdc25c, Cdc2) and some apoptosis-related proteins (e.g. Bax, Bad, Bcl-2, Bcl-xl). Importantly, 12P significantly reduced the growth of xenograft tumours of A549 cells in vivo (tumour inhibitory rate of 12P: 84.2%), without any loss of body weight. Taken together, these in vitro and in vivo results suggested that 12P may become a promising lead compound for the development of new anticancer drugs. Copyright © 2015 Elsevier Inc. All rights reserved.

  12. Tanshinone IIA combined with adriamycin inhibited malignant biological behaviors of NSCLC A549 cell line in a synergistic way.

    PubMed

    Xie, Jun; Liu, Jia-Hui; Liu, Heng; Liao, Xiao-Zhong; Chen, Yuling; Lin, Mei-Gui; Gu, Yue-Yu; Liu, Tao-Li; Wang, Dong-Mei; Ge, Hui; Mo, Sui-Lin

    2016-11-18

    The study was designed to develop a platform to verify whether the extract of herbs combined with chemotherapy drugs play a synergistic role in anti-tumor effects, and to provide experimental evidence and theoretical reference for finding new effective sensitizers. Inhibition of tanshinone IIA and adriamycin on the proliferation of A549, PC9 and HLF cells were assessed by CCK8 assays. The combination index (CI) was calculated with the Chou-Talalay method, based on the median-effect principle. Migration and invasion ability of A549 cells were determined by wound healing assay and transwell assay. Flow cytometry was used to detect the cell apoptosis and the distribution of cell cycles. TUNEL staining was used to detect the apoptotic cells. Immunofluorescence staining was used to detect the expression of Cleaved Caspase-3. Western blotting was used to detect the proteins expression of relative apoptotic signal pathways. CDOCKER module in DS 2.5 was used to detect the binding modes of the drugs and the proteins. Both tanshinone IIA and adriamycin could inhibit the growth of A549, PC9, and HLF cells in a dose- and time-dependent manner, while the proliferative inhibition effect of tanshinone IIA on cells was much weaker than that of adriamycin. Different from the cancer cells, HLF cells displayed a stronger sensitivity to adriamycin, and a weaker sensitivity to tanshinone IIA. When tanshinone IIA combined with adriamycin at a ratio of 20:1, they exhibited a synergistic anti-proliferation effect on A549 and PC9 cells, but not in HLF cells. Tanshinone IIA combined with adriamycin could synergistically inhibit migration, induce apoptosis and arrest cell cycle at the S and G2 phases in A549 cells. Both groups of the single drug treatment and the drug combination up-regulated the expressions of Cleaved Caspase-3 and Bax, but down-regulated the expressions of VEGF, VEGFR2, p-PI3K, p-Akt, Bcl-2, and Caspase-3 protein. Compared with the single drug treatment groups, the drug

  13. Pigment Epithelial-derived Factor (PEDF)-triggered Lung Cancer Cell Apoptosis Relies on p53 Protein-driven Fas Ligand (Fas-L) Up-regulation and Fas Protein Cell Surface Translocation*

    PubMed Central

    Li, Lei; Yao, Ya-Chao; Fang, Shu-Huan; Ma, Cai-Qi; Cen, Yi; Xu, Zu-Min; Dai, Zhi-Yu; Li, Cen; Li, Shuai; Zhang, Ting; Hong, Hong-Hai; Qi, Wei-Wei; Zhou, Ti; Li, Chao-Yang; Yang, Xia; Gao, Guo-Quan

    2014-01-01

    Pigment epithelium-derived factor (PEDF), a potent antiangiogenesis agent, has recently attracted attention for targeting tumor cells in several types of tumors. However, less is known about the apoptosis-inducing effect of PEDF on human lung cancer cells and the underlying molecular events. Here we report that PEDF has a growth-suppressive and proapoptotic effect on lung cancer xenografts. Accordingly, in vitro, PEDF apparently induced apoptosis in A549 and Calu-3 cells, predominantly via the Fas-L/Fas death signaling pathway. Interestingly, A549 and Calu-3 cells are insensitive to the Fas-L/Fas apoptosis pathway because of the low level of cell surface Fas. Our results revealed that, in addition to the enhancement of Fas-L expression, PEDF increased the sensitivity of A549 and Calu-3 cells to Fas-L-mediated apoptosis by triggering the translocation of Fas protein to the plasma membrane in a p53- and FAP-1-dependent manner. Similarly, the up-regulation of Fas-L by PEDF was also mediated by p53. Furthermore, peroxisome proliferator-activated receptor γ was determined to be the upstream regulator of p53. Together, these findings uncover a novel mechanism of tumor cell apoptosis induced by PEDF and provide a potential therapeutic strategy for tumors that are insensitive to Fas-L/Fas-dependent apoptosis because of a low level of cell surface Fas. PMID:25225287

  14. [Effect of ginseng rare ginsenoside components combined with paclitaxel on A549 lung cancer].

    PubMed

    Yang, Lei; Zhang, Zhen-Hai; Jia, Xiao-Bin

    2018-04-01

    Traditional Chinese medicine combined with anticancer drugs is a new direction of clinical cancer therapy in recent years. In this study, the optimal ratio of ginseng rare ginsenoside components and paclitaxel was optimized by MTT method, and the proliferative, apoptotic and anti-tumor effects of lung cancer A549 cells were investigated. It was found that the inhibitory effect on the proliferation of lung cancer A549 cells was the same as that on paclitaxel when the ratio of rare ginseng rare ginsenoside components to paclitaxel was 4∶6. Further studies showed that the combined therapy significantly increased the inductive effect of apoptosis in A549 cells, and up-regulated the expression of caspase-3 protein and down-regulated the ratio of Bcl-2/Bax. The tumor-bearing mice model showed that the combination therapy of ginseng rare ginsenoside components and paclitaxel could significantly inhibit the growth of tumor and alleviate the toxic and side effects of paclitaxel on liver. A multi-component system of ginseng rare ginsenoside components-paclitaxel was established in this paper. The proliferation and growth of lung cancer A549 cells were inhibited by paclitaxel-induced apoptosis, the dosage of paclitaxel and the toxicity of paclitaxel were reduced, and the effect of anti-lung cancer was enhanced, which provided a theoretical basis for later studies and clinical application. Copyright© by the Chinese Pharmaceutical Association.

  15. Mycobacterium bovis BCG promotes tumor cell survival from tumor necrosis factor-α-induced apoptosis.

    PubMed

    Holla, Sahana; Ghorpade, Devram Sampat; Singh, Vikas; Bansal, Kushagra; Balaji, Kithiganahalli Narayanaswamy

    2014-09-11

    Increased incidence of lung cancer among pulmonary tuberculosis patients suggests mycobacteria-induced tumorigenic response in the host. The alveolar epithelial cells, candidate cells that form lung adenocarcinoma, constitute a niche for mycobacterial replication and infection. We thus explored the possible mechanism of M. bovis Bacillus Calmette-Guérin (BCG)-assisted tumorigenicity in type II epithelial cells, human lung adenocarcinoma A549 and other cancer cells. Cancer cell lines originating from lung, colon, bladder, liver, breast, skin and cervix were treated with tumor necrosis factor (TNF)-α in presence or absence of BCG infection. p53, COP1 and sonic hedgehog (SHH) signaling markers were determined by immunoblotting and luciferase assays, and quantitative real time PCR was done for p53-responsive pro-apoptotic genes and SHH signaling markers. MTT assays and Annexin V staining were utilized to study apoptosis. Gain- and loss-of-function approaches were used to investigate the role for SHH and COP1 signaling during apoptosis. A549 xenografted mice were used to validate the contribution of BCG during TNF-α treatment. Here, we show that BCG inhibits TNF-α-mediated apoptosis in A549 cells via downregulation of p53 expression. Substantiating this observation, BCG rescued A549 xenografts from TNF-α-mediated tumor clearance in nude mice. Furthermore, activation of SHH signaling by BCG induced the expression of an E3 ubiquitin ligase, COP1. SHH-driven COP1 targeted p53, thereby facilitating downregulation of p53-responsive pro-apoptotic genes and inhibition of apoptosis. Similar effects of BCG could be shown for HCT116, T24, MNT-1, HepG2 and HELA cells but not for HCT116 p53(-/-) and MDA-MB-231 cells. Our results not only highlight possible explanations for the coexistence of pulmonary tuberculosis and lung cancer but also address probable reasons for failure of BCG immunotherapy of cancers.

  16. Proteomic response to 5,6-dimethylxanthenone 4-acetic acid (DMXAA, vadimezan) in human non-small cell lung cancer A549 cells determined by the stable-isotope labeling by amino acids in cell culture (SILAC) approach

    PubMed Central

    Pan, Shu-Ting; Zhou, Zhi-Wei; He, Zhi-Xu; Zhang, Xueji; Yang, Tianxin; Yang, Yin-Xue; Wang, Dong; Qiu, Jia-Xuan; Zhou, Shu-Feng

    2015-01-01

    5,6-Dimethylxanthenone 4-acetic acid (DMXAA), also known as ASA404 and vadimezan, is a potent tumor blood vessel-disrupting agent and cytokine inducer used alone or in combination with other cytotoxic agents for the treatment of non-small cell lung cancer (NSCLC) and other cancers. However, the latest Phase III clinical trial has shown frustrating outcomes in the treatment of NSCLC, since the therapeutic targets and underlying mechanism for the anticancer effect of DMXAA are not yet fully understood. This study aimed to examine the proteomic response to DMXAA and unveil the global molecular targets and possible mechanisms for the anticancer effect of DMXAA in NSCLC A549 cells using a stable-isotope labeling by amino acids in cell culture (SILAC) approach. The proteomic data showed that treatment with DMXAA modulated the expression of 588 protein molecules in A549 cells, with 281 protein molecules being up regulated and 306 protein molecules being downregulated. Ingenuity pathway analysis (IPA) identified 256 signaling pathways and 184 cellular functional proteins that were regulated by DMXAA in A549 cells. These targeted molecules and signaling pathways were mostly involved in cell proliferation and survival, redox homeostasis, sugar, amino acid and nucleic acid metabolism, cell migration, and invasion and programed cell death. Subsequently, the effects of DMXAA on cell cycle distribution, apoptosis, autophagy, and reactive oxygen species (ROS) generation were experimentally verified. Flow cytometric analysis showed that DMXAA significantly induced G1 phase arrest in A549 cells. Western blotting assays demonstrated that DMXAA induced apoptosis via a mitochondria-dependent pathway and promoted autophagy, as indicated by the increased level of cytosolic cytochrome c, activation of caspase 3, and enhanced expression of beclin 1 and microtubule-associated protein 1A/1B-light chain 3 (LC3-II) in A549 cells. Moreover, DMXAA significantly promoted intracellular ROS

  17. Agarol, an ergosterol derivative from Agaricus blazei, induces caspase-independent apoptosis in human cancer cells.

    PubMed

    Shimizu, Takamitsu; Kawai, Junya; Ouchi, Kenji; Kikuchi, Haruhisa; Osima, Yoshiteru; Hidemi, Rikiishi

    2016-04-01

    Agaricus blazei (A. blazei) is a mushroom with many biological effects and active ingredients. We purified a tumoricidal substance from A. blazei, an ergosterol derivative, and named it 'Agarol'. Cytotoxic effects of Agarol were determined by the MTT assay using A549, MKN45, HSC-3, and HSC-4 human carcinoma cell lines treated with Agarol. Apoptosis was detected by flow cytometry analysis. Reactive oxygen species (ROS) levels and mitochondria membrane potential (∆ψm) were also determined by flow cytometry. Western blot analysis was used to quantify the expression of apoptosis-related proteins. Agarol predominantly induced apoptosis in two p53-wild cell lines (A549 and MKN45) compared to the other p53-mutant cell lines (HSC-3 and HSC-4). Further mechanistic studies revealed that induction of apoptosis is associated with increased generation of ROS, reduced ∆ψm, release of apoptosis-inducing factor (AIF) from the mitochondria to the cytosol, upregulation of Bax, and downregulation of Bcl-2. Caspase-3 activities did not increase, and z-VAD-fmk, a caspase inhibitor, did not inhibit the Agarol-induced apoptosis. These findings indicate that Agarol induces caspase-independent apoptosis in human carcinoma cells through a mitochondrial pathway. The in vivo anticancer activity of Agarol was confirmed in a xenograft murine model. This study suggests a molecular mechanism by which Agarol induces apoptosis in human carcinoma cells and indicates the potential use of Agarol as an anticancer agent.

  18. Dual‑sensitive HRE/Egr1 promoter regulates Smac overexpression and enhances radiation‑induced A549 human lung adenocarcinoma cell death under hypoxia.

    PubMed

    Li, Chang-Feng; Chen, Li-Bo; Li, Dan-Dan; Yang, Lei; Zhang, Bao-Gang; Jin, Jing-Peng; Zhang, Ying; Zhang, Bin

    2014-08-01

    The aim of this study was to construct an expression vector carrying the hypoxia/radiation dual‑sensitive chimeric hypoxia response element (HRE)/early growth response 1 (Egr‑1) promoter in order to overexpress the therapeutic second mitochondria‑derived activator of caspases (Smac). Using this expression vector, the present study aimed to explore the molecular mechanism underlying radiotherapy‑induced A549 human lung adenocarcinoma cell death and apoptosis under hypoxia. The plasmids, pcDNA3.1‑Egr1‑Smac (pE‑Smac) and pcDNA3.1‑HRE/Egr-1‑Smac (pH/E‑Smac), were constructed and transfected into A549 human lung adenocarcinoma cells using the liposome method. CoCl2 was used to chemically simulate hypoxia, followed by the administration of 2 Gy X‑ray irradiation. An MTT assay was performed to detect cell proliferation and an Annexin V‑fluorescein isothiocyanate apoptosis detection kit was used to detect apoptosis. Quantitative polymerase chain reaction and western blot analyses were used for the detection of mRNA and protein expression, respectively. Infection with the pE‑Smac and pH/E‑Smac plasmids in combination with radiation and/or hypoxia was observed to enhance the expression of Smac. Furthermore, Smac overexpression was found to enhance the radiation‑induced inhibition of cell proliferation and promotion of cycle arrest and apoptosis. The cytochrome c/caspase‑9/caspase‑3 pathway was identified to be involved in this regulation of apoptosis. Plasmid infection in combination with X‑ray irradiation was found to markedly induce cell death under hypoxia. In conclusion, the hypoxia/radiation dual‑sensitive chimeric HRE/Egr‑1 promoter was observed to enhance the expression of the therapeutic Smac, as well as enhance the radiation‑induced inhibition of cell proliferation and promotion of cycle arrest and apoptosis under hypoxia. This apoptosis was found to involve the mitochondrial pathway.

  19. The enhancing effect of genistein on apoptosis induced by trichostatin A in lung cancer cells with wild type p53 genes is associated with upregulation of histone acetyltransferase

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Wu, Tzu-Chin; Lin, Yi-Chin; Chen, Hsiao-Ling

    Genistein has been shown to enhance the antitumor activity of trichostatin A (TSA) in human lung carcinoma A549 cells. However, whether the combined treatment exerts the same effect in other lung cancer cells is unclear. In the present study we first compared the enhancing effect of genistein on the antitumor effect of TSA in ABC-1, NCI-H460 (H460) and A549 cells. Second, we investigated whether the effects of genistein are associated with increased histone/non-histone protein acetylation. We found that the enhancing effect of genistein on cell-growth-arrest in ABC-1 cells (p53 mutant) was less than in A549 and H460 cells. Genistein enhancedmore » TSA induced apoptosis in A549 and H460 cells rather than in ABC-1 cells. After silencing p53 expression in A549 and H460 cells, the enhancing effect of genistein was diminished. In addition, genistein increased TSA-induced histone H3/H4 acetylation in A549 and H460 cells. Genistein also increased p53 acetylation in H460 cells. The inhibitor of acetyltransferase, anacardic acid, diminished the enhancing effect of genistein on all TSA-induced histone/p53 acetylation and apoptosis. Genistein in combination with TSA increased the expression of p300 protein, an acetyltransferase, in A549 and NCI-H460 cells. Furthermore, we demonstrated that genistein also enhanced the antitumor effect of genistein in A549-tumor-bearing mice. Taken together, these results suggest that the enhancing effects of genistein on TSA-induced apoptosis in lung cancer cells were p53-dependent and were associated with histone/non-histone protein acetylation. - Highlights: • Genistein enhances the antitumor effect of TSA through p53-associated pathways. • Genistein enhances TSA-induced histone acetylation commonly. • An acetyltransferase inhibitor diminishes the antitumor effect of genistein + TSA. • TSA in combination with genistein increases the expression of p300. • Genistein given by i.p. injection increases the antitumor effect of TSA in

  20. Nicotine prevents the apoptosis induced by menadione in human lung cancer cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zhang Tao; Lu Heng; Shang Xuan

    Approximately 50% of long-term cigarette smokers die prematurely from the adverse effects of smoking, including on lung cancer and other illnesses. Nicotine is a main component in tobacco and has been implicated as a potential factor in the pathogenesis of human lung cancer. However, the mechanism of nicotine action in the development of lung cancer remains largely unknown. In the present study, we designed a nicotine-apoptosis system, by pre-treatment of nicotine making lung cancer cell A549 to be in a physiological nicotine environment, and observed that nicotine promoted cell proliferation and prevented the menadione-induced apoptosis, and exerts its role ofmore » anti-apoptosis by shift of apoptotic stage induced by menadione from late apoptotic stage to early apoptotic stage, in which NF-{kappa}B was up-regulated. Interference analysis of NF-{kappa}B in A549 cells showed that knock down of NF-{kappa}B resulted in apoptosis promotion and counteracted the protective effect of nicotine. The findings suggest that nicotine has potential effect in lung cancer genesis, especially in patients with undetectable early tumor development and development of specific NF-{kappa}B inhibitors would represent a potentially exciting new pharmacotherapy for tobacco-related lung cancer.« less

  1. TRIM25 is associated with cisplatin resistance in non-small-cell lung carcinoma A549 cell line via downregulation of 14-3-3σ.

    PubMed

    Qin, Xia; Qiu, Feng; Zou, Zhen

    2017-11-04

    Lung cancer, in particular, non-small cell lung cancer (NSCLC), is the leading cause of cancer-related mortality. Cis-Diamminedichloroplatinum (cisplatin, CDDP) as first-line chemotherapy for NSCLC, but resistance occurs frequently. We previously reported that Tripartite motif protein 25 (TRIM25) was highly expressed in cisplatin-resistant human lung adenocarcinoma A549 cells (A549/CDDP) in comparison with its parental A549 cells. Herein, we take a further step to demonstrate the association of TRIM25 and cisplatin resistance and also the underlying mechanisms. Knockdown of TRIM25 by RNA interference in A549/CDDP cells decreased half maximal inhibitory concentration (IC 50 ) values and promoted apoptosis in response to cisplatin, whereas overexpression of TRIM25 had opposite effects. More importantly, we found that concomitant knockdown of 14-3-3σ and TRIM25 absolutely reversed the decreased MDM2, increased p53, increased cleaved-Capsese3 and decreased IC 50 value induced by knockdown of TRIM25 individually, suggesting that TRIM25 mediated cisplatin resistance primarily through downregulation of 14-3-3σ. Our results indicate that TRIM25 is associated with cisplatin resistance and 14-3-3σ-MDM2-p53 signaling pathway is involved in this process, suggesting targeting TRIM25 may be a potential strategy for the reversal of cisplatin resistance. Copyright © 2017 Elsevier Inc. All rights reserved.

  2. Depleted aldehyde dehydrogenase 1A1 (ALDH1A1) reverses cisplatin resistance of human lung adenocarcinoma cell A549/DDP.

    PubMed

    Wei, Yunyan; Wu, Shuangshuang; Xu, Wei; Liang, Yan; Li, Yue; Zhao, Weihong; Wu, Jianqing

    2017-01-01

    Cisplatin is the standard first-line chemotherapeutic agent for the treatment of non-small cell lung cancer (NSCLC). However, resistance to chemotherapy has been a major obstacle in the management of NSCLC. Aldehyde dehydrogenase 1A1 (ALDH1A1) overexpression has been observed in a variety of cancers, including lung cancer. The purpose of this study was to investigate the effect of ALDH1A1 expression on cisplatin resistance and explore the mechanism responsible. Reverse transcriptase-PCR was applied to measure the messenger RNA expression of ALDH1A1, while Western blot assay was employed to evaluate the protein expression of ALDH1A1, B-cell lymphoma 2, Bcl-2-like protein 4, phospho-protein kinase B (p-AKT) and AKT. A short hairpin RNA was used to knockdown ALDH1A1 expression. A 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay was used to determine the effect of ALDH1A1 decrease on cell viability. The cell apoptotic rate was tested using flow cytometry assay. ALDH1A1 is overexpressed in cisplatin resistant cell line A549/DDP, compared with A549. ALDH1A1 depletion significantly decreased A549/DDP proliferation, increased apoptosis, and reduced cisplatin resistance. In addition, the phosphoinositide 3-kinase (PI3K) / AKT pathway is activated in A549/DDP, and ALDH1A1 knockdown reduced the phosphorylation level of AKT. Moreover, the combination of ALDH1A1-short hairpin RNA and PI3K/AKT pathway inhibitor LY294002 markedly inhibited cell viability, enhanced apoptotic cell death, and increased cisplatin sensitivity. These results suggest that ALDH1A1 depletion could reverse cisplatin resistance in human lung cancer cell line A549/DDP, and may act as a potential target for the treatment of lung cancers resistant to cisplatin. © 2016 The Authors. Thoracic Cancer published by China Lung Oncology Group and John Wiley & Sons Australia, Ltd.

  3. Magnolol induces apoptosis via caspase-independent pathways in non-small cell lung cancer cells.

    PubMed

    Tsai, Jong-Rung; Chong, Inn-Wen; Chen, Yung-Hsiang; Hwang, Jhi-Jhu; Yin, Wei-Hsian; Chen, Hsiu-Lin; Chou, Shah-Hwa; Chiu, Chien-Chih; Liu, Po-Len

    2014-04-01

    Magnolol, a hydroxylated biphenyl agent isolated from herbal planet Magnolia officinalis, is a component of traditional Asian herbal teas. It has been reported to have anti-microbial, anti-inflammatory, and anti-cancer activity. Non-small cell lung cancer (NSCLC) cell lines (A549, H441 and H520) and normal human bronchial epithelial cells (HBECs) were used to evaluate the cytotoxic effect of magnolol. We show that magnolol inhibited cellular proliferation, increased DNA fragmentation, and decreased mitochondrial membrane potential in all NSCLC cells, but had no cytotoxic effect on HBECs. Magnolol triggered the release of pro-apoptotic proteins: Bid, Bax and cytochrome c from mitochondria, but did not activate the caspase-3, -8, and -9, suggesting that magnolol induces apoptosis of NSCLC cell lines via a caspase-independent pathway. The caspase-independent pathway is mediated through the activation of nuclear translocation of apoptosis-inducing factor, endonuclease G and cleaved poly(ADP-ribose) polymerase, which played important roles in mediating cell death. Furthermore, magnolol inhibited PI3K/AKT and ERK1/2 activity, but up-regulated p38 and JNK activity in A549 cell lines. The results of this study provided a basis for understanding and developing magnolol as a novel treatment of NSCLC.

  4. Trichostatin A-induced apoptosis is mediated by Kruppel-like factor 4 in ovarian and lung cancer.

    PubMed

    Zohre, Sadeghi; Kazem, Nejati-Koshki; Abolfazl, Akbarzadeh; Mohammad, Rahmati-Yamchi; Aliakbar, Movassaghpour; Effat, Alizadeh; Zahra, Davoudi; Hassan, Dariushnejad; Nosratollah, Zarghami

    2014-01-01

    The istone deacetylase (HDAC) inhibitor trichostatin A (TSA) is known to mediate the regulation of gene expression and anti proliferation activity in cancer cells. Kruppel-like factor 4 (klf4) is a zinc finger- containing transcription factor of the SP/KLF family, that is expressed in a variety of tissues and regulates cell proliferation, differentiation, tumorigenesis, and apoptosis. It may either either function as a tumor suppressor or an oncogene depending on genetic context of tumors. In this study, we tested the possibility that TSA may increase klf4 expression and cancer cell growth inhibition and apoptosis in SKOV-3 and A549 cells. The cytotoxicity of TSA was determined using the MTT assay test, while klf4 gene expression was assessed by real time PCR and to ability of TSA to induce apoptosis using a Vybrant Apoptosis Assay kit. Our results showed that TSA exerted dose and time dependent cytotoxicity effect on SKOV-3 and A549 cells. Moreover TSA up-regulated klf4 expression. Flow cytometric analysis demonstrated that apoptosis was increased after TSA treatment. Taken together, this study showed that TSA increased klf4 expression in SKOV3 and A549 cell lines, consequently, klf4 may played a tumor-suppressor role by increasing both cell growth inhibition and apoptosis. This study sheds light on the details of molecular mechanisms of HDACI-induced cell cycle arrest and apoptosis.

  5. SiRNA/DOX lodeded chitosan based nanoparticles: Development, Characterization and in vitro evaluation on A549 lung cancer cell line.

    PubMed

    Seifi-Najmi, M; Hajivalili, M; Safaralizadeh, R; Sadreddini, S; Esmaeili, S; Razavi, R; Ahmadi, M; Mikaeili, H; Baradaran, B; Shams-Asenjan, K; Yousefi, M

    2016-09-30

    High-mobility group AT-hook2 (HMGA2), involved in epithelial mesenchymal transition (EMT) process, has a pivotal role in lung cancer metastasis. Lung cancer therapy with HMGA2 suppressing small interfering RNA (siRNA) has been introduced recently while doxorubicin (DOX) has been used as a frequent cancer chemotherapy agent. Both reagents have been faced with obstacles in clinic which make them ineffective. NanoParticles (NPs) provided a platform for efficient co delivery of the anticancer drugs. The aim of this study was production and in vitro characterization of different pharmacological groups (siRNA, DOX or siRNA-DOX) of carboxymethyl dextran thrimethyl chitosan nanoparticles (CMDTMChiNPs) on cytotoxicity, gene expression, apoptosis and migration of metastatic lung cancer cell line (A-549). CMDTMChiNPs were synthesized and encapsulated with siRNA, DOX or siRNA-DOX. Then the effects of HMGA2 siRNA and DOX co delivery was assessed in A549 viability and target genes (HMGA2, Ecadherin, vimentin and MMP9) by MTT and real time PCR, respectively. In addition capability of apoptosis induction and anti-migratory features of formulated NPs were analyzed by flowcytometry and wound healing assays. SiRNA-DOX-CMDTM ChiNPs approximate size were 207±5 with poly dispersity index (PDI) and zeta potential of 0.4 and 16.3±0.3, respectively. NPs loaded with DOX and siRNA were the most efficient drug formulations in A549 cell cytotoxicity, altering of EMT markers, apoptosis induction and migration inhibition. Generally our results showed that co delivery of HMGA2 siRNA and DOX by novel designed CMDTMChiNPs is a new therapeutic approach with great potential efficiency for lung cancer treatment.

  6. Inhibition of NF-kappaB-mediated transcription and induction of apoptosis in human breast cancer cells by epoxypseudoisoeugenol-2-methyl butyrate.

    PubMed

    Ma, Guoyi; Tabanca, Nurhayat; Husnu Can Baser, K; Kirimer, Nese; Pasco, David S; Khan, Ikhlas A; Khan, Shabana I

    2009-03-01

    Breast cancer is one of the most prevalent woman cancers. Genomic instability, accumulative mutations, and subsequent changes in intracellular signaling cascades play key roles in the development of human breast cancers. Activation of nuclear factor-kappaB (NF-kappaB) has been implicated in oncogenesis of breast cancers and is known to be associated with resistance to anticancer agents and apoptosis. Blocking NF-kappaB signaling may represent a therapeutic strategy in breast cancer therapy. The objective of this study is to investigate the in vitro effects of epoxypseudoisoeugenol-2-methyl butyrate (EPB), a phenylpropranoid isolated from Pimpinella corymbosa, on the activation of NF-kappaB, cell growth, cell cycle progression and apoptosis in MCF-7 (estrogen-dependent) and BT-549 (estrogen-independent) breast cancer cells. Transcriptional activity of NF-kappaB was measured by cell based reporter gene assay. Cell proliferation was determined by MTT assay. Cell cycle analysis was carried out by flow cytometry and apoptosis was observed by DAPI staining assy. EPB inhibited the NF-kappaB-mediated transcription activity induced by tumor necrosis factor-alpha (TNF-alpha) and phorbol myristate acetate (PMA) in MCF-7 cells. EPB also inhibited constitutive NF-kappaB transcriptional activity in BT-549 cells. EPB inhibited the proliferation of both MCF-7 and BT-549 cells in a concentration- and time-dependent manner. EPB induced cell cycle arrest in G(1)/G(0) phase and apoptosis in both MCF-7 and BT 549 cells. These in vitro results indicated that EPB has a potential for use against both hormone-dependent and hormone-independent breast cancers and its effects seem to be mediated by inhibiting the NF-kappaB activity.

  7. Cytotoxic Effects of 24-Methylenecyloartanyl Ferulate on A549 Nonsmall Cell Lung Cancer Cells through MYBBP1A Up-Regulation and AKT and Aurora B Kinase Inhibition.

    PubMed

    Doello, Sofia; Liang, Zhibin; Cho, Il Kyu; Kim, Jung Bong; Li, Qing X

    2018-04-11

    Lung cancer is the second most prevalent cancer. Nonsmall cell lung cancer (NSCLC) is the most common type of lung cancer. The low efficacy in current chemotherapies impels us to find new alternatives to prevent or treat NSCLC. Rice bran oil is cytotoxic to A549 cells, a NSCLC cell line. Here, we identified 24-methylenecyloartanyl ferulate (24-mCAF) as the main component responsible for the cytotoxicity in A549 cells. An iTRAQ-based quantitative proteomics analysis revealed that 24-mCAF inhibits cell proliferation and activates cell death and apoptosis. 24-mCAF induces up-regulation of Myb binding protein 1A (MYBBP1A), a tumor suppressor that halts cancer progression. 24-mCAF inhibits the activity of AKT and Aurora B kinase, two Ser/Thr kinases involved in MYBBP1A regulation and that represent important targets in NSCLC. This study provides the first insight of the effect of 24-mCAF, the main component of rice bran oil, on A459 cells at the cellular and molecular levels.

  8. Suppression of A549 cell proliferation and metastasis by calycosin via inhibition of the PKC‑α/ERK1/2 pathway: An in vitro investigation.

    PubMed

    Cheng, Xu-Dong; Gu, Jun-Fei; Yuan, Jia-Rui; Feng, Liang; Jia, Xiao-Bin

    2015-12-01

    The migration and invasion of lung cancer cells into the extracellular matrix contributes to the high mortality rates of lung cancer. The protein kinase C (PKC) and downstream signaling pathways are important in the invasion and migration of lung cancer cells. Calycosin (Cal), an effector chemical from Astragalus has been reported to affect the recurrence and metastasis of cancer cells via the regulation of the protein expression of matrix metalloproteinases (MMPs). The inhibition of Cal on the migration and invasion of A549 cells was investigated in the present study. Cell viability and apoptosis assays were performed using MTT and flow cytometric analyses. A wound healing assay and Transwell invasion assay were performed to evaluate the effect of Cal on A549 cell migration and invasion. Invasion‑associated proteins, including MMP‑2, MMP‑9, E‑cadherin (E‑cad), integrin β1, PKC‑α and extracellular signal‑regulated kinase 1/2 (ERK1/2) were detected using western blotting. In addition, PKC‑α inhibitor, AEB071, and ERK1/2 inhibitor, PD98059, were used to determine the association between the suppression of PKC‑α /ERK1/2 and invasion, MMP‑2, MMP‑9, E‑cad and integrin β1. Cal was observed to suppress cell proliferation and induce apoptosis. There were significant differences between the phorbol‑12‑myristate‑13‑acetate (TPA)‑induced A549 cells treated with Cal and the untreated cells in the rates of migration and invasion. The levels of MMP‑2, MMP‑9, E‑cad and integrin β1 in the TPA‑induced A549 cells changed markedly, compared with the untreated cells. In addition, the suppression of Cal was affected by the PKC inhibitor, AEB071, an ERK1/2 inhibitor, PD98059. The results of the present study indicated that Cal inhibited the proliferation, adhesion, migration and invasion of the TPA‑induced A549 cells. The Cal‑induced repression of PKC‑α/ERK1/2, increased the expression of E‑Cad and inhibited the expression

  9. A hybrid of coumarin and phenylsulfonylfuroxan induces caspase-dependent apoptosis and cytoprotective autophagy in lung adenocarcinoma cells.

    PubMed

    Wang, Qian; Guo, Yalan; Jiang, Shanshan; Dong, Mengxue; Kuerban, Kudelaidi; Li, Jiyang; Feng, Meiqing; Chen, Ying; Ye, Li

    2018-01-15

    Lung adenocarcinoma is the most primary histologic subtype of non-small cell lung cancer (NSCLC). Compound 8b, a novel coumarin derivative with phenylsulfonylfuroxan group, shows significant antiproliferation activity against lung adenocarcinoma cell with low toxicity. This study aims to uncover the potential of compound 8b in relation to apoptosis as well as autophagy induction in lung adenocarcinoma cells. The cytotoxicity and apoptosis of A549 and H1299 cells induced by compound 8b were detected by MTT, microscope and western blot analysis. Autophagy was determined by TEM, confocal microscopy and western blot analysis. Akt/mTOR and Erk signaling pathway were also examined by western blot analysis. First, significant growth inhibition and caspase-dependent apoptosis were observed in compound 8b-treated A549 and H1299 cells. Then, we confirmed compound 8b-induced autophagy by autophagosomes formation, upregulated expression of autophagy-related protein LC3-II and autophagic flux. Importantly, abolishing autophagy using inhibitors and ATG5 siRNA enhanced the cytotoxicity of compound 8b, indicating the cytoprotective role of autophagy in lung adenocarcinoma. Further mechanistic investigations suggested that Akt/mTOR and Erk signaling pathways contributed to autophagy induction by compound 8b. This results demonstrate that compound 8b induces caspase-dependent apoptosis as well as cytoprotective autophagy in lung adenocarcinoma cells, which may provide scientific evidence for developing this furoxan-based NO-releasing coumarin derivative as a potential anti-lung adenocarcinoma therapeutic agents. Copyright © 2017 Elsevier GmbH. All rights reserved.

  10. Development of drug-loaded chitosan hollow nanoparticles for delivery of paclitaxel to human lung cancer A549 cells.

    PubMed

    Jiang, Jie; Liu, Ying; Wu, Chao; Qiu, Yang; Xu, Xiaoyan; Lv, Huiling; Bai, Andi; Liu, Xuan

    2017-08-01

    In this study, biodegradable chitosan hollow nanospheres (CHN) were fabricated using polystyrene nanospheres (PS) as templates. CHN were applied to increase the solubility of poorly water-soluble drugs. The lung cancer drug paclitaxel (PTX), which is used as a model drug, was loaded into CHN by the adsorption equilibrium method. The drug-loaded sample (PTX-CHN) offered sustained PTX release and good bioavailability. The state characterization of PTX by differential scanning calorimetry (DSC), X-ray diffraction (XRD) and Fourier transform infrared spectroscopy (FTIR) showed that the PTX absorbed into CHN existed in an amorphous state. An in vitro toxicity experiment indicated that CHN were nontoxic as carriers of poorly water-soluble drugs. The PTX-CHN produced a marked inhibition of lung cancer A549 cells proliferation and encouraged apoptosis. A cell uptake experiment indicated that PTX-CHN was successfully taken up by lung cancer A549 cells. Furthermore, a degradation experiment revealed that CHN were readily biodegradable. These findings state clearly that CHN can be regarded as promising biomaterials for lung cancer treatment.

  11. Dihydroartemisinin Inhibits Glucose Uptake and Cooperates with Glycolysis Inhibitor to Induce Apoptosis in Non-Small Cell Lung Carcinoma Cells

    PubMed Central

    Gao, Jing; Luo, Xian-yang; Liu, Yu; Li, Ning; Li, Chun-lei; Chen, Yu-qiang; Yu, Xiu-yi; Jiang, Jie

    2015-01-01

    Despite recent advances in the therapy of non-small cell lung cancer (NSCLC), the chemotherapy efficacy against NSCLC is still unsatisfactory. Previous studies show the herbal antimalarial drug dihydroartemisinin (DHA) displays cytotoxic to multiple human tumors. Here, we showed that DHA decreased cell viability and colony formation, induced apoptosis in A549 and PC-9 cells. Additionally, we first revealed DHA inhibited glucose uptake in NSCLC cells. Moreover, glycolytic metabolism was attenuated by DHA, including inhibition of ATP and lactate production. Consequently, we demonstrated that the phosphorylated forms of both S6 ribosomal protein and mechanistic target of rapamycin (mTOR), and GLUT1 levels were abrogated by DHA treatment in NSCLC cells. Furthermore, the upregulation of mTOR activation by high expressed Rheb increased the level of glycolytic metabolism and cell viability inhibited by DHA. These results suggested that DHA-suppressed glycolytic metabolism might be associated with mTOR activation and GLUT1 expression. Besides, we showed GLUT1 overexpression significantly attenuated DHA-triggered NSCLC cells apoptosis. Notably, DHA synergized with 2-Deoxy-D-glucose (2DG, a glycolysis inhibitor) to reduce cell viability and increase cell apoptosis in A549 and PC-9 cells. However, the combination of the two compounds displayed minimal toxicity to WI-38 cells, a normal lung fibroblast cell line. More importantly, 2DG synergistically potentiated DHA-induced activation of caspase-9, -8 and -3, as well as the levels of both cytochrome c and AIF of cytoplasm. However, 2DG failed to increase the reactive oxygen species (ROS) levels elicited by DHA. Overall, the data shown above indicated DHA plus 2DG induced apoptosis was involved in both extrinsic and intrinsic apoptosis pathways in NSCLC cells. PMID:25799586

  12. Dihydroartemisinin inhibits glucose uptake and cooperates with glycolysis inhibitor to induce apoptosis in non-small cell lung carcinoma cells.

    PubMed

    Mi, Yan-jun; Geng, Guo-jun; Zou, Zheng-zhi; Gao, Jing; Luo, Xian-yang; Liu, Yu; Li, Ning; Li, Chun-lei; Chen, Yu-qiang; Yu, Xiu-yi; Jiang, Jie

    2015-01-01

    Despite recent advances in the therapy of non-small cell lung cancer (NSCLC), the chemotherapy efficacy against NSCLC is still unsatisfactory. Previous studies show the herbal antimalarial drug dihydroartemisinin (DHA) displays cytotoxic to multiple human tumors. Here, we showed that DHA decreased cell viability and colony formation, induced apoptosis in A549 and PC-9 cells. Additionally, we first revealed DHA inhibited glucose uptake in NSCLC cells. Moreover, glycolytic metabolism was attenuated by DHA, including inhibition of ATP and lactate production. Consequently, we demonstrated that the phosphorylated forms of both S6 ribosomal protein and mechanistic target of rapamycin (mTOR), and GLUT1 levels were abrogated by DHA treatment in NSCLC cells. Furthermore, the upregulation of mTOR activation by high expressed Rheb increased the level of glycolytic metabolism and cell viability inhibited by DHA. These results suggested that DHA-suppressed glycolytic metabolism might be associated with mTOR activation and GLUT1 expression. Besides, we showed GLUT1 overexpression significantly attenuated DHA-triggered NSCLC cells apoptosis. Notably, DHA synergized with 2-Deoxy-D-glucose (2DG, a glycolysis inhibitor) to reduce cell viability and increase cell apoptosis in A549 and PC-9 cells. However, the combination of the two compounds displayed minimal toxicity to WI-38 cells, a normal lung fibroblast cell line. More importantly, 2DG synergistically potentiated DHA-induced activation of caspase-9, -8 and -3, as well as the levels of both cytochrome c and AIF of cytoplasm. However, 2DG failed to increase the reactive oxygen species (ROS) levels elicited by DHA. Overall, the data shown above indicated DHA plus 2DG induced apoptosis was involved in both extrinsic and intrinsic apoptosis pathways in NSCLC cells.

  13. BAG3 Overexpression and Cytoprotective Autophagy Mediate Apoptosis Resistance in Chemoresistant Breast Cancer Cells.

    PubMed

    Das, Chandan Kanta; Linder, Benedikt; Bonn, Florian; Rothweiler, Florian; Dikic, Ivan; Michaelis, Martin; Cinatl, Jindrich; Mandal, Mahitosh; Kögel, Donat

    2018-03-01

    Target-specific treatment modalities are currently not available for triple-negative breast cancer (TNBC), and acquired chemotherapy resistance is a primary obstacle for the treatment of these tumors. Here we employed derivatives of BT-549 and MDA-MB-468 TNBC cell lines that were adapted to grow in the presence of either 5-Fluorouracil, Doxorubicin or Docetaxel in an aim to identify molecular pathways involved in the adaptation to drug-induced cell killing. All six drug-adapted BT-549 and MDA-MB-468 cell lines displayed cross resistance to chemotherapy and decreased apoptosis sensitivity. Expression of the anti-apoptotic co-chaperone BAG3 was notably enhanced in two thirds (4/6) of the six resistant lines simultaneously with higher expression of HSP70 in comparison to parental controls. Doxorubicin-resistant BT-549 (BT-549 r DOX 20 ) and 5-Fluorouracil-resistant MDA-MB-468 (MDA-MB-468 r 5-FU 2000 ) cells were chosen for further analysis with the autophagy inhibitor Bafilomycin A1 and lentiviral depletion of ATG5, indicating that enhanced cytoprotective autophagy partially contributes to increased drug resistance and cell survival. Stable lentiviral BAG3 depletion was associated with a robust down-regulation of Mcl-1, Bcl-2 and Bcl-xL, restoration of drug-induced apoptosis and reduced cell adhesion in these cells, and these death-sensitizing effects could be mimicked with the BAG3/Hsp70 interaction inhibitor YM-1 and by KRIBB11, a selective transcriptional inhibitor of HSF-1. Furthermore, BAG3 depletion was able to revert the EMT-like transcriptional changes observed in BT-549 r DOX 20 and MDA-MB-468 r 5-FU 2000 cells. In summary, genetic and pharmacological interference with BAG3 is capable to resensitize TNBC cells to treatment, underscoring its relevance for cell death resistance and as a target to overcome therapy resistance of breast cancer. Copyright © 2018 The Authors. Published by Elsevier Inc. All rights reserved.

  14. Predictive role of computer simulation in assessing signaling pathways of crizotinib-treated A549 lung cancer cells.

    PubMed

    Xia, Pu; Mou, Fei-Fei; Wang, Li-Wei

    2012-01-01

    Non-small-cell lung cancer (NSCLC) is a leading cause of cancer deaths worldwide. Crizotinib has been approved by the U.S. Food and Drug Administration for the treatment of patients with advanced NSCLC. However, understanding of mechanisms of action is still limited. In our studies, we confirmed crizotinib-induced apoptosis in A549 lung cancer cells. In order to assess mechanisms, small molecular docking technology was used as a preliminary simulation of signaling pathways. Interesting, our results of experiments were consistent with the results of computer simulation. This indicates that small molecular docking technology should find wide use for its reliability and convenience.

  15. Osthole induces lung cancer cell apoptosis through inhibition of inhibitor of apoptosis family proteins

    PubMed Central

    Xu, Xiao-Man; Zhang, Man-Li; Zhang, Yi; Zhao, Li

    2016-01-01

    In the present study, we investigated the effects and mechanisms of Osthole on the apoptosis of non-small cell lung cancer (NSCLC) cells and its synergistic effect with Embelin. Our results revealed that treatment with both Osthole and Embelin inhibited cell proliferation. Notably, combination treatment of Osthole and Embelin inhibited cell proliferation more significantly compared with monotherapy. In addition, morphological analysis and Annexin V/propidium iodide analysis revealed that the combination of Osthole and Embelin enhanced their effect on cell apoptosis. We further examined the effect of Osthole on the expression of inhibitor of apoptosis protein (IAP) family proteins. That treatment of A549 lung cancer cells with various concentrations of Osthole was observed to decrease the protein expression of X-chromosome-encoded IAP, c-IAP1, c-IAP2 and Survivin, and increase Smac expression in a dose-dependent manner. Furthermore, it was noted that Osthole or Embelin alone increased the expression of BAX, caspase-3, caspase-9, cleaved caspase-3 and cleaved caspase-9, and decreased Bcl-2 levels following treatment. Osthole and Embelin combination treatment had a synergistic effect on the regulation of these proteins. In conclusion, our study demonstrated that Osthole inhibited proliferation and induced the apoptosis of lung cancer cells via IAP family proteins in a dose-dependent manner. Osthole enhances the antitumor effect of Embelin, indicating that combination of Osthole and Embelin has potential clinical significance in the treatment of NSCLC. PMID:27895730

  16. Ferrous glycinate regulates cell energy metabolism by restrictinghypoxia-induced factor-1α expression in human A549 cells.

    PubMed

    Kuo, Yung-Ting; Jheng, Jhong-Huei; Lo, Mei-Chen; Chen, Wei-Lu; Wang, Shyang-Guang; Lee, Horng-Mo

    2018-06-04

    Iron or oxygen regulates the stability of hypoxia inducible factor-1α (HIF-1α). We investigated whether ferrous glycinate would affect HIF-1α accumulation, aerobic glycolysis and mitochondrial energy metabolism in human A549 lung cancer cells. Incubation of A549 cells with ferrous glycinate decreased the protein levels of HIF-1α, which was abrogated by proteosome inhibitor, or prolyl hydroxylase inhibitor. The addition of ferrous glycinate decreased protein levels of glucose transporter-1, hexokinase-2, and lactate dehydrogenase A, and decreased pyruvate dehydrogenase kinase-1 (PDK-1) and pyruvate dehydrogenase (PDH) phosphorylation in A549 cells. Ferrous glycinate also increased the expression of the mitochondrial transcription factor A (TFAM), and the mitochondrial protein, cytochrome c oxidase (COX-IV). Silencing of HIF-1α expression mimicked the effects of ferrous glycinate on PDK-1, PDH, TFAM and COX-IV in A549 cells. Ferrous glycinate increased mitochondrial membrane potential and ATP production in A549 cells. These results suggest that ferrous glycinate may reverse Warburg effect through down regulating HIF-1α in A549 cells.

  17. Withaferin A induces mitochondrial-dependent apoptosis in non-small cell lung cancer cells via generation of reactive oxygen species.

    PubMed

    Liu, Xi; Chen, Lei; Liang, Tao; Tian, Xiao-Dong; Liu, Yang; Zhang, Tao

    2017-01-01

    Withaferin A (WA) is a bioactive lactone, isolated from natural sources, mainly found in Withania somnifera, and was known to be highly effective against a variety of tumor cells both in vitro and in vivo. Accumulating experimental evidence suggests the involvement of reactive oxygen species (ROS) in WA-mediated cytotoxicity against cancer cells. Hence, the purpose of this study was to investigate the effect of WA in non-small cell lung cancer (NSCLC) cells and also the role of ROC in WA-mediated cytotoxicity. In the present study we investigated the cytotoxic potential of WA against NSCLC cell line A549 and also highlighted the mechanism of cytotoxicity of this compound. Non-carcinoma WI-38 and PBMC cell lines were used as controls. WA treatment resulted in a dose-dependent cytotoxicity in A549 cells, while the non-carcinoma cells WI-38 and PBMC were unaffected. Further experimental approaches revealed that ROS plays a major role in WAinduced apoptosis in NSCLC cells. WA induces oxidative damage to NSCLC cells with minimum toxicity to normal cells.

  18. Inhibition of Raf-MEK-ERK and Hypoxia pathways by Phyllanthus prevents metastasis in human lung (A549) cancer cell line

    PubMed Central

    2013-01-01

    Background Lung cancer constitutes one of the malignancies with the greatest incidence and mortality rates with 1.6 million new cases and 1.4 million deaths each year. Prognosis remains poor due to deleterious development of multidrug resistance resulting in less than 15% lung cancer patients reaching five years survival. We have previously shown that Phyllanthus induced apoptosis in conjunction with its antimetastastic action. In the current study, we aimed to determine the signaling pathways utilized by Phyllanthus to exert its antimetastatic activities. Methods Cancer 10-pathway reporter array was performed to screen the pathways affected by Phyllanthus in lung carcinoma cell line (A549) to exert its antimetastatic effects. Results from this array were then confirmed with western blotting, cell cycle analysis, zymography technique, and cell based ELISA assay for human total iNOS. Two-dimensional gel electrophoresis was subsequently carried out to study the differential protein expressions in A549 after treatment with Phyllanthus. Results Phyllanthus was observed to cause antimetastatic activities by inhibiting ERK1/2 pathway via suppression of Raf protein. Inhibition of this pathway resulted in the suppression of MMP2, MMP7, and MMP9 expression to stop A549 metastasis. Phyllanthus also inhibits hypoxia pathway via inhibition of HIF-1α that led to reduced VEGF and iNOS expressions. Proteomic analysis revealed a number of proteins downregulated by Phyllanthus that were involved in metastatic processes, including invasion and mobility proteins (cytoskeletal proteins), transcriptional proteins (proliferating cell nuclear antigen; zinc finger protein), antiapoptotic protein (Bcl2) and various glycolytic enzymes. Among the four Phyllanthus species tested, P. urinaria showed the greatest antimetastatic activity. Conclusions Phyllanthus inhibits A549 metastasis by suppressing ERK1/2 and hypoxia pathways that led to suppression of various critical proteins for A549

  19. Inhibition of Raf-MEK-ERK and hypoxia pathways by Phyllanthus prevents metastasis in human lung (A549) cancer cell line.

    PubMed

    Lee, Sau Har; Jaganath, Indu Bala; Manikam, Rishya; Sekaran, Shamala Devi

    2013-10-20

    Lung cancer constitutes one of the malignancies with the greatest incidence and mortality rates with 1.6 million new cases and 1.4 million deaths each year. Prognosis remains poor due to deleterious development of multidrug resistance resulting in less than 15% lung cancer patients reaching five years survival. We have previously shown that Phyllanthus induced apoptosis in conjunction with its antimetastastic action. In the current study, we aimed to determine the signaling pathways utilized by Phyllanthus to exert its antimetastatic activities. Cancer 10-pathway reporter array was performed to screen the pathways affected by Phyllanthus in lung carcinoma cell line (A549) to exert its antimetastatic effects. Results from this array were then confirmed with western blotting, cell cycle analysis, zymography technique, and cell based ELISA assay for human total iNOS. Two-dimensional gel electrophoresis was subsequently carried out to study the differential protein expressions in A549 after treatment with Phyllanthus. Phyllanthus was observed to cause antimetastatic activities by inhibiting ERK1/2 pathway via suppression of Raf protein. Inhibition of this pathway resulted in the suppression of MMP2, MMP7, and MMP9 expression to stop A549 metastasis. Phyllanthus also inhibits hypoxia pathway via inhibition of HIF-1α that led to reduced VEGF and iNOS expressions. Proteomic analysis revealed a number of proteins downregulated by Phyllanthus that were involved in metastatic processes, including invasion and mobility proteins (cytoskeletal proteins), transcriptional proteins (proliferating cell nuclear antigen; zinc finger protein), antiapoptotic protein (Bcl2) and various glycolytic enzymes. Among the four Phyllanthus species tested, P. urinaria showed the greatest antimetastatic activity. Phyllanthus inhibits A549 metastasis by suppressing ERK1/2 and hypoxia pathways that led to suppression of various critical proteins for A549 invasion and migration.

  20. SU-F-T-677: Synergistic Effect(s) of Clotrimazole On Radiation Cell Survival of A549 Lung Cancer Cells in Glucose Vs. Galactose Media

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Boss, G; Tambasco, M; Garakani, M

    Purpose: In order to determine the synergistic effect of clotrimazole on radiosensitivity of A549 lung cancer cells, and the effect of oxidative pathways on modulating radiosensitivity, we studied how these cells survived under varying amounts of radiation and clotrimazole as well ass when glucose was switched for galactose media. Methods: The glucose media was used to determine the presence of any synergistic effect of clotrimazole on radiation using values of radiation and clotrimazole concentrations, varying from 0 – 8 Gy and 0 – 20 µM, respectively. As a galactose diet is known to activate oxidative pathways, which do not relymore » on hexokinase II (HK2), all trials were repeated using galactose media to determine the extent that HK2 unbinding from the mitochondrial membrane plays a role in modulating the observed radiosensitivity. An apoptosis vs. necrosis assay was implemented to find out the modality by which cell death occurred. An intracellular lactate assay was performed to exhibit the extent of anaerobic glycolysis. Results: After running the primary experiments, it was found that in glucose media, the cancer cells showed higher cell kill when clotrimazole was added to the media, followed by the cells being irradiated. Conclusion: Given the preliminary results it is validated that under higher concentrations of clotrimazole, in glucose media, A549 lung cancer cells exhibit a lower amount of survival. While all results have not yet been gathered. We anticipate that in galactose media the A549 cells will exhibit this effect to a much smaller degree, if at all.« less

  1. Copper(II) complexes with naringenin and hesperetin: cytotoxic activity against A 549 human lung adenocarcinoma cells and investigation on the mode of action.

    PubMed

    Tamayo, Lenka V; Gouvea, Ligiane R; Sousa, Anna C; Albuquerque, Ronniel M; Teixeira, Sarah Fernandes; de Azevedo, Ricardo Alexandre; Louro, Sonia R W; Ferreira, Adilson Kleber; Beraldo, Heloisa

    2016-02-01

    Copper(II) complexes [Cu(H2O)2 (L1)(phen)](ClO4) (1) and [Cu(H2O)(L2)(phen)](ClO4) (2) (HL1 = naringenin; HL2 = hesperetin) were obtained, in which an anionic flavonoid ligand is attached to the metal center along with 1,10-phenanthroline (phen) as co-ligand. Complexes (1) and (2) were assayed for their cytotoxic activity against A549 lung carcinoma and against normal lung fibroblasts (LL-24) and human umbilical vein endothelial cells (HUVEC). We found IC50 = 16.42 µM (1) and IC50 = 5.82 µM (2) against A549 tumor cells. Complexes (1) and (2) exhibited slight specificity, being more cytotoxic against malignant than against non-malignant cells. 1 and 2 induced apoptosis on A549 cells in a mitochondria-independent pathway, and showed antioxidant activity. The antioxidant effect of the complexes could possibly improve their apoptotic action, most likely by a PI3K-independent reduction of autophagy. Complexes (1) and (2) interact in vitro with calf thymus DNA by an intercalative binding mode. EPR data indicated that 1 and 2 interact with human serum albumin (HSA) forming mixed ligand species.

  2. A549 Cells: Lung Carcinoma Cell Line for Adenovirus | NCI Technology Transfer Center | TTC

    Cancer.gov

    Scientists at the National Cancer Institute have developed a cell line designated A549 that was derived from explanted cultures of human lung cancer tissue. The A549 cell line has been tested under the guidance of the United States Food and Drug Administration (FDA) so, under current Good Manufacturing Practices (GMP), these cells may be suitable for use in manufacturing constructs for use in clinical trials. The National Cancer Institute seeks parties to non-exclusively license this research material.

  3. [Overexpression of Keap1 inhibits the cell proliferation and metastasis and overcomes the drug resistance in human lung cancer A549 cells].

    PubMed

    Weng, X; Yan, Y Y; Tong, Y H; Fan, Y; Zeng, J M; Wang, L L; Lin, N M

    2016-06-23

    To investigate the effect of Keap1-Nrf2 pathway on cell proliferation, metastasis and drug resistance of human lung cancer A549 cell line. A549-Keap1 cell line, constantly expressing wild type Keap1, was established by lentiviral transfection. Real-time RT-PCR and western blot were used to determine the expression of Nrf2 and its target gene in A549 cells. Sulforhodamine B (SRB) assay, flow cytometry, colony formation assay, transwell assay, and cell wound-healing assay were performed to explore the effect of wild type Keap1 expression on the proliferation, invasion, migration and drug resistance of A549 cells. Over-expressed Keap1 decreased the expression of Nrf2 protein and the mRNA level of its downstream target genes and inhibited the ability of cell proliferation and clone formation of A549 cells. Keap1 overexpression induced G0/G1 phase arrest. The percentage of A549-Keap1 cells in G0/G1 phase was significantly higher than that of A549-GFP cells (80.2±5.9)% vs. (67.1±0.9%)(P<0.05). Compared with the invasive A549-Keap1 cells (156.33±17.37), the number of invasive A549-GFP cells was significantly higher (306.67±22.19) in a high power field. Keap1 overexpression significantly enhanced the sensitivity of A549 cells to carboplatin and gemcitabine (P<0.01). The IC50s of carboplatin in A549-Keap1 and A549-GFP cells were (52.1±3.3) μmol/L and (107.8±12.9) μmol/L, respectively. The IC50s of gemcitabine in A549-Keap1 and A549-GFP cells were (6.8±1.2) μmol/L and (9.9±0.5) μmol/L, respectively. Keap1 overexpression significantly inhibits the expression of Nrf2 and its downstream target genes, suppresses tumor cell proliferation and metastasis, and enhances the sensitivity of A549 cells to anticancer drugs.

  4. MicroRNA-9 functions as a tumor suppressor and enhances radio-sensitivity in radio-resistant A549 cells by targeting neuropilin 1.

    PubMed

    Xiong, Kai; Shao, Li Hong; Zhang, Hai Qin; Jin, Linlin; Wei, Wei; Dong, Zhuo; Zhu, Yue Quan; Wu, Ning; Jin, Shun Zi; Xue, Li Xiang

    2018-03-01

    Radiotherapy is commonly used to treat lung cancer but may not kill all cancer cells, which may be attributed to the radiotherapy resistance that often occurs in non-small cell lung cancer (NSCLC). At present, the molecular mechanism of radio-resistance remains unclear. Neuropilin 1 (NRP1), a co-receptor for vascular endothelial growth factor (VEGF), was demonstrated to be associated with radio-resistance of NSCLC cells via the VEGF-phosphoinositide 3-kinase-nuclear factor-κB pathway in our previous study. It was hypothesized that certain microRNAs (miRs) may serve crucial functions in radio-sensitivity by regulating NRP1. Bioinformatics predicted that NRP1 was a potential target of miR-9, and this was validated by luciferase reporter assays. Functionally, miR-9-transfected A549 cells exhibited a decreased proliferation rate, increased apoptosis rate and attenuated migratory and invasive abilities. Additionally, a high expression of miR-9 also significantly enhanced the radio-sensitivity of A549 cells in vitro and in vivo . These data improve understanding of the mechanisms of cell radio-resistance, and suggest that miR-9 may be a molecular target for the prediction of radio-sensitivity in NSCLC.

  5. Emodin induces apoptosis of lung cancer cells through ER stress and the TRIB3/NF-κB pathway.

    PubMed

    Su, Jin; Yan, Yan; Qu, Jingkun; Xue, Xuewen; Liu, Zi; Cai, Hui

    2017-03-01

    Emodin is a phytochemical with potent anticancer activities against various human malignant cancer types, including lung cancer; however, the molecular mechanisms underlying the effects of emodin remain unclear. In the present study, the A549 and H1299 human non-small lung cancer cell lines were treated with emodin and the induced molecular effects were investigated. Changes in cell viability were evaluated by MTT assay, Hoechst staining was used to indicate the apoptotic cells, and western blotting was utilized to assess endoplasmic reticulum (ER) stress and signaling changes. RNA interference was also employed to further examine the role of tribbles homolog 3 (TRIB3) in the emodin-induced apoptosis of lung cancer cells. Emodin was found to reduce the viability of lung cancer cells and induce apoptosis in a concentration-dependent manner. Emodin-induced apoptosis was impaired by inhibition of ER stress using 4-phenylbutyrate (4-PBA). ER stress and TRIB3/nuclear factor-κB signaling was activated in emodin-treated lung cancer cells. Emodin-induced apoptosis was reduced by TRIB3 knockdown in A549 cells, whereas ER stress was not reduced. In vivo assays verified the significance of these results, revealing that emodin inhibited lung cancer growth and that the inhibitory effects were reduced by inhibition of ER stress with 4-PBA. In conclusion, the results suggest that TRIB3 signaling is associated with emodin-induced ER stress-mediated apoptosis in lung cancer cells.

  6. Aptamer based electrochemical sensor for detection of human lung adenocarcinoma A549 cells

    NASA Astrophysics Data System (ADS)

    Sharma, Rachna; Varun Agrawal, Ved; Sharma, Pradeep; Varshney, R.; Sinha, R. K.; Malhotra, B. D.

    2012-04-01

    We report results of the studies relating to development of an aptamer-based electrochemical biosensor for detection of human lung adenocarcinoma A549 cells. The aminated 85-mer DNA aptamer probe specific for the A549 cells has been covalently immobilized onto silane self assembled monolayer (SAM) onto ITO surface using glutaraldehyde as the crosslinker. The results of cyclic voltammetry and differential pulse voltammetry studies reveal that the aptamer functionalized bioelectrode can specifically detect lung cancer cells in the concentration range of 103 to 107 cells/ml with detection limit of 103 cells/ml within 60 s. The specificity studies of the bioelectrode have been carried out with control KB cells. No significant change in response is observed for control KB cells as compared to that of the A549 target cells.

  7. Jolkinolide A and Jolkinolide B Inhibit Proliferation of A549 Cells and Activity of Human Umbilical Vein Endothelial Cells.

    PubMed

    Shen, Lei; Zhang, Shan-Qiang; Liu, Lei; Sun, Yu; Wu, Yu-Xuan; Xie, Li-Ping; Liu, Ji-Cheng

    2017-01-14

    BACKGROUND Jolkinolide A (JA) and Jolkinolide B (JB) are diterpenoids extracted from the roots of Euphorbia fischeriana Steud and have been shown to have anti-tumor activity. However, their effects on the ability of tumor cells to invade blood vessels and metastasize remain largely unknown. Investigations into the effects of JA and JB on the angiogenesis of tumor tissues may facilitate the identification of new natural drugs with anti-tumor growth and metastasis activities. MATERIAL AND METHODS We used different concentrations of JA and JB (20 μg/ml, 40 μg/ml, 60 μg/ml, 80 μg/ml, and 100 μg/ml) to stimulate A549 cells and then studied the effects on the growth and metastasis of lung cancers. In addition, we used conditional media from A549 cells (A549-CM) stimulated by either JA or JB in different concentrations to culture human umbilical vein endothelial cells (HUVECs). RESULTS We found that both JA and JB significantly inhibited the Akt-STAT3-mTOR signaling pathway and reduced the expression of VEGF in A549 cells, but JB exhibited more significant inhibitory effects than JA. The JB-stimulated A549 cell conditional media had a greater inhibitory effect on the proliferation and migration of HUVECs than did the conditional media of JA-stimulated A549 cells. This effect gradually increased with increasing concentrations of either type of Jolkinolide. CONCLUSIONS Our results suggest that JA and JB inhibited VEGF expression in A549 cells through the inhibition of the Akt-STAT3-mTOR signaling pathway, and directly inhibited the proliferation and migration of HUVECs. These findings are of great significance for the development of new plant-derived chemotherapy agents for the treatment of cancer.

  8. Jolkinolide A and Jolkinolide B Inhibit Proliferation of A549 Cells and Activity of Human Umbilical Vein Endothelial Cells

    PubMed Central

    Shen, Lei; Zhang, Shan-Qiang; Liu, Lei; Sun, Yu; Wu, Yu-Xuan; Xie, Li-Ping; Liu, Ji-Cheng

    2017-01-01

    Background Jolkinolide A (JA) and Jolkinolide B (JB) are diterpenoids extracted from the roots of Euphorbia fischeriana Steud and have been shown to have anti-tumor activity. However, their effects on the ability of tumor cells to invade blood vessels and metastasize remain largely unknown. Investigations into the effects of JA and JB on the angiogenesis of tumor tissues may facilitate the identification of new natural drugs with anti-tumor growth and metastasis activities. Material/Methods We used different concentrations of JA and JB (20 μg/ml, 40 μg/ml, 60 μg/ml, 80 μg/ml, and 100 μg/ml) to stimulate A549 cells and then studied the effects on the growth and metastasis of lung cancers. In addition, we used conditional media from A549 cells (A549-CM) stimulated by either JA or JB in different concentrations to culture human umbilical vein endothelial cells (HUVECs). Results We found that both JA and JB significantly inhibited the Akt-STAT3-mTOR signaling pathway and reduced the expression of VEGF in A549 cells, but JB exhibited more significant inhibitory effects than JA. The JB-stimulated A549 cell conditional media had a greater inhibitory effect on the proliferation and migration of HUVECs than did the conditional media of JA-stimulated A549 cells. This effect gradually increased with increasing concentrations of either type of Jolkinolide. Conclusions Our results suggest that JA and JB inhibited VEGF expression in A549 cells through the inhibition of the Akt-STAT3-mTOR signaling pathway, and directly inhibited the proliferation and migration of HUVECs. These findings are of great significance for the development of new plant-derived chemotherapy agents for the treatment of cancer. PMID:28087861

  9. [Construction of BAD Lentivirus Vector and Its Effect on Proliferation in A549 Cell Lines].

    PubMed

    Huang, Na; He, Yan-qi; Zhu, Jing; Li, Wei-min

    2015-05-01

    To construct the recombinant lentivirus expressing vector BAD (Bcl-2-associated death protein) gene and to study its effect on A549 cell proliferation. The BAD gene was amplified from plasmid pAV-MCMV-BAD-GFP by PCR. The purified BAD gene fragment was inserted into a lentivirus vector (pLVX-IRES-ZsGreen 1), and the insertion was identified by PCR, restriction endonuclease analysis and DNA sequencing. A549 cells were then transfected with the packaged recombinant lentivirus, and resistant cell clones were selected with flow cytometry. The expression of BAD in A549 cell lines stably transduction with a lentivirus was examined using Western blot. The effect of BAD overexpression on proliferation of A549 cells was evaluated by using CCK-8 kit. Restriction enzyme digestion and DNA sequencing showed that the full-length BAD gene (507 bp) had been successfully subcloned into the lentiviral vector to result in the recombinant vector pLVX-IRES-ZsGreen 1. Monoclonal cell lines BAD-A549 was produced after transfection with the recombinant lentivirus and selected with flow cytometry. Stable expression of BAD protein was verified by Western blot. In vitro, the OD value in BAD group was significantly lower than that of control groups from 120-144 h (P<0. 05). A549 cell lines stably transduced with a lentivirus expressing the BAD gene had been successfully generated. In vitro, BAD overexpression significantly inhibited A549 cells proliferation.

  10. Metuzumab enhanced chemosensitivity and apoptosis in non-small cell lung carcinoma

    PubMed Central

    Feng, Fei; Wang, Bin; Sun, Xiuxuan; Zhu, Yumeng; Tang, Hao; Nan, Gang; Wang, Lijuan; Wu, Bo; Huhe, Muren; Liu, Shuangshuang; Diao, Tengyue; Hou, Rong; Zhang, Yang; Zhang, Zheng

    2017-01-01

    ABSTRACT Targeted therapeutics is used as an alternative treatment of non-small cell lung cancer (NSCLC); however, treatment effect is far from being satisfactory, and therefore identification of new targets is needed. We have previously shown that metuzumab inhibit tumor growth in vivo. The present study was performed to investigate the anti-tumor efficacy of metuzumab combined with gemcitabine and cisplatin (GP), paclitaxel and cisplatin (TP) or navelbine and cisplatin (NP) regimens in multiple NSCLC cell lines. Our results demonstrate that, in comparison to single agent metuzumab or GP treated cells, metuzumab combined with GP display inhibitory effects on tumor growth. Furthermore, we found that metuzumab elevated the sensitivity of cell lines to gemcitabine, which was identified by MTT assay. Flow cytometric analysis showed that metuzumab combined with gemcitabine (GEM) treatment led to an obvious G1 arrest and an elevated apoptosis in A549, NCI-H460 and NCI-H520 cells. Western blot analysis also demonstrated a significantly reduced level of cyclin D1, Bcl-2, and an obviously increase level of Bax and full-length caspase-3 in A549, NCI-H460 and NCI-H520 cells treated with metuzumab/gemcitabine combination in comparison with single agent treated cells. In addition, metuzumab/gemcitabine treated A549, NCI-H460 and NCI-H520 cells also demonstrated a significantly increase in deoxycytidine kinase (dCK) protein level compared with single agent metuzumab or gemcitabine treated cells. Xenograft models also demonstrated that this metuzumab/gemcitabine combination led to upregulation of dCK. Taken together, the mechanisms of metuzumab combined with GP repress tumor growth were that the combined treatment significantly inhibited the tumor cell proliferation, apoptosis and cell cycle in vitro and in vivo and at least partially by induction of dCK expression. Our results suggested that metuzumab could significantly enhance chemosensitivity of human NSCLC cells to

  11. Mesenchymal stem cells promote cell invasion and migration and autophagy-induced epithelial-mesenchymal transition in A549 lung adenocarcinoma cells.

    PubMed

    Luo, Dan; Hu, Shiyuan; Tang, Chunlan; Liu, Guoxiang

    2018-03-01

    Mesenchymal stem cells (MSCs) are recruited into the tumour microenvironment and promote tumour growth and metastasis. Tumour microenvironment-induced autophagy is considered to suppress primary tumour formation by impairing migration and invasion. Whether these recruited MSCs regulate tumour autophagy and whether autophagy affects tumour growth are controversial. Our data showed that MSCs promote autophagy activation, reactive oxygen species production, and epithelial-mesenchymal transition (EMT) as well as increased migration and invasion in A549 cells. Decreased expression of E-cadherin and increased expression of vimentin and Snail were observed in A549 cells cocultured with MSCs. Conversely, MSC coculture-mediated autophagy positively promoted tumour EMT. Autophagy inhibition suppressed MSC coculture-mediated EMT and reduced A549 cell migration and invasion slightly. Furthermore, the migratory and invasive abilities of A549 cells were additional increased when autophagy was further enhanced by rapamycin treatment. Taken together, this work suggests that microenvironments containing MSCs can promote autophagy activation for enhancing EMT; MSCs also increase the migratory and invasive abilities of A549 lung adenocarcinoma cells. Mesenchymal stem cell-containing microenvironments and MSC-induced autophagy signalling may be potential targets for blocking lung cancer cell migration and invasion. Copyright © 2018 John Wiley & Sons, Ltd.

  12. Wnt5a Increases Properties of Lung Cancer Stem Cells and Resistance to Cisplatin through Activation of Wnt5a/PKC Signaling Pathway

    PubMed Central

    Yang, Jiali; Zhang, Kangjian; Wu, Jing; Shi, Juan; Xue, Jing; Li, Jing; Zhu, Yongzhao; Wei, Jun

    2016-01-01

    The development of chemoresistance to cisplatin regimens causes a poor prognosis in patients with advanced NSCLC. The role of noncanonical Wnt signaling in the regulation of properties of lung cancer stem cells and chemoresistance was interrogated, by accessing capacities of cell proliferation, migration, invasion, and clonogenicity as well as the apoptosis in A549 cell lines and cisplatin-resistant A549 cells treated with Wnt5a conditional medium or protein kinase C (PKC) inhibitor GF109203X. Results showed that the noncanonical Wnt signaling ligand, Wnt5a, could promote the proliferation, migration, invasion, and colony formation in A549 lung adenocarcinoma cells and cisplatin-resistant A549/DDP cells and increase the fraction of ALDH-positive cell in A549/DDP cells. An exposure of cells to Wnt5a led to a significant reduction of A549/DDP cell apoptosis but not A549 cells. An addition of GF109203X could both strikingly increase the baseline apoptosis and resensitize the Wnt5a-inhibited cell apoptosis. Interestingly, an inhibition of Wnt/PKC signaling pathway could reduce properties of lung cancer stem cells, promote cell apoptosis, and resensitize cisplatin-resistant cells to cisplatin via a caspase/AIF-dependent pathway. These data thus suggested that the Wnt5a could promote lung cancer cell mobility and cisplatin-resistance through a Wnt/PKC signaling pathway and a blockage of this signaling may be an alternative therapeutic strategy for NSCLC patients with resistance to chemotherapies. PMID:27895670

  13. A novel alkaloid, evodiamine causes nuclear localization of cytochrome-c and induces apoptosis independent of p53 in human lung cancer cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Mohan, Vijay; Agarwal, Rajesh; Singh, Rana P., E-mail: ranaps@hotmail.com

    Lung cancer is the most frequently diagnosed malignancy that contributes to high proportion of deaths globally among patients who die due to cancer. Chemotherapy remains the common mode of treatment for lung cancer patients though with limited success. We assessed the biological effects and associated molecular changes of evodiamine, a plant alkaloid, on human lung cancer A549 and H1299 cells along with other epithelial cancer and normal lung SAEC cells. Our data showed that 20–40 μM evodiamine treatment for 24–48 h strongly (up to 73%, P < 0.001) reduced the growth and survival of these cancer cells. However, it also moderately inhibited growth and survivalmore » of SAEC cells. A strong inhibition (P < 0.001) was observed on clonogenicity of A549 cells. Further, evodiamine increased (4-fold) mitochondrial membrane depolarization with 6-fold increase in apoptosis and a slight increase in Bax/Bcl-2 ratio. It increased the cytochrome-c release from mitochondria into the cytosol as well as nucleus. Cytosolic cytochrome-c activated cascade of caspase-9 and caspase-3 intrinsic pathway, however, DR5 and caspase-8 extrinsic pathway was also activated which could be due to nuclear cytochrome-c. Pan-caspase inhibitor (z-VAD.fmk) partially reversed evodiamine induced apoptosis. An increase in p53 as well as its serine 15 phosphorylation was also observed. Pifithrin-α, a p53 inhibitor, slightly inhibited growth of A549 cells and under p53 inhibitory condition evodiamine-induced apoptosis could not be reversed. Together these findings suggest that evodiamine is a strong inducer of apoptosis in lung epithelial cancer cells independent of their p53 status and that could involve both intrinsic as well as extrinsic pathway of apoptosis. Thus evodiamine could be a potential anticancer agent against lung cancer. - Highlights: • Evodiamine, a novel plant alkaloid, relatively selectively inhibited growth and survival of human lung cancer cells. • Increased cancer

  14. Destruxin B Isolated from Entomopathogenic Fungus Metarhizium anisopliae Induces Apoptosis via a Bcl-2 Family-Dependent Mitochondrial Pathway in Human Nonsmall Cell Lung Cancer Cells

    PubMed Central

    Wu, Chun-Chi; Chen, Tzu-Hsiu; Liu, Bing-Lan; Wu, Li-Chen; Chen, Yung-Ching; Tzeng, Yew-Min; Hsu, Shih-Lan

    2013-01-01

    Destruxin B, isolated from entomopathogenic fungus Metarhizium anisopliae, is one of the cyclodepsipeptides with insecticidal and anticancer activities. In this study, destruxin B was extracted and purified by ion-exchange chromatography, silica gel chromatography, and semipreparative high-performance liquid chromatography. The potential anticancer effects and molecular mechanisms of destruxin B in human nonsmall cell lung cancer cell lines were characterized. Our results showed that destruxin B induced apoptotic cell death in A549 cells. This event was accompanied by the activation of caspase-2, -3, and -9. Moreover, destruxin B increased the expression level of proapoptotic molecule, PUMA, while decreased antiapoptotic molecule Mcl-1. Additionally, the translocation of Bax from cytosol to mitochondrial membrane was observed upon destruxin B treatment. Knockdown of Bax by shRNA effectively attenuated destruxin-B-triggered apoptosis in A549 cells. Interestingly, similar toxic effects and underlying mechanisms including caspase activation, upregulation of PUMA, and downregulation of Mcl-1 were also observed in a p53-null lung cancer H1299 cell line upon destruxin B treatment. Taken together, our findings suggest that destruxin-B-induced apoptosis in human nonsmall cell lung cancer cells is via a Bcl-2 family-dependent mitochondrial pathway. PMID:24204395

  15. Sepia ink oligopeptide induces apoptosis and growth inhibition in human lung cancer cells

    PubMed Central

    Zhou, Guoren; Xie, Peng; Ye, Jinjun

    2017-01-01

    Sepia ink oligopeptide (SIO), as a tripeptide extracted from Sepia ink, could be used as an inducer of apoptosis in human prostate cancer cells. We designed a cyclo-mimetic peptide of SIO by introducing a disulfide bond to stabilize the native peptide into beta turn structure, and produced a peptide with higher cell permeability and stability. Through labeling an FITC to the N-terminus of the peptide, the cell permeability was examined. Stabilized peptide showed enhanced cellular uptake than linear tripeptide as indicated by flow cytometry and cell fluorescent imaging. The high intracellular delivery of stable SIO could more efficiently inhibit cell proliferation and induce apoptosis. Furthermore, the expression of the anti-apoptotic protein Bcl-2 was down-regulated, whereas pro-apoptotic proteins P53 and caspase-3 were up-regulated by stable SIO. In conclusion, our study is the first to use stable SIO to induce apoptosis in two lung cancer cells A549 and H1299. PMID:28423568

  16. Apoptosis-inducing activity of HPLC fraction from Voacanga globosa (Blanco) Merr. on the human colon carcinoma cell.

    PubMed

    Acebedo, Alvin Resultay; Amor, Evangeline Cancio; Jacinto, Sonia Donaldo

    2014-01-01

    Voacanga globosa (Blanco), a plant endemic to the Philippines, is traditionally used especially by indigenous people of Bataan in the treatment of ulcers, wounds and tumorous growths. This study aimed to provide scientific evidence to therapeutic properties by determining cytotoxic and pro-apoptotic activity of HPLC fractions from leaves on HCT116 human colon carcinoma and A549 human lung carcinoma cell lines. Ethanolic extraction was performed on V globosa leaves followed by hexane and ethyl acetate partitioning. Silica gel column chromatography and high performance liquid chromatography (HPLC) produced MP1, MP2 and MP3 fractions. Cytotoxic activity of the fractions was determined through MTT assay against the cancer cell lines HCT116 and A549 and the non-cancer AA8 Chinese hamster ovarian cell line. Pro-apoptotic activities of the most active fractions were further assessed through DAPI staining, TUNEL assay and JC-1 mitochondrial membrane potential assay with HCT116 cells. While the MP1 fraction exerted no significant activity against all cell lines tested, MP2 and MP3 fractions demonstrated high toxicity against HCT116 and A549 cells. The MP3 fraction induced formation of apoptotic bodies, condensed DNA and other morphological changes consistent with apoptosis of HCT116 cells and TUNEL assay showed significant increase in DNA fragmentation over time. In these cells, the MP3 fraction also induced mitochondrial membrane destabilization, which is generally associated with the beginning of apoptosis. Phytochemical analysis demonstrated the presence only of saponins and terpenoids in the MP3 fraction. The results indicate that the MP3 fraction exerts cytotoxic activity on HCT116 cells via induction of apoptosis triggered by loss of mitochondrial membrane potential crucial for cell survival.

  17. Chlorella sorokiniana induces mitochondrial-mediated apoptosis in human non-small cell lung cancer cells and inhibits xenograft tumor growth in vivo.

    PubMed

    Lin, Ping-Yi; Tsai, Ching-Tsan; Chuang, Wan-Ling; Chao, Ya-Hsuan; Pan, I-Horng; Chen, Yu-Kuo; Lin, Chi-Chen; Wang, Bing-Yen

    2017-02-01

    Lung cancer is one of the leading causes of cancer related deaths worldwide. Marine microalgae are a source of biologically active compounds and are widely consumed as a nutritional supplement in East Asian countries. It has been reported that Chlorella or Chlorella extracts have various beneficial pharmacological compounds that modulate immune responses; however, no studies have investigated the anti-cancer effects of Chlorella sorokiniana (CS) on non-small cell lung cancer (NSCLC). In this study, we evaluated the anti-cancer effects of CS in two human NSCLC cell lines (A549 and CL1-5 human lung adenocarcinoma cells), and its effects on tumor growth in a subcutaneous xenograft tumor model. We also investigated the possible molecular mechanisms governing the pharmacological function of CS. Our results showed that exposure of the two cell lines to CS resulted in a concentration-dependent reduction in cell viability. In addition, the percentage of apoptotic cells increased in a dose-dependent manner, suggesting that CS might induce apoptosis in human NSCLC cells. Western blot analysis revealed that exposure to CS resulted in increased protein expression of the cleaved/activated forms of caspase-3, caspase-9, and PARP, except caspase-8. ZDEVD (caspase-3 inhibitor) and Z-LEHD (caspase-9 inhibitor) were sufficient at preventing apoptosis in both A549 and CL1-5 cells, proving that CS induced cell death via the mitochondria-mediated apoptotic pathway. Exposure of A549 and CL1-5 cells to CS for 24 h resulted in decreased expression of Bcl-2 protein and increased expression of Bax protein as well as decreased expression of two IAP family proteins, survivin and XIAP. We demonstrated that CS induces mitochondrial-mediated apoptosis in NSCLC cells via downregulation of Bcl-2, XIAP and survivin. In addition, we also found that the tumors growth of subcutaneous xenograft in vivo was markedly inhibited after oral intake of CS.

  18. Curcumin induced autophagy anticancer effects on human lung adenocarcinoma cell line A549

    PubMed Central

    Liu, Furong; Gao, Song; Yang, Yuxuan; Zhao, Xiaodan; Fan, Yameng; Ma, Wenxia; Yang, Danrong; Yang, Aimin; Yu, Yan

    2017-01-01

    To investigate the anticancer effects of curcumin-induced autophagy and its effects on the human lung adenocarcinoma A549 cell line, inverted phase contrast microscopy was used to observe alterations to the cytomorphology of cells. An MTT assay was used to measure cell viability. Autophagy was detected using acridine orange (AO) staining and 3-methyladenine (3-MA) was used as an autophagy-specific inhibitor. Dose- and time-dependent A549 cell viability inhibition was observed following curcumin treatment. A dose-dependent increase in the red fluorescent structures in A549 cells was identified following curcumin treatment for 48 h through AO staining. In addition, the activation of autophagy was determined through changes in the number of autophagic vesicles (AVs; fluorescent particles) infected with monodansylcadaverine (MDC). The fluorescence intensity and density of AVs in the curcumin-treated groups were higher at 48 h compared with the control group. Finally, the MTT assay demonstrated that the survival rates of the curcumin-treated cells were increased when pretreated with 3-MA for 3 h, indicating that the inhibitory effect of curcumin on A549 cells is reduced following the inhibition of autophagy. Furthermore, AO and MDC staining confirmed that 3-MA does inhibit the induction of autophagy. Thus, it was hypothesized that the induction of autophagy is partially involved in the reduction of cell viability observed following curcumin treatment. The anticancer effects of curcumin on A549 cells can be reduced using autophagy inhibitors. This suggests a possible cancer therapeutic application of curcumin through the activation of autophagy. These findings have improved the understanding of the mechanism underlying the anticancer property of curcumin. PMID:28928819

  19. Curcumin induced autophagy anticancer effects on human lung adenocarcinoma cell line A549.

    PubMed

    Liu, Furong; Gao, Song; Yang, Yuxuan; Zhao, Xiaodan; Fan, Yameng; Ma, Wenxia; Yang, Danrong; Yang, Aimin; Yu, Yan

    2017-09-01

    To investigate the anticancer effects of curcumin-induced autophagy and its effects on the human lung adenocarcinoma A549 cell line, inverted phase contrast microscopy was used to observe alterations to the cytomorphology of cells. An MTT assay was used to measure cell viability. Autophagy was detected using acridine orange (AO) staining and 3-methyladenine (3-MA) was used as an autophagy-specific inhibitor. Dose- and time-dependent A549 cell viability inhibition was observed following curcumin treatment. A dose-dependent increase in the red fluorescent structures in A549 cells was identified following curcumin treatment for 48 h through AO staining. In addition, the activation of autophagy was determined through changes in the number of autophagic vesicles (AVs; fluorescent particles) infected with monodansylcadaverine (MDC). The fluorescence intensity and density of AVs in the curcumin-treated groups were higher at 48 h compared with the control group. Finally, the MTT assay demonstrated that the survival rates of the curcumin-treated cells were increased when pretreated with 3-MA for 3 h, indicating that the inhibitory effect of curcumin on A549 cells is reduced following the inhibition of autophagy. Furthermore, AO and MDC staining confirmed that 3-MA does inhibit the induction of autophagy. Thus, it was hypothesized that the induction of autophagy is partially involved in the reduction of cell viability observed following curcumin treatment. The anticancer effects of curcumin on A549 cells can be reduced using autophagy inhibitors. This suggests a possible cancer therapeutic application of curcumin through the activation of autophagy. These findings have improved the understanding of the mechanism underlying the anticancer property of curcumin.

  20. β, β-Dimethylacrylshikonin induces mitochondria-dependent apoptosis of human lung adenocarcinoma cells in vitro via p38 pathway activation

    PubMed Central

    Wang, Hai-bing; Ma, Xiao-qiong

    2015-01-01

    Aim: β, β-Dimethylacrylshikonin (DMAS) is an anticancer compound extracted from the roots of Lithospermum erythrorhizon. In the present study, we investigated the effects of DMAS on human lung adenocarcinoma cells in vitro and explored the mechanisms of its anti-cancer action. Methods: Human lung adenocarcinoma A549 cells were tested. Cell viability was assessed using an MTT assay, and cell apoptosis was evaluated with flow cytometry and DAPI staining. The expression of the related proteins was detected using Western blotting. The mitochondrial membrane potential was measured using a JC-1 kit, and subcellular distribution of cytochrome c was analyzed using immunofluorescence staining. Results: Treatment of A549 cells with DMAS suppressed the cell viability in dose- and time-dependent manners (the IC50 value was 14.22 and 10.61 μmol/L, respectively, at 24 and 48 h). DMAS (7.5, 10, and 15 μmol/L) dose-dependently induced apoptosis, down-regulated cIAP-2 and XIAP expression, and up-regulated Bax and Bak expression in the cells. Furthermore, DMAS resulted in loss of mitochondrial membrane potential and release of cytochrome c in the cells, and activated caspase-9, caspase-8, and caspase-3, and subsequently cleaved PARP, which was abolished by pretreatment with Z-VAD-FMK, a pan-caspase inhibitor. DMAS induced sustained p38 phosphorylation in the cells, while pretreatment with SB203580, a specific p38 inhibitor, blocked DMAS-induced p38 activation and apoptosis. Conclusion: DMAS inhibits the growth of human lung adenocarcinoma A549 cells in vitro via activation of p38 signaling pathway. PMID:25434989

  1. [The effect and mechanism of vinorelbine on cisplatin resistance of human lung cancer cell line A549/DDP].

    PubMed

    Qi, Chunsheng; Gao, Sen; Li, Huiqiang; Gao, Weizhen

    2014-02-01

    Drug resistance is a major obstacle on lung cancer treatment and Vinorelbine is an effective drug to inhibition of tumor proliferation and metastasis. In this study, we investigated the effect and mechanism of Vinorelbine on reversing the cisplatin resistance of human lung cancer A549/DDP cell line. With 1 μmol/L and 5 μmol/L Vinorelbine treatment, MTS assay was employed to determine the effect of the cisplatin sensitivity of tumor cells, flow cytometry to determine the apoptosis rate and change of Rh-123 content; Western blot to determine the expression of MDR1, Bcl-2, surviving, PTEN, caspase-3/8 and phosphorylation level of Akt (p-Akt); Real-time PCR was to determine the mRNA expression of MDR1, Bcl-2, survivin and PTEN. Finally the transcriptional activities of NF-κB, Twist and Snail were determined by reporter gene system. With 1 μmol/L and 5 μmol/L Vinorelbine treatment, the sensitivity of cancer cells to cisplatin was increased by 1.91- and 2.54- folds respectively, flow cytometry showed that the content of Rh-123 was elevated 1.93- and 2.95- folds and apoptosis rate was increased 2.25- and 3.82- folds, Western blot showed that the expression of multidrug resistance related proteins MDR, Bcl-2 and survivin were downregulated, caspase-3/8 and PTEN was upregulated, phosphorylation of Akt was downregulated as well, real-time assay showed that the mRNA expression of MDR1 was downregulated 43.5% and 25.8%, Bcl-2 was downregulated 57.3% and 34.1%, survivin was downregulated 37.6% and 12.4%, PTEN was upregulated 183.4% and 154.2%, the transcriptional activities of NF-κB was downregulated 53.2% and 34.5%, Twist was downregulated 61.4% and 33.5%, and Snail was downregulated 57.8% and 18.7%. Vinorelbine treatment led to increase of cisplatin sensitivity of A549/DDP cells and the mechanisms included the regulation of PTEN/AKT/NF-κB signal pathway to decreased drug resistance gene expression and increased pro-apoptosis gene expression.

  2. Ac-SDKP suppresses epithelial-mesenchymal transition in A549 cells via HSP27 signaling.

    PubMed

    Deng, Haijing; Yang, Fang; Xu, Hong; Sun, Yue; Xue, Xinxin; Du, Shipu; Wang, Xiaojun; Li, Shifeng; Liu, Yan; Wang, Ruimin

    2014-08-01

    The synthetic tetrapeptide N-acetyl-seryl-aspartyl-lysyl-proline (Ac-SDKP) has been shown to be a modulator of molecular aspects of the fibrosis pathway. This study reveals that Ac-SDKP exerts an anti-fibrotic effect on human type II alveolar epithelial cells (A549), which are a source of myofibroblasts once exposed to TGF-β1, by decreasing the expression of heat shock protein 27 (HSP27). We used A549 cells in vitro to detect morphological evidence of epithelial-mesenchymal transition (EMT) by phase-contrast microscopy. Immunocytochemical and western blot analysis determined the distributions of cytokeratin 8 (CK8), α-smooth muscle actin (α-SMA), and SNAI1. Confocal laser scanning microscopy revealed a colocalization of HSP27 and SNAI1 on TGF-β1-induced A549 cells. These results also demonstrated that A549 cells became spindle-like when exposed to TGF-β1. Coincident with these morphological changes, expression levels of CK8 and E-cad decreased, while those of vimentin and α-SMA increased. This process was accompanied by increases in levels of HSP27, SNAI1, and type I and type III collagen. In vitro transfection experiments demonstrated that the inhibition of HSP27 in cultured A549 cells could decrease the expression of SNAI1 and α-SMA while increasing the expression of E-cad. A noticeable reduction in collagen types I and III was also evident. Our results found that Ac-SDKP inhibited the transition of cultured A549 cells to myofibroblasts and attenuated collagen synthesis through modulating the expression of HSP27. Copyright © 2014 Elsevier Inc. All rights reserved.

  3. Mitochondria-targeted platinum(II) complexes induce apoptosis-dependent autophagic cell death mediated by ER-stress in A549 cancer cells.

    PubMed

    Wang, Feng-Yang; Tang, Xiao-Ming; Wang, Xia; Huang, Ke-Bin; Feng, Hai-Wen; Chen, Zhen-Feng; Liu, You-Nian; Liang, Hong

    2018-06-09

    Agents with multiple modes of tumor cell death can be effective chemotherapeutic drugs. One example of a bimodal chemotherapeutic approach is an agent that can induce both apoptosis and autophagic death. Thus far, no clinical anticancer drug has been shown to simultaneously induce both these pathways. Mono-functional platinum complexes are potent anticancer drug candidates which act through mechanisms distinct from cisplatin. Here, we describe the synthesis and characterize of two mono-functional platinum complexes containing 8-substituted quinoline derivatives as ligands, [PtL 1 Cl]Cl [L 1  = (Z)-1-(pyridin-2-yl)-N-(quinolin-8-ylmethylene) methanamine] (Mon-Pt-1) and [PtL 2 Cl]Cl [L 2  = (Z)-2-(pyridin-2-yl)-N-(quinolin-8-ylmethylene) ethanamine] (Mon-Pt-2). In comparison to cisplatin, Mon-Pt-2 exhibited a greater in vitro cytotoxicity, was more effective in resistant cells and elicited a better anticancer effect. Mechanistic experiments indicate that Mon-Pt-2 mainly accumulates in mitochondria, and stimulates significant TrxR inhibition ROS release and an ER stress response, mediated by mitochondrial dysfunction, ultimately resulting in a simultaneous induction of apoptosis and autophagy. Importantly, compared to cisplatin, Mon-Pt-2 exhibits lower acute toxicity and better anticancer activity in a murine tumor model. To the best of our knowledge, Mon-Pt-2 is the first mono-functional platinum complex inducing pro-death autophagy and apoptosis of cancer cells. Copyright © 2018 The Authors. Published by Elsevier Masson SAS.. All rights reserved.

  4. Activation of AIFM2 enhances apoptosis of human lung cancer cells undergoing toxicological stress.

    PubMed

    Lu, Jun; Chen, Jian; Xu, Nianjun; Wu, Jun; Kang, Yani; Shen, Tingting; Kong, Hualei; Ma, Chao; Cheng, Ming; Shao, Zhifeng; Xu, Ling; Zhao, Xiaodong

    2016-09-06

    Application of cisplatin (DDP) for treating lung cancer is restricted due to its toxicity and lung cancer's drug resistance. In this study, we examined the effect of Jinfukang (JFK), an effective herbal medicine against lung cancer, on DDP-induced cytotoxicity in lung cancer cells. Morphologically, we observed that JFK increases DDP-induced pro-apoptosis in A549 cells in a synergistic manner. Transcriptome profiling analysis indicated that the combination of JFK and DDP regulates genes involved in apoptosis-related signaling pathways. Moreover, we found that the combination of JFK and DDP produces synergistic pro-apoptosis effect in other lung cancer cell lines, such as NCI-H1975, NCI-H1650, and NCI-H2228. Particularly, we demonstrated that AIFM2 is activated by the combined treatment of JFK and DDP and partially mediates the synergistic pro-apoptosis effect. Collectively, this study not only offered the first evidence that JFK promotes DDP-induced cytotoxicity, and activation of AIFM2 enhances apoptosis of human lung cancer cells undergoing toxicological stress, but also provided a novel insight for improving cytotoxicity by combining JFK with DDP to treat lung cancer cells. Copyright © 2016 Elsevier Ireland Ltd. All rights reserved.

  5. A novel synthetic analog of militarin, MA-1 induces mitochondrial dependent apoptosis by ROS generation in human lung cancer cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Yoon, Deok Hyo; Lim, Mi-Hee; Lee, Yu Ran

    A synthetic Militarin analog-1[(2R,3R,4R,5R)-1,6-bis(4-(2,4,4-trimethylpentan-2-yl)phenoxy) hexane-2,3,4,5-tetraol] is a novel derivative of constituents from Cordyceps militaris, which has been used to treat a variety of chronic diseases including inflammation, diabetes, hyperglycemia and cancers. Here, we report for the first time the synthesis of Militarin analog-1 (MA-1) and the apoptotic mechanism of MA-1 against human lung cancer cell lines. Treatment with MA-1 significantly inhibited the viability of 3 human lung cancer cell lines. The inhibition of viability and growth in MA-1-treated A549 cells with an IC{sub 50} of 5 μM were mediated through apoptosis induction, as demonstrated by an increase in DNA fragmentation,more » sub-G{sub 0}/G{sub 1}-DNA fraction, nuclear condensation, and phosphatidylserine exposure. The apoptotic cell death caused mitochondrial membrane permeabilization through regulation of expression of the Bcl-2 family proteins, leading to cytochrome c release in a time-dependent manner. Subsequently, the final stage of apoptosis, activation of caspase-9/-3 and cleavage of poly (ADP ribose) polymerase, was induced. Furthermore, A549 lung cancer cells were more responsive to MA-1 than a bronchial epithelial cell line (BEAS-2B), involving the rapid generation of reactive oxygen species (ROS), c-Jun N-terminal kinase (JNK) and p38 mitogen-activated protein kinase (MAPK) activation. The pharmacological inhibition of ROS generation and JNK/p38 MAPK exhibited attenuated DNA fragmentation in MA-1-induced apoptosis. Oral administration of MA-1 also retarded growth of A549 orthotopic xenografts. In conclusion, the present study indicates that the new synthetic derivative MA-1 triggers mitochondrial apoptosis through ROS generation and regulation of MAPKs and may be a potent therapeutic agent against human lung cancer. - Highlights: • We report a novel synthesized derivative, militarin analog-1 (MA-1). • MA-1-induced cancer cell death was

  6. Cytotoxic and genotoxic effects of defence secretion of Ulomoides dermestoides on A549 cells.

    PubMed

    Crespo, Rosana; Villaverde, M Luciana; Girotti, Juan R; Güerci, Alba; Juárez, M Patricia; de Bravo, Margarita G

    2011-06-14

    Ulomoides dermestoides (Fairmaire, 1893) is a cosmopolitan tenebrionid beetle reared by Argentine people who consume them alive as an alternative medicine in the treatment of different illnesses such as asthma, Parkinson's, diabetes, arthritis, HIV and specially cancer. To evaluate the cytotoxicity and DNA damage of the major volatile components released by Ulomoides dermestoides on human lung carcinoma epithelial cell line A549. The defence compounds of Ulomoides dermestoides were extracted with dichloromethane and analyzed and quantified by capillary gas chromatography. The toxicity effects of the beetle's extract against A549 cell line were evaluated. Cytotoxicity was evaluated by MTT test and Trypan blue assay and genotoxicity was evaluated by the comet assay. The synthetic compounds, individually or combined, were also tested in A549 cells and normal mononuclear human cells. The defence compounds of Ulomoides dermestoides extracted with dichloromethane (methyl-1,4-benzoquinones, ethyl-1,4-benzoquinones and 1-pentadecene as major components) showed cytotoxic activity on A549 cells demonstrated by MTT test and Trypan blue assay, with IC(50) values of 0.26equivalent/ml and 0.34equivalent/ml, respectively (1equivalent=amount of components extracted per beetle). The inhibition of A549 cell proliferation with the synthetic blend (1,4-benzoquinone and 1-pentadecene) or 1,4-benzoquinone alone was similar to that obtained with the insect extract. 1-Pentadecene showed no inhibitory effect. Low doses of insect extract or synthetic blend (0.15equivalent/ml) inhibited mononuclear cell proliferation by 72.2±2.7% and induced significant DNA damage both in tumor and mononuclear cells. Results of this study demonstrated that defence compounds of Ulomoides dermestoides reduced cell viability and induced DNA damage. We also concluded that the insect benzoquinones are primarily responsible for inducing cytotoxicity and genotoxicity in culture cells. Copyright © 2011 Elsevier

  7. Curcumin inhibits interferon-{alpha} induced NF-{kappa}B and COX-2 in human A549 non-small cell lung cancer cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Lee, Jeeyun; Im, Young-Hyuck; Jung, Hae Hyun

    2005-08-26

    The A549 cells, non-small cell lung cancer cell line from human, were resistant to interferon (IFN)-{alpha} treatment. The IFN-{alpha}-treated A549 cells showed increase in protein expression levels of NF-{kappa}B and COX-2. IFN-{alpha} induced NF-{kappa}B binding activity within 30 min and this increased binding activity was markedly suppressed with inclusion of curcumin. Curcumin also inhibited IFN-{alpha}-induced COX-2 expression in A549 cells. Within 10 min, IFN-{alpha} rapidly induced the binding activity of a {gamma}-{sup 32}P-labeled consensus GAS oligonucleotide probe, which was profoundly reversed by curcumin. Taken together, IFN-{alpha}-induced activations of NF-{kappa}B and COX-2 were inhibited by the addition of curcumin in A549more » cells.« less

  8. COX-2 and PPAR-γ confer cannabidiol-induced apoptosis of human lung cancer cells.

    PubMed

    Ramer, Robert; Heinemann, Katharina; Merkord, Jutta; Rohde, Helga; Salamon, Achim; Linnebacher, Michael; Hinz, Burkhard

    2013-01-01

    The antitumorigenic mechanism of cannabidiol is still controversial. This study investigates the role of COX-2 and PPAR-γ in cannabidiol's proapoptotic and tumor-regressive action. In lung cancer cell lines (A549, H460) and primary cells from a patient with lung cancer, cannabidiol elicited decreased viability associated with apoptosis. Apoptotic cell death by cannabidiol was suppressed by NS-398 (COX-2 inhibitor), GW9662 (PPAR-γ antagonist), and siRNA targeting COX-2 and PPAR-γ. Cannabidiol-induced apoptosis was paralleled by upregulation of COX-2 and PPAR-γ mRNA and protein expression with a maximum induction of COX-2 mRNA after 8 hours and continuous increases of PPAR-γ mRNA when compared with vehicle. In response to cannabidiol, tumor cell lines exhibited increased levels of COX-2-dependent prostaglandins (PG) among which PGD(2) and 15-deoxy-Δ(12,14)-PGJ(2) (15d-PGJ(2)) caused a translocation of PPAR-γ to the nucleus and induced a PPAR-γ-dependent apoptotic cell death. Moreover, in A549-xenografted nude mice, cannabidiol caused upregulation of COX-2 and PPAR-γ in tumor tissue and tumor regression that was reversible by GW9662. Together, our data show a novel proapoptotic mechanism of cannabidiol involving initial upregulation of COX-2 and PPAR-γ and a subsequent nuclear translocation of PPAR-γ by COX-2-dependent PGs.

  9. Multidimensional effects of biologically synthesized silver nanoparticles in Helicobacter pylori, Helicobacter felis, and human lung (L132) and lung carcinoma A549 cells.

    PubMed

    Gurunathan, Sangiliyandi; Jeong, Jae-Kyo; Han, Jae Woong; Zhang, Xi-Feng; Park, Jung Hyun; Kim, Jin-Hoi

    2015-01-01

    Silver nanoparticles (AgNPs) are prominent group of nanomaterials and are recognized for their diverse applications in various health sectors. This study aimed to synthesize the AgNPs using the leaf extract of Artemisia princeps as a bio-reductant. Furthermore, we evaluated the multidimensional effect of the biologically synthesized AgNPs in Helicobacter pylori, Helicobacter felis, and human lung (L132) and lung carcinoma (A549) cells. UV-visible (UV-vis) spectroscopy confirmed the synthesis of AgNPs. X-ray diffraction (XRD) indicated that the AgNPs are specifically indexed to a crystal structure. The results from Fourier transform infrared spectroscopy (FTIR) indicate that biomolecules are involved in the synthesis and stabilization of AgNPs. Dynamic light scattering (DLS) studies showed the average size distribution of the particle between 10 and 40 nm, and transmission electron microscopy (TEM) confirmed that the AgNPs were significantly well separated and spherical with an average size of 20 nm. AgNPs caused dose-dependent decrease in cell viability and biofilm formation and increase in reactive oxygen species (ROS) generation and DNA fragmentation in H. pylori and H. felis. Furthermore, AgNPs induced mitochondrial-mediated apoptosis in A549 cells; conversely, AgNPs had no significant effects on L132 cells. The results from this study suggest that AgNPs could cause cell-specific apoptosis in mammalian cells. Our findings demonstrate that this environmentally friendly method for the synthesis of AgNPs and that the prepared AgNPs have multidimensional effects such as anti-bacterial and anti-biofilm activity against H. pylori and H. felis and also cytotoxic effects against human cancer cells. This report describes comprehensively the effects of AgNPs on bacteria and mammalian cells. We believe that biologically synthesized AgNPs will open a new avenue towards various biotechnological and biomedical applications in the near future.

  10. Multidimensional effects of biologically synthesized silver nanoparticles in Helicobacter pylori, Helicobacter felis, and human lung (L132) and lung carcinoma A549 cells

    NASA Astrophysics Data System (ADS)

    Gurunathan, Sangiliyandi; Jeong, Jae-Kyo; Han, Jae Woong; Zhang, Xi-Feng; Park, Jung Hyun; Kim, Jin-Hoi

    2015-02-01

    Silver nanoparticles (AgNPs) are prominent group of nanomaterials and are recognized for their diverse applications in various health sectors. This study aimed to synthesize the AgNPs using the leaf extract of Artemisia princeps as a bio-reductant. Furthermore, we evaluated the multidimensional effect of the biologically synthesized AgNPs in Helicobacter pylori, Helicobacter felis, and human lung (L132) and lung carcinoma (A549) cells. UV-visible (UV-vis) spectroscopy confirmed the synthesis of AgNPs. X-ray diffraction (XRD) indicated that the AgNPs are specifically indexed to a crystal structure. The results from Fourier transform infrared spectroscopy (FTIR) indicate that biomolecules are involved in the synthesis and stabilization of AgNPs. Dynamic light scattering (DLS) studies showed the average size distribution of the particle between 10 and 40 nm, and transmission electron microscopy (TEM) confirmed that the AgNPs were significantly well separated and spherical with an average size of 20 nm. AgNPs caused dose-dependent decrease in cell viability and biofilm formation and increase in reactive oxygen species (ROS) generation and DNA fragmentation in H. pylori and H. felis. Furthermore, AgNPs induced mitochondrial-mediated apoptosis in A549 cells; conversely, AgNPs had no significant effects on L132 cells. The results from this study suggest that AgNPs could cause cell-specific apoptosis in mammalian cells. Our findings demonstrate that this environmentally friendly method for the synthesis of AgNPs and that the prepared AgNPs have multidimensional effects such as anti-bacterial and anti-biofilm activity against H. pylori and H. felis and also cytotoxic effects against human cancer cells. This report describes comprehensively the effects of AgNPs on bacteria and mammalian cells. We believe that biologically synthesized AgNPs will open a new avenue towards various biotechnological and biomedical applications in the near future.

  11. The enriched fraction of Vernonia cinerea L. induces apoptosis and inhibits multi-drug resistance transporters in human epithelial cancer cells.

    PubMed

    Appadath Beeran, Asmy; Maliyakkal, Naseer; Rao, Chamallamudi Mallikarjuna; Udupa, Nayanabhirama

    2014-12-02

    Vernonia cinerea Less. (VC) of the family Asteraceaes is considered as the sacred plant; 'Dasapushpam' which is ethnopharmacologically significant to the people of Kerala in India. In fact, VC has been used in the traditional system of medicine (Ayurveda) for the treatment of various ailments including cancer. Cytotoxicity of the ethanolic extract of VC (VC-ET), petroleum ether fraction (VC-PET), dichloromethane fraction (VC-DCM), n-butyl alcohol fraction (VC-BT), and rest fraction (VC-R) was evaluated in cervical carcinoma (HeLa), lung adenocarcinoma (A549), breast cancer (MCF-7), and colon carcinoma (Caco-2) cells using Sulforhodamine B (SRB) assay. The apoptotic effects of VC-DCM were assessed in cancer cells using Annexin V assay. The effects of VC-DCM on multi-drug resistance (MDR) transporters in HeLa, A549, MCF-7, and Caco-2 cells were evaluated using flow cytometry based functional assays. Similarly, drug uptake in cancer cells and sensitization of cancer cells towards chemotherapeutic drugs in the presence of VC-DCM were studied using Daunorubicin (DNR) accumulation assay and SRB assay, respectively. Cytotoxicity assay revealed that the enriched fraction of VC (VC-DCM) possessed dose-dependent cytotoxic effects in human epithelial cancer cells (HeLa, A549, MCF-7, and Caco-2). Further, treatment of cancer cells (HeLa, A549, MCF-7, and Caco-2) with VC-DCM led to a significant increase in both early and late apoptosis, indicating the induction of apoptosis. Interestingly, VC-DCM significantly inhibited functional activity of MDR transporters (ABC-B1 and ABC-G2), enhanced DNR-uptake in cancer cells, and sensitized cancer cells towards chemotherapeutic drug-mediated cytotoxicity, thus indicating the ability of VC-DCM to reverse MDR in cancer and enhance the cytotoxic effects of anticancer drugs. A methodological investigation on the anti-cancer properties of Vernonia cinerea Less. (VC) revealed that an enriched fraction of VC (VC-DCM) possessed cytotoxic

  12. Cisplatin resistance in non-small cell lung cancer cells is associated with an abrogation of cisplatin-induced G2/M cell cycle arrest

    PubMed Central

    Kalayda, Ganna V.; Mannewitz, Mareike; Cinatl, Jindrich; Rothweiler, Florian; Michaelis, Martin; Saafan, Hisham; Ritter, Christoph A.; Jaehde, Ulrich

    2017-01-01

    The efficacy of cisplatin-based chemotherapy in cancer is limited by the occurrence of innate and acquired drug resistance. In order to better understand the mechanisms underlying acquired cisplatin resistance, we have compared the adenocarcinoma-derived non-small cell lung cancer (NSCLC) cell line A549 and its cisplatin-resistant sub-line A549rCDDP2000 with regard to cisplatin resistance mechanisms including cellular platinum accumulation, DNA-adduct formation, cell cycle alterations, apoptosis induction and activation of key players of DNA damage response. In A549rCDDP2000 cells, a cisplatin-induced G2/M cell cycle arrest was lacking and apoptosis was reduced compared to A549 cells, although equitoxic cisplatin concentrations resulted in comparable platinum-DNA adduct levels. These differences were accompanied by changes in the expression of proteins involved in DNA damage response. In A549 cells, cisplatin exposure led to a significantly higher expression of genes coding for proteins mediating G2/M arrest and apoptosis (mouse double minute 2 homolog (MDM2), xeroderma pigmentosum complementation group C (XPC), stress inducible protein (SIP) and p21) compared to resistant cells. This was underlined by significantly higher protein levels of phosphorylated Ataxia telangiectasia mutated (pAtm) and p53 in A549 cells compared to their respective untreated control. The results were compiled in a preliminary model of resistance-associated signaling alterations. In conclusion, these findings suggest that acquired resistance of NSCLC cells against cisplatin is the consequence of altered signaling leading to reduced G2/M cell cycle arrest and apoptosis. PMID:28746345

  13. Mangiferin induces apoptosis in human ovarian adenocarcinoma OVCAR3 cells via the regulation of Notch3

    PubMed Central

    Zou, Bingyu; Wang, Hailian; Liu, Yilong; Qi, Ping; Lei, Tiantian; Sun, Minghan; Wang, Yi

    2017-01-01

    Ovarian cancer is the most lethal gynecological malignancy in the world. Our previous studies showed that mangiferin, purified from plant source, possessed anti-neoplasm effect on human lung adenocarcinoma A549 cells. This study aimed to determine the apoptosis-inducing effect of mangiferin on human ovarian carcinoma OVCAR3 cells. By in vitro studies, we found mangiferin significantly inhibited viability of OVCAR3 cells, and remarkably increased the sensitivity of OVCAR3 cells to cisplatin. In addition, the activation of caspase-dependent apoptosis was observed in mangiferin treated ovarian cancer cells. Importantly, we observed an obviously downregulated Notch expression after mangiferin treatment, indicating the crucial role of Notch in mangiferin mediated apoptosis. In contrast, overexpression of Notch3 abrogated the apoptosis-inducing efficacy of mangiferin, further demonstrating that mangiferin induced apoptosis via Notch pathway. Furthermore, OVCAR3 cell xenograft models revealed that mangiferin treatment inhibited tumor growth and expanded survival of tumor xenograft mice. Based on these results, we concluded that mangiferin could significantly inhibit the proliferation and induce apoptosis in OVCAR3 cells. Our study also suggested the anti-neoplasm effect of mangiferin might be via the regulation of Notch3. Taken together, by targeting cell apoptosis pathways and enhancing the response to cisplatin treatment, mangiferin may represent a potential new drug for the treatment of human ovarian cancer. PMID:28714011

  14. Mangiferin induces apoptosis in human ovarian adenocarcinoma OVCAR3 cells via the regulation of Notch3.

    PubMed

    Zou, Bingyu; Wang, Hailian; Liu, Yilong; Qi, Ping; Lei, Tiantian; Sun, Minghan; Wang, Yi

    2017-09-01

    Ovarian cancer is the most lethal gynecological malignancy in the world. Our previous studies showed that mangiferin, purified from plant source, possessed anti-neoplasm effect on human lung adenocarcinoma A549 cells. This study aimed to determine the apoptosis-inducing effect of mangiferin on human ovarian carcinoma OVCAR3 cells. By in vitro studies, we found mangiferin significantly inhibited viability of OVCAR3 cells, and remarkably increased the sensitivity of OVCAR3 cells to cisplatin. In addition, the activation of caspase-dependent apoptosis was observed in mangiferin treated ovarian cancer cells. Importantly, we observed an obviously downregulated Notch expression after mangiferin treatment, indicating the crucial role of Notch in mangiferin mediated apoptosis. In contrast, overexpression of Notch3 abrogated the apoptosis-inducing efficacy of mangiferin, further demonstrating that mangiferin induced apoptosis via Notch pathway. Furthermore, OVCAR3 cell xenograft models revealed that mangiferin treatment inhibited tumor growth and expanded survival of tumor xenograft mice. Based on these results, we concluded that mangiferin could significantly inhibit the proliferation and induce apoptosis in OVCAR3 cells. Our study also suggested the anti-neoplasm effect of mangiferin might be via the regulation of Notch3. Taken together, by targeting cell apoptosis pathways and enhancing the response to cisplatin treatment, mangiferin may represent a potential new drug for the treatment of human ovarian cancer.

  15. 4-Nitroquinoline-1-oxide effects human lung adenocarcinoma A549 cells by regulating the expression of POLD4

    PubMed Central

    HUANG, QIN-MIAO; ZENG, YI-MING; ZHANG, HUA-PING; LV, LIANG-CHAO; YANG, DONG-YONG; LIN, HUI-HUANG

    2016-01-01

    The aim of the present study was to explore the expression of POLD4 in human lung adenocarcinoma A549 cells under 4-nitroquinoline-1-oxide (4NQO) stimulation to investigate the role of POLD4 in smoking-induced lung cancer. The lung cancer A549 cell line was treated with 4NQO, with or without MG132 (an inhibitor of proteasome activity), and subsequently the POLD4 level was determined by western blot analysis. Secondly, the cell sensitivity to 4NQO and Taxol was determined when the POLD4 expression level was downregulated by siRNA. The POLD4 protein levels in the A549 cells decreased following treatment with 4NQO; however, MG132 could reverse this phenotype. Downregulation of the POLD4 expression by siRNA enhanced A549 cell sensitivity to 4NQO, but not to Taxol. In conclusion, 4NQO affects human lung adenocarcinoma A549 cells by regulating the expression of POLD4. PMID:26998273

  16. Effects of Nrf2 knockdown on the properties of irradiated cell conditioned medium from A549 human lung cancer cells.

    PubMed

    Yoshino, Hironori; Murakami, Kanna; Nawamaki, Mikoto; Kashiwakura, Ikuo

    2018-05-01

    The nuclear factor erythroid 2-related factor 2 (Nrf2) plays an important role in cellular defense against oxidative stress. Recent studies have demonstrated that Nrf2 is a useful target for cancer treatment, including radiation therapy. Ionizing radiation affects, not only the irradiated cells, but also the non-irradiated neighboring cells, and this effect is known as radiation-induced bystander effect. Upon exposure to radiation, the irradiated cells transmit signals to the non-irradiated cells via gap junctions or soluble factors. These signals in turn cause biological effects, such as a decrease in the clonogenic potential and cell death, in the non-irradiated neighboring cells. Nrf2 inhibition enhances cellular radiosensitivity. However, whether this modification of radiosensitivity by Nrf2 inhibition affects the radiation-induced bystander effects is unknown. In this study, we prepared an Nrf2 knockdown human lung cancer cell A549 and investigated whether the effects of irradiated cell conditioned medium (ICCM) on cell growth and cell death induction of non-irradiated cells vary depending on the Nrf2 knockdown. We found that Nrf2 knockdown resulted in a decrease in the cell growth and an increase in the radiosensitivity of A549 cells. When non-irradiated A549 cells were transfected with control siRNA and treated with ICCM, no significant difference was observed in the cell growth and proportion of Annexin V + dead cells between ICCM from non-irradiated cells and that from 2 or 8 Gy-irradiated cells. Similarly, no significant difference was observed in the cell growth and cell death induction upon treatment with ICCM in the Nrf2 knockdown A549 cells. Taken together, these results suggest that Nrf2 knockdown decreases cell growth and enhances the radiosensitivity of A549 cells; however, it does not alter the effect of ICCM on cell growth.

  17. Sepia Ink Oligopeptide Induces Apoptosis of Lung Cancer Cells via Mitochondrial Pathway.

    PubMed

    Wang, Xiaohua; Chen, Cheng; Zhou, Guoren; Ye, Jinjun; Yin, Rong; Feng, Dongjie; Zhang, Shuai; Wang, Xiaojun; Zhao, Xin; Zhang, Zhi

    2018-01-01

    Our previous study suggested the anti-tumor activity of sepia ink oligopeptide (SIO). Here we sought to investigate the underlying molecular mechanism. Cell proliferation was evaluated by cell counting kit-8 (CCK-8) assay. Cell apoptosis was determined by Annexin V/Propidium Iodide (PI) staining. The mitochondria pathway was characterized by quantification of Bcl-2, Bax, Caspase-9 and Cyto-C. The death receptor pathway was analyzed by determinement of Fas, Caspase-8 and NIK. The endoplasmic reticulum (ER)-dependent pathway was determined by measurement the expression of CHOP, Caspase-12, GRP78 and Calpain. The associated gene expression was quantified by RT-PCR and protein level was determined by immunoblotting. We demonstrated treatment with structurally modified SIO (CSIO, 5 µM) significantly inhibited cell proliferation and induced apoptosis in lung cancer cell line A549. The mitochondrial pathway, death receptor pathway and ER stress induced apoptosis were stimulated upon CSIO treatment. The administration with respective inhibitors including midiv-1 (50 µM for 2 h), PDTC (20 µM PDTC for 30 min) and ALLN (20 mM ALLN for 5 h) readily reversed the apoptosis inducing effect of CSIO. Our data demonstrates that CSIO is capable of induction apoptosis in lung cancer cell line, which is mediated by all three classical apoptotic pathways. Our results warrant further in vivo investigations of the anti-tumor potential of CSIO. © 2018 The Author(s). Published by S. Karger AG, Basel.

  18. Evaluation of permeability alteration and epithelial-mesenchymal transition induced by transforming growth factor-β1 in A549, NCI-H441, and Calu-3 cells: Development of an in vitro model of respiratory epithelial cells in idiopathic pulmonary fibrosis.

    PubMed

    Togami, Kohei; Yamaguchi, Kotaro; Chono, Sumio; Tada, Hitoshi

    2017-07-01

    Idiopathic pulmonary fibrosis (IPF) is a lethal lung disease, which is accompanied by changes in lung structure. With regard to treatment, aerosolized drugs administered intrapulmonarily are rapidly distributed into the plasma and do not remain in the lungs due to damage to the alveolar epithelium that occurs from pulmonary fibrosis. In this study, we sought to develop an in vitro model of respiratory epithelial cells in IPF for the evaluation of the intrapulmonary distribution of aerosolized drugs. We investigated transforming growth factor (TGF)-β 1 -induced epithelial-mesenchymal transition (EMT) and permeability alteration in A549, NCI-H441, and Calu-3 cell monolayers. After TGF-β 1 treatment of A549, NCI-H441, and Calu-3 cells, EMT markers including E-cadherin and vimentin and tight junction proteins including claudins-1, -3, and -5 were stained using immunofluorescence methods and detected using immunoblotting methods. Transport experiments were performed using TGF-β 1 -treated cell monolayers and fluorescein isothiocyanate dextrans (FD; 4.4, 10, and 70kDa). In addition, TGF-β 1 -induced apoptosis and necrosis were evaluated by flow cytometry using Annexin V and ethidium homodimer III, respectively. In NCI-H441 cells, incomplete EMT, destruction of claudins-1 and -3, and enhancement of FD permeability were caused by TGF-β 1 treatment. In A549 cells, complete EMT occurred but was not adequate for transport experiments because of low transepithelial electrical resistance. Whereas in Calu-3 cells, no changes were observed. TGF-β 1 -induced apoptosis and necrosis were not observed in any of the cell lines. Incomplete EMT and permeability enhancement were observed in the alveolar epithelium of IPF. Therefore, our results indicate that TGF-β 1 -treated NCI-H441 cell monolayers may serve as a useful in vitro model of respiratory epithelial cells for IPF. Copyright © 2017 Elsevier Inc. All rights reserved.

  19. Antimony trichloride induces a loss of cell viability via reactive oxygen species-dependent autophagy in A549 cells.

    PubMed

    Zhao, Xinyuan; Xing, Fengjun; Cong, Yewen; Zhuang, Yin; Han, Muxi; Wu, Zhiqiang; Yu, Shali; Wei, Haiyan; Wang, Xiaoke; Chen, Gang

    2017-12-01

    Antimony (Sb) is one of the most prevalent heavy metals and frequently leads to biological toxicity. Although autophagy is believed to be involved in metal-associated cytotoxicity, there is no evidence of its involvement following exposure. Moreover, the underlying mechanism of autophagy remains unclear. In this study, treatment with antimony trichloride caused autophagy in a dose- and time-dependent manner in A549 cells but did not affect the level of Atg5 or Atg7 mRNA expression. Furthermore, Sb enhanced autophagic flux while upregulating p62 gene and protein levels. The classic mechanistic target of rapamycin (mTOR) pathway is not involved in Sb-induced autophagy. However, Sb-induced autophagy and the upregulation of p62 were inhibited by treatment with the antioxidant N-acetylcysteine (NAC). Subsequent analyses demonstrated that the inhibition of autophagy protected A549 cells from a loss of cell viability, while the activation of autophagy by rapamycin had the opposite effect. These data suggest that reactive oxygen species-dependent autophagy mediates Sb-stimulated cell viability loss in A549 cells. Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. Green tea extract induces protective autophagy in A549 non-small lung cancer cell line.

    PubMed

    Izdebska, Magdalena; Klimaszewska-Wiśniewska, Anna; Hałas, Marta; Gagat, Maciej; Grzanka, Alina

    2015-12-31

    For many decades, polyphenols, including green tea extract catechins, have been reported to exert multiple anti-tumor activities. However, to date the mechanisms of their action have not been completely elucidated. Thus, the aim of this study was to assess the effect of green tea extract on non-small lung cancer A549 cells. A549 cells following treatment with GTE were analyzed using the inverted light and fluorescence microscope. In order to evaluate cell sensitivity and cell death, the MTT assay and Tali image-based cytometer were used, respectively. Ultrastructural alterations were assessed using a transmission electron microscope. The obtained data suggested that GTE, even at the highest dose employed (150 μM), was not toxic to A549 cells. Likewise, the treatment with GTE resulted in only a very small dose-dependent increase in the population of apoptotic cells. However, enhanced accumulation of vacuole-like structures in response to GTE was seen at the light and electron microscopic level. Furthermore, an increase in the acidic vesicular organelles and LC3-II puncta formation was observed under the fluorescence microscope, following GTE treatment. The analysis of the functional status of autophagy revealed that GTE-induced autophagy may provide self-protection against its own cytotoxicity, since we observed that the blockage of autophagy by bafilomycin A1 decreased the viability of A549 cells and potentiated necrotic cell death induction in response to GTE treatment. Collectively, our results revealed that A549 cells are insensitive to both low and high concentrations of the green tea extract, probably due to the induction of cytoprotective autophagy. These data suggest that a potential utility of GTE in lung cancer therapy may lie in its synergistic combinations with drugs or small molecules that target autophagy, rather than in monotherapy.

  1. A549 lung epithelial cells grown as three-dimensional aggregates: alternative tissue culture model for Pseudomonas aeruginosa pathogenesis.

    PubMed

    Carterson, A J; Höner zu Bentrup, K; Ott, C M; Clarke, M S; Pierson, D L; Vanderburg, C R; Buchanan, K L; Nickerson, C A; Schurr, M J

    2005-02-01

    A three-dimensional (3-D) lung aggregate model was developed from A549 human lung epithelial cells by using a rotating-wall vessel bioreactor to study the interactions between Pseudomonas aeruginosa and lung epithelial cells. The suitability of the 3-D aggregates as an infection model was examined by immunohistochemistry, adherence and invasion assays, scanning electron microscopy, and cytokine and mucoglycoprotein production. Immunohistochemical characterization of the 3-D A549 aggregates showed increased expression of epithelial cell-specific markers and decreased expression of cancer-specific markers compared to their monolayer counterparts. Immunohistochemistry of junctional markers on A549 3-D cells revealed that these cells formed tight junctions and polarity, in contrast to the cells grown as monolayers. Additionally, the 3-D aggregates stained positively for the production of mucoglycoprotein while the monolayers showed no indication of staining. Moreover, mucin-specific antibodies to MUC1 and MUC5A bound with greater affinity to 3-D aggregates than to the monolayers. P. aeruginosa attached to and penetrated A549 monolayers significantly more than the same cells grown as 3-D aggregates. Scanning electron microscopy of A549 cells grown as monolayers and 3-D aggregates infected with P. aeruginosa showed that monolayers detached from the surface of the culture plate postinfection, in contrast to the 3-D aggregates, which remained attached to the microcarrier beads. In response to infection, proinflammatory cytokine levels were elevated for the 3-D A549 aggregates compared to monolayer controls. These findings suggest that A549 lung cells grown as 3-D aggregates may represent a more physiologically relevant model to examine the interactions between P. aeruginosa and the lung epithelium during infection.

  2. TXNIP mediates the differential responses of A549 cells to sodium butyrate and sodium 4-phenylbutyrate treatment.

    PubMed

    Jin, Xuefang; Wu, Nana; Dai, Juji; Li, Qiuxia; Xiao, XiaoQiang

    2017-02-01

    Sodium butyrate (NaBu) and sodium 4-phenylbutyrate (4PBA) have promising futures in cancer treatment; however, their underlying molecular mechanisms are not clearly understood. Here, we show A549 cell death induced by NaBu and 4PBA are not the same. NaBu treatment induces a significantly higher level of A549 cell death than 4PBA. A gene expression microarray identified more than 5000 transcripts that were altered (>1.5-fold) in NaBu-treated A549 cells, but fewer than 2000 transcripts that were altered in 4PBA. Moreover, more than 100 cell cycle-associated genes were greatly repressed by NaBu, but slightly repressed by 4PBA; few genes were significantly upregulated only in 4PBA-treated cells. Gene expression was further validated by other experiments. Additionally, A549 cells that were treated with these showed changes in glucose consumption, caspase 3/7 activation and histone modifications, as well as enhanced mitochondrial superoxide production. TXNIP was strongly induced by NaBu (30- to 40-fold mRNA) but was only slightly induced by 4PBA (two to fivefold) in A549 cells. TXNIP knockdown by shRNA in A549 cells significantly attenuated caspase 3/7 activation and restored cell viability, while TXNIP overexpression significantly increased caspase 3/7 activation and cell death only in NaBu-treated cells. Moreover, TXNIP also regulated NaBu- but not 4PBA-induced H4K5 acetylation and H3K4 trimethylation, possibly by increasing WDR5 expression. Finally, we demonstrated that 4PBA induced a mitochondrial superoxide-associated cell death, while NaBu did so mainly through a TXNIP-mediated pathway. The above data might benefit the future clinic application. © 2016 The Authors. Cancer Medicine published by John Wiley & Sons Ltd.

  3. Modulation of DNA damage response and induction of apoptosis mediates synergism between doxorubicin and a new imidazopyridine derivative in breast and lung cancer cells.

    PubMed

    El-Awady, Raafat A; Semreen, Mohammad H; Saber-Ayad, Maha M; Cyprian, Farhan; Menon, Varsha; Al-Tel, Taleb H

    2016-01-01

    DNA damage response machinery (DDR) is an attractive target of cancer therapy. Modulation of DDR network may alter the response of cancer cells to DNA damaging anticancer drugs such as doxorubicin. The aim of the present study is to investigate the effects of a newly developed imidazopyridine (IAZP) derivative on the DDR after induction of DNA damage in cancer cells by doxorubicin. Cytotoxicity sulphrhodamine-B assay showed a weak anti-proliferative effect of IAZP alone on six cancer cell lines (MCF7, A549, A549DOX11, HepG2, HeLa and M8) and a normal fibroblast strain. Combination of IAZP with doxorubicin resulted in synergism in lung (A549) and breast (MCF7) cancer cells but neither in the other cancer cell lines nor in normal fibroblasts. Molecular studies revealed that synergism is mediated by modulation of DNA damage response and induction of apoptosis. Using constant-field gel electrophoresis and immunofluorescence detection of γ-H2AX foci, IAZP was shown to inhibit the repair of doxorubicin-induced DNA damage in A549 and MCF7 cells. Immunoblot analysis showed that IAZP suppresses the phosphorylation of the ataxia lelangiectasia and Rad3 related (ATR) protein, which is an important player in the response of cancer cells to chemotherapy-induced DNA damage. Moreover, IAZP augmented the doxorubicin-induced degradation of p21, activation of p53, CDK2, caspase 3/7 and phosphorylation of Rb protein. These effects enhanced doxorubicin-induced apoptosis in both cell lines. Our results indicate that IAZP is a promising agent that may enhance the cytotoxic effects of doxorubicin on some cancer cells through targeting the DDR. It is a preliminary step toward the clinical application of IAZP in combination with anticancer drugs and opens the avenue for the development of compounds targeting the DDR pathway that might improve the therapeutic index of anticancer drugs and enhance their cure rate. Copyright © 2015 Elsevier B.V. All rights reserved.

  4. [Effects of icotinib hydrochloride on the proliferation and apoptosis of human lung cancer cell lines].

    PubMed

    Ma, Li; Han, Xiao-hong; Wang, Shuai; Wang, Jian-fei; Shi, Yuan-kai

    2012-09-25

    To explore the effects of icotinib on the proliferation and apoptosis of various lung cancer cell lines. Human lung cancer cell lines HCC827, H1650, H1975, A549 and human epidermal cancer cell line A431 were treated in vitro with icotinib or gefitinib at a concentration gradient of 0 - 40 µmol/L. Their proliferation effects were analyzed by the thiazolyl blue (MTT) assay and the apoptotic effects detected by flow cytometer. The downstream signaling proteins were detected by Western blot. The median inhibitory concentrations (IC(50)) of icotinib for A431 and HCC827 cell lines were (0.04 ± 0.02) and (0.15 ± 0.06) µmol/L respectively. No significant differences existed between the inhibitions of gefitinib and icotinib on A431, HCC827, H1650, H1975 and A549 cell lines (all P > 0.05). Compared with H1650, H1975 and A549 cell lines, icotinib significantly inhibited A431 (P = 0.009, 0.005 and 0.000) and HCC827 (P = 0.001, 0.001 and 0.000) cell lines. And it lowered the expressions of p-AKT, p-ERK and survivin protein expression through the inhibited activity of p-EGFR protein. Icotinib can arrest the proliferation of lung adenocarcinoma cells with EGFR mutation or over-expression by inhibiting the signal pathways of AKT-ERK and survivin.

  5. Platycodin D induced apoptosis and autophagy in PC-12 cells through mitochondrial dysfunction pathway

    NASA Astrophysics Data System (ADS)

    Zeng, Chuan-Chuan; Zhang, Cheng; Yao, Jun-Hua; Lai, Shang-Hai; Han, Bing-Jie; Li, Wei; Tang, Bing; Wan, Dan; Liu, Yun-Jun

    2016-11-01

    In this article, the in vitro cytotoxicity of platycodin D was evaluated in human PC-12, SGC-7901, BEL-7402, HeLa and A549 cancer cell lines. PC-12 cells were sensitive to platycodin D treatment, with an IC50 value of 13.5 ± 1.2 μM. Morphological and comet assays showed that platycodin D effectively induced apoptosis in PC-12 cells. Platycodin D increased the levels of reactive oxygen species (ROS) and induced a decrease in mitochondrial membrane potential. Platycodin D induced cell cycle arrest at the G0/G1 phase in the PC-12 cell line. Platycodin D can induce autophagy. In addition, platycodin D can down-regulate the expression of Bcl-2 and Bcl-x, and up-regulate the levels of Bid protein in the PC-12 cells. The results demonstrated that platycodin D induced PC-12 cell apoptosis through a ROS-mediated mitochondrial dysfunction pathway.

  6. Tetrandrine suppresses lung cancer growth and induces apoptosis, potentially via the VEGF/HIF-1α/ICAM-1 signaling pathway

    PubMed Central

    Chen, Zhuo; Zhao, Liang; Zhao, Feng; Yang, Guanghai; Wang, Jian Jun

    2018-01-01

    The present study investigated the effect of tetrandrine on lung cancer cell growth and apoptosis, and its possible underlying molecular mechanism. A549 human lung cancer cells were incubated with between 2.5 and 10 µM tetrandrine for 12, 24 and 48 h, following which the effect of tetrandrine on cell viability and apoptosis were assessed using an MTT assay and flow cytometry. ELISA and western blotting were used to analyze VEGF activity, and the expression of poly (ADP-ribose) polymerase (PARP), phosphorylated protein kinase B (Akt), Bcl-2-associated X protein (Bax), hypoxia inducible factor (HIF)-1α and inter-cellular adhesion molecule-1 (ICAM-1). Tetrandrine effectively suppressed the growth of and induced apoptosis in A549 lung cancer cells. The expression of PARP, Bax, intercellular adhesion molecule-1 (ICAM-1) and vascular endothelial growth factor (VEGF) was significantly upregulated, and the phosphorylation of Akt and expression of HIF-1α was significantly suppressed in A549 lung cancer cells. Therefore, tetrandrine may suppress cell viability and induce apoptosis via the VEGF/HIF-1α/ICAM-1 signaling pathway. PMID:29849794

  7. Discovery of a Novel Anti-Cancer Agent Targeting Both Topoisomerase I & II as Well as Telomerase Activities in Human Lung Adenocarcinoma A549 Cells In Vitro and In Vivo: Cinnamomum verum Component Cuminaldehyde.

    PubMed

    Chen, Ta-Wei; Tsai, Kuen-Daw; Yang, Shu-Mei; Wong, Ho-Yiu; Liu, Yi-Heng; Cherng, Jonathan; Chou, Kuo-Shen; Wang, Yang-Tz; Cuizon, Janise; Cherng, Jaw-Ming

    2016-01-01

    Cinnamomum verum is used to make the spice cinnamon and has been used for more than 5000 years by both of the two most ancient forms of medicine in the words: Ayurveda and traditional Chinese herbal medicines for various applications such as adenopathy, rheumatism, dermatosis, dyspepsia, stroke, tumors, elephantiasis, trichomonas, yeast, and virus infections. We evaluated the anticancer effect of cuminaldehyde (CuA), a constituent of the bark of the plant, and its underlying molecular biomarkers associated with carcinogenesis in human lung adenocarcinoma A549 cells. The results show that cuminaldehyde suppressed proliferation and induced apoptosis as indicated by mitochondrial membrane potential loss, activation of caspase 3 and 9, increase in annexin V+PI+ cells, and morphological characteristics of apoptosis, including blebbing of plasma membrane, nuclear condensation, fragmentation, apoptotic body formation, and comet with elevated tail intensity and moment. In addition, cuminaldehyde also induced lysosomal vacuolation with increased volume of acidic compartments (VAC), suppressions of both topoisomerase I & II as well as telomerase activities in a dose-dependent manner. Further study reveals the growth-inhibitory effect of cuminaldehyde was also evident in a nude mice model. Taken together, the data suggest that the growth-inhibitory effect of cuminaldehyde against A549 cells is accompanied by downregulations of proliferative control involving apoptosis, both topoisomerase I & II as well as telomerase activities, together with an upregulation of lysosomal vacuolation and VAC. Similar effects (including all of the above-mentioned effects) were found in other cell lines, including human lung squamous cell carcinoma NCI-H520 and colorectal adenocarcinoma COLO 205 (results not shown). Our data suggest that cuminaldehyde could be a potential agent for anticancer therapy.

  8. Microemulsion-based synergistic dual-drug codelivery system for enhanced apoptosis of tumor cells.

    PubMed

    Qu, Ding; Ma, Yihua; Sun, Wenjie; Chen, Yan; Zhou, Jing; Liu, Congyan; Huang, Mengmeng

    2015-01-01

    A microemulsion-based synergistic dual-drug codelivery system was developed for enhanced cell apoptosis by transporting coix seed oil and etoposide into A549 (human lung carcinoma) cells simultaneously. Results obtained by dynamic light scattering showed that an etoposide (VP16)-loaded coix seed oil microemulsion (EC-ME) delivery system had a small size around 35 nm, a narrow polydispersity index, and a slightly negative surface charge. The encapsulating efficiency and total drug loading rate were 97.01% and 45.48%, respectively, by high-performance liquid chromatography. The release profiles at various pH values showed an obvious pH-responsive difference, with the accumulated amount of VP16 released at pH 4.5 (and pH 5.5) being 2.7-fold higher relative to that at pH 7.4. Morphologic alteration (particle swelling) associated with a mildly acidic pH environment was found on transmission electron microscopy. In the cell study, the EC-ME system showed a significantly greater antiproliferative effect toward A549 cells in comparison with free VP16 and the mixture of VP16 and coix seed oil. The half-maximal inhibitory concentration of the EC-ME system was 3.9-fold and 10.4-fold lower relative to that of free VP16 and a mixture of VP16 and coix seed oil, respectively. Moreover, fluorescein isothiocyanate and VP16 (the green fluorescent probe and entrapped drug, respectively) were efficiently internalized into the cells by means of coix seed oil microemulsion through intuitive observation and quantitative measurement. Importantly, an EC-ME system containing 20 μg/mL of VP16 showed a 3.3-fold and 3.5-fold improvement in induction of cell apoptosis compared with the VP-16-loaded microemulsion and free VP16, respectively. The EC-ME combination strategy holds promise as an efficient drug delivery system for induction of apoptosis and treatment of lung cancer.

  9. Formononetin suppresses the proliferation of human non-small cell lung cancer through induction of cell cycle arrest and apoptosis

    PubMed Central

    Yang, Yi; Zhao, Yi; Ai, Xinghao; Cheng, Baijun; Lu, Shun

    2014-01-01

    Formononetin is a novel herbal isoflavonoid isolated from Astragalus membranaceus and possesses antitumorigenic properties. In the present study, we investigated the anti-proliferative effects of formononetin on human non-small cell lung cancer (NSCLC), and further elucidated the molecular mechanism underlying the anti-tumor property. MTT assay showed that formononetin treatment significantly inhibited the proliferation of two NSCLC cell lines including A549 and NCI-H23 in a time- and dose-dependent manner. Flow cytometric analysis demonstrated that formononetin induced G1-phase cell cycle arrest and promoted cell apoptosis in NSCLC cells. On the molecular level, we observed that exposure to formononetin altered the expression levels of cell cycle arrest-associated proteins p21, cyclin A and cyclin D1. Meanwhile, the apoptosis-related proteins cleaved caspase-3, bax and bcl-2 were also changed following treatment with formononetin. In addition, the expression level of p53 was dose-dependently upregulated after administration with formononetin. We also found that formononetin treatment increased the phosphorylation of p53 at Ser15 and Ser20 and enhances its transcriptional activity in a dose-dependent manner. Collectively, these results demonstrated that formononetin might be a potential chemopreventive drug for lung cancer therapy through induction of cell cycle arrest and apoptosis in NSCLC cells. PMID:25674209

  10. Formononetin suppresses the proliferation of human non-small cell lung cancer through induction of cell cycle arrest and apoptosis.

    PubMed

    Yang, Yi; Zhao, Yi; Ai, Xinghao; Cheng, Baijun; Lu, Shun

    2014-01-01

    Formononetin is a novel herbal isoflavonoid isolated from Astragalus membranaceus and possesses antitumorigenic properties. In the present study, we investigated the anti-proliferative effects of formononetin on human non-small cell lung cancer (NSCLC), and further elucidated the molecular mechanism underlying the anti-tumor property. MTT assay showed that formononetin treatment significantly inhibited the proliferation of two NSCLC cell lines including A549 and NCI-H23 in a time- and dose-dependent manner. Flow cytometric analysis demonstrated that formononetin induced G1-phase cell cycle arrest and promoted cell apoptosis in NSCLC cells. On the molecular level, we observed that exposure to formononetin altered the expression levels of cell cycle arrest-associated proteins p21, cyclin A and cyclin D1. Meanwhile, the apoptosis-related proteins cleaved caspase-3, bax and bcl-2 were also changed following treatment with formononetin. In addition, the expression level of p53 was dose-dependently upregulated after administration with formononetin. We also found that formononetin treatment increased the phosphorylation of p53 at Ser15 and Ser20 and enhances its transcriptional activity in a dose-dependent manner. Collectively, these results demonstrated that formononetin might be a potential chemopreventive drug for lung cancer therapy through induction of cell cycle arrest and apoptosis in NSCLC cells.

  11. Anti-inflammatory effects of embelin in A549 cells and human asthmatic airway epithelial tissues.

    PubMed

    Lee, In-Seung; Cho, Dong-Hyuk; Kim, Ki-Suk; Kim, Kang-Hoon; Park, Jiyoung; Kim, Yumi; Jung, Ji Hoon; Kim, Kwanil; Jung, Hee-Jae; Jang, Hyeung-Jin

    2018-02-01

    Allergic asthma is the most common type in asthma, which is defined as a chronic inflammatory disease of the lung. In this study, we investigated whether embelin (Emb), the major component of Ardisia japonica BL. (AJB), exhibits anti-inflammatory effects on allergic asthma via inhibition of NF-κB activity using A549 cells and asthmatic airway epithelial tissues. Inflammation was induced in A549 cells, a human airway epithelial cell line, by IL-1β (10 ng/ml) treatment for 4 h. The effects of Emb on NF-κB activity and COX-2 protein expression in inflamed airway epithelial cells and human asthmatic airway epithelial tissues were analyzed via western blot. The secretion levels of NF-κB-mediated cytokines/chemokines, including IL-4, 6, 9, 13, TNF-α and eotaxin, were measured by a multiplex assay. Emb significantly blocked NF-κB activity in IL-1β-treated A549 cells and human asthmatic airway epithelial tissues. COX-2 expression was also reduced in both IL-1β-treated A549 cells and asthmatic tissues Emb application. Emb significantly reduced the secretion of IL-4, IL-6 and eotaxin in human asthmatic airway epithelial tissues by inhibiting activity of NF-κB. The results of this study suggest that Emb may be used as an anti-inflammatory agent via inhibition of NF-κB and related cytokines.

  12. Cytotoxicity and apoptosis of ovarian and breast cancer cell lines induced by OVS1 monoclonal antibody and paclitaxel.

    PubMed

    Moongkarndi, Primchanien; Kaslungka, Sineenart; Kosem, Nuttavut; Junnu, Sarawut; Jongsomboonkusol, Suna; Theptaranon, Yodsaward; Neungton, Neelobol

    2003-03-01

    OVS1 monoclonal antibody (MAb) produced against ovarian cancer is currently used to identify mucinous cystadenocarcinoma antigen as a tumor marker secreted in serum. The potential of OVS1 MAb in ovarian cancer treatment was studied by evaluating the induction of cytotoxicity and apoptosis of SKOV3 ovarian cancer and BT549 breast cancer cell lines induced by OVS1. Paclitaxel, an antitumor drug, was used as positive control and applied as a combined drug together with OVS1 MAb. OVS1 MAb and paclitaxel were found by MTT assay to induce cytotoxicity against both cell lines. The ED50 of OVS1 MAb were 26.25 and 25.00 microg/ml and of paclitaxel were 21.88 and 9.20 nM against SKOV3 and BT549 cell lines, respectively. The quantitative amount of cells determined by fluorimetric assay was correlated to the results of the MTT assay. The combined application of OVS1 MAb and paclitaxel on these two cell lines resulted in a greater cytotoxicity than observed by either agent alone. OVS1 MAb and paclitaxel applied against both cell lines induced the morphological changes of apoptotic cell death at 24 hours visualized by two color fluorescence dyes, Ho33342 and propidium iodide. Combination of the two substances enhanced the rate of apoptosis compared to either OVS1 MAb or paclitaxel given alone. DNA fragmentation was detected in an agarose gel electrophoresis after treating cells with OVS1 MAb and paclitaxel at 24 hours. These findings on the induction of cytotoxicity and apoptosis by OVS1 MAb on cancer cell lines have implications on the potential application of OVS1 MAb for clinical therapy.

  13. GAS5 modulated autophagy is a mechanism modulating cisplatin sensitivity in NSCLC cells.

    PubMed

    Zhang, N; Yang, G-Q; Shao, X-M; Wei, L

    2016-06-01

    In this study, we investigated the association between lncRNA GAS5 and cisplatin (DDP) resistance in NSCLC and further studied the regulative effect of GAS5 on autophagy and DDP resistance. GAS5 expression in cancerous and adjacent normal tissues from 15 NSCLC patients received neoadjuvant chemotherapy and the following surgery were measured using qRT-PCR analysis. GAS5 gain-and-loss study was performed using A549 and A549/DDP cells as an in-vitro model to investigate the effect of GAS5 on autophagy and cisplatin sensitivity. NSCLC tissues had a substantially lower expression of GAS5 than adjacent normal tissues. The NSCLC tissues from patients with progressive disease (PD) had even lower GAS5 expression. GAS5 knockdown increased DDP IC50 of A549 cells, while GAS5 overexpression decreased DDP IC50 of A549/DDP cells. A549/DDP cells had significantly higher basal autophagy than A549 cells. GAS5 knockdown resulted in decreased autophagy in A549 cells, while GAS5 overexpression led to increased autophagy in A549/DDP cells. Treatment with 3-MA, an autophagy inhibitor, significantly decreased DDP IC50 and promoted DDP-induced cell apoptosis in A549 cells. In addition, 3-MA also partly reversed the effect of GAS5 knockdown. In A549/DDP cells, GAS5 showed the similar effect as 3-MA in reducing DPP IC50 and promoting DDP-induced apoptosis and also presented synergic effect with 3-MA. GAS5 downregulation is associated with cisplatin resistance in NSCLC. GAS5 can inhibit autophagy and therefore enhance cisplatin sensitivity in NSCLC cells.

  14. 5-demethyltangeretin inhibits human nonsmall cell lung cancer cell growth by inducing G2/M cell cycle arrest and apoptosis.

    PubMed

    Charoensinphon, Noppawat; Qiu, Peiju; Dong, Ping; Zheng, Jinkai; Ngauv, Pearline; Cao, Yong; Li, Shiming; Ho, Chi-Tang; Xiao, Hang

    2013-12-01

    Tangeretin (TAN) and 5-demethyltangeretin (5DT) are two closely related polymethoxyflavones found in citrus fruits. We investigated growth inhibitory effects on three human nonsmall cell lung cancer (NSCLC) cells. Cell viability assay demonstrated that 5DT inhibited NSCLC cell growth in a time- and dose-dependent manner, and IC50 s of 5DT were 79-fold, 57-fold, and 56-fold lower than those of TAN in A549, H460, and H1299 cells, respectively. Flow cytometry analysis showed that 5DT induced extensive G2/M cell cycle arrest and apoptosis in NSCLC cells, while TAN at tenfold higher concentrations did not. The apoptosis induced by 5DT was further confirmed by activation of caspase-3 and cleavage of PARP. Moreover, 5DT dose-dependently upregulated p53 and p21(Cip1/Waf1), and downregulated Cdc-2 (Cdk-1) and cyclin B1. HPLC analysis revealed that the intracellular levels of 5DT in NSCLC cells were 2.7-4.9 fold higher than those of TAN after the cells were treated with 5DT or TAN at the same concentration. Our results demonstrated that 5DT inhibited NSCLC cell growth by inducing G2/M cell cycle arrest and apoptosis. These effects were much stronger than those produced by TAN, which is partially due to the higher intracellular uptake of 5DT than TAN. © 2013 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  15. Salvianolic acid A reverses cisplatin resistance in lung cancer A549 cells by targeting c-met and attenuating Akt/mTOR pathway.

    PubMed

    Tang, Xia-Li; Yan, Li; Zhu, Ling; Jiao, De-Min; Chen, Jun; Chen, Qing-Yong

    2017-09-01

    Drug resistance is one of the leading causes of chemotherapy failure in non-small cell lung cancer (NSCLC) treatment. The purpose of this study was to investigate the role of c-met in human lung cancer cisplatin resistance cell line (A549/DDP) and the reversal mechanism of salvianolic acid A (SAA), a phenolic active compound extracted from Salvia miltiorrhiza. In this study, we found that A549/DDP cells exert up-regulation of c-met by activating the Akt/mTOR signaling pathway. We also show that SAA could increase the chemotherapeutic efficacy of cisplatin, suggesting a synergistic effect of SAA and cisplatin. Moreover, we revealed that SAA enhanced sensitivity to cisplatin in A549/DDP cells mainly through suppression of the c-met/AKT/mTOR signaling pathway. Knockdown of c-met revealed similar effects as that of SAA in A549/DDP cells. In addition, SAA effectively prevented multidrug resistance associated protein1 (MDR1) up-regulation in A549/DDP cells. Taken together, our results indicated that SAA suppressed c-met expression and enhanced the sensitivity of lung adenocarcinoma A549 cells to cisplatin through AKT/mTOR signaling pathway. Copyright © 2017 The Authors. Production and hosting by Elsevier B.V. All rights reserved.

  16. Acrolein preferentially damages nucleolus eliciting ribosomal stress and apoptosis in human cancer cells.

    PubMed

    Wang, Hsiang-Tsui; Chen, Tzu-Ying; Weng, Ching-Wen; Yang, Chun-Hsiang; Tang, Moon-Shong

    2016-12-06

    Acrolein (Acr) is a potent cytotoxic and DNA damaging agent which is ubiquitous in the environment and abundant in tobacco smoke. Acr is also an active cytotoxic metabolite of the anti-cancer drugs cyclophosphamide and ifosfamide. The mechanisms via which Acr exerts its anti-cancer activity and cytotoxicity are not clear. In this study, we found that Acr induces cytotoxicity and cell death in human cancer cells with different activities of p53. Acr preferentially binds nucleolar ribosomal DNA (rDNA) to form Acr-deoxyguanosine adducts, and induces oxidative damage to both rDNA and ribosomal RNA (rRNA). Acr triggers ribosomal stress responses, inhibits rRNA synthesis, reduces RNA polymerase I binding to the promoter of rRNA gene, disrupts nucleolar integrity, and impairs ribosome biogenesis and polysome formation. Acr causes an increase in MDM2 levels and phosphorylation of MDM2 in A549 and HeLa cells which are p53 active and p53 inactive, respectively. It enhances the binding of ribosomal protein RPL11 to MDM2 and reduces the binding of p53 and E2F-1 to MDM2 resulting in stabilization/activation of p53 in A549 cells and degradation of E2F-1 in A549 and HeLa cells. We propose that Acr induces ribosomal stress which leads to activation of MDM2 and RPL11-MDM2 binding, consequently, activates p53 and enhances E2F-1 degradation, and that taken together these two processes induce apoptosis and cell death.

  17. Multifunctional polyamidoamine-modified selenium nanoparticles dual-delivering siRNA and cisplatin to A549/DDP cells for reversal multidrug resistance.

    PubMed

    Zheng, Wenjing; Cao, Chengwen; Liu, Yanan; Yu, Qianqian; Zheng, Chuping; Sun, Dongdong; Ren, Xiaofan; Liu, Jie

    2015-01-01

    Multidrug resistance (MDR) is a major barrier against effective cancer treatment. Dual-delivering a therapeutic small interfering RNA (siRNA) and chemotherapeutic agents has been developed to reverse drug resistance in tumor cells. In this study, amine-terminated generation 5 polyamidoamine (PAMAM) dendrimers (G5.NH2)-modified selenium nanoparticles (G5@Se NP) were synthesized for the systemic dual-delivery of mdr1 siRNA and cisplatin (cis-diamminedichloroplatinum-(II), DDP), which was demonstrated to enhance siRNA loading, releasing efficiency and gene-silencing efficacy. When the mdr1 siRNA was conjugated with G5@Se NP via electrostatic interaction, a significant down-regulation of P-glycoprotein and multidrug resistance-associated protein expression was observed; G5@Se-DDP-siRNA arrested A549/DDP cells at G1 phase and led to enhanced cytotoxicity in A549/DDP cells through induction of apoptosis involving the AKT and ERK signaling pathways. Interestingly, G5@Se-DDP NP were much less reactive than DDP in the reactions with both MT and GSH, indicating that loading of DDP in a nano-delivery system could effectively prevent cell detoxification. Furthermore, animal studies demonstrated that the new delivery system of G5@Se-DDP-siRNA significantly enhanced the anti-tumor effect on tumor-bearing nude mice, with no appreciable abnormality in the major organs. These results suggest that G5@Se NP could be a potential platform to combine chemotherapy and gene therapy technology in the treatment of human disease. Copyright © 2014 Acta Materialia Inc. Published by Elsevier Ltd. All rights reserved.

  18. Beta sitosterol and Daucosterol (phytosterols identified in Grewia tiliaefolia) perturbs cell cycle and induces apoptotic cell death in A549 cells.

    PubMed

    Rajavel, Tamilselvam; Mohankumar, Ramar; Archunan, Govindaraju; Ruckmani, Kandasamy; Devi, Kasi Pandima

    2017-06-13

    Lung cancer is the leading cause of cancer related deaths both in developed and developing countries. Since majority of the existing therapeutic methods harms both normal and malignant cells, a viable alternative is to switch into safe and beneficial traditional medicinal plants. Hence the present study was framed to identify selective anti-lung cancer agents from the medicinal plant Grewia tiliaefolia (GT). Cell viability experiments showed that benzene extract of GT (BGT) leaf effectively inhibited the growth of A549 cells, while being non-toxic to normal human lung L132 and PBMC cells. Ames and comet assays demonstrated that BGT is of non-mutagenic and non-genotoxic nature in untransformed cells. The hematological and histopathological profiles of the in vivo acute and sub-acute toxicity studies demonstrated that BGT is safe and tolerable. Importantly, western blot analysis and Annexin V-FITC staining confirmed that BGT promotes mitochondrial dependent apoptotic cell death in A549 cells by arresting cell cycle at G2/M phase. Bio-assay guided fractionation revealed the presence of phytosteols (β-sitosterol and daucosterol) which significantly inhibited the growth of A549 cells both alone and in combination. This study warrants that these phytosterols in alone or in combination can be considered as safe and potential drug candidates for lung cancer treatment.

  19. Inhibition of PDGFR by CP-673451 induces apoptosis and increases cisplatin cytotoxicity in NSCLC cells via inhibiting the Nrf2-mediated defense mechanism.

    PubMed

    Yang, Yang; Deng, Yanchao; Chen, Xiangcui; Zhang, Jiahao; Chen, Yueming; Li, Huachao; Wu, Qipeng; Yang, Zhicheng; Zhang, Luyong; Liu, Bing

    2018-05-29

    Platelet-derived growth factor receptors (PDGFRs) are abundantly expressed by stromal cells in the non-small cell lung cancer (NSCLC) microenvironment, and in a subset of cancer cells, usually with their overexpression and/or activating mutation. However, the effect of PDGFR inhibition on lung cancer cells themselves has been largely neglected. In this study, we investigated the anticancer activity of CP-673451, a potent and selective inhibitor of PDGFRβ, on NSCLC cell lines (A549 and H358) and the potential mechanism. The results showed that inhibition of PDGFRβ by CP-673451 induced a significant increase in cell apoptosis, accompanied by ROS accumulation. However, CP-673451 exerted less cytotoxicity in normal lung epithelial cell line BEAS-2B cells determined by MTT and apoptosis assay. Elimination of ROS by NAC reversed the CP-673451-induced apoptosis in NSCLC cells. Furthermore, CP-673451 down-regulated the expression of nuclear factor erythroid 2-related factor 2 (Nrf2) probably through inhibition of PI3K/Akt pathway. Rescue of Nrf2 activity counteracted the effects of CP-673451 on cell apoptosis and ROS accumulation. Silencing PDGFRβ expression by PDGFRβ siRNA exerted similar effects with CP-673451 in A549 cells, and when PDGFRβ was knockdowned by PDGFRβ siRNA, CP-673451 produced no additional effects on cell viability, ROS and GSH production, Nrf2 expression as well as PI3K/Akt pathway activity. Specifically, Nrf2 plays an indispensable role in NSCLC cell sensitivity to platinum-based treatments and we found that combination of CP-673451 and cisplatin produced a synergistic anticancer effect and substantial ROS production in vitro. Therefore, these results clearly demonstrate the effectiveness of inhibition of PDGFRβ against NSCLC cells and strongly suggest that CP-673451 may be a promising adjuvant chemotherapeutic drug. Copyright © 2018 Elsevier B.V. All rights reserved.

  20. Combined treatment of curcumin and small molecule inhibitors suppresses proliferation of A549 and H1299 human non-small-cell lung cancer cells.

    PubMed

    Lin, Hui-Ping; Kuo, Li-Kuo; Chuu, Chih-Pin

    2012-01-01

    Curcumin (diferuloylmethane) is a phenolic compound present in turmeric and is ingested daily in many parts of the world. Curcumin has been reported to cause inhibition on proliferation and induction of apoptosis in many human cancer cell lines, including non-small cell lung cancer cells (NSCLC). However, the clinical application of curcumin is restricted by its low bioavailability. In this report, it was observed that combined treatment of a low dosage of curcumin (5-10 µM) with a low concentration (0.1-2.5 µM) of small molecule inhibitors, including AG1478, AG1024, PD173074, LY294002 and caffeic acid phenethyl ester (CAPE) increased the growth inhibition in two human NSCLC cell lines: A549 and H1299 cells. The observation suggested that combined treatment of a low dosage of curcumin with inhibitors against epidermal growth factor receptor (EGFR), insulin-like growth factor 1 (IGF-1R), fibroblast growth factors receptor (FGFR), phosphatidylinositol 3-kinases (PI3K) or NF-κB signaling pathway may be a potential adjuvant therapy beneficial to NSCLC patients. Copyright © 2011 John Wiley & Sons, Ltd.

  1. Activation of interferon regulatory factor-3 via toll-like receptor 3 and immunomodulatory functions detected in A549 lung epithelial cells exposed to misplaced U1-snRNA.

    PubMed

    Sadik, Christian D; Bachmann, Malte; Pfeilschifter, Josef; Mühl, Heiko

    2009-08-01

    U1-snRNA is an integral part of the U1 ribonucleoprotein pivotal for pre-mRNA splicing. Toll-like receptor (TLR) signaling has recently been associated with immunoregulatory capacities of U1-snRNA. Using lung A549 epithelial/carcinoma cells, we report for the first time on interferon regulatory factor (IRF)-3 activation initiated by endosomally delivered U1-snRNA. This was associated with expression of the IRF3-inducible genes interferon-beta (IFN-beta), CXCL10/IP-10 and indoleamine 2,3-dioxygenase. Mutational analysis of the U1-snRNA-activated IFN-beta promoter confirmed the crucial role of the PRDIII element, previously proven pivotal for promoter activation by IRF3. Notably, expression of these parameters was suppressed by bafilomycin A(1), an inhibitor of endosomal acidification, implicating endosomal TLR activation. Since resiquimod, an agonist of TLR7/8, failed to stimulate A549 cells, data suggest TLR3 to be of prime relevance for cellular activation. To assess the overall regulatory potential of U1-snRNA-activated epithelial cells on cytokine production, co-cultivation with peripheral blood mononuclear cells (PBMC) was performed. Interestingly, A549 cells activated by U1-snRNA reinforced phytohemagglutinin-induced interleukin-10 release by PBMC but suppressed that of tumor necrosis factor-alpha, indicating an anti-inflammatory potential of U1-snRNA. Since U1-snRNA is enriched in apoptotic bodies and epithelial cells are capable of performing efferocytosis, the present data in particular connect to immunobiological aspects of apoptosis at host/environment interfaces.

  2. Propolis extracts from the northern region of Thailand suppress cancer cell growth through induction of apoptosis pathways.

    PubMed

    Khacha-Ananda, Supakit; Tragoolpua, Khajornsak; Chantawannakul, Panuwan; Tragoolpua, Yingmanee

    2016-12-01

    The continual increase in mortality rates and number of cancer cases is a matter of serious concern in developing countries. The incorporation of natural products into classical cancer treatment approaches is a promising direction. The mechanisms of A549 and HeLa cancer cell death induction by ethanolic extracts of propolis samples from Phayao, Chiang Mai, and Nan provinces in northern Thailand were investigated in this study. The propolis extract from Chiang Mai showed the highest antioxidant activity and the greatest total phenolic content. The propolis extract from Nan also exhibited the highest total flavonoid content. The proliferation of A549 and HeLa cells grown in the presence of the propolis extracts was suppressed in a dose- and time-dependent manner. Moreover, treatment of both cancer cells with the propolis extracts showed DNA fragmentation and significantly increased the number of the apoptotic cells. On A549 cells, the extrinsic and intrinsic pathways of caspase enzymes were activated by the propolis extracts from Phayao and Chiang Mai. In the case of the propolis extract from Nan, the mechanisms involved apoptosis on the A549 cells were caspase-independent pathway. The extrinsic pathway of the caspase enzyme was triggered by all of the propolis extracts on HeLa cells. Finally, oral administration of the propolis granule produced from the propolis extract from Nan resulted in extended survival of tumour-bearing mice. Therefore, propolis extracts from the northern region of Thailand demonstrated pharmacological properties, both antioxidant and anticancer activities. From these findings, it is evident that propolis extracts can be considered as a naturally obtained agent extremely useful in cancer treatment.

  3. Garcinol from Garcinia indica Downregulates Cancer Stem-like Cell Biomarker ALDH1A1 in Nonsmall Cell Lung Cancer A549 Cells through DDIT3 Activation.

    PubMed

    Wang, Jinhan; Wang, Liwen; Ho, Chi-Tang; Zhang, Kunsheng; Liu, Qiang; Zhao, Hui

    2017-05-10

    Nonsmall cell lung cancer (NSCLC) is the predominant type of lung cancer. Patients with NSCLC show high mortality rates because of failure to clean up cancer stem cells (CSCs). The anticancer activity of phytochemical garcinol has been identified in various cancer cell models. However, the effect of garcinol on NSCLC cell lines is still lacking. Of the NSCLC cell lines we tested, A549 cells were the most sensitive to garcinol. Interestingly, Aldehyde Dehydrogenase 1 Family Member A1 (ALDH1A1) was preferentially expressed in A549 cells and downregulated by the addition of garcinol. We also found that garcinol enriched DNA damage-inducible transcript 3 (DDIT3) and then altered DDIT3-CCAAT-enhancer-binding proteins beta (C/EBPβ) interaction resulting in a decreased binding of C/EBPβ to the endogenous ALDH1A1 promoter. Furthermore, garcinol's inhibition of ALDH1A1 was identified in a xenograft mice model. Garcinol repressed ALDH1A1 transcription in A549 cells through alterations in the interaction between DDIT3 and C/EBPβ. Garcinol could be a potential dietary phytochemical candidate for NSCLCs patients whose tumors harbored high ALDH1A1 expression.

  4. Radiosensitization of tumour cell lines by the polyphenol Gossypol results from depressed double-strand break repair and not from enhanced apoptosis.

    PubMed

    Kasten-Pisula, Ulla; Windhorst, Sabine; Dahm-Daphi, Jochen; Mayr, Georg; Dikomey, Ekkehard

    2007-06-01

    New drugs are needed to increase the efficiency of radiotherapy in order to improve the therapeutic outcome of tumour patients. In this respect, the polyphenol Gossypol might be of interest, because of its effect on apoptosis and DNA repair, which is either mediated directly or indirectly via the inositol phosphate metabolism. It was investigated, whether these effects result in enhanced radiosensitivity of tumour cells. Tumour cell lines investigated: A549, FaDu, H1299, MCF7 and Du145. Cell cycle distribution was determined by FACS analysis, apoptosis was measured by DAPI staining and caspase3/7 activity. Double-strand breaks (DSB) were investigated via gammaH2AX-foci and cell survival by colony formation assay. The level of inositol phosphates was determined by HPLC, protein expression by Western blot. In A549 cells, Gossypol at concentrations 1microM strongly affects proliferation with only a modest arrest in the G1-phase, but with no increase in the fraction of apoptotic cells or the number of additional DSB. Additional DSB were only seen in FaDu cells, where Gossypol (2microM) was extremely toxic with a plating efficiency <0.002. When combined with irradiation, incubation with Gossypol (1-2microM) was found to result in an enhanced radiosensitivity with, however, a substantial variation. While there was a strong radiosensitization for FaDu and Du145 cells, there was an intermediate response for A549 cells, but almost no effect for H1299 and MCF7 cells. This sensitization was not caused from an elevated rate of apoptosis, but primarily resulted from reduced DSB repair capacity. The reduction in DSB repair could be ascribed neither to changes in the level of repair proteins relevant for non-homologous end-joining (Ku70, Ku80, DNA-PKcs) nor to changes in the level of higher phosphorylated inositols, whereby the latter were even found to be enhanced by Gossypol. For some tumour cell lines treatment with low concentrations of Gossypol can be used to inhibit DSB

  5. The ethanol extract of Scutellaria baicalensis and the active compounds induce cell cycle arrest and apoptosis including upregulation of p53 and Bax in human lung cancer cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Gao Jiayu; Morgan, Winston A.; Sanchez-Medina, Alberto

    2011-08-01

    Despite a lack of scientific authentication, Scutellaria baicalensis is clinically used in Chinese medicine as a traditional adjuvant to chemotherapy of lung cancer. In this study, cytotoxicity assays demonstrated that crude ethanolic extracts of S. baicalensis were selectively toxic to human lung cancer cell lines A549, SK-LU-1 and SK-MES-1 compared with normal human lung fibroblasts. The active compounds baicalin, baicalein and wogonin did not exhibit such selectivity. Following exposure to the crude extracts, cellular protein expression in the cancer cell lines was assessed using 2D gel electrophoresis coupled with MALDI-TOF-MS/Protein Fingerprinting. The altered protein expression indicated that cell growth arrestmore » and apoptosis were potential mechanisms of cytotoxicity. These observations were supported by PI staining cell cycle analysis using flow cytometry and Annexin-V apoptotic analysis by fluorescence microscopy of cancer cells treated with the crude extract and pure active compounds. Moreover, specific immunoblotting identification showed the decreased expression of cyclin A results in the S phase arrest of A549 whereas the G{sub 0}/G{sub 1} phase arrest in SK-MES-1 cells results from the decreased expression of cyclin D1. Following treatment, increased expression in the cancer cells of key proteins related to the enhancement of apoptosis was observed for p53 and Bax. These results provide further insight into the molecular mechanisms underlying the clinical use of this herb as an adjuvant to lung cancer therapy. - Research Highlights: > Scutellaria baicalensis is a clinical adjuvant to lung cancer chemotherapy in China. > Scutellaria ethanol extracts selectively toxic to A549, SK-LU-1 and SK-MES-1. > Baicalin, baicalein and wogonin were toxic to all lung cancer cell lines. > Proteomics identified increased p53 and BAX in response to Scutellaria extracts.« less

  6. Effects of tanshinone nanoemulsion and extract on inhibition of lung cancer cells A549

    NASA Astrophysics Data System (ADS)

    Lee, W. D.; Liang, Y. J.; Chen, B. H.

    2016-12-01

    Danshen (Salvia miltiorrhiza), a Chinese medicinal herb, consists of several functional components including tanshinones responsible for prevention of several chronic diseases. This study intends to prepare tanshinone extract and nanoemulsion from danshen and determine their inhibition effect on lung cancer cells A549. A highly stable tanshinone nanoemulsion composed of Capryol 90, Tween 80, ethanol and deionized water with the mean particle size of 14.2 nm was successfully prepared. Tanshinone nanoemulsion was found to be more effective in inhibiting A549 proliferation than tanshinone extract. Both nanoemulsion and extract could penetrate into cytoplasm through endocytosis, with the former being more susceptible than the latter. A dose-dependent response in up-regulation of p-JNK, p53 and p21 and down-regulation of CDK2, cyclin D1 and cyclin E1 expressions was observed with the cell cycle arrested at G0/G1 phase. The cellular microcompartment change of A549 was also investigated. The study demonstrated that tanshinone nanoemulsion may be used as a botanic drug for treatment of lung cancer.

  7. Migration-stimulating factor (MSF) is over-expressed in non-small cell lung cancer and promotes cell migration and invasion in A549 cells over-expressing MSF

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Deng, Xuefeng, E-mail: dengxfdoctor@hotmail.com; Department of Cardio-thoracic Surgery, Affiliated Hospital of Academy of Military Medical Sciences; Ma, Qunfeng

    Migration-stimulating factor (MSF), an oncofetal truncated isoform of fibronectin, is a potent stimulator of cell invasion. However, its distribution and motogenic role in non-small cell lung cancer (NSCLC) have never been identified. In this study, real-time PCR and immunohistochemical staining (IHC) were performed to detect MSF mRNA and protein levels in tumor tissues and matched adjacent tumor-free tissues. Furthermore, to examine the effect of MSF on invasiveness, MSF was upregulated in A549 cells. The invasiveness and viability of A549 cells were then determined using a transwell migration assay and the 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) viability assays, respectively. The expression level ofmore » MSF in NSCLC tissue was markedly higher than in matched adjacent tumor-free tissue. Additionally, the level of MSF protein expression in stage III and IV NSCLC samples was higher than in stage I and II NSCLC samples. More importantly, we also demonstrated that migration and invasion of A549 cells increased substantially after upregulating MSF, although proliferation remained unchanged. Meanwhile, we found no correlation between increasing motility and invasiveness of MSF-overexpressing cells and expression levels and activities of matrix metalloprotease MMP-2 and MMP-9. Our current study shows that MSF plays a role in migration and invasion of A549 cells and suggests that MSF may be a potential biomarker of NSCLC progression. - Highlights: • MSF expression was upregulated in NSCLC and correlated with TNM stages. • MSF may be a new biomarker for NSCLC progression. • MSF promoted migration and invasion in A549 cells, independent of MMP-2/MMP-9 expression.« less

  8. The investigation of ceranib-2 on apoptosis and drug interaction with carboplatin in human non small cell lung cancer cells in vitro.

    PubMed

    Yildiz-Ozer, Merve; Oztopcu-Vatan, Pinar; Kus, Gokhan

    2018-02-01

    Ceramide is found to be involved in inhibition of cell division and induction of apoptosis in certain tumour cells. Ceranib-2 is an agent that increases ceramide levels by inhibiting ceramidase in cancer cells. Therefore, we aimed to investigate the effects of ceranib-2 on cell survival, apoptosis and interaction with carboplatin in human non-small cell lung cancer cells. The cytotoxic effect of ceranib-2 (1-100 µM) was determined by MTT assay in human lung adenocarcinoma (A549) and large cell lung carcinoma (H460) cells. Carboplatin (1-100 µM) and lung bronchial epithelial cells (BEAS-2B) were used as positive controls. Morphological and ultrastructural changes were analysed by light microscope and TEM. Apoptotic/necrotic cell death and acid ceramidase activity were analysed by ELISA. Combination effects of ceranib-2 and carboplatin were investigated by MTT. The expression levels of CASP3, CASP9, BAX and BCL-2 were examined by qRT-PCR. The IC 50 of ceranib-2 was determined as 22 μM in A549 cells and 8 μM in H460 cells for 24 h. Morphological changes and induction of DNA fragmentation have revealed apoptotic effects of ceranib-2 in both cell lines. Ceranib-2 and carboplatin has shown synergism in combined treatment at 10 and 25 μM doses in H460 cells for 24 h. Ceranib-2 inhibited acid ceramidase activity by 44% at 25 µM in H460 cells. Finally, CASP3, CASP9 and BAX expressions were increased while BCL-2 expression was reduced in both cells. Our results obtained some preliminary results about the cytotoxic and apoptotic effects of ceranib-2 for the first time in NSCLC cell lines.

  9. Involvement of lysosomal dysfunction in silver nanoparticle-induced cellular damage in A549 human lung alveolar epithelial cells.

    PubMed

    Miyayama, Takamitsu; Matsuoka, Masato

    2016-01-01

    While silver nanoparticles (AgNPs) are widely used in consumer and medical products, the mechanism by which AgNPs cause pulmonary cytotoxicity is not clear. AgNP agglomerates are found in endo-lysosomal structures within the cytoplasm of treated cells. In this study, the functional role of lysosomes in AgNP-induced cellular damage was examined in A549 human lung alveolar epithelial cells. We evaluated the intracellular distribution of AgNPs, lysosomal pH, cellular viability, Ag dissolution, and metallothionein (MT) mRNA levels in AgNP-exposed A549 cells that were treated with bafilomycin A1, the lysosomal acidification inhibitor. Exposure of A549 cells to citrate-coated AgNPs (20 nm diameter) for 24 h induced cellular damage and cell death at 100 and 200 μg Ag/ml, respectively. Confocal laser microscopic examination of LysoTracker-stained cells showed that AgNPs colocalized with lysosomes and their agglomeration increased in a dose-dependent manner (50-200 μg Ag/ml). In addition, the fluorescence signals of LysoTracker were reduced following exposure to AgNPs, suggesting the elevation of lysosomal pH. Treatment of A549 cells with 200 nM bafilomycin A1 and AgNPs (50 μg Ag/ml) further reduced the fluorescence signals of LysoTracker. AgNP-induced cell death was also increased by bafilomycin A1 treatment. Finally, treatment with bafilomycin A1 suppressed the dissolution of Ag and decreased the mRNA expression levels of MT-I and MT-II following exposure to AgNPs. The perturbation of lysosomal pH by AgNP exposure may play a role in AgNP agglomeration and subsequent cellular damage in A549 cells.

  10. Mycobacterium tuberculosis infection causes different levels of apoptosis and necrosis in human macrophages and alveolar epithelial cells.

    PubMed

    Danelishvili, Lia; McGarvey, Jeffery; Li, Yong-Jun; Bermudez, Luiz E

    2003-09-01

    Mycobacterium tuberculosis interacts with macrophages and epithelial cells in the alveolar space of the lung, where it is able to invade and replicate in both cell types. M. tuberculosis-associated cytotoxicity to these cells has been well documented, but the mechanisms of host cell death are not well understood. We examined the induction of apoptosis and necrosis of human macrophages (U937) and type II alveolar epithelial cells (A549) by virulent (H37Rv) and attenuated (H37Ra) M. tuberculosis strains. Apoptosis was determined by both enzyme-linked immunosorbent assay (ELISA) and TdT-mediated dUTP nick end labelling (TUNEL) assay, whereas necrosis was evaluated by the release of lactate dehydrogenase (LDH). Both virulent and attenuated M. tuberculosis induced apoptosis in macrophages; however, the attenuated strain resulted in significantly more apoptosis than the virulent strain after 5 days of infection. In contrast, cytotoxicity of alveolar cells was the result of necrosis, but not apoptosis. Although infection with M. tuberculosis strains resulted in apoptosis of 14% of the cells on the monolayer, cell death associated with necrosis was observed in 59% of alveolar epithelial cells after 5 days of infection. Infection with M. tuberculosis suppressed apoptosis of alveolar epithelial cells induced by the kinase inhibitor, staurosporine. Because our findings suggest that M. tuberculosis can modulate the apoptotic response of macrophages and epithelial cells, we carried out an apoptosis pathway-specific cDNA microarray analysis of human macrophages and alveolar epithelial cells. Whereas the inhibitors of apoptosis, bcl-2 and Rb, were upregulated over 2.5-fold in infected (48 h) alveolar epithelial cells, the proapoptotic genes, bad and bax, were downregulated. The opposite was observed when U937 macrophages were infected with M. tuberculosis. Upon infection of alveolar epithelial cells with M. tuberculosis, the generation of apoptosis, as determined by the

  11. Effect of microRNA-135a on Cell Proliferation, Migration, Invasion, Apoptosis and Tumor Angiogenesis Through the IGF-1/PI3K/Akt Signaling Pathway in Non-Small Cell Lung Cancer.

    PubMed

    Zhou, Yufei; Li, Shaoxia; Li, Jiangtao; Wang, Dongfeng; Li, Quanxing

    2017-01-01

    This study explored the ability of microRNA-135a (miR-135a) to influence cell proliferation, migration, invasion, apoptosis and tumor angiogenesis through the IGF-1/PI3K/Akt signaling pathway in non-small cell lung cancer (NSCLC). NSCLC tissues and adjacent normal tissues were collected from 138 NSCLC patients. Quantitative real-time polymerase chain reaction (qRT-PCR) was used to detect the expression levels of miR-135a and IGF-1, PI3K, Akt, VEGF, bFGF and IL-8 mRNA; western blotting was used to determine the expression levels of IGF-1, PI3K and Akt protein; and enzyme-linked immunosorbent assay (ELISA) was used to analyze the expression levels of VEGF, bFGF and IL-8 protein. Human NSCLC cell lines (A549, H460, and H1299) and the human bronchial epithelial cell line (HBE) were selected. A549 cells were assigned to blank, negative control (NC), miR-135a mimics, miR-135a inhibitors, IGF-1 siRNA and miR-135a inhibitors + IGF-1 siRNA groups. The following were performed: an MTT assay to assess cell proliferation, a scratch test to detect cell migration, a Transwell assay to measure cell invasion, and a flow cytometry to analyze cell apoptosis. The expression level of miR-135a was lower while those of IGF-1, PI3K and Akt mRNA were higher in NSCLC tissues than in the adjacent normal tissues. Dual-luciferase reporter assay indicated IGF-1 as a target of miR-135a. The in vitro results showed that compared with the blank group, cell proliferation, migration and invasion were suppressed, mRNA and protein levels of IGF-1, PI3K, Akt, VEGF, bFGF and IL-8 were reduced, and cell apoptosis was enhanced in the miR-135a mimics and IGF-1 siRNA groups. Compared with the IGF-1 siRNA group, cells in the miR-135a inhibitors + IGF-1 siRNA group demonstrated increased cell proliferation, migration and invasion, elevated mRNA and protein levels of IGF-1, PI3K, Akt, VEGF, bFGF and IL-8 and reduced cell apoptosis. These findings indicated that miR-135a promotes cell apoptosis and inhibits

  12. Gamma Irradiation Upregulates B-cell Translocation Gene 2 to Attenuate Cell Proliferation of Lung Cancer Cells Through the JNK and NF-κB Pathways.

    PubMed

    Wang, Peihe; Cai, Yuanyuan; Lin, Dongju; Jiang, Yingxiao

    2017-08-07

    Gamma ray can promote cancer cell apoptosis and cell cycle arrest. It is often used in the clinical treatment of tumors, including lung cancer. In this study, we aimed to explore the role of gamma ray treatment and its correlation with BTG2 in cell proliferation, apoptosis, and cell cycle arrest regulation in a lung cancer cell line. A549 cell viability, apoptosis rate, and cell cycle were investigated after gamma ray treatment. We then used siRNA for BTG2 to detect the effect of BTG2 knockdown on the progress of gamma ray-treated lung cancer cells. Finally, we investigated the signaling pathway by which gamma ray might regulate BTG2. We found that gamma ray inhibited A549 cell viability and promoted apoptosis and cell cycle arrest, while BTG2 knockdown could relieve the effect caused by gamma ray on A549 cells. Moreover, we confirmed that the effect of BTG2 partly depends on p53 expression and gamma ray-promoting BTG2 expression through the JNK/NF-κB signaling pathway. Our study assessed the possible mechanism of gamma ray in tumor treatment and also investigated the role of BTG2 in gamma ray therapy. All these findings might give a deep understanding of the effect of gamma ray on the progression of lung cancer involving BTG2.

  13. Synergistic Antitumor Effect of Oligogalacturonides and Cisplatin on Human Lung Cancer A549 Cells.

    PubMed

    Huang, Cian-Song; Huang, Ai-Chun; Huang, Ping-Hsiu; Lo, Diana; Wang, Yuh-Tai; Wu, Ming-Chang

    2018-06-14

    Cisplatin (DPP), a clinically potent antineoplastic agent, is limited by its severe adverse effects. The aim of this study was to investigate the effect of oligogalacturonides (OGA) and DDP on human lung cancer A549 cells. The combined use of OGA and DDP had a synergistic effect on the growth inhibition of A549 cells, changed the cell cycle distribution, and enhanced apoptotic response, especially in sequential combination treatment group of DDP 12 h + OGA 12 h. Western blot analyses showed that the combination treatment of OGA and DDP upregulated Bax, p53, and Caspase-3 and downregulated Bcl-2 proteins. More importantly, DDP-induced toxicity was attenuated by OGA and DDP combination treatment in normal HEK293 cells. Our data suggests that the combined use of OGA from natural sources and DDP could be an important new adjuvant therapy for lung cancer as well as offer important insights for reducing kidney toxicity of DDP and delaying the development of DDP resistance.

  14. Enhanced expression of PKM2 associates with the biological properties of cancer stem cells from A549 human lung cancer cells.

    PubMed

    Guo, Chang-Ying; Yan, Chen; Luo, Lan; Goto, Shinji; Urata, Yoshishige; Xu, Jian-Jun; Wen, Xiao-Ming; Kuang, Yu-Kang; Tou, Fang-Fang; Li, Tao-Sheng

    2017-04-01

    Cancer cells express the M2 isoform of glycolytic enzyme pyruvate kinase (PKM2) for favoring the survival under a hypoxic condition. Considering the relative low oxygen microenvironment in stem cell niche, we hypothesized that an enhanced PKM2 expression associates with the biological properties of cancer stem cells. We used A549 human lung cancer cell line and surgical resected lung cancer tissue samples from patients for experiments. We confirmed the co-localization of PKM2 and CD44, a popular marker for cancer stem cells in lung cancer tissue samples from patients. The expression of PKM2 was clearly observed in approximately 80% of the A549 human lung cancer cells. Remarkably, enhanced expression of PKM2 was specially observed in these cells that also positively expressed CD44. Downregulation of PKM2 in CD44+ cancer stem cells by siRNA significantly impaired the potency for spheroid formation, decreased the cell survival under fetal bovine serum deprivation and hypoxic conditions, but increased their sensitivity to anti-cancer drug of cisplatin and γ-ray. The enhanced expression of PKM2 seems to associate with the biological properties of cancer stem cells from A549 human lung cancer cells. Selective targeting of PKM2 may provide a new strategy for cancer therapy, especially for patients with therapeutic resistance.

  15. Knockdown of Immature Colon Carcinoma Transcript 1 Inhibits Proliferation and Promotes Apoptosis of Non–Small Cell Lung Cancer Cells

    PubMed Central

    He, Jiantao; Zhang, Shenghui; Yang, Qingbo; Wang, Bo; Liu, Zhiyu; Wu, Xintian

    2016-01-01

    Non–small cell lung cancer, as the most frequent type lung cancer, has lower survival rate of 5 years, despite improvements in surgery and chemotherapy. Previous studies showed immature colon carcinoma transcript 1 is closely related to tumorigenesis of human cancer cells. In the present study, we found immature colon carcinoma transcript 1 was overexpressed in lung cancer tissues using Oncomine database mining, and the biological effect of immature colon carcinoma transcript 1 was investigated in non–small cell lung cancer cell lines 95D and A549. Lentivirus-mediated RNA interference was used to knock down immature colon carcinoma transcript 1 expression in 95D and A549 cells in vitro, and the knockdown efficiency was determined using quantitative real-time polymerase chain reaction and Western blot assay. Knockdown of immature colon carcinoma transcript 1 significantly suppressed non–small cell lung cancer cell proliferation and colony formation ability confirmed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and colony formation assay. Flow cytometry was applied to measure cell cycle arrest, and the result showed the cell cycle arrested in G2/M phase in 95D cells and arrested in G0/G1 phase in A549 cells. Furthermore, we measured the levels of cell cycle–associated proteins by Western blot analysis and found immature colon carcinoma transcript 1–mediated cell proliferation inhibition appeared due to downregulation of cell cycle activator cyclin D1 and upregulation of cell cycle inhibitor p21. In addition, immature colon carcinoma transcript 1 silencing significantly induced non–small cell lung cancer cell apoptosis by annexin V/7-amino-actinomycin D double-staining assay. All our data suggest that immature colon carcinoma transcript 1 may play an important role for non–small cell lung cancer cell proliferation and could be a potential molecular target for diagnosing and treating human non–small cell lung cancer. PMID:27413166

  16. Irisin-mediated protective effect on LPS-induced acute lung injury via suppressing inflammation and apoptosis of alveolar epithelial cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Shao, Lei; Jinan Central Hospital Affiliated to Shandong University, Jinan 250012; Meng, Di

    It is considered that the essence of acute lung injury (ALI) is an excessive and uncontrolled inflammatory response in lung, of which mainly is attributed to the release of inflammatory mediators. Recent studies demonstrated that irisin, which is a metabolism associated factor after physical exercise could suppression of inflammation by regulating cellular signaling pathways, however, the underlying molecular mechanism remains to be determined. The present study aimed to reveal the potential mechanism responsible for the anti-inflammatory effects of irisin on LPS-induced acute lung injury in mice and in A549 cells. The results of histopathological changes showed that irisin ameliorated the lungmore » injury that was induced by LPS in time- and dose-dependent manner. QRT-PCR assays demonstrated that irisin suppressed the production of IL-1β, IL-6, MCP-1 and TNF-α, and western blot assays demonstrated that irisin suppressed apoptosis of ALI. The expression of caspase-3 and Bax were decreased and Bcl-2 was increased by irisin administration. Further study was conducted on nuclear factor (NF)-κB and mitogen-activated protein kinase (MAPK) using pathways using western blots. The results showed that irisin inhibited reduced LPS-induced activation of MAPK and NF-κB signaling. All results indicated that irisin has protective effect on LPS-induced ALI in mice and in A549 cells. Thus, irisn related with physical exercise may be a potential therapy for the treatment of pulmonary inflammation. - Highlights: • Irisin inhibited the inflammation reactivity of cells and pathological changes of LPS-induced lung injury in mice. • Irisin inhibited mRNA expression of inflammatory cytokines induced by LPS in A549 cells. • Irisin inhibited apoptosis induced by LPS in the injured lung. • Irisin reduced LPS-induced activation of MAPK and NF-κB signaling pathways.« less

  17. Induction of reactive oxygen species-stimulated distinctive autophagy by chelerythrine in non-small cell lung cancer cells.

    PubMed

    Tang, Zheng-Hai; Cao, Wen-Xiang; Wang, Zhao-Yu; Lu, Jia-Hong; Liu, Bo; Chen, Xiuping; Lu, Jin-Jian

    2017-08-01

    Chelerythrine (CHE), a natural benzo[c]phenanthridine alkaloid, shows anti-cancer effect through a number of mechanisms. Herein, the effect and mechanism of the CHE-induced autophagy, a type II programmed cell death, in non-small cell lung cancer (NSCLC) cells were studied for the first time. CHE induced cell viability decrease, colony formation inhibition, and apoptosis in a concentration-dependent manner in NSCLC A549 and NCI-H1299 cells. In addition, CHE triggered the expression of phosphatidylethanolamine-modified microtubule-associated protein light-chain 3 (LC3-II). The CHE-induced expression of LC3-II was further increased in the combination treatment with chloroquine (CQ), an autophagy inhibitor, and large amounts of red-puncta were observed in the CHE-treated A549 cells with stable expression of mRFP-EGFP-LC3, indicating that CHE induces autophagy flux. Silence of beclin 1 reversed the CHE-induced expression of LC3-II. Inhibition of autophagy remarkably reversed the CHE-induced cell viability decrease and apoptosis in NCI-H1299 cells but not in A549 cells. Furthermore, CHE triggered reactive oxygen species (ROS) generation in both cell lines. A decreased level of ROS through pretreatment with N-acetyl-L-cysteine reversed the CHE-induced cell viability decrease, apoptosis, and autophagy. Taken together, CHE induced distinctive autophagy in A549 (accompanied autophagy) and NCI-H1299 (pro-death autophagy) cells and a decreased level of ROS reversed the effect of CHE in NSCLC cells in terms of cell viability, apoptosis, and autophagy. Copyright © 2017 The Authors. Published by Elsevier B.V. All rights reserved.

  18. Deactivation of cisplatin-resistant human lung/ovary cancer cells with pyropheophorbide-α methyl ester-photodynamic therapy.

    PubMed

    Qian, Guanhua; Wang, Li; Zheng, Xueling; Yu, Tinghe

    2017-12-02

    The aim of this study was to determine whether photodynamic therapy (PDT) alone or combined with cisplatin (DDP), can deactivate cisplatin-resistant cancer cells. Human cancer cell lines A549 and SKOV3, and chemoresistant sublines A549/DDP and SKOV3/DDP, were subjected to PDT, DDP, or PDT combined with DDP. Cell viability and apoptosis were analyzed, and then intracellular reactive oxygen species (ROS) and proteins related to apoptosis were determined. PDT caused cell death, and PDT combined with DDP led to the highest percentage of dead cells in 4 cell lines; similar results were detected in ROS; a quantification evaluation manifested that the combined effect was addition. DDP increased the percentage of apoptotic cells, and the ROS level in A549 and SKOV3 cells, which was not observed in A549/DDP and SKOV3/DDP cells. Western blot revealed an increase of caspase 3 and Bax, and a decrease of Bcl-2, demonstrating the occurrence of apoptosis. The data suggest that PDT can efficiently deactivate resistant cells and enhance the action of DDP against resistant cancer cells.

  19. Flavonoids and Tannins from Smilax china L. Rhizome Induce Apoptosis Via Mitochondrial Pathway and MDM2-p53 Signaling in Human Lung Adenocarcinoma Cells.

    PubMed

    Fu, San; Yang, Yanfang; Liu, Dan; Luo, Yan; Ye, Xiaochuan; Liu, Yanwen; Chen, Xin; Wang, Song; Wu, Hezhen; Wang, Yuhang; Hu, Qiwei; You, Pengtao

    2017-01-01

    In vitro evidence indicates that Smilax china L. rhizome (SCR) can inhibit cell proliferation. Therefore, in the present study, we analyzed the effects in vitro of SCR extracts on human lung adenocarcinoma A549 cells. Our results showed that A549 cell growth was inhibited in a dose- and time-dependent manner after treatment with SCR extracts. Total flavonoids and total tannins from SCR induced A549 apoptosis in a dose-dependent manner, as shown by our flow cytometry analysis, which was consistent with the alterations in nuclear morphology we observed. In addition, the total apoptotic rate induced by total tannins was higher than the rate induced by total flavonoids at the same dose. Cleaved-caspase-3 protein levels in A549 cells after treatment with total flavonoids or total tannins were increased in a dose-dependent manner, followed by the activation of caspase-8 and caspase-9, finally triggering to PARP cleavage. Furthermore, total flavonoids and total tannins increased the expression of Bax, decreased the expression of Bcl-2, and promoted cytochrome [Formula: see text] release. Moreover, MDM2 and p-MDM2 proteins were decreased, while p53 and p-p53 proteins were increased, both in a dose-dependent manner, after A549 treatment with total flavonoids and total tannins. Finally, cleaved-caspase-3 protein levels in the total flavonoids or total tannins-treated H1299 (p53 null) and p53-knockdown A549 cells were increased. Our results indicated that total flavonoids and total tannins from SCR exerted a remarkable effect in reducing A549 growth through their action on mitochondrial pathway and disruption of MDM2-p53 balance. Hence, our findings demonstrated a potential application of total flavonoids and total tannins from SCR in the treatment of human lung adenocarcinoma.

  20. Effects of quercetin on CDK4 mRNA and protein expression in A549 cells infected by H1N1

    PubMed Central

    WAN, QIAOFENG; WANG, HAO; LIN, YUAN; GU, LIGANG; HAN, MEI; YANG, ZHIWEI; ZHANG, YANLI; MA, RUI; WANG, LI; WANG, ZHISHENG

    2013-01-01

    This study was conducted to investigate the effects of quercetin on the expression of cyclin-dependent kinase (CDK4) mRNA and protein in A549 lung epithelial tumor cells infected by H1N1. First, the Thiazolyl Blue Tetrazolium Bromide (MTT) method was used to determine H1N1 virulence, quercetin cytotoxicity and inhibition of the cytopathic effect of H1N1 on A549 cells by quercetin. Subsequently, 100 TCID50 H1N1 was used to infect A549 cells for 2 h prior to culture in maintenance media containing 10 mg/l quercetin. After 4, 12, 24 and 48 h of culture, the cells were collected and total RNA and protein were extracted. Fluorescent quantitative polymerase chain reaction and western blot analysis were then performed to assess the expression of CDK4 mRNA and protein. The experiment demonstrated that the TCID50 of H1N1 in A549 cells was 10−4.75, the maximum non-toxic concentration of quercetin in A549 cells was 30–60 mg/l and the minimum effective concentration of quercetin for the inhibition of the H1N1 cytopathic effect on A549 cells was 10 mg/l. The results indicated that quercetin may significantly inhibit CDK4 mRNA and protein overexpression caused by H1N1 within 4–48 h. In conclusion, quercetin may protect against H1N1 infection by effectively reducing the mRNA and protein expression of CDK4 caused by H1N1 infection. PMID:24649026

  1. Ghrelin promotes human non-small cell lung cancer A549 cell proliferation through PI3K/Akt/mTOR/P70S6K and ERK signaling pathways.

    PubMed

    Zhu, Jianhua; Yao, Jianfeng; Huang, Rongfu; Wang, Yueqin; Jia, Min; Huang, Yan

    2018-04-06

    Ghrelin is a gastric acyl-peptide that plays an important role in cell proliferation. In the present study, we explored the role of ghrelin in A549cell proliferation and the possible molecular mechanisms. We found that ghrelin promotes A549cell proliferation, knockdown of the growth hormone secretagogue receptor (GHSR) attenuated A549cell proliferation caused by ghrelin. Ghrelin induced the rapid phosphorylation of phosphatidylinositol 3-kinase (PI3K), Akt, ERK, mammalian target of rapamycin (mTOR) and P70S6K. PI3K inhibitor (LY 294002), ERK inhibitor (PD98059) and mTOR inhibitor (Rapamycin) inhibited ghrelin-induced A549cell proliferation. Moreover, GHSR siRNA inhibited phosphorylation of PI3K, Akt, ERK, mTOR and P70S6K induced by ghrelin. Akt and mTOR/P70S6K phosphorylation was inhibited by LY 294002 but not by PD98059. These results indicate that ghrelin promotes A549cell proliferation via GHSR-dependent PI3K/Akt/mTOR/P70S6K and ERK signaling pathways. Copyright © 2018 Elsevier Inc. All rights reserved.

  2. Fisetin inhibits the growth and migration in the A549 human lung cancer cell line via the ERK1/2 pathway.

    PubMed

    Wang, Junjian; Huang, Shaoxiang

    2018-03-01

    Lung cancer is the most prevalent malignant tumor type in the developed world and the discovery of novel anti-tumor drugs is a research hotspot. Fisetin, a naturally occurring flavonoid, has been reported to have anti-cancer effects in multiple tumor types. The present study found that fisetin inhibited the growth and migration of non-small cell lung cancer in vitro . MTT, wound-healing, cell-matrix adhesion and Transwell assays were performed and demonstrated that fisetin suppressed proliferation, migration, adhesion and invasion, respectively. Flow cytometric analysis indicated that fisetin induced apoptosis in the A549 cell line by decreasing the expression of c-myc, cyclin-D1, cyclooxygenase-2, B cell lymphoma-2, CXC chemokine receptor type 4, cluster of differentiation 44 and metalloproteinase-2/9, increasing the expression of cyclin dependent kinase inhibitor (CDKN) 1A/B, CDKN2D and E-cadherin and increasing the activity of caspase-3/9 via targeting the extracellular signal-regulated kinase signaling pathway. The results provided comprehensive evidence for the anti-tumor effects of fisetin in non-small cell lung cancer in vitro , which may provide a novel approach for clinical treatment.

  3. Fisetin inhibits the growth and migration in the A549 human lung cancer cell line via the ERK1/2 pathway

    PubMed Central

    Wang, Junjian; Huang, Shaoxiang

    2018-01-01

    Lung cancer is the most prevalent malignant tumor type in the developed world and the discovery of novel anti-tumor drugs is a research hotspot. Fisetin, a naturally occurring flavonoid, has been reported to have anti-cancer effects in multiple tumor types. The present study found that fisetin inhibited the growth and migration of non-small cell lung cancer in vitro. MTT, wound-healing, cell-matrix adhesion and Transwell assays were performed and demonstrated that fisetin suppressed proliferation, migration, adhesion and invasion, respectively. Flow cytometric analysis indicated that fisetin induced apoptosis in the A549 cell line by decreasing the expression of c-myc, cyclin-D1, cyclooxygenase-2, B cell lymphoma-2, CXC chemokine receptor type 4, cluster of differentiation 44 and metalloproteinase-2/9, increasing the expression of cyclin dependent kinase inhibitor (CDKN) 1A/B, CDKN2D and E-cadherin and increasing the activity of caspase-3/9 via targeting the extracellular signal-regulated kinase signaling pathway. The results provided comprehensive evidence for the anti-tumor effects of fisetin in non-small cell lung cancer in vitro, which may provide a novel approach for clinical treatment. PMID:29467859

  4. JA, a new type of polyunsaturated fatty acid isolated from Juglans mandshurica Maxim, limits the survival and induces apoptosis of heptocarcinoma cells.

    PubMed

    Gao, Xiu-Li; Lin, Hua; Zhao, Wei; Hou, Ya-Qin; Bao, Yong-Li; Song, Zhen-Bo; Sun, Lu-Guo; Tian, Shang-Yi; Liu, Biao; Li, Yu-Xin

    2016-03-01

    Juglans mandshurica Maxim (Juglandaceae) is a famous folk medicine for cancer treatment and some natural compounds isolated from it have been studied extensively. Previously we isolated a type of ω-9 polyunsaturated fatty acid (JA) from the bark of J. mandshurica, however little is known about its activity and the underlying mechanisms. In this study, we studied anti-tumor activity of JA on several human cancer cell lines. Results showed that JA is cytotoxic to HepG2, MDA-MB-231, SGC-7901, A549 and Huh7 cells at a concentration exerting minimal toxic effects on L02 cells. The selective toxicity of JA was better than other classical anti-cancer drugs. Further investigation indicated that JA could induce cell apoptosis, characterized by chromatin condensation, DNA fragmentation and activation of the apoptosis-associated proteins such as Caspase-3 and PARP-1. Moreover, we investigated the cellular apoptosis pathway involved in the apoptosis process in HepG2 cells. We found that proteins involved in mitochondrion (cleaved-Caspase-9, Apaf-1, HtrA2/Omi, Bax, and Mitochondrial Bax) and endocytoplasmic reticulum (XBP-1s, GRP78, cleaved-Caspase-7 and cleaved-Caspase-12) apoptotic pathways were up-regulated when cells were treated by JA. In addition, a morphological change in the mitochondrion was detected. Furthermore, we found that JA could inhibit DNA synthesis and induce G2/M cell cycle arrest. The expression of G2-to-M transition related proteins, such as CyclinB1 and phosphorylated-CDK1, were reduced. In contrast, the G2-to-M inhibitor p21 was increased in JA-treated cells. Overall, our results suggest that JA can induce mitochondrion- and endocytoplasmic reticulum-mediated apoptosis, and G2/M phase arrest in HepG2 cells, making it a promising therapeutic agent against hepatoma.

  5. BCI induces apoptosis via generation of reactive oxygen species and activation of intrinsic mitochondrial pathway in H1299 lung cancer cells.

    PubMed

    Shin, Jong-Woon; Kwon, Sae-Bom; Bak, Yesol; Lee, Sang-Ku; Yoon, Do-Young

    2018-03-28

    The compound (E)-2-benzylidene-3-(cyclohexylamino)-2,3-dihydro-1H-inden-1-one (BCI) is known as an inhibitor of dual specific phosphatase 1/6 and mitogen-activated protein kinase. However, its precise anti-lung cancer mechanism remains unknown. In this study, the effects of BCI on the viability of non-small cell lung cancer cell lines NCI-H1299, A549, and NCI-H460 were evaluated. We confirmed that BCI significantly inhibited the viability of p53(-) NCI-H1299 cells as compared to NCI-H460 and A549 cells, which express wild-type p53. Furthermore, BCI treatment increased the level of cellular reactive oxygen species and pre-treatment of cells with N-acetylcysteine markedly attenuated BCI-mediated apoptosis of NCI-H1299 cells. BCI induced cellular morphological changes, inhibited viability, and produced reactive oxygen species in NCI-H1299 cells in a dose-dependent manner. BCI induced processing of caspase-9, caspase-3, and poly ADP-ribose polymerase as well as the release of cytochrome c from the mitochondria into the cytosol. In addition, BCI downregulated Bcl-2 expression and enhanced Bax expression in a dose-dependent manner in NCI-H1299 cells. However, BCI failed to modulate the expression of the death receptor and extrinsic factor caspase-8 and Bid, a linker between the intrinsic and extrinsic apoptotic pathways in NCI-H1299 cells. Thus, BCI induces apoptosis via generation of reactive oxygen species and activation of the intrinsic pathway in NCI-H1299 cells.

  6. Anti-lung cancer potential of pure esteric-glycoside condurangogenin A against nonsmall-cell lung cancer cells in vitro via p21/p53 mediated cell cycle modulation and DNA damage-induced apoptosis.

    PubMed

    Sikdar, Sourav; Mukherjee, Avinaba; Khuda-Bukhsh, Anisur Rahman

    2015-05-01

    Marsdenia condurango (condurango) is a tropical woody vine native to South America. Our earlier study was limited to evaluation of anti-cancer potentials of crude condurango extract and its glycoside-rich components in vitro on lung cancer. This study aims at evaluating the effect of the single isolated active ingredient condurangogenin A (ConA; C32H42O7) on A549, H522 and H460-nonsmall-cell lung cancer cells. ConA was isolated by column chromatography and analyzed by mass spectroscopy, Fourier transform infrared spectroscopy and proton-nuclear magnetic resonance. diphenyltetrazolium bromide assays were conducted on three cell-types using 6%-alcohol as control. Critical studies on cellular morphology, cell-cycle regulation, reactive oxygen species, mitochondrial membrane potential, and DNA-damage were made, and expressions of related signaling markers studied. As IC50 doses of ConA proved to be too high and toxic to both A549 and H522 cells, all experimental studies were carried out on H460 cells with the IC50 dose (32 μg/ml - 24 h). Cellular morphology revealed typical apoptotic features after ConA treatment. At early treatment hours (2 h-12 h), maximum cells were arrested at G0/G1 phase that could be correlated with reduced level of cyclin D1-CDK with p21 up-regulation. At 18 h - 24 h, sub G0/G1 cell population was increased gradually, as revealed from cytochrome-c release and caspase-3 activation, further confirming the apoptosis-inducing ability of ConA at later phases. Gradual increase of TUNEL-positive cells with significant modulation of mitochondria-dependent apoptotic markers at longer time-points would establish apoptosis-induction property of ConA, indicating its potential as a strong candidate for anti-cancer drug formulation. Further studies are warranted against other types of cancer cells and animal models before its possible human use.

  7. Anti-lung cancer potential of pure esteric-glycoside condurangogenin A against nonsmall-cell lung cancer cells in vitro via p21/p53 mediated cell cycle modulation and DNA damage-induced apoptosis

    PubMed Central

    Sikdar, Sourav; Mukherjee, Avinaba; Khuda-Bukhsh, Anisur Rahman

    2015-01-01

    Background: Marsdenia condurango (condurango) is a tropical woody vine native to South America. Our earlier study was limited to evaluation of anti-cancer potentials of crude condurango extract and its glycoside-rich components in vitro on lung cancer. Objective: This study aims at evaluating the effect of the single isolated active ingredient condurangogenin A (ConA; C32H42O7) on A549, H522 and H460-nonsmall-cell lung cancer cells. Materials and Methods: ConA was isolated by column chromatography and analyzed by mass spectroscopy, Fourier transform infrared spectroscopy and proton-nuclear magnetic resonance. diphenyltetrazolium bromide assays were conducted on three cell-types using 6%-alcohol as control. Critical studies on cellular morphology, cell-cycle regulation, reactive oxygen species, mitochondrial membrane potential, and DNA-damage were made, and expressions of related signaling markers studied. Results: As IC50 doses of ConA proved to be too high and toxic to both A549 and H522 cells, all experimental studies were carried out on H460 cells with the IC50 dose (32 μg/ml − 24 h). Cellular morphology revealed typical apoptotic features after ConA treatment. At early treatment hours (2 h-12 h), maximum cells were arrested at G0/G1 phase that could be correlated with reduced level of cyclin D1-CDK with p21 up-regulation. At 18 h − 24 h, sub G0/G1 cell population was increased gradually, as revealed from cytochrome-c release and caspase-3 activation, further confirming the apoptosis-inducing ability of ConA at later phases. Gradual increase of TUNEL-positive cells with significant modulation of mitochondria-dependent apoptotic markers at longer time-points would establish apoptosis-induction property of ConA, indicating its potential as a strong candidate for anti-cancer drug formulation. Conclusion: Further studies are warranted against other types of cancer cells and animal models before its possible human use. PMID:26109778

  8. Development of a transmission alpha particle dosimetry technique using A549 cells and a Ra-223 source for targeted alpha therapy.

    PubMed

    Al Darwish, R; Staudacher, A H; Li, Y; Brown, M P; Bezak, E

    2016-11-01

    In targeted radionuclide therapy, regional tumors are targeted with radionuclides delivering therapeutic radiation doses. Targeted alpha therapy (TAT) is of particular interest due to its ability to deliver alpha particles of high linear energy transfer within the confines of the tumor. However, there is a lack of data related to alpha particle distribution in TAT. These data are required to more accurately estimate the absorbed dose on a cellular level. As a result, there is a need for a dosimeter that can estimate, or better yet determine the absorbed dose deposited by alpha particles in cells. In this study, as an initial step, the authors present a transmission dosimetry design for alpha particles using A549 lung carcinoma cells, an external alpha particle emitting source (radium 223; Ra-223) and a Timepix pixelated semiconductor detector. The dose delivery to the A549 lung carcinoma cell line from a Ra-223 source, considered to be an attractive radionuclide for alpha therapy, was investigated in the current work. A549 cells were either unirradiated (control) or irradiated for 12, 1, 2, or 3 h with alpha particles emitted from a Ra-223 source positioned below a monolayer of A549 cells. The Timepix detector was used to determine the number of transmitted alpha particles passing through the A549 cells and DNA double strand breaks (DSBs) in the form of γ-H2AX foci were examined by fluorescence microscopy. The number of transmitted alpha particles was correlated with the observed DNA DSBs and the delivered radiation dose was estimated. Additionally, the dose deposited was calculated using Monte Carlo code SRIM. Approximately 20% of alpha particles were transmitted and detected by Timepix. The frequency and number of γ-H2AX foci increased significantly following alpha particle irradiation as compared to unirradiated controls. The equivalent dose delivered to A549 cells was estimated to be approximately 0.66, 1.32, 2.53, and 3.96 Gy after 12, 1, 2, and 3 h

  9. Apigenin inhibits cell proliferation, migration, and invasion by targeting Akt in the A549 human lung cancer cell line.

    PubMed

    Zhou, Zhongping; Tang, Miaomiao; Liu, Yi; Zhang, Zhuyi; Lu, Rongzhu; Lu, Jian

    2017-04-01

    Apigenin (APG), a widely distributed flavonoid in vegetables and fruits, with low toxicity, and a nonmutagenic characteristic, has been reported to have many targets. Evidence indicates that APG can inhibit the proliferation, migration, invasion, and metastasis of some tumor cells, but the mechanism, specifically in lung cancer, is unclear. The phosphoinositide 3-kinase (PI3K)/Akt signaling pathway regulates a diverse set of cellular functions relevant to the growth and progression of lung cancer, including proliferation, survival, migration, and invasion. Our results showed that APG exerted anti-proliferation, anti-migration, and anti-invasion effects in A549 human lung cancer cells by targeting the PI3K/Akt signaling pathway. 3-(4, 5-dimethylthiszol-2-yl)-2, 5-diphenytetrazolium bromide assay and colony formation assay showed that APG suppressed cell proliferation in a dose-dependent and time-dependent manner. Cell motility and invasiveness were assayed using a wound healing and Transwell assay, suggesting that APG inhibited the migration and invasion of A549 cells. Western blot analyses were carried out to examine the Akt signaling pathways. The results confirmed that APG decreased Akt expression and its activation. Then, cells were transfected with Akt-active and Akt-DN plasmids separately. The migration and invasion of A549 cells were significantly changed, constitutively activating Akt or knocking down Akt, indicating that APG can suppress the migration and invasion of lung cancer cells by modulating the PI3K/Akt signaling pathway. Furthermore, the results indicated that APG not only suppressed phosphorylation of Akt, thereby preventing its activation, but also inhibited its downstream gene expression of matrix metalloproteinases-9, glycogen synthase kinase-3β, and HEF1. Together, APG is a new inhibitor of Akt in lung cancer and a potential natural compound for cancer chemoprevention.

  10. Protein alkylation, transcriptional responses and cytochrome c release during acrolein toxicity in A549 cells: influence of nucleophilic culture media constituents.

    PubMed

    Thompson, Colin A; Burcham, Philip C

    2008-06-01

    Acrolein is a toxic combustion product that elicits apoptotic and/or necrotic cell death depending on the conditions under which exposure occurs. As a strong electrophile, side-reactions with nucleophilic media constituents seem likely to accompany study of its toxicity in vitro, but these reactions are poorly characterized. We have thus examined the effect of media composition on the toxicity of acrolein in A549 cells. Cells were exposed to acrolein in either Dulbecco's buffered saline (DBS) or F12 supplemented with various concentrations of fetal bovine serum. Cell viability was assessed using the MTT assay, while heme oxygenase-1 (HO-1) and cytoplasmic cytochrome c were measured as respective markers of transcriptional response and apoptosis. Protein damage was evaluated using the protein carbonyl assay. Compared to F12 media (with or without serum), maximal cell death as evaluated using the MTT assay, as well as adduction of intracellular proteins, occurred when cells were exposed to acrolein in DBS. In contrast, cytochrome c release was maximal in cells exposed to acrolein in serum-containing F12, conditions which inhibited protein modification and overt cell death. These findings highlight the need for careful attention to experimental conditions when conducting in vitro toxicological studies of reactive substances.

  11. Simultaneous targeting of ATM and Mcl-1 increases cisplatin sensitivity of cisplatin-resistant non-small cell lung cancer.

    PubMed

    Zhang, Fuquan; Shen, Mingjing; Yang, Li; Yang, Xiaodong; Tsai, Ying; Keng, Peter C; Chen, Yongbing; Lee, Soo Ok; Chen, Yuhchyau

    2017-08-03

    Development of cisplatin-resistance is an obstacle in non-small cell lung cancer (NSCLC) therapeutics. To investigate which molecules are associated with cisplatin-resistance, we analyzed expression profiles of several DNA repair and anti-apoptosis associated molecules in parental (A549P and H157P) and cisplatin-resistant (A549CisR and H157CisR) NSCLC cells. We detected constitutively upregulated nuclear ATM and cytosolic Mcl-1 molcules in cisplatin-resistant cells compared with parental cells. Increased levels of phosphorylated ATM (p-ATM) and its downstream molecules, CHK2, p-CHK2, p-53, and p-p53 were also detected in cisplatin-resistant cells, suggesting an activation of ATM signaling in these cells. Upon inhibition of ATM and Mcl-1 expression/activity using specific inhibitors of ATM and/or Mcl-1, we found significantly enhanced cisplatin-cytotoxicity and increased apoptosis of A549CisR cells after cisplatin treatment. Several A549CisR-derived cell lines, including ATM knocked down (A549CisR-siATM), Mcl-1 knocked down (A549CisR-shMcl1), ATM/Mcl-1 double knocked down (A549CisR-siATM/shMcl1) as well as scramble control (A549CisR-sc), were then developed. Higher cisplatin-cytotoxicity and increased apoptosis were observed in A549CisR-siATM, A549CisR-shMcl1, and A549CisR-siATM/shMcl1 cells compared with A549CisR-sc cells, and the most significant effect was shown in A549CisR-siATM/shMcl1 cells. In in vivo mice studies using subcutaneous xenograft mouse models developed with A549CisR-sc and A549CisR-siATM/shMcl1 cells, significant tumor regression in A549CisR-siATM/shMcl1 cells-derived xenografts was observed after cisplatin injection, but not in A549CisR-sc cells-derived xenografts. Finally, inhibitor studies revealed activation of Erk signaling pathway was most important in upregulation of ATM and Mcl-1 molcules in cisplatin-resistant cells. These studies suggest that simultaneous blocking of ATM/Mcl-1 molcules or downstream Erk signaling may recover the

  12. Picfeltarraenin IA inhibits lipopolysaccharide-induced inflammatory cytokine production by the nuclear factor-κB pathway in human pulmonary epithelial A549 cells.

    PubMed

    Shi, Rong; Wang, Qing; Ouyang, Yang; Wang, Qian; Xiong, Xudong

    2016-02-01

    The present study aimed to investigate the effect of picfeltarraenin IA (IA) on respiratory inflammation by analyzing its effect on interleukin (IL)-8 and prostaglandin E2 (PGE2) production. The expression of cyclooxygenase 2 (COX2) in human pulmonary adenocarcinoma epithelial A549 cells in culture was also examined. Human pulmonary epithelial A549 cells and the human monocytic leukemia THP-1 cell line were used in the current study. Cell viability was measured using a methylthiazol tetrazolium assay. The production of IL-8 and PGE2 was investigated using an enzyme-linked immunosorbent assay. The expression of COX2 and nuclear factor-κB (NF-κB)-p65 was examined using western blot analysis. Treatment with lipopolysaccharide (LPS; 10 µg/ml) resulted in the increased production of IL-8 and PGE2, and the increased expression of COX2 in the A549 cells. Furthermore, IA (0.1-10 µmol/l) significantly inhibited PGE2 production and COX2 expression in cells with LPS-induced IL-8, in a concentration-dependent manner. The results suggested that IA downregulates LPS-induced COX2 expression, and inhibits IL-8 and PGE2 production in pulmonary epithelial cells. Additionally, IA was observed to suppress the expression of COX2 in THP-1 cells, and also to regulate the expression of COX2 via the NF-κB pathway in the A549 cells, but not in the THP-1 cells. These results indicate that IA regulates LPS-induced cytokine release in A549 cells via the NF-κB pathway.

  13. Taspine derivative 12k suppressed A549 cell migration through the Wnt/β-catenin and EphrinB2 signaling pathway.

    PubMed

    Dai, Bingling; Ma, Yujiao; Yang, Tianfeng; Wang, Wenjie; Zhang, Yanmin

    2017-03-01

    12k, a taspine derivative, has been demonstrated to have the potent anti-tumor activity in lung cancer and colorectal cancer. The study aims to further explore the underlying mechanisms of 12k on A549 cell migration in vitro. Our data demonstrated that 12k negatively regulated Wnt signaling pathway by suppressing the phosphorylation of LRP5/6, and inhibiting the expression and nuclear translocation of β-catenin. 12k was shown to downregulate MMP3 and MMP7 expression which regulated by β-catenin interacts with TCF/LEF in the nucleus, and effectively impaired the related migration protein expression of MMP2 and MMP9 in A549 cells. In addition, 12k repressed the EphrinB2 and its PDZ protein, impairing the VEGFR2 and VEGFR3 expression in A549 cells, as well as inhibited the downstream of VEGFR2 included PI3K/AKT/mTOR and ERK/MAPK signaling pathways. Taken together, our findings revealed that 12k suppressed migration of A549 cells through the Wnt/β-catenin signaling pathway and EphrinB2 related signaling pathway. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  14. Diabetes and renal tubular cell apoptosis

    PubMed Central

    Habib, Samy L

    2013-01-01

    Apoptosis contributes to the development of diabetic nephropathy, but the mechanism by which high glucose induces apoptosis is not fully understood. Apoptosis of tubular epithelial cells is a major feature of diabetic kidney disease, and hyperglycemia triggers the generation of free radicals and oxidant stress in tubular cells. Hyperglycemia and high glucose in vitro also lead to apoptosis, a form of programmed cell death. High glucose similar to those seen with hyperglycemia in people with diabetes mellitus, lead to accelerated apoptosis, a form of programmed cell death characterized by cell shrinkage, chromatin condensation and DNA fragmentation, in variety of cell types, including renal proximal tubular epithelial cells. PMID:23593533

  15. Diabetes and renal tubular cell apoptosis.

    PubMed

    Habib, Samy L

    2013-04-15

    Apoptosis contributes to the development of diabetic nephropathy, but the mechanism by which high glucose induces apoptosis is not fully understood. Apoptosis of tubular epithelial cells is a major feature of diabetic kidney disease, and hyperglycemia triggers the generation of free radicals and oxidant stress in tubular cells. Hyperglycemia and high glucose in vitro also lead to apoptosis, a form of programmed cell death. High glucose similar to those seen with hyperglycemia in people with diabetes mellitus, lead to accelerated apoptosis, a form of programmed cell death characterized by cell shrinkage, chromatin condensation and DNA fragmentation, in variety of cell types, including renal proximal tubular epithelial cells.

  16. Long Term Culture of the A549 Cancer Cell Line Promotes Multilamellar Body Formation and Differentiation towards an Alveolar Type II Pneumocyte Phenotype

    PubMed Central

    Cooper, James Ross; Abdullatif, Muhammad Bilal; Burnett, Edward C.; Kempsell, Karen E.; Conforti, Franco; Tolley, Howard; Collins, Jane E.; Davies, Donna E.

    2016-01-01

    Pulmonary research requires models that represent the physiology of alveolar epithelium but concerns with reproducibility, consistency and the technical and ethical challenges of using primary or stem cells has resulted in widespread use of continuous cancer or other immortalized cell lines. The A549 ‘alveolar’ cell line has been available for over four decades but there is an inconsistent view as to its suitability as an appropriate model for primary alveolar type II (ATII) cells. Since most work with A549 cells involves short term culture of proliferating cells, we postulated that culture conditions that reduced proliferation of the cancer cells would promote a more differentiated ATII cell phenotype. We examined A549 cell growth in different media over long term culture and then used microarray analysis to investigate temporal regulation of pathways involved in cell cycle and ATII differentiation; we also made comparisons with gene expression in freshly isolated human ATII cells. Analyses indicated that long term culture in Ham’s F12 resulted in substantial modulation of cell cycle genes to result in a quiescent population of cells with significant up-regulation of autophagic, differentiation and lipidogenic pathways. There were also increased numbers of up- and down-regulated genes shared with primary cells suggesting adoption of ATII characteristics and multilamellar body (MLB) development. Subsequent Oil Red-O staining and Transmission Electron Microscopy confirmed MLB expression in the differentiated A549 cells. This work defines a set of conditions for promoting ATII differentiation characteristics in A549 cells that may be advantageous for studies with this cell line. PMID:27792742

  17. Anacardic acid, a histone acetyltransferase inhibitor, modulates LPS-induced IL-8 expression in a human alveolar epithelial cell line A549

    PubMed Central

    Takizawa, Hajime

    2013-01-01

    Objective and design: The histone acetylation processes, which are believed to play a critical role in the regulation of many inflammatory genes, are reversible and regulated by histone acetyltransferases (HATs), which promote acetylation, and histone deacetylases (HDACs), which promote deacetylation. We studied the effects of lipopolysaccharide (LPS) on histone acetylation and its role in the regulation of interleukin (IL)-8 expression.  Material: A human alveolar epithelial cell line A549 was used in vitro. Methods: Histone H4 acetylation at the IL-8 promoter region was assessed by a chromatin immunoprecipitation (ChIP) assay. The expression and production of IL-8 were evaluated by quantitative polymerase chain reaction and specific immunoassay. Effects of a HDAC inhibitor, trichostatin A (TSA), and a HAT inhibitor, anacardic acid, were assessed.  Results: Escherichia coli-derived LPS showed a dose- and time-dependent stimulatory effect on IL-8 protein production and mRNA expression in A549 cells in vitro. LPS showed a significant stimulatory effect on histone H4 acetylation at the IL-8 promoter region by ChIP assay. Pretreatment with TSA showed a dose-dependent stimulatory effect on IL-8 release from A549 cells as compared to LPS alone. Conversely, pretreatment with anacardic acid inhibited IL-8 production and expression in A549 cells.  Conclusion: These data suggest that LPS-mediated proinflammatory responses in the lungs might be modulated via changing chromatin remodeling by HAT inhibition. PMID:24627774

  18. A potential oral anticancer drug candidate, Moringa oleifera leaf extract, induces the apoptosis of human hepatocellular carcinoma cells

    PubMed Central

    JUNG, IL LAE; LEE, JU HYE; KANG, SE CHAN

    2015-01-01

    It has previously been reported that cold water-extracts of Moringa oleifera leaf have anticancer activity against various human cancer cell lines, including non-small cell lung cancer. In the present study, the anticancer activity of M. oleifera leaf extracts was investigated in human hepatocellular carcinoma HepG2 cells. By the analysis of apoptotic signals, including the induction of caspase or poly(ADP-ribose) polymerase cleavage, and the Annexin V and terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling assays, it was demonstrated that M. oleifera leaf extracts induce the apoptosis of HepG2 cells. In the hollow fiber assay, oral administration of the leaf extracts significantly reduced (44–52%) the proliferation of the HepG2 cells and A549 non-small cell lung cancer cells. These results support the potential of soluble extracts of M. oleifera leaf as orally administered therapeutics for the treatment of human liver and lung cancers. PMID:26622717

  19. Apoptosis-Dependent and Apoptosis-Independent Functions Bim in Prostate Cancer Cells

    DTIC Science & Technology

    2004-03-01

    AD_ Award Number: DAMD17-03-1-0146 TITLE: Apoptosis-Dependent and Apoptosis-Independent Functions of Bim in Prostate Cancer Cells PRINCIPAL...FUNDING NUMBERS Apoptosis-Dependent and Apoptosis-Independent Functions of DAMD17-03-1-0146 Bim in Prostate Cancer Cells 6. A UTHORs) Junwei Liu, M.D...extended cell survival have been implicated in prostate cancer (PCa) development and progression. We recently found that Bim , a BH3-only pro

  20. Cytotoxicity and apoptotic activities of alpha-, gamma- and delta-tocotrienol isomers on human cancer cells.

    PubMed

    Lim, Su-Wen; Loh, Hwei-San; Ting, Kang-Nee; Bradshaw, Tracey D; Zeenathul, Nazariah A

    2014-12-06

    Tocotrienols, especially the gamma isomer was discovered to possess cytotoxic effects associated with the induction of apoptosis in numerous cancers. Individual tocotrienol isomers are believed to induce dissimilar apoptotic mechanisms in different cancer types. This study was aimed to compare the cytotoxic potency of alpha-, gamma- and delta-tocotrienols, and to explore their resultant apoptotic mechanisms in human lung adenocarcinoma A549 and glioblastoma U87MG cells which are scarcely researched. The cytotoxic effects of alpha-, gamma- and delta-tocotrienols in both A549 and U87MG cancer cells were first determined at the cell viability and morphological aspects. DNA damage types were then identified by comet assay and flow cytometric study was carried out to support the incidence of apoptosis. The involvements of caspase-8, Bid, Bax and mitochondrial membrane permeability (MMP) in the execution of apoptosis were further expounded. All tocotrienols inhibited the growth of A549 and U87MG cancer cells in a concentration- and time-dependent manner. These treated cancer cells demonstrated some hallmarks of apoptotic morphologies, apoptosis was further confirmed by cell accumulation at the pre-G1 stage. All tocotrienols induced only double strand DNA breaks (DSBs) and no single strand DNA breaks (SSBs) in both treated cancer cells. Activation of caspase-8 leading to increased levels of Bid and Bax as well as cytochrome c release attributed by the disruption of mitochondrial membrane permeability in both A549 and U87MG cells were evident. This study has shown that delta-tocotrienol, in all experimental approaches, possessed a higher efficacy (shorter induction period) and effectiveness (higher induction rate) in the execution of apoptosis in both A549 and U87MG cancer cells as compared to alpha- and gamma-tocotrienols. Tocotrienols in particular the delta isomer can be an alternative chemotherapeutic agent for treating lung and brain cancers.

  1. 6-Shogaol, an active constituent of dietary ginger, induces autophagy by inhibiting the AKT/mTOR pathway in human non-small cell lung cancer A549 cells.

    PubMed

    Hung, Jen-Yu; Hsu, Ya-Ling; Li, Chien-Te; Ko, Ying-Chin; Ni, Wen-Chiu; Huang, Ming-Shyan; Kuo, Po-Lin

    2009-10-28

    This study is the first study to investigate the anticancer effect of 6-shogaol in human non-small cell lung cancer A549 cells. 6-Shogaol inhibited cell proliferation by inducing autophagic cell death, but not, predominantly, apoptosis. Pretreatment of cells with 3-methyladenine (3-MA), an autophagy inhibitor, suppressed 6-shogaol mediated antiproliferation activity, suggesting that induction of autophagy by 6-shogaol is conducive to cell death. We also found that 6-shogaol inhibited survival signaling through the AKT/mTOR signaling pathway by blocking the activation of AKT and downstream targets, including the mammalian target of rapamycin (mTOR), forkhead transcription factors (FKHR) and glycogen synthase kinase-3beta (GSK-3beta). Phosphorylation of both of mTOR's downstream targets, p70 ribosomal protein S6 kinase (p70S6 kinase) and 4E-BP1, was also diminished. Overexpression of AKT by AKT cDNA transfection decreased 6-shogaol mediated autophagic cell death, supporting inhibition of AKT beneficial to autophagy. Moreover, reduction of AKT expression by siRNA potentiated 6-shogaol's effect, also supporting inhibition of AKT beneficial to autophagy. Taken together, these findings suggest that 6-shogaol may be a promising chemopreventive agent against human non-small cell lung cancer.

  2. Middle Infrared Radiation Induces G2/M Cell Cycle Arrest in A549 Lung Cancer Cells

    PubMed Central

    Huang, Hsuan-Cheng; Tsai, Shang-Ru; Juan, Hsueh-Fen; Lee, Si-Chen

    2013-01-01

    There were studies investigating the effects of broadband infrared radiation (IR) on cancer cell, while the influences of middle-infrared radiation (MIR) are still unknown. In this study, a MIR emitter with emission wavelength band in the 3–5 µm region was developed to irradiate A549 lung adenocarcinoma cells. It was found that MIR exposure inhibited cell proliferation and induced morphological changes by altering the cellular distribution of cytoskeletal components. Using quantitative PCR, we found that MIR promoted the expression levels of ATM (ataxia telangiectasia mutated), ATR (ataxia-telangiectasia and Rad3-related and Rad3-related), TP53 (tumor protein p53), p21 (CDKN1A, cyclin-dependent kinase inhibitor 1A) and GADD45 (growth arrest and DNA-damage inducible), but decreased the expression levels of cyclin B coding genes, CCNB1 and CCNB2, as well as CDK1 (Cyclin-dependent kinase 1). The reduction of protein expression levels of CDC25C, cyclin B1 and the phosphorylation of CDK1 at Thr-161 altogether suggest G2/M arrest occurred in A549 cells by MIR. DNA repair foci formation of DNA double-strand breaks (DSB) marker γ-H2AX and sensor 53BP1 was induced by MIR treatment, it implies the MIR induced G2/M cell cycle arrest resulted from DSB. This study illustrates a potential role for the use of MIR in lung cancer therapy by initiating DSB and blocking cell cycle progression. PMID:23335992

  3. 4-Methoxychalcone Enhances Cisplatin-Induced Oxidative Stress and Cytotoxicity by Inhibiting the Nrf2/ARE-Mediated Defense Mechanism in A549 Lung Cancer Cells

    PubMed Central

    Lim, Juhee; Lee, Sung Ho; Cho, Sera; Lee, Ik-Soo; Kang, Bok Yun; Choi, Hyun Jin

    2013-01-01

    Nuclear factor erythroid 2-related factor 2 (Nrf2) is a key transcriptional regulator for the protection of cells against oxidative and xenobiotic stresses. Recent studies have demonstrated that high constitutive expression of Nrf2 is observed in many types of cancer cells showing resistance to anti-cancer drugs, suggesting that the suppression of overexpressed Nrf2 could be an attractive therapeutic strategy to overcome cancer drug resistance. In the present study, we aimed to find small molecule compounds that enhance the sensitivity of tumor cells to cisplatin induced cytotoxicity by suppressing Nrf2-mediated defense mechanism. A549 lung cancer cells were shown to be more resistant to the anti-cancer drug cisplatin than HEK293 cells, with higher Nrf2 signaling activity; constitutively high amounts of Nrf2-downstream target proteins were observed in A549 cells. Among the three chalcone derivatives 4-methoxy-chalcone (4-MC), hesperidin methylchalcone, and neohesperidin dihydrochalcone, 4-MC was found to suppress transcriptional activity of Nrf2 in A549 cells but to activate it in HEK293 cells. 4-MC was also shown to down-regulate expression of Nrf2 and the downstream phase II detoxifying enzyme NQO1 in A549 cells. The PI3K/Akt pathway was found to be involved in the 4-MC-induced inhibition of Nrf2/ARE activity in A549 cells. This inhibition of Nrf2 signaling results in the accelerated generation of reactive oxygen species and exacerbation of cytotoxicity in cisplatin-treated A549 cells. Taken together, these results suggest that the small molecule compound 4-MC could be used to enhance the sensitivity of tumor cells to the therapeutic effect of cisplatin through the regulation of Nrf2/ARE signaling. PMID:24046186

  4. 4-methoxychalcone enhances cisplatin-induced oxidative stress and cytotoxicity by inhibiting the Nrf2/ARE-mediated defense mechanism in A549 lung cancer cells.

    PubMed

    Lim, Juhee; Lee, Sung Ho; Cho, Sera; Lee, Ik-Soo; Kang, Bok Yun; Choi, Hyun Jin

    2013-10-01

    Nuclear factor erythroid 2-related factor 2 (Nrf2) is a key transcriptional regulator for the protection of cells against oxidative and xenobiotic stresses. Recent studies have demonstrated that high constitutive expression of Nrf2 is observed in many types of cancer cells showing resistance to anti-cancer drugs, suggesting that the suppression of overexpressed Nrf2 could be an attractive therapeutic strategy to overcome cancer drug resistance. In the present study, we aimed to find small molecule compounds that enhance the sensitivity of tumor cells to cisplatin induced cytotoxicity by suppressing Nrf2-mediated defense mechanism. A549 lung cancer cells were shown to be more resistant to the anti-cancer drug cisplatin than HEK293 cells, with higher Nrf2 signaling activity; constitutively high amounts of Nrf2-downstream target proteins were observed in A549 cells. Among the three chalcone derivatives 4-methoxy-chalcone (4-MC), hesperidin methylchalcone, and neohesperidin dihydrochalcone, 4-MC was found to suppress transcriptional activity of Nrf2 in A549 cells but to activate it in HEK293 cells. 4-MC was also shown to down-regulate expression of Nrf2 and the downstream phase II detoxifying enzyme NQO1 in A549 cells. The PI3K/Akt pathway was found to be involved in the 4-MC-induced inhibition of Nrf2/ARE activity in A549 cells. This inhibition of Nrf2 signaling results in the accelerated generation of reactive oxygen species and exacerbation of cytotoxicity in cisplatin-treated A549 cells. Taken together, these results suggest that the small molecule compound 4-MC could be used to enhance the sensitivity of tumor cells to the therapeutic effect of cisplatin through the regulation of Nrf2/ARE signaling.

  5. [Combined effects of interferon γ and γ ray irradiation on A549 cells in vitro].

    PubMed

    Xia, Hui; Zhang, Yi-ming; Yu, Chang-hai; Zhang, Wen; Zhang, Bao-shi; Fang, Fang

    2012-02-07

    To define the role of interferon-γ on radiotherapy of lung cancer and explore a new way to clinical treatment. A549 cells were exposed to γ ray with or without IFN-γ co-treatment. MTT assay was performed to evaluate cell viability. Western blot was used to observe the expression of P53 protein. The results showed that co-treatment of IFN-γ decreased the cell viability significantly compared with the γ ray irradiation group (71.4% ± 2.1% vs 44.1% ± 3.1%, n = 7, P < 0.01). In addition, the expression of P53 protein also increased significantly after co-treatment (P < 0.01); Furthermore, the cell cycle was changed obviously in co-treatment group compared with γ ray irradiation group, S phase increased (12.9% vs 20.9%, n = 5, P < 0.05) and also blocked the G2/M phase (28.8% vs 38.9%, n = 5, P < 0.05). The results suggested that γ ray irradiation combined with IFN-γ can increase the efficiency of radiotherapy on A549 cells and there is much broad prospect in the clinical treatment of lung cancer.

  6. Mineral fiber-mediated activation of phosphoinositide-specific phospholipase c in human bronchoalveolar carcinoma-derived alveolar epithelial A549 cells.

    PubMed

    Loreto, Carla; Carnazza, Maria Luisa; Cardile, Venera; Libra, Massimo; Lombardo, Laura; Malaponte, Grazia; Martinez, Giuseppina; Musumeci, Giuseppe; Papa, Veronica; Cocco, Lucio

    2009-02-01

    Given the role of phosphoinositide-specific phospholipase C (PLC) isozymes in the control of cell growth and differentiation we were prompted to analyze the expression of some of these PLC in human bronchoalveolar carcinoma-derived alveolar epithelial A549 cells. The effects of several fluoro-edenite fibers were compared with those of tremolite, a member of the calcic amphibole group of asbestos that originates from Calabria (Italy), and crocidolite, that, due to its high toxicity, is one of the most studied asbestos amphiboles. Our data show an increased expression of both PLC beta1 and PLC gamma1 in A549 cells treated with asbestos-like fibers, hinting at a role of PLC signalling in those cancerous cells.

  7. Encapsulation in lipid-core nanocapsules overcomes lung cancer cell resistance to tretinoin.

    PubMed

    Schultze, Eduarda; Ourique, Aline; Yurgel, Virginia Campello; Begnini, Karine Rech; Thurow, Helena; de Leon, Priscila Marques Moura; Campos, Vinicius Farias; Dellagostin, Odir Antônio; Guterres, Silvia R; Pohlmann, Adriana R; Seixas, Fabiana Kömmling; Beck, Ruy Carlos Ruver; Collares, Tiago

    2014-05-01

    Tretinoin is a retinoid derivative that has an antiproliferative effect on several kinds of tumours. Human lung adenocarcinoma epithelial cell lines (A549) exhibit a profound resistance to the effects of tretinoin. Nanocarriers seem to be a good alternative to overcomecellular resistance to drugs. The aim of this study was to test whether tretinoin-loaded lipid-core nanocapsules exert anantitumor effect on A549 cells. A549 cells were incubated with free tretinoin (TTN), blank nanocapsules (LNC) and tretinoin-loaded lipid-core nanocapsules (TTN-LNC). Data from evaluation of DNA content and Annexin V binding assay by flow cytometry showed that TTN-LNC induced apoptosis and cell cycle arrest at the G1-phase while TTN did not. TTN-LNC showed higher cytotoxic effects than TTN on A549 cells evaluated by MTT and LIVE/DEAD cell viability assay. Gene expression profiling identified up-regulated expression of gene p21 by TTN-LNC, supporting the cell cycle arrest effect. These results showed for the first time that TTN-LNC are able to overcome the resistance of adenocarcinoma cell line A549 to treatment with TTN by inducing apoptosis and cell cycle arrest, providing support for their use in applications in lung cancer therapy. Copyright © 2014 Elsevier B.V. All rights reserved.

  8. Lycium europaeum fruit extract: antiproliferative activity on A549 human lung carcinoma cells and PC12 rat adrenal medulla cancer cells and assessment of its cytotoxicity on cerebellum granule cells.

    PubMed

    Ghali, Wafa; Vaudry, David; Jouenne, Thierry; Marzouki, Mohamed Nejib

    2015-01-01

    Cancer is a major worldwide health problem and one of the leading causes of death either in developed or developing countries. Plant extracts and derivatives have always been used for various disease treatments and many anticancer agents issued from plants and vegetables are clinically recognized and used all over the world. Lycium europaeum (Solanaceae) also called "wolfberry" was known since ancient times in the Mediterranean area as a medicinal plant and used in several traditional remedies. The Lycium species capacity of reducing the incidence of cancer and also of halting or reserving the growth of cancer was reported by traditional healers. In this study, the antiproliferative capacity, protective properties, and antioxidant activity of the hydro-alcoholic fruit extract of Lycium europaeum were investigated. Results showed that Lycium extract exhibits the ability to reduce cancer cell viability, inhibits proliferation, and induces apoptosis in A549 human lung cancer cells and PC12 rat adrenal medulla cancer cells, in a concentration- and time-dependent manner. Cytotoxic effect on normal rat cerebellum granule cells was assessed to be nonsignificant. Results also showed that Lycium fruit extract protected lipids, proteins, and DNA against oxidative stress damages induced by H2O2 via scavenging reactive oxygen species.

  9. Increased levels of the long noncoding RNA, HOXA-AS3, promote proliferation of A549 cells.

    PubMed

    Zhang, Hongyue; Liu, Ying; Yan, Lixin; Zhang, Min; Yu, Xiufeng; Du, Wei; Wang, Siqi; Li, Qiaozhi; Chen, He; Zhang, Yafeng; Sun, Hanliang; Tang, Zhidong; Zhu, Daling

    2018-06-13

    Many long noncoding RNAs (lncRNAs) have been identified as powerful regulators of lung adenocarcinoma (LAD). However, the role of HOXA-AS3, a novel lncRNA, in LAD is largely unknown. In this study, we showed that HOXA-AS3 was significantly upregulated in LAD tissues and A549 cells. After knockdown of HOXA-AS3, cell proliferation, migration, and invasion were inhibited. Xenografts derived from A549 cells transfected with shRNA/HOXA-AS3 had significantly lower tumor weights and smaller tumor volumes. We also demonstrated that HOXA-AS3 increased HOXA6 mRNA stability by forming an RNA duplex. In addition, HOXA6 promoted cell proliferation, migration, and invasion in vitro. Using a RNA pull-down assay, we found that HOXA-AS3 bonded with NF110, which regulated the cell localization of HOXA-AS3. Moreover, histone acetylation was involved in upregulation of HOXA-AS3. These results demonstrate that HOXA-AS3 was activated in LAD and supported cancer cell progression. Therefore, inhibition of HOXA-AS3 could be an effective targeted therapy for patients with LAD.

  10. Evaluation of whole cigarette smoke induced oxidative stress in A549 and BEAS-2B cells.

    PubMed

    Zhang, Shimin; Li, Xiang; Xie, Fuwei; Liu, Kejian; Liu, Huimin; Xie, Jianping

    2017-09-01

    Cigarette smoke is a complex and oxidative aerosol. Previous researches on the hazards of cigarette smoke mainly focused on the adverse bioeffects induced by its condensates or gas vapor phase, which ignored the dynamic processes of smoking and the cigarette smoke aging. To overcome these disadvantages, we performed air-liquid interface exposure of whole smoke, which used native and unmodified smoke and ensured the exposure similar to physiological inhalation. Our results indicated that whole cigarette smoke induced lung epithelial cells (A549) and bronchial epithelial cells (BEAS-2B) damages in cytotoxicity assays (methyl thiazoly tetrazolium and neutral red uptake assays). In addition, A549 and BEAS-2B cells showed oxidative damages in whole smoke exposure, with concentration change of several biomarkers (reduced and oxidized glutathione, malondialdehyde, 4-hydroxyhydroxy-2-nonenal, extracellular superoxide dismutase, and 8-hydroxyl deoxyguanosine). These results indicate that whole smoke-induced oxidative stress occurs in two different kinds of cells at air-liquid interface. Copyright © 2017 Elsevier B.V. All rights reserved.

  11. EGCG reverses human neutrophil elastase-induced migration in A549 cells by directly binding to HNE and by regulating α1-AT

    PubMed Central

    Xiaokaiti, Yilixiati; Wu, Haoming; Chen, Ya; Yang, Haopeng; Duan, Jianhui; Li, Xin; Pan, Yan; Tie, Lu; Zhang, Liangren; Li, Xuejun

    2015-01-01

    Lung carcinogenesis is a complex process that occurs in unregulated inflammatory environment. EGCG has been extensively investigated as a multi-targeting anti-tumor and anti-inflammatory compound. In this study, we demonstrated a novel mechanism by which EGCG reverses the neutrophil elastase-induced migration of A549 cells. We found that neutrophil elastase directly triggered human adenocarcinoma A549 cell migration and that EGCG suppressed the elevation of tumor cell migration induced by neutrophil elastase. We observed that EGCG directly binds to neutrophil elastase and inhibits its enzymatic activity based on the CDOCKER algorithm, MD stimulation by GROMACS, SPR assay and elastase enzymatic activity assay. As the natural inhibitor of neutrophil elastase, α1-antitrypsin is synthesized in tumor cells. We further demonstrated that the expression of α1-antitrypsin was up-regulated after EGCG treatment in neutrophil elastase-treated A549 cells. We preliminarily discovered that the EGCG-mediated induction of α1-antitrypsin expression might be correlated with the regulatory effect of EGCG on the PI3K/Akt pathway. Overall, our results suggest that EGCG ameliorates the neutrophil elastase-induced migration of A549 cells. The mechanism underlying this effect may include two processes: EGCG directly binds to neutrophil elastase and inhibits its enzymatic activity; EGCG enhances the expression of α1-antitrypsin by regulating the PI3K/AKT pathway. PMID:26177797

  12. EGCG reverses human neutrophil elastase-induced migration in A549 cells by directly binding to HNE and by regulating α1-AT

    NASA Astrophysics Data System (ADS)

    Xiaokaiti, Yilixiati; Wu, Haoming; Chen, Ya; Yang, Haopeng; Duan, Jianhui; Li, Xin; Pan, Yan; Tie, Lu; Zhang, Liangren; Li, Xuejun

    2015-07-01

    Lung carcinogenesis is a complex process that occurs in unregulated inflammatory environment. EGCG has been extensively investigated as a multi-targeting anti-tumor and anti-inflammatory compound. In this study, we demonstrated a novel mechanism by which EGCG reverses the neutrophil elastase-induced migration of A549 cells. We found that neutrophil elastase directly triggered human adenocarcinoma A549 cell migration and that EGCG suppressed the elevation of tumor cell migration induced by neutrophil elastase. We observed that EGCG directly binds to neutrophil elastase and inhibits its enzymatic activity based on the CDOCKER algorithm, MD stimulation by GROMACS, SPR assay and elastase enzymatic activity assay. As the natural inhibitor of neutrophil elastase, α1-antitrypsin is synthesized in tumor cells. We further demonstrated that the expression of α1-antitrypsin was up-regulated after EGCG treatment in neutrophil elastase-treated A549 cells. We preliminarily discovered that the EGCG-mediated induction of α1-antitrypsin expression might be correlated with the regulatory effect of EGCG on the PI3K/Akt pathway. Overall, our results suggest that EGCG ameliorates the neutrophil elastase-induced migration of A549 cells. The mechanism underlying this effect may include two processes: EGCG directly binds to neutrophil elastase and inhibits its enzymatic activity; EGCG enhances the expression of α1-antitrypsin by regulating the PI3K/AKT pathway.

  13. EGCG reverses human neutrophil elastase-induced migration in A549 cells by directly binding to HNE and by regulating α1-AT.

    PubMed

    Xiaokaiti, Yilixiati; Wu, Haoming; Chen, Ya; Yang, Haopeng; Duan, Jianhui; Li, Xin; Pan, Yan; Tie, Lu; Zhang, Liangren; Li, Xuejun

    2015-07-16

    Lung carcinogenesis is a complex process that occurs in unregulated inflammatory environment. EGCG has been extensively investigated as a multi-targeting anti-tumor and anti-inflammatory compound. In this study, we demonstrated a novel mechanism by which EGCG reverses the neutrophil elastase-induced migration of A549 cells. We found that neutrophil elastase directly triggered human adenocarcinoma A549 cell migration and that EGCG suppressed the elevation of tumor cell migration induced by neutrophil elastase. We observed that EGCG directly binds to neutrophil elastase and inhibits its enzymatic activity based on the CDOCKER algorithm, MD stimulation by GROMACS, SPR assay and elastase enzymatic activity assay. As the natural inhibitor of neutrophil elastase, α1-antitrypsin is synthesized in tumor cells. We further demonstrated that the expression of α1-antitrypsin was up-regulated after EGCG treatment in neutrophil elastase-treated A549 cells. We preliminarily discovered that the EGCG-mediated induction of α1-antitrypsin expression might be correlated with the regulatory effect of EGCG on the PI3K/Akt pathway. Overall, our results suggest that EGCG ameliorates the neutrophil elastase-induced migration of A549 cells. The mechanism underlying this effect may include two processes: EGCG directly binds to neutrophil elastase and inhibits its enzymatic activity; EGCG enhances the expression of α1-antitrypsin by regulating the PI3K/AKT pathway.

  14. High-Throughput Quantitative Proteomic Analysis of Dengue Virus Type 2 Infected A549 Cells

    PubMed Central

    Chiu, Han-Chen; Hannemann, Holger; Heesom, Kate J.; Matthews, David A.; Davidson, Andrew D.

    2014-01-01

    Disease caused by dengue virus is a global health concern with up to 390 million individuals infected annually worldwide. There are no vaccines or antiviral compounds available to either prevent or treat dengue disease which may be fatal. To increase our understanding of the interaction of dengue virus with the host cell, we analyzed changes in the proteome of human A549 cells in response to dengue virus type 2 infection using stable isotope labelling in cell culture (SILAC) in combination with high-throughput mass spectrometry (MS). Mock and infected A549 cells were fractionated into nuclear and cytoplasmic extracts before analysis to identify proteins that redistribute between cellular compartments during infection and reduce the complexity of the analysis. We identified and quantified 3098 and 2115 proteins in the cytoplasmic and nuclear fractions respectively. Proteins that showed a significant alteration in amount during infection were examined using gene enrichment, pathway and network analysis tools. The analyses revealed that dengue virus infection modulated the amounts of proteins involved in the interferon and unfolded protein responses, lipid metabolism and the cell cycle. The SILAC-MS results were validated for a select number of proteins over a time course of infection by Western blotting and immunofluorescence microscopy. Our study demonstrates for the first time the power of SILAC-MS for identifying and quantifying novel changes in cellular protein amounts in response to dengue virus infection. PMID:24671231

  15. PET imaging of apoptosis in tumor-bearing mice and rabbits after paclitaxel treatment with 18F-Labeled recombinant human His10-annexin V

    PubMed Central

    Qin, Haidong; Zhang, Ming-Rong; Xie, Lin; Hou, Yanjie; Hua, Zichun; Hu, Minjin; Wang, Zizheng; Wang, Feng

    2015-01-01

    Monitoring response to chemo- or radiotherapy is of great importance in clinical practice. Apoptosis imaging serves as a very useful tool for the early evaluation of tumor response. The goal of this study was PET imaging of apoptosis with 18F-labeled recombinant human annexin V linked with 10 histidine tag (18F-rh-His10-annexin V) in nude mice bearing an A549 tumor and rabbits bearing a VX2 lung cancer after paclitaxel therapy. 18F-rh-His10-annexin V was prepared by conjugation of rh-His10-annexin V with N-succinimidyl 4-[18F]fluorobenzoate. Biodistribution was determined in mice by the dissection method and small-animal PET. Single-dose paclitaxel (175 mg/m2) was used to induce apoptosis in A549 and VX2 tumor models. 18F-rh-His10-annexin V was injected into A549 mice and VX rabbits to acquire dynamic and static PET images 72 h after paclitaxel treatment. The uptake of 18F-rh-His10-annexin V in apoptotic cells 4 h after induction was 6.45±0.52 fold higher than that in non-induced cells. High focal uptake of 18F-rh-His10-annexin V was visualized in A549 (SUVmax: 0.35±0.13) and VX2 (0.41±0.23) tumor models after paclitaxel treatment, whereas lower uptake was found in the corresponding tumors before treatment (A549 SUVmax: 0.04±0.02; VX2: 0.009±0.002). The apoptotic index was 75.61±11.56% in the treated VX2 cancer, much higher than that in the untreated VX2 (8.03±2.81%). This study demonstrated the feasibility of 18F-rh-His10-annexin V for the detection of apoptosis after chemotherapy in A549 and VX2 tumor models. PMID:25625024

  16. In- and ex-vivo molecular imaging of apoptosis to assess sensitivity of non-small cell lung cancer to EGFR inhibitors using probe-based confocal laser endomicroscopy.

    PubMed

    Guisier, Florian; Bohn, Pierre; Patout, Maxime; Piton, Nicolas; Farah, Insaf; Vera, Pierre; Thiberville, Luc; Salaün, Mathieu

    2017-01-01

    Prediction of treatment outcome of non-small cell lung cancer (NSCLC) with EGFR inhibitors on the basis of the genetic analysis of the tumor can be incorrect in case of rare or complex mutations, bypass molecular activation pathways, or pharmacodynamic variations. The aim of this study was to develop an ex vivo and in vivo real-time quantitative imaging test for EGFR inhibitors sensitivity assessment. Erlotinib resistant (A549, H460, H1975), insensitive (H1650) and hypersensitive (HCC827) cell lines were injected subcutaneously in Nude mice. Tumor xenografts from mice treated with Erlotinib were imaged ex vivo and in vivo using probe-based confocal laser endomicroscopy (pCLE) and NucView 488 Caspase 3 substrate, a fluorescent probe specific for the activated caspase 3. Assessment of apoptosis at 24h post treatment, both ex vivo in explanted tumor xenografts and in vivo, showed a significant difference between resistant cell lines (A549, H460 and H1975) and insensitive (H1650) or hypersensitive (HCC827) ones (p<0.05 for ex vivo imaging, p≤0.02 for in vivo imaging). There was also a significant difference between insensitive and hypersensitive cell lines, both ex vivo (p<0.05) and in vivo (p = 0.01). Real-time in vivo and ex vivo assessment of apoptosis using pCLE differentiates resistant from sensitive NSCLC xenografts to Erlotinib.

  17. Compound K induced apoptosis via endoplasmic reticulum Ca2+ release through ryanodine receptor in human lung cancer cells.

    PubMed

    Shin, Dong-Hyun; Leem, Dong-Gyu; Shin, Ji-Sun; Kim, Joo-Il; Kim, Kyung-Tack; Choi, Sang Yoon; Lee, Myung-Hee; Choi, Jung-Hye; Lee, Kyung-Tae

    2018-04-01

    Extended endoplasmic reticulum (ER) stress may initiate apoptotic pathways in cancer cells, and ER stress has been reported to possibly increase tumor death in cancer therapy. We previously reported that caspase-8 played an important role in compound K-induced apoptosis via activation of caspase-3 directly or indirectly through Bid cleavage, cytochrome c release, and caspase-9 activation in HL-60 human leukemia cells. The mechanisms leading to apoptosis in A549 and SK-MES-1 human lung cancer cells and the role of ER stress have not yet been understood. The apoptotic effects of compound K were analyzed using flow cytometry, and the changes in protein levels were determined using Western blot analysis. The intracellular calcium levels were monitored by staining with Fura-2/AM and Fluo-3/AM. Compound K-induced ER stress was confirmed through increased phosphorylation of eIF2α and protein levels of GRP78/BiP, XBP-1S, and IRE1α in human lung cancer cells. Moreover, compound-K led to the accumulation of intracellular calcium and an increase in m-calpain activities that were both significantly inhibited by pretreatment either with BAPTA-AM (an intracellular Ca 2+ chelator) or dantrolene (an RyR channel antagonist). These results were correlated with the outcome that compound K induced ER stress-related apoptosis through caspase-12, as z-ATAD-fmk (a specific inhibitor of caspase-12) partially ameliorated this effect. Interestingly, 4-PBA (ER stress inhibitor) dramatically improved the compound K-induced apoptosis. Cell survival and intracellular Ca 2+ homeostasis during ER stress in human lung cancer cells are important factors in the induction of the compound K-induced apoptotic pathway.

  18. Bromelain-induced apoptosis in GI-101A breast cancer cells.

    PubMed

    Dhandayuthapani, Sivanesan; Perez, Honey Diaz; Paroulek, Alexandra; Chinnakkannu, Panneerselvam; Kandalam, Umadevi; Jaffe, Mark; Rathinavelu, Appu

    2012-04-01

    Bromelain is a proteolytic enzyme extracted from the stems and the immature fruits of pineapple that was found to be antitumorigenic in different in vitro models. Bromelain has been reported to promote apoptosis, particularly in breast cancer cells, with the up-regulation of c-Jun N-terminal kinase and p38 kinase. Our study was designed to determine if bromelain could induce apoptosis in GI-101A breast cancer cells. GI-101A cells were treated with increasing concentrations of bromelain for 24 hours. The effect of bromelain for inducing cell death via activation of the apoptosis mechanism in GI-101A cells was further determined by using caspase-9 and caspase-3 assays along with the M30-Apoptosense assay to measure cytokeratin 18 (CK18) levels in the cytoplasm of the cultured cancer cells. A dose-dependent increase in the activities of caspase-9 and caspase-3 coinciding with elevation of CK18 levels was found in bromelain-treated cells compared with control cells. Furthermore, the apoptosis induction by bromelain was confirmed by DNA fragmentation analysis and 4,6'-diamino-2-phenylindole dihydrochloride fluorescence staining of the nucleus. Our results indicate an increase in apoptosis-related cell death in breast cancer cells with increasing concentrations of bromelain.

  19. Neferine augments therapeutic efficacy of cisplatin through ROS- mediated non-canonical autophagy in human lung adenocarcinoma (A549 cells).

    PubMed

    Kalai Selvi, Sivalingam; Vinoth, Amirthalingam; Varadharajan, Thiyagarajan; Weng, Ching Feng; Vijaya Padma, Viswanadha

    2017-05-01

    Combination of dietary components with chemotherapy drugs is an emerging new strategy for cancer therapy to increase antitumor responses. Neferine, major bisbenzylisoquinoline alkaloid isolated from the seed embryo of Nelumbo nucifera (Lotus). In the present study, we investigated the efficacy of the combinatorial regimen of neferine and cisplatin compared to cisplatin high dose in human lung adenocarcinoma (A549) cells. Co-treatment with neferine enhanced cisplatin-induced autophagy in A549 cells was accompanied by Acidic vesicular accumulation (AVO), enhanced generation of reactive oxygen species (ROS) and depletion of intracellular glutathione (GSH), down regulation of PI3K/AKT/mTOR pathway, conversion of LC3B-I to LC3B-II. This enhanced autophagy developed via a non-canonical mechanism that did not require Beclin-1, PI3KCIII. In conclusion, these results suggest that neferine enhances cisplatin -induced autophagic cancer cell death through downregulation of PI3K/Akt/mTOR signaling pro-survival pathway and ROS- mediated Beclin-1 and PI3K CIII independent autophagy in human lung adenocarcinoma (A549 cells). Copyright © 2017 Elsevier Ltd. All rights reserved.

  20. ROS-dependent Atg4 upregulation mediated autophagy plays an important role in Cd-induced proliferation and invasion in A549 cells.

    PubMed

    Lv, Wei; Sui, Linlin; Yan, Xiaona; Xie, Huaying; Jiang, Liping; Geng, Chengyan; Li, Qiujuan; Yao, Xiaofeng; Kong, Ying; Cao, Jun

    2018-01-05

    Cadmium (Cd) is a toxic heavy metal that is widely used in industry and agriculture. In this study the role of autophagy in Cd-induced proliferation, migration and invasion was investigated in A549 cells. Exposure to Cd (2 μM) significantly increased reactive oxygen species (ROS) production, induced autophagy and enhanced cell growth, migration and invasion in A549 cells. Western blot analysis showed that the expression of autophagy-related proteins, LC3-II, Beclin-1 and Atg4 and invasion-related protein MMP-9 were upregulated in Cd-treated cells. N-acetyl cysteine (NAC) markedly prevented Cd-induced proliferation of A549 cells and the increasing protein level of LC3-II and Atg4. Blocking Atg4 expression by siRNA strongly reduced Beclin-1 and LC3-II protein expression and the number of autophagosome positive cells induced by Cd. Furthermore, Atg4 siRNA increased the number of cells at G0/G1 phase, reduced the number of S and G2/M phase cells, and inhibited Cd-induced cell growth significantly compared with that of Cd-treated Control siRNA cells. 3-MA pretreatment increased the percentage of G0/G1 phase cells, decreased S phase and G2/M phase percentage, and inhibited Cd-induced cell growth remarkably compared with that of only Cd-treated cells. Knocking down Atg4 reduced the number of cells that migrated and invaded through the Matrigel matrix significantly and led to a significant decrease of MMP-9 expression. In addition, in lung tissues of Cd-treated BALB/c mice, the increased expression of LC3-II, Beclin-1 and Atg4 were observed. Taken together, our results demonstrated that ROS-dependent Atg4-mediated autophagy plays an important role in Cd-induced cell growth, migration and invasion in A549 cells. Copyright © 2017 Elsevier B.V. All rights reserved.

  1. Copper doping enhanced the oxidative stress-mediated cytotoxicity of TiO2 nanoparticles in A549 cells.

    PubMed

    Ahmad, J; Siddiqui, M A; Akhtar, M J; Alhadlaq, H A; Alshamsan, A; Khan, S T; Wahab, R; Al-Khedhairy, A A; Al-Salim, A; Musarrat, J; Saquib, Q; Fareed, M; Ahamed, M

    2018-05-01

    Physicochemical properties of titanium dioxide nanoparticles (TiO 2 NPs) can be tuned by doping with metals or nonmetals. Copper (Cu) doping improved the photocatalytic behavior of TiO 2 NPs that can be applied in various fields such as environmental remediation and nanomedicine. However, interaction of Cu-doped TiO 2 NPs with human cells is scarce. This study was designed to explore the role of Cu doping in cytotoxic response of TiO 2 NPs in human lung epithelial (A549) cells. Characterization data demonstrated the presence of both TiO 2 and Cu in Cu-doped TiO 2 NPs with high-quality lattice fringes without any distortion. The size of Cu-doped TiO 2 NPs (24 nm) was lower than pure TiO 2 NPs (30 nm). Biological results showed that both pure and Cu-doped TiO 2 NPs induced cytotoxicity and oxidative stress in a dose-dependent manner. Low mitochondrial membrane potential and higher caspase-3 enzyme (apoptotic markers) activity were also observed in A549 cells exposed to pure and Cu-doped TiO 2 NPs. We further observed that cytotoxicity caused by Cu-doped TiO 2 NPs was higher than pure TiO 2 NPs. Moreover, antioxidant N-acetyl cysteine effectively prevented the reactive oxygen species generation, glutathione depletion, and cell viability reduction caused by Cu-doped TiO 2 NPs. This is the first report showing that Cu-doped TiO 2 NPs induced cytotoxicity and oxidative stress in A549 cells. This study warranted further research to explore the role of Cu doping in toxicity mechanisms of TiO 2 NPs.

  2. Synergistic Antiproliferative Effects of a New Cucurbitacin B Derivative and Chemotherapy Drugs on Lung Cancer Cell Line A549.

    PubMed

    Marostica, Lucas Lourenço; Silva, Izabella Thaís; Kratz, Jadel Müller; Persich, Lara; Geller, Fabiana Cristina; Lang, Karen Luise; Caro, Miguel Soriano Balparda; Durán, Fernando Javier; Schenkel, Eloir Paulo; Simões, Cláudia Maria Oliveira

    2015-10-19

    Nonsmall cell lung cancer (NSCLC) represents an important cause of mortality worldwide due to its aggressiveness and growing resistance to currently available therapy. Cucurbitacins have emerged as novel potential anticancer agents showing strong antiproliferative effects and can be promising candidates for combined treatments with clinically used anticancer agents. This study investigates the synergistic antiproliferative effects of a new semisynthetic derivative of cucurbitacin B (DACE) with three chemotherapy drugs: cisplatin (CIS), irinotecan (IRI), and paclitaxel (PAC) on A549 cells. The most effective combinations were selected for studies of the mechanism of action. Using an in silico tool, DACE seems to act by a different mechanism of action when compared with that of different classes of drugs already used in clinical settings. DACE also showed potent synergic effects with drugs, and the most potent combinations induced G2/M cell cycle arrest by modulating survivin and p53 expression, disruption of F-actin cytoskeleton, and cell death by apoptosis. These treatments completely inhibited the clonogenic potential and did not reduce the proliferation of nontumoral lung cells (MRC-5). DACE also showed relevant antimigratory and anti-invasive effects, and combined treatments modulated cell migration signaling pathways evolved with metastasis progression. The effects of DACE associated with drugs was potentiated by the oxidant agent l-buthionine-sulfoximine (BSO), and attenuated by N-acetilcysteine (NAC), an antioxidant agent. The antiproliferative effects induced by combined treatments were attenuated by a pan-caspase inhibitor, indicating that the effects of these treatments are dependent on caspase activity. Our data highlight the therapeutic potential of DACE used in combination with known chemotherapy drugs and offer important insights for the development of more effective and selective therapies against lung cancer.

  3. X-irradiation of human bronchial cancer cells causes the bystander effects in normal bronchial cells in vitro.

    PubMed

    Konopacka, M; Rogoliński, J

    2010-01-01

    Using X radiation commonly used in radiotherapy of cancers we investigated bystander interactions between human cells: irradiated A549 bronchial carcinoma human cells and non irradiated BEAS-2B normal bronchial epithelial cells. Non irradiated cells were incubated in medium transferred from irradiated A549 cells (ICM-irradiation conditioned medium) for 48h and next the chromosomal damage and apoptosis were estimated. Conditioned medium collected from irradiated cancer cells induced in non irradiated cells of the same line as well as in BEAS-2B normal cells genetic changes such as micronuclei, chromatid and chromosomal breaks and condensation of chromatin characteristic for processes of apoptosis. Addition of only 1% of conditioned medium to fresh medium was sufficient to induction of bystander response to normal bronchial cells. The presented results in this study could have implications for human radiation risk and in evaluating the secondary effects of radiotherapy.

  4. SIRT6 knockout cells resist apoptosis initiation but not progression: a computational method to evaluate the progression of apoptosis.

    PubMed

    Domanskyi, Sergii; Nicholatos, Justin W; Schilling, Joshua E; Privman, Vladimir; Libert, Sergiy

    2017-11-01

    Apoptosis is essential for numerous processes, such as development, resistance to infections, and suppression of tumorigenesis. Here, we investigate the influence of the nutrient sensing and longevity-assuring enzyme SIRT6 on the dynamics of apoptosis triggered by serum starvation. Specifically, we characterize the progression of apoptosis in wild type and SIRT6 deficient mouse embryonic fibroblasts using time-lapse flow cytometry and computational modelling based on rate-equations and cell distribution analysis. We find that SIRT6 deficient cells resist apoptosis by delaying its initiation. Interestingly, once apoptosis is initiated, the rate of its progression is higher in SIRT6 null cells compared to identically cultured wild type cells. However, SIRT6 null cells succumb to apoptosis more slowly, not only in response to nutrient deprivation but also in response to other stresses. Our data suggest that SIRT6 plays a role in several distinct steps of apoptosis. Overall, we demonstrate the utility of our computational model to describe stages of apoptosis progression and the integrity of the cellular membrane. Such measurements will be useful in a broad range of biological applications.

  5. Anti-invasive effect of Cyclamen pseudibericum extract on A549 non-small cell lung carcinoma cells via inhibition of ZEB1 mediated by miR-200c.

    PubMed

    Karagur, Ege Riza; Ozay, Cennet; Mammadov, Ramazan; Akca, Hakan

    2018-06-01

    Scientists are increasingly focusing attention on natural products of plant origin for use as agents in cancer protection and treatment. Cyclamen L. tuber extracts contain saponin glycosides that have been shown to have anti-cancer and other biological activities. The epithelial-to-mesenchymal transition (EMT) is thought to enhance malignant tumour progress. The transcriptional repressor zinc-finger E-box binding homeobox 1 (ZEB1) is an important inducer of EMT in different human tumours and has recently been shown to boost invasion by tumour cells. In this study, we investigated the effects of endemic Cyclamen pseudibericum (CP) saponin-rich tuber extract on the capacity of non-small cell lung cancer line A549 cells to proliferate, invade and migrate and also examined the expression levels of several invasion-migration-related microRNAs (miRNAs) to identify those which directly targeted ZEB1. The cytotoxicity effect of the CP extract on the A549 cancer cells was determined by the luminometric method. The half-minimal (50%) inhibitory concentration dose in the A549 cells was determined to be 41.64 ± 2.35 µg/mL. Using the Matrigel invasion chamber system and the wound healing assay we observed that the CP extract suppressed the invasion and migration capacity of A549 cells, respectively. The expression of miRNAs in A549 cells was evaluated by real-time PCR. Our data showed that overexpression of miRNA miR-200c hindered the EMT by increasing the expression of E-cadherin and decreasing the expression of both N-cadherin and vimentin through the direct targeting of ZEB1. These findings suggest that the saponin-rich tuber extract of CP may have considerable anti-cancer properties in lung cancer. Further studies are required to examine in detail the molecular-based mechanism involved in the EMT process of the extract along with isolation and identification of active saponin components.

  6. Effects of 2-methoxyestradiol on apoptosis and HIF-1α and HIF-2α expression in lung cancer cells under normoxia and hypoxia.

    PubMed

    Aquino-Gálvez, Arnoldo; González-Ávila, Georgina; Delgado-Tello, Javier; Castillejos-López, Manuel; Mendoza-Milla, Criselda; Zúñiga, Joaquín; Checa, Marco; Maldonado-Martínez, Héctor Aquiles; Trinidad-López, Axel; Cisneros, José; Torres-Espíndola, Luz María; Hernández-Jiménez, Claudia; Sommer, Bettina; Cabello-Gutiérrez, Carlos; Gutiérrez-González, Luis H

    2016-01-01

    Hypoxic tumor cells are known to be more resistant to conventional chemotherapy and radiation than normoxic cells. However, the effects of 2-methoxyestradiol (2-ME), an anti-angiogenic, antiproliferative and pro-apoptotic drug, on hypoxic lung cancer cells are unknown. The aim of the present study was to compare the effects of 2-ME on cell growth, apoptosis, hypoxia-inducible factor 1α (HIF-1α) and HIF-2α gene and protein expression in A549 cells under normoxic and hypoxic conditions. To establish the optimal 2-ME concentration with which to carry out the apoptosis assay and to examine mRNA and protein expression of HIFs, cell growth analysis was carried out through N-hexa-methylpararosaniline staining assays in A549 cell cultures treated with one of five different 2-ME concentrations at different times under normoxic or hypoxic growth conditions. The 2-ME concentration of 10 mM at 72 h was selected to perform all further experiments. Apoptotic cells were analyzed by flow cytometry. Western blotting was used to determine HIF-1α and HIF-2α protein expression in total cell extracts. Cellular localization of HIF-1α and HIF-2α was assessed by immunocytochemistry. HIF-1α and HIF-2α gene expression was determined by real-time PCR. A significant increase in the percentage of apoptosis was observed when cells were treated with 2-ME under a normoxic but not under hypoxic conditions (p=0.006). HIF-1α and HIF-2α protein expression levels were significantly decreased in cells cultured under hypoxic conditions and treated with 2-ME (p<0.001). Furthermore, 2-ME decreased the HIF-1α and HIF-2α nuclear staining in cells cultured under hypoxia. The HIF-1α and HIF-2α mRNA levels were significantly lower when cells were exposed to 2-ME under normoxia and hypoxia. Our results suggest that 2-ME could have beneficial results when used with conventional chemotherapy in an attempt to lower the invasive and metastatic processes during cancer development due to its effects on

  7. Paclitaxel and the dietary flavonoid fisetin: a synergistic combination that induces mitotic catastrophe and autophagic cell death in A549 non-small cell lung cancer cells.

    PubMed

    Klimaszewska-Wisniewska, Anna; Halas-Wisniewska, Marta; Tadrowski, Tadeusz; Gagat, Maciej; Grzanka, Dariusz; Grzanka, Alina

    2016-01-01

    The use of the dietary polyphenols as chemosensitizing agents to enhance the efficacy of conventional cytostatic drugs has recently gained the attention of scientists and clinicians as a plausible approach for overcoming the limitations of chemotherapy (e.g. drug resistance and cytotoxicity). The aim of this study was to investigate whether a naturally occurring diet-based flavonoid, fisetin, at physiologically attainable concentrations, could act synergistically with clinically achievable doses of paclitaxel to produce growth inhibitory and/or pro-death effects on A549 non-small cell lung cancer cells, and if it does, what mechanisms might be involved. The drug-drug interactions were analyzed based on the combination index method of Chou and Talalay and the data from MTT assays. To provide some insights into the mechanism underlying the synergistic action of fisetin and paclitaxel, selected morphological, biochemical and molecular parameters were examined, including the morphology of cell nuclei and mitotic spindles, the pattern of LC3-II immunostaining, the formation of autophagic vacuoles at the electron and fluorescence microscopic level, the disruption of cell membrane asymmetry/integrity, cell cycle progression and the expression level of LC3-II, Bax, Bcl-2 and caspase-3 mRNA. Here, we reported the first experimental evidence for the existence of synergism between fisetin and paclitaxel in the in vitro model of non-small cell lung cancer. This synergism was, at least partially, ascribed to the induction of mitotic catastrophe. The switch from the cytoprotective autophagy to the autophagic cell death was also implicated in the mechanism of the synergistic action of fisetin and paclitaxel in the A549 cells. In addition, we revealed that the synergism between fisetin and paclitaxel was cell line-specific as well as that fisetin synergizes with arsenic trioxide, but not with mitoxantrone and methotrexate in the A549 cells. Our results provide rationale for

  8. Pirfenidone inhibits TGF-β1-induced over-expression of collagen type I and heat shock protein 47 in A549 cells

    PubMed Central

    2012-01-01

    Background Pirfenidone is a novel anti-fibrotic and anti-inflammatory agent that inhibits the progression of fibrosis in animal models and in patients with idiopathic pulmonary fibrosis (IPF). We previously showed that pirfenidone inhibits the over-expression of collagen type I and of heat shock protein (HSP) 47, a collagen-specific molecular chaperone, in human lung fibroblasts stimulated with transforming growth factor (TGF)-β1 in vitro. The increased numbers of HSP47-positive type II pneumocytes as well as fibroblasts were also diminished by pirfenidone in an animal model of pulmonary fibrosis induced by bleomycin. The present study evaluates the effects of pirfenidone on collagen type I and HSP47 expression in the human alveolar epithelial cell line, A549 cells in vitro. Methods The expression of collagen type I, HSP47 and E-cadherin mRNAs in A549 cells stimulated with TGF-β1 was evaluated by Northern blotting or real-time PCR. The expression of collagen type I, HSP47 and fibronectin proteins was assessed by immunocytochemical staining. Results TGF-β1 stimulated collagen type I and HSP47 mRNA and protein expression in A549 cells, and pirfenidone significantly inhibited this process. Pirfenidone also inhibited over-expression of the fibroblast phenotypic marker fibronectin in A549 cells induced by TGF-β1. Conclusion We concluded that the anti-fibrotic effects of pirfenidone might be mediated not only through the direct inhibition of collagen type I expression but also through the inhibition of HSP47 expression in alveolar epithelial cells, which results in reduced collagen synthesis in lung fibrosis. Furthermore, pirfenidone might partially inhibit the epithelial-mesenchymal transition. PMID:22694981

  9. Curcumin sensitizes human lung cancer cells to apoptosis and metastasis synergistically combined with carboplatin

    PubMed Central

    Kang, Ji Ho; Kang, Hye Seon; Kim, In Kyoung; Lee, Hwa Young; Ha, Jick Hwan; Yeo, Chang Dong; Kang, Hyun Hui; Moon, Hwa Sik

    2015-01-01

    Although carboplatin is one of the standard chemotherapeutic agents for non-small cell lung cancer (NSCLC), it has limited therapeutic efficacy due to activation of a survival signaling pathway and the induction of multidrug resistance. Curcumin, a natural compound isolated from the plant Curcuma longa, is known to sensitize tumors to different chemotherapeutic agents. The aim of this study is to evaluate whether curcumin can chemosensitize lung cancer cells to carboplatin and to analyze the signaling pathway underlying this synergism. We investigated the synergistic effect of both agents on cell proliferation, apoptosis, invasion, migration, and expression of related signaling proteins using the human NSCLC cell line, A549. A549 cell was treated with different concentrations of curcumin and carboplatin alone and in combination. Combined treatment with curcumin and carboplatin inhibited tumor cell growth, migration, and invasion compared with either drug alone. Matrix metalloproteinase (MMP)-2 and MMP-9 were more efficiently downregulated by co-treatment than by each treatment alone. mRNA and protein expression of caspase-3 and caspase-9 and proapoptotic genes was increased in cells treated with a combination of curcumin and carboplatin, whereas expression of the antiapoptotic Bcl-2 gene was suppressed. Co-treatment of both agents substantially suppressed NF-κB activation and increased expression of p53. Phosphorylation of Akt, a protein upstream of NF-κB, was reduced, resulting in inhibition of the degradation of inhibitor of κB(IκBα), whereas the activity of extracellular signal-regulated kinase (ERK1/2) was enhanced. Our study demonstrated that the synergistic antitumor activity of curcumin combined with carboplatin is mediated by multiple mechanisms involving suppression of NF-κB via inhibition of the Akt/IKKα pathway and enhanced ERK1/2 activity. Based on this mechanism, curcumin has potential as a chemosensitizer for carboplatin in the treatment of

  10. Study on Inhibitory Effect of MaiMenDong Decoction and WeiJing Decoction Combination with Cisplatin on NCI-A549 Xenograft in Nude Mice and Its Mechanism.

    PubMed

    Xiong, Fei; Jiang, Miao; Chen, Meijuan; Wang, Xiaoxia; Zhang, Shiping; Zhou, Jing; Li, Ke; Sheng, Yan; Yin, Lian; Tang, Yuping; Ye, Lihong; Wu, Mianhua; Fu, Haian; Zhang, Xu

    2017-01-01

    MaiMenDong Decoction and WeiJing Decoction (Jin formula) is a traditional Chinese medication that consists of 8 medicinal plants, which recorded in the classical TCM literature Jin Kui Yao Lue and has been utilized in the treatment of lung diseases for hundreds of years in China. The present study aimed to determine the anti-tumor activity and the underlying mechanisms of Jin formula combined with cisplatin in the treatment of non-small cell lung cancer (NSCLC). Xenograft model of NCI-A549 was established in Balb/c nude mice. Five groups, including normal, MOCK, Jin, cisplatin (DDP), and Jin+DDP were included in the study. We found that Jin formula ameliorated the body weight loss caused by DDP 15 days after drug administration. Moreover, the combination of Jin with DDP enhanced the anti-tumor function of DDP. Microarray analysis showed that Jin suppressed gene expression of certain pathways which regulating cell cycle and apoptosis. Furthermore, DDP mainly decreased the gene expression level of angiogenesis associated factors, such as VEGFA, TGF-β and MMP-1. Moreover, co-treatment with Jin and DDP not only down-regulated Bcl-2 and E2F1, but also decreased the expression of MYC, MET, and MCAM. In addition, co-formula decreased the levels of p-AKT (thr308) and p-PTEN, increased Bax/Bcl-2 value, and resulted in apoptosis of tumor cells. Taken together , Jin+DDP significantly inhibited the growth of A549 cell transplanted solid tumor with slight side effect compared to the treatment by DDP only, and had a better effect than the Jin group. The mechanisms may be mainly associated with inactivation of PI3K/AKT pathway and apoptosis induction.

  11. Resistance to etoposide-induced apoptosis in a Burkitt's lymphoma cell line.

    PubMed

    Zhao, E G; Song, Q; Cross, S; Misko, I; Lees-Miller, S P; Lavin, M F

    1998-08-31

    Burkitt's lymphoma cells that vary in their phenotypic characteristics show significantly different degrees of susceptibility to radiation-induced apoptosis. Propensity to undergo apoptosis is reflected in the degradation of substrates such as DNA-dependent protein kinase but the status of bcl-2, c-myc and p53 has been uninformative. In this study, we have focused on 2 Epstein-Barr virus (EBV)-associated Burkitt's cell lines, one (WW2) susceptible and the other (BL29) resistant to etoposide-induced apoptosis. Differences in expression of BHRF1, an EBV gene that is homologous to the Bcl-2 proto-oncogene and known to inhibit apoptosis, or changes in apoptosis inhibitory proteins (IAPs), did not appear to account for the difference in susceptibility in the 2 cell lines. Cytoplasmic extracts from etoposide-treated WW2 cells caused apoptotic changes in nuclei isolated from either BL29 or WW2 cells, whereas extracts from BL29 cells failed to do so. In addition, extracts from etoposide-treated WW2 cells degraded the catalytic subunit of DNA-dependent protein kinase (DNA-PKcs), an important indicator of apoptosis, but this protein was resistant to degradation by BL29 extracts. It appears likely that caspase 3 (CPP32) is involved in this degradation since it was activated only in the apoptosis susceptible cells and the pattern of cleavage of DNA-PKcs was similar to that reported previously with recombinant caspase 3. As observed previously, addition of caspase 3 to nuclei failed to induce morphological changes indicative of apoptosis, but addition of caspase 3 to nuclei in the presence of extract from the resistant cells led to apoptotic changes. We conclude that resistance to apoptosis in BL29 cells is due to a failure of etoposide to activate upstream effectors of caspase activity.

  12. Effective deactivation of A549 tumor cells in vitro and in vivo by RGD-decorated chitosan-functionalized single-walled carbon nanotube loading docetaxel.

    PubMed

    Li, Bin; Zhang, Xiao-Xue; Huang, Hao-Yan; Chen, Li-Qing; Cui, Jing-Hao; Liu, Yanli; Jin, Hehua; Lee, Beom-Jin; Cao, Qing-Ri

    2018-05-30

    This study aims to construct and evaluate RGD-decorated chitosan (CS)-functionalized pH-responsive single-walled carbon nanotube (SWCNT) carriers using docetaxel (DTX) as a model anticancer drug. DTX was loaded onto SWCNT via π-π stacking interaction (SWCNT-DTX), followed by the non-covalent conjugation of RGD-decorated CS to SWCNT-DTX to prepare RGD-CS-SWCNT-DTX. The RGD-CS-SWCNT-DTX showed significantly higher drug release than the pure drug, giving higher release rate at pH 5.0 (68%) than pH 7.4 (49%). The RGD-CS-SWCNT-DTX could significantly inhibit the growth of A549 tumor cells in vitro, and the uptake amount of A549 cells was obviously higher than that of MCF-7 cells. Meanwhile, the cellular uptake of RGD-CS-SWCNT-DTX was higher than that of CS-SWCNT-DTX in A549 cells, mainly through clathrin and caveolae-mediated endocytosis. The RGD-CS-SWCNT-DTX significantly inhibited tumor growth of A549 cell-bearing nude mice through active tumor-targeting ability. Furthermore, no pathological changes were found in tissues and organs. The result demonstrated that RGD-CS-SWCNT-DTX displayed high drug loading, pH-responsive drug release, remarkable antitumor effect in vitro and in vivo, and also good safety to animal body. Copyright © 2018 Elsevier B.V. All rights reserved.

  13. Imiquimod activates p53-dependent apoptosis in a human basal cell carcinoma cell line.

    PubMed

    Huang, Shi-Wei; Chang, Shu-Hao; Mu, Szu-Wei; Jiang, Hsin-Yi; Wang, Sin-Ting; Kao, Jun-Kai; Huang, Jau-Ling; Wu, Chun-Ying; Chen, Yi-Ju; Shieh, Jeng-Jer

    2016-03-01

    The tumor suppressor p53 controls DNA repair, cell cycle, apoptosis, autophagy and numerous other cellular processes. Imiquimod (IMQ), a synthetic toll-like receptor (TLR) 7 ligand for the treatment of superficial basal cell carcinoma (BCC), eliminates cancer cells by activating cell-mediated immunity and directly inducing apoptosis and autophagy in cancer cells. To evaluate the role of p53 in IMQ-induced cell death in skin cancer cells. The expression, phosphorylation and subcellular localization of p53 were detected by real-time PCR, luciferase reporter assay, cycloheximide chase analysis, immunoblotting and immunocytochemistry. Using BCC/KMC1 cell line as a model, the upstream signaling of p53 activation was dissected by over-expression of TLR7/8, the addition of ROS scavenger, ATM/ATR inhibitors and pan-caspase inhibitor. The role of p53 in IMQ-induced apoptosis and autophagy was assessed by genetically silencing p53 and evaluated by a DNA content assay, immunoblotting, LC3 puncta detection and acridine orange staining. IMQ induced p53 mRNA expression and protein accumulation, increased Ser15 phosphorylation, promoted nuclear translocation and up-regulated its target genes in skin cancer cells in a TLR7/8-independent manner. In BCC/KMC1 cells, the induction of p53 by IMQ was achieved through increased ROS production to stimulate the ATM/ATR-Chk1/Chk2 axis but was not mediated by inducing DNA damage. The pharmacological inhibition of ATM/ATR significantly suppressed IMQ-induced p53 activation and apoptosis. Silencing of p53 significantly decreased the IMQ-induced caspase cascade activation and apoptosis but enhanced autophagy. Mutant p53 skin cancer cell lines were more resistant to IMQ-induced apoptosis than wildtype p53 skin cancer cell lines. IMQ induced ROS production to stimulate ATM/ATR pathways and contributed to p53-dependent apoptosis in a skin basal cell carcinoma cell line BCC/KMC1. Copyright © 2015 Japanese Society for Investigative Dermatology

  14. Extract from Nandina domestica inhibits lipopolysaccharide-induced cyclooxygenase-2 expression in human pulmonary epithelial A549 cells.

    PubMed

    Ueki, Takuro; Akaishi, Tatsuhiro; Okumura, Hidenobu; Abe, Kazuho

    2012-01-01

    Extract from fruits of Nandina domestica THUNBERG (NDE) has been used to improve cough and breathing difficulty in Japan for many years. To explore whether NDE may alleviate respiratory inflammation, we investigated its effect on expression of cyclooxygenase-2 (COX-2) and production of prostaglandin E₂ (PGE₂) in human pulmonary epithelial A549 cells in culture. Treatment with lipopolysaccharide (LPS; 6 µg/mL) resulted in an increase of COX-2 expression and PGE₂ production in A549 cells. Both the LPS-induced COX-2 expression and PGE₂ production were significantly inhibited by NDE (1-10 µg/mL) in a concentration-dependent manner. NDE did not affect COX-1 expression nor COX activity. These results suggest that NDE downregulates LPS-induced COX-2 expression and inhibits PGE₂ production in pulmonary epithelial cells. Furthermore, higenamine and nantenine, two major constituents responsible for tracheal relaxing effect of NDE, did not mimic the inhibitory effect of NDE on LPS-induced COX-2 expression in A549 cells. To identify active constituent(s) of NDE responsible for the anti-inflammatory effect, NDE was introduced in a polyaromatic absorbent resin column and stepwise eluted to yield water fraction, 20% methanol fraction, 40% methanol fraction, 99.8% methanol fraction, and 99.5% acetone fraction. However, none of these five fractions alone inhibited LPS-induced COX-2 expression. On the other hand, exclusion of water fraction from NDE abolished the inhibitory effect of NDE on LPS-induced COX-2 expression. These results suggest that constituent(s) present in water fraction is required but not sufficient for the anti-inflammatory activity of NDE, which may result from interactions among multiple constituents.

  15. Inhibition of TNF-alpha-induced NF-kappaB activation and IL-8 release in A549 cells with the proteasome inhibitor MG-132.

    PubMed

    Fiedler, M A; Wernke-Dollries, K; Stark, J M

    1998-08-01

    The working hypothesis of the studies described herein was that inhibition of proteasome-mediated IkappaB degradation would inhibit TNF-alpha-induced nuclear factor-kappaB (NF-kappaB) activation, interleukin-8 (IL-8) gene transcription, and IL-8 protein release in A549 cells. Mutational analysis of the 5' flanking region of the IL-8 gene confirmed that an intact NF-kappaB site is necessary for TNF-alpha-induced IL-8 gene transcription. The addition of TNF-alpha to A549 cells resulted in rapid loss of IkappaB from the cytoplasm of cells, associated with a corresponding increase in NF-kappaB-binding activity in nuclear extracts from the cells. However, pretreatment of the cells with the proteasome inhibitor N-cbz-Leu-Leu-leucinal (MG-132, 10 microM) reversed the effects of TNF-alpha on IL-8 release from A549 cells (as determined with an enzyme-linked immunosorbent assay [ELISA]) and on IL-8 gene transcription (as determined with reporter-gene assays). MG-132 reversed the effects of TNF-alpha on IkappaB degradation as determined by Western blot analysis. IkappaB phosphorylation and ubiquination were not altered by MG-132, which implies that the effects of MG-132 were secondary to proteasome inhibition. MG-132 also reversed the increase in NF-kappaB binding in nuclear extracts from TNF-alpha-treated cells. These studies show that inhibition of proteasome-mediated IkappaB degradation results in inhibition of TNF-alpha induced IL-8 production in A549 cells by limiting NF-kappaB-mediated gene transcription.

  16. Aspartame-induced apoptosis in PC12 cells.

    PubMed

    Horio, Yukari; Sun, Yongkun; Liu, Chuang; Saito, Takeshi; Kurasaki, Masaaki

    2014-01-01

    Aspartame is an artificial sweetner added to many low-calorie foods. The safety of aspartame remains controversial even though there are many studies on its risks. In this study, to understand the physiological effects of trace amounts of artificial sweetners on cells, the effects of aspartame on apoptosis were investigated using a PC12 cell system. In addition, the mechanism of apoptosis induced by aspartame in PC12 cells and effects on apoptotic factors such as cytochrome c, apoptosis-inducing factor, and caspase family proteins were studied by Western blotting and RT-PCR. Aspartame-induced apoptosis in PC12 cells in a dose-dependent manner. In addition, aspartame exposure increased the expressions of caspases 8 and 9, and cytochrome c. These results indicate that aspartame induces apoptosis mainly via mitochondrial pathway involved in apoptosis due to oxigen toxicity. Copyright © 2013 Elsevier B.V. All rights reserved.

  17. Decreased levels of irisin, a skeletal muscle cell-derived myokine, are related to emphysema associated with chronic obstructive pulmonary disease.

    PubMed

    Sugiyama, Yukari; Asai, Kazuhisa; Yamada, Kazuhiro; Kureya, Yuko; Ijiri, Naoki; Watanabe, Tetsuya; Kanazawa, Hiroshi; Hirata, Kazuto

    2017-01-01

    Cigarette smoking-induced oxidant-antioxidant imbalance is a factor that contributes to the pathogenesis of COPD through epithelial cell apoptosis. Irisin is a skeletal muscle cell-derived myokine associated with physical activity. Irisin is also known to decrease oxidant-induced apoptosis in patients with diabetes mellitus. However, the correlation between irisin and emphysema in COPD and its role in epithelial cell apoptosis remains unknown. Forty patients with COPD were enrolled in this study. Pulmonary function tests and measurements of the percentage of low-attenuation area on high-resolution computed tomography images were performed, and the results were evaluated for correlation with serum irisin levels. The effect of irisin on cigarette-smoke extract-induced A549 cell apoptosis and the expression of Nrf2, a transcription factor for antioxidants, was also examined in vitro. Serum irisin levels were significantly correlated with lung diffusing capacity for carbon monoxide divided by alveolar volume ( r =0.56, P <0.01) and percentage of low-attenuation area ( r =-0.79, P <0.01). Moreover, irisin significantly enhanced Nrf2 expression ( P <0.05) and reduced cigarette-smoke extract-induced A549 cell apoptosis ( P <0.05). Decreased serum irisin levels are related to emphysema in patients with COPD and involved in epithelial apoptosis, resulting in emphysema. Irisin could be a novel treatment for emphysema in patients with COPD.

  18. Role of {alpha}{sub v}{beta}{sub 5} integrin receptor in endocytosis of crocidolite and its effect on intracellular glutathione levels in human lung epithelial (A549) cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Pande, Priyadarshini; Mosleh, Tariq A.; Aust, Ann E.

    Crocidolite, containing 27% iron by weight, is the most carcinogenic form of asbestos. Crocidolite fibers are endocytized by {alpha}{sub v}{beta}{sub 5} integrin receptors in rabbit pleural mesothelial cells. We show here that crocidolite fibers are endocytized in human lung epithelial (A549) cells and in primary small airway epithelial (SAEC) cells. Presence of the integrin {alpha}{sub v}{beta}{sub 5} blocking antibody, P1F6, significantly reduced the uptake of crocidolite fibers in A549 cells. Thus, the integrin {alpha}{sub v}{beta}{sub 5} receptor is involved in endocytosis of crocidolite fibers in A549 cells as well. Previously, it has been observed that asbestos fibers lead to changesmore » in the intracellular redox environment, i.e. a marked decrease in intracellular glutathione concentrations and an increase in the extracellular glutathione in A549 cells. In addition, the decrease in intracellular glutathione was found to be largely independent of iron present on the surface of the fiber. A549 cells were treated with crocidolite in the presence of endocytosis inhibitor cytochalasin D. Our data indicate that, upon preventing endocytosis, we were able to reverse the decrease in total intracellular glutathione. The decrease in total intracellular glutathione could also be prevented in the presence of the monoclonal antibody P1F6. Thus, we observed that endocytosis of crocidolite fibers via integrin {alpha}{sub v}{beta}{sub 5} receptor is linked to the marked decrease in total intracellular glutathione in A549 cells.« less

  19. Cellular uptake and toxic effects of fine and ultrafine metal-sulfate particles in human A549 lung epithelial cells.

    PubMed

    Könczöl, Mathias; Goldenberg, Ella; Ebeling, Sandra; Schäfer, Bianca; Garcia-Käufer, Manuel; Gminski, Richard; Grobéty, Bernard; Rothen-Rutishauser, Barbara; Merfort, Irmgard; Gieré, Reto; Mersch-Sundermann, Volker

    2012-12-17

    Ambient airborne particulate matter is known to cause various adverse health effects in humans. In a recent study on the environmental impacts of coal and tire combustion in a thermal power station, fine crystals of PbSO(4) (anglesite), ZnSO(4)·H(2)O (gunningite), and CaSO(4) (anhydrite) were identified in the stack emissions. Here, we have studied the toxic potential of these sulfate phases as particulates and their uptake in human alveolar epithelial cells (A549). Both PbSO(4) and CaSO(4) yielded no loss of cell viability, as determined by the WST-1 and NR assays. In contrast, a concentration-dependent increase in cytotoxicity was observed for Zn sulfate. For all analyzed sulfates, an increase in the production of reactive oxygen species (ROS), assessed by the DCFH-DA assay and EPR, was observed, although to a varying extent. Again, Zn sulfate was the most active compound. Genotoxicity assays revealed concentration-dependent DNA damage and induction of micronuclei for Zn sulfate and, to a lower extent, for CaSO(4), whereas only slight effects could be found for PbSO(4). Moreover, changes of the cell cycle were observed for Zn sulfate and PbSO(4). It could be shown further that Zn sulfate increased the nuclear factor kappa-B (NF-κB) DNA binding activity and activated JNK. During our TEM investigations, no effect on the appearance of the A549 cells exposed to CaSO(4) compared to the nonexposed cells was observed, and in our experiments, only one CaSO(4) particle was detected in the cytoplasm. In the case of exposure to Zn sulfate, no particles were found in the cytoplasm of A549 cells, but we observed a concentration-dependent increase in the number and size of dark vesicles (presumably zincosomes). After exposure to PbSO(4), the A549 cells contained isolated particles as well as agglomerates both in vesicles and in the cytoplasm. Since these metal-sulfate particles are emitted into the atmosphere via the flue gas of coal-fired power stations, they may be

  20. Inducing cell death in vitro in cancer cells by targeted delivery of cytochrome c via a transferrin conjugate

    PubMed Central

    Delgado, Yamixa; Sharma, Rohit Kumar; Sharma, Shweta; Guzmán, Solimar Liz Ponce De León; Tinoco, Arthur D.; Griebenow, Kai

    2018-01-01

    One of the major drawbacks of many of the currently used cancer drugs are off-target effects. Targeted delivery is one method to minimize such unwanted and detrimental events. To actively target lung cancer cells, we have developed a conjugate of the apoptosis inducing protein cytochrome c with transferrin because the transferrin receptor is overexpressed by many rapidly dividing cancer cells. Cytochrome c and transferrin were cross-linked with a redox sensitive disulfide bond for the intra-cellular release of the protein upon endocytosis by the transferrin receptor. Confocal results demonstrated the cellular uptake of the cytochrome c-transferrin conjugate by transferrin receptor overexpressing A549 lung cancer cells. Localization studies further validated that this conjugate escaped the endosome. Additionally, an in vitro assay showed that the conjugate could induce apoptosis by activating caspase-3. The neo-conjugate not only maintained an IC50 value similar to the well known drug cisplatin (50 μM) in A549 cancer cells but also was nontoxic to the normal lung (MRC5) cells. Our neo-conjugate holds promise for future development to target cancers with enhanced transferrin receptor expression. PMID:29649293

  1. Preprocessing with Photoshop Software on Microscopic Images of A549 Cells in Epithelial-Mesenchymal Transition.

    PubMed

    Ren, Zhou-Xin; Yu, Hai-Bin; Shen, Jun-Ling; Li, Ya; Li, Jian-Sheng

    2015-06-01

    To establish a preprocessing method for cell morphometry in microscopic images of A549 cells in epithelial-mesenchymal transition (EMT). Adobe Photoshop CS2 (Adobe Systems, Inc.) was used for preprocessing the images. First, all images were processed for size uniformity and high distinguishability between the cell and background area. Then, a blank image with the same size and grids was established and cross points of the grids were added into a distinct color. The blank image was merged into a processed image. In the merged images, the cells with 1 or more cross points were chosen, and then the cell areas were enclosed and were replaced in a distinct color. Except for chosen cellular areas, all areas were changed into a unique hue. Three observers quantified roundness of cells in images with the image preprocess (IPP) or without the method (Controls), respectively. Furthermore, 1 observer measured the roundness 3 times with the 2 methods, respectively. The results between IPPs and Controls were compared for repeatability and reproducibility. As compared with the Control method, among 3 observers, use of the IPP method resulted in a higher number and a higher percentage of same-chosen cells in an image. The relative average deviation values of roundness, either for 3 observers or 1 observer, were significantly higher in Controls than in IPPs (p < 0.01 or 0.001). The values of intraclass correlation coefficient, both in Single Type or Average, were higher in IPPs than in Controls both for 3 observers and 1 observer. Processed with Adobe Photoshop, a chosen cell from an image was more objective, regular, and accurate, creating an increase of reproducibility and repeatability on morphometry of A549 cells in epithelial to mesenchymal transition.

  2. Inhibitory Effect of Kurarinone on Growth of Human Non-small Cell Lung Cancer: An Experimental Study Both in Vitro and in Vivo Studies

    PubMed Central

    Yang, Jie; Chen, Hao; Wang, Qiang; Deng, Shihao; Huang, Mi; Ma, Xinhua; Song, Ping; Du, Jingwen; Huang, Yun; Wen, Yanzhang; Ren, Yongshen; Yang, Xinzhou

    2018-01-01

    Kurarinone, a flavonoid isolated from Sophora flavescens Aiton, has been reported to have significant antitumor activity. However, the cytotoxic activity of kurarinone against non-small cell lung cancer (NSCLC) cells is still under explored. In our study, we have evaluated the inhibitory effects of kurarinone on the growth of NSCLC both in vivo and in vitro as well as the molecular mechanisms underlying kurarinone-induced A549 cell apoptosis. The results showed that kurarinone effectively inhibited the proliferation of A549 cells with little toxic effects on human bronchial epithelial cell line BEAS-2B. FASC examination and Hoechst 33258 staining assay showed that kurarinone dose-dependently provoked A549 cells apoptosis. Mechanistically, kurarinone significantly decreased the ratio of Bcl-2/Bax, thereby causing the activation of caspase 9 and caspase 3, and reduced the expression of Grp78, which led to relieve the inhibition of caspase-12 and caspase-7, as well as suppressing the activity of AKT. Meanwhile, modeling results from the Surflex-Dock program suggested that residue Ser473 of Akt is a potential binding site for kurarinone. In vivo, kurarinone inhibited the growth of A549 xenograft mouse models without apparent signs of toxicity. Our study indicated that kurarinone has the potential effects of anti-NSCLC, implemented through activating mitochondria apoptosis signaling pathway, as well as repressing the activity of endoplasmic reticulum pathway and AKT in A549 cells. PMID:29628889

  3. MicroRNA regulatory networks reflective of polyhexamethylene guanidine phosphate-induced fibrosis in A549 human alveolar adenocarcinoma cells.

    PubMed

    Shin, Da Young; Jeong, Mi Ho; Bang, In Jae; Kim, Ha Ryong; Chung, Kyu Hyuck

    2018-05-01

    Polyhexamethylene guanidine phosphate (PHMG-phosphate), an active component of humidifier disinfectant, is suspected to be a major cause of pulmonary fibrosis. Fibrosis, induced by recurrent epithelial damage, is significantly affected by epigenetic regulation, including microRNAs (miRNAs). The aim of this study was to investigate the fibrogenic mechanisms of PHMG-phosphate through the profiling of miRNAs and their target genes. A549 cells were treated with 0.75 μg/mL PHMG-phosphate for 24 and 48 h and miRNA microarray expression analysis was conducted. The putative mRNA targets of the miRNAs were identified and subjected to Gene Ontology analysis. After exposure to PHMG-phosphate for 24 and 48 h, 46 and 33 miRNAs, respectively, showed a significant change in expression over 1.5-fold compared with the control. The integrated analysis of miRNA and mRNA microarray results revealed the putative targets that were prominently enriched were associated with the epithelial-mesenchymal transition (EMT), cell cycle changes, and apoptosis. The dose-dependent induction of EMT by PHMG-phosphate exposure was confirmed by western blot. We identified 13 putative EMT-related targets that may play a role in PHMG-phosphate-induced fibrosis according to the Comparative Toxicogenomic Database. Our findings contribute to the comprehension of the fibrogenic mechanism of PHMG-phosphate and will aid further study on PHMG-phosphate-induced toxicity. Copyright © 2018 Elsevier B.V. All rights reserved.

  4. Effect of functionalized and non-functionalized nanodiamond on the morphology and activities of antioxidant enzymes of lung epithelial cells (A549).

    PubMed

    Solarska-Ściuk, Katarzyna; Gajewska, Agnieszka; Glińska, Sława; Michlewska, Sylwia; Balcerzak, Łucja; Jamrozik, Agnieszka; Skolimowski, Janusz; Burda, Květoslava; Bartosz, Grzegorz

    2014-10-05

    The development of nanotechnology opens up new ways for biomedical applications of unmodified and modified diamond nanoparticles which are one of the most popular nanomaterials used in biology, biotechnology, medicine, cosmetics and engineering. They have been applied as diagnostic and therapeutic agents because they can be targeted to and localized in cells causing apoptosis and necrosis. The problem of biocompatibility of nanodiamonds at higher concentrations is thus of primary importance. The first step in the modification of DNPs is usually the introduction of hydrogen groups, which can bind other functional groups. The basic method to introduce -OH groups onto nanoparticles is the Fenton reaction. The aim of this study was to compare the effect of unmodified nanodiamond particles and nanoparticles modified by introduction of -OH groups and etoposide onto their surface reaction on human non-small lung cancer cells. A549 cells were incubated with 2-100μg/ml nanopowders and at 0.6-24μg/ml etoposide in the DMEM medium. We observed a decrease of cells viability and generation of reactive oxygen/ nitrogen species in the cells after incubation, estimated by oxidation of H2DCF-DA and DAF-FM-DA. Modified detonation nanoparticles affected also the cellular content of glutathione and activities of main antioxidant enzymes (glutathione peroxidase, glutathione reductase, glutathione S-transferase, superoxide dismutase and catalase). The results of TEM microscopy show changes in cell morphology. These data demonstrate that modified nanoparticles induce oxidative stress in the target cells. Copyright © 2014 Elsevier Ireland Ltd. All rights reserved.

  5. Curcumin improves the efficacy of cisplatin by targeting cancer stem-like cells through p21 and cyclin D1-mediated tumour cell inhibition in non-small cell lung cancer cell lines

    PubMed Central

    BAHARUDDIN, PUTERI; SATAR, NAZILAH; FAKIRUDDIN, KAMAL SHAIK; ZAKARIA, NORASHIKIN; LIM, MOON NIAN; YUSOFF, NARAZAH MOHD; ZAKARIA, ZUBAIDAH; YAHAYA, BADRUL HISHAM

    2016-01-01

    Natural compounds such as curcumin have the ability to enhance the therapeutic effectiveness of common chemotherapy agents through cancer stem-like cell (CSC) sensitisation. In the present study, we showed that curcumin enhanced the sensitivity of the double-positive (CD166+/EpCAM+) CSC subpopulation in non-small cell lung cancer (NSCLC) cell lines (A549 and H2170) to cisplatin-induced apoptosis and inhibition of metastasis. Our results revealed that initial exposure of NSCLC cell lines to curcumin (10–40 µM) markedly reduced the percentage of viability to an average of ~51 and ~54% compared to treatment with low dose cisplatin (3 µM) with only 94 and 86% in both the A549 and H2170 cells. Moreover, sensitisation of NSCLC cell lines to curcumin through combined treatment enhanced the single effect induced by low dose cisplatin on the apoptosis of the double-positive CSC subpopulation by 18 and 20% in the A549 and H2170 cells, respectively. Furthermore, we found that curcumin enhanced the inhibitory effects of cisplatin on the highly migratory CD166+/EpCAM+ subpopulation, marked by a reduction in cell migration to 9 and 21% in the A549 and H2170 cells, respectively, indicating that curcumin may increase the sensitivity of CSCs to cisplatin-induced migratory inhibition. We also observed that the mRNA expression of cyclin D1 was downregulated, while a substantial increased in p21 expression was noted, followed by Apaf1 and caspase-9 activation in the double-positive (CD166+/EpCAM+) CSC subpopulation of A549 cells, suggested that the combined treatments induced cell cycle arrest, therefore triggering CSC growth inhibition via the intrinsic apoptotic pathway. In conclusion, we provided novel evidence of the previously unknown therapeutic effects of curcumin, either alone or in combination with cisplatin on the inhibition of the CD166+/EpCAM+ subpopulation of NSCLC cell lines. This finding demonstrated the potential therapeutic approach of using curcumin that may

  6. Phosphorylation of p53 at serine 15 in A549 pulmonary epithelial cells exposed to vanadate: Involvement of ATM pathway

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Suzuki, Katsura; Inageda, Kiyoshi; Nishitai, Gen

    2007-04-01

    When A549 cells were exposed to sodium metavanadate (NaVO{sub 3}), the pentavalent species of vanadium (vanadate), phosphorylation of p53 protein at Ser15 was found in a time (8-48 h)- and dose (10-200 {mu}M)-dependent manner. After the incubation with 50 or 100 {mu}M NaVO{sub 3} for 48 h, accumulation of p53 protein was accompanied with Ser15 phosphorylation. Among serines in p53 protein immunoprecipitated from A549 cells treated with 100 {mu}M NaVO{sub 3} for 48 h, only Ser15 was markedly phosphorylated. Treatment with other vanadate compounds, sodium orthovanadate (Na{sub 3}VO{sub 4}) and ammonium metavanadate (NH{sub 4}VO{sub 3}), also induced Ser15 phosphorylation andmore » accumulation of p53 protein. While phosphorylation of extracellular signal-regulated protein kinase (ERK) was found in cells treated with NaVO{sub 3}, treatment with U0126 did not suppress Ser15 phosphorylation. On the other hand, treatment with wortmannin or caffeine, the inhibitors to phosphatidylinositol 3-kinase related kinases (PIKKs), suppressed both NaVO{sub 3}-induced Ser15 phosphorylation and accumulation of p53 protein. The silencing of ataxia telangiectasia mutated (ATM) expression using short-interference RNA resulted in the marked suppression of Ser15 phosphorylation in A549 cells exposed to NaVO{sub 3}. However, treatment with antioxidants such as catalase and N-acetylcysteine did not suppress NaVO{sub 3}-induced Ser15 phosphorylation. Transcriptional activation of p53 and DNA fragmentation in A549 cells treated with NaVO{sub 3} were suppressed only slightly by S15A mutation, suggesting that Ser15 phosphorylation is not essential for these responses. The present results showed that vanadate induces the phosphorylation of p53 at Ser15 depending on ATM, one of the members of PIKK family, in this human pulmonary epithelial cell line.« less

  7. MicroRNA-196b Inhibits Cell Growth and Metastasis of Lung Cancer Cells by Targeting Runx2.

    PubMed

    Bai, Xiaoxue; Meng, Lin; Sun, Huijie; Li, Zhuo; Zhang, Xiufang; Hua, Shucheng

    2017-01-01

    Lung cancer is one of the most common causes of cancer related deaths worldwide. The role of several microRNAs (miRNAs) including miR-196b in different cancers has already been established. The study was aimed to explore the role of miR-196b in lung cancer and its possible underlying mechanism. Human lung cancer cell line A549 was transfected with miR-196b mimic, miR-196b inhibitor and corresponding controls. Then cell viability, migration, invasion, and apoptosis of A549 lung cancer cells either with overexpression or with suppression of miR-196b were estimated sequentially. Next, dual luciferase activity assay was performed to clarify whether Runx2 was a direct target of miR-196b. Finally, the expressions of main factors associated with epithelial mesenchymal transition (EMT), PI3K/AKT/GSK3β, Smad, and JNK pathways were detected by western blot. MiR-196b expression was significantly decreased in A549, H1650 and H1299 cell lines compared with in WI-38 and HEL-1 cell lines. Overexpression of miR-196b suppressed cell viability, migration, invasion, and induced apoptosis as well as inhibited TGF-β induced EMT process in A549 cells. In addition, Runx2 was a putative target of miR-196b, and Runx2 silence remarkably increased cell apoptosis and abolished the promotive effects of miR-196b suppression on cell viability, migration and invasion. Finally, miR-196b also mediated its action by inactivation of PI3K/AKT/GSK3β, Smad, and JNK pathways by down-regulation of Runx2. MiR-196b functions as a tumor suppressor that inhibited cell growth and metastasis of lung cancer cells by targeting Runx2. These findings provided further evidences for treatment of lung cancer. The Author(s). Published by S. Karger AG, Basel.

  8. Anticancer effects of β-elemene with hyperthermia in lung cancer cells

    PubMed Central

    Wu, Zhibing; Wang, Ting; Zhang, Yanmei; Zheng, Zhishuang; Yu, Shuhuan; Jing, Saisai; Chen, Sumei; Jiang, Hao; Ma, Shenglin

    2017-01-01

    β-elemene is a novel, plant-derived anticancer drug, which has been used to target multiple solid tumor types. Hyperthermia is an adjuvant therapeutic modality to treat cancer. However, the underlying mechanisms associated with the efficacy of these two treatments are largely unknown. The aim of the present study was to evaluate the effects of β-elemene combined with hyperthermia in lung cancer cell lines. An MTT assay was used to determine cell viability. The cell cycle and apoptosis were analyzed using flow cytometry. The morphology of cells during apoptosis was determined using a transmission electron microscope. The expression levels of P21, survivin, caspase-9, B-cell lymphoma 2 (Bcl-2) and Bcl-2-like protein 4 (Bax) mRNA were detected using quantitative polymerase chain reaction. β-elemene with hyperthermia treatment significantly inhibited the viability and increased the apoptosis rate of A549 cells compared with β-elemene treatment alone (P<0.01), and significantly decreased the proportion of cells in S phase compared with the control (P<0.01). Morphological observation using transmission electron microscopy indicated cross-sectional features of apoptosis: Chromatin condensation, reduced integrity of the plasma membrane, increased cellular granularity, nuclear collapse and the formation of apoptotic bodies. β-elemene with hyperthermia treatment significantly promoted P21 and Bax mRNA expression (P<0.01) and significantly decreased caspase-9, Bcl-2 and survivin mRNA expression (P<0.01) in A549 cells. In conclusion, β-elemene with hyperthermia has a significant inhibitory effect on A549 cells. This occurs through reducing S phase and inducing apoptosis, via an increase in P21 and Bax expression and a decrease in caspase-9, Bcl-2 and survivin expression. PMID:28588670

  9. [Effect of silencing Bmi-1 expression in reversing cisplatin resistance in lung cancer cells and its mechanism].

    PubMed

    Mao, Nan; He, Guansheng; Rao, Jinjun; Lv, Lin

    2014-06-01

    To investigate the effect of silencing Bmi-1 expression in reversing cisplatin resistance in human lung cancer cells and explore the possible mechanisms. Cisplatin-resistant A549/DDP cells with small interference RNA (siRNA)-mediated Bmi-1 expression silencing were examined for cisplatin sensitivity using MTT assay and alterations in cell cycle distribution and apoptosis with flow cytometry, and the changes in cell senescence was assessed using β-galactosidase staining. The protein expressions of Bmi-1, P14(ARF), P16(INK4a), P53, P21, Rb and ubi-H2AK119 in the cells were determined with Western blotting. A549/DDP cells showed significantly higher Bmi-1 expression than A549 cells. After siRNA-mediated Bmi-1 silencing, A549/DDP cells showed significantly enhanced cisplatin sensitivity with an increased IC50 from 40.3±4.1 µmol/L to 18.3±2.8 µmol/L (P<0.01) and increased cell percentage in G0/G1 phase from (48.9±2.3)% to (78.7±7.6)% (P<0.01). Silencing Bmi-1 did not cause significant changes in the cell apoptosis rate but induced obvious senescence phenotype in A549/DDP cells with down-regulated expression of ubi-H2AK119 and up-regulated expressions of P14(ARF), P16(INK4a), P53, P21 and Rb. Silencing Bmi-1 by RNA interference can induce cell senescence and resensitize A549/DDP cells to cisplatin possibly by regulating INK4a/ARF/Rb senescence pathway.

  10. Ap4A induces apoptosis in human cultured cells.

    PubMed

    Vartanian, A; Alexandrov, I; Prudowski, I; McLennan, A; Kisselev, L

    1999-07-30

    Diadenosine oligophosphates (Ap(n)A) have been proposed as intracellular and extracellular signaling molecules in animal cells. The ratio of diadenosine 5',5'''-P1,P3-triphosphate to diadenosine 5',5'''-P1,P4-tetraphosphate (Ap3A/Ap4A) is sensitive to the cellular status and alters when cultured cells undergo differentiation or are treated with interferons. In cells undergoing apoptosis induced by DNA topoisomerase II inhibitor VP16, the concentration of Ap3A decreases significantly while that of Ap4A increases. Here, we have examined the effects of exogenously added Ap3A and Ap4A on apoptosis and morphological differentiation. Penetration of Ap(n)A into cells was achieved by cold shock. Ap4A at 10 microM induced programmed cell death in human HL60, U937 and Jurkat cells and mouse VMRO cells and this effect appeared to require Ap4A breakdown as hydrolysis-resistant analogues of Ap4A were inactive. On its own, Ap3A induced neither apoptosis nor cell differentiation but did display strong synergism with the protein kinase C activators 12-deoxyphorbol-13-O-phenylacetate and 12-deoxyphorbol-13-O-phenylacetate-20-acetate in inducing differentiation of HL60 cells. We propose that Ap4A and Ap3A are physiological antagonists in determination of the cellular status: Ap4A induces apoptosis whereas Ap3A is a co-inductor of differentiation. In both cases, the mechanism of signal transduction remains unknown.

  11. miR-181a and miR-630 regulate cisplatin-induced cancer cell death.

    PubMed

    Galluzzi, Lorenzo; Morselli, Eugenia; Vitale, Ilio; Kepp, Oliver; Senovilla, Laura; Criollo, Alfredo; Servant, Nicolas; Paccard, Caroline; Hupé, Philippe; Robert, Thomas; Ripoche, Hugues; Lazar, Vladimir; Harel-Bellan, Annick; Dessen, Philippe; Barillot, Emmanuel; Kroemer, Guido

    2010-03-01

    MicroRNAs (miRNA) are noncoding RNAs that regulate multiple cellular processes, including proliferation and apoptosis. We used microarray technology to identify miRNAs that were upregulated by non-small cell lung cancer (NSCLC) A549 cells in response to cisplatin (CDDP). The corresponding synthetic miRNA precursors (pre-miRNAs) per se were not lethal when transfected into A549 cells yet affected cell death induction by CDDP, C2-ceramide, cadmium, etoposide, and mitoxantrone in an inducer-specific fashion. Whereas synthetic miRNA inhibitors (anti-miRNAs) targeting miR-181a and miR-630 failed to modulate the response of A549 to CDDP, pre-miR-181a and pre-miR-630 enhanced and reduced CDDP-triggered cell death, respectively. Pre-miR-181a and pre-miR-630 consistently modulated mitochondrial/postmitochondrial steps of the intrinsic pathway of apoptosis, including Bax oligomerization, mitochondrial transmembrane potential dissipation, and the proteolytic maturation of caspase-9 and caspase-3. In addition, pre-miR-630 blocked early manifestations of the DNA damage response, including the phosphorylation of the ataxia-telangiectasia mutated (ATM) kinase and of two ATM substrates, histone H2AX and p53. Pharmacologic and genetic inhibition of p53 corroborated the hypothesis that pre-miR-630 (but not pre-miR-181a) blocks the upstream signaling pathways that are ignited by DNA damage and converge on p53 activation. Pre-miR-630 arrested A549 cells in the G0-G1 phase of the cell cycle, correlating with increased levels of the cell cycle inhibitor p27(Kip1) as well as with reduced proliferation rates and resulting in greatly diminished sensitivity of A549 cells to the late S-G2-M cell cycle arrest mediated by CDDP. Altogether, these results identify miR-181a and miR-630 as novel modulators of the CDDP response in NSCLC.

  12. HMGA2 upregulation mediates Cd-induced migration and invasion in A549 cells and in lung tissues of mice.

    PubMed

    Luo, Huiyuan; Li, Zhiguo; Ge, Hong; Mei, Dan; Zhao, Lian; Jiang, Liping; Geng, Chengyan; Li, Qiujuan; Yao, Xiaofeng; Cao, Jun

    2017-11-01

    Cadmium (Cd) is a toxic metal widely found in a number of environmental matrices, and it induces serious adverse effects in various organs and tissues. In this study, the role of high mobility group A2 (HMGA2) in promoting migration and invasion in Cd-treated A549 cells and lung tissues of mice was investigated. Our findings showed that exposure to Cd (2 μM) for 48 h or subcutaneous injection of Cd daily for 6 weeks significantly enhanced the expression of matrix metalloproteinase-9 (MMP-9), matrix metalloproteinase-2 (MMP-2), phosphorylated focal adhesion kinase (p-FAK), and HMGA2 in A549 cells or lung tissues of mice. In A549 cells, HMGA2 knockdown significantly decreased expression of MMP-9, MMP-2 and p-FAK and inhibited the migration and invasion compared to that of only Cd-treated cultures. Overexpression of HMGA2 in HEK-293T cells increased expression of MMP-9, MMP-2 and p-FAK and enhanced the migration and invasion compared with the empty vector transfection group. In conclusion, upregulation of HMGA2 plays an important role in Cd-enhanced migration and invasion. Suppressing HMGA2 expression might have potential values in prevention of Cd-resulted toxicities. Copyright © 2017 Elsevier B.V. All rights reserved.

  13. Silica nanoparticles and biological dispersants: genotoxic effects on A549 lung epithelial cells

    NASA Astrophysics Data System (ADS)

    Brown, David M.; Varet, Julia; Johnston, Helinor; Chrystie, Alison; Stone, Vicki

    2015-10-01

    Silica nanoparticle exposure could be intentional (e.g. medical application or food) or accidental (e.g. occupational inhalation). On entering the body, particles become coated with specific proteins depending on the route of entry. The ability of silica particles of different size and charge (non-functionalized 50 and 200 nm and aminated 50 and 200 nm) to cause genotoxic effects in A549 lung epithelial cells was investigated. Using the modified comet assay and the micronucleus assay, we examined the effect of suspending the particles in different dispersion media [RPMI or Hanks' balanced salt solution (HBSS), supplemented with bovine serum albumin (BSA), lung lining fluid (LLF) or serum] to determine if this influenced the particle's activity. Particle characterisation suggested that the particles were reasonably well dispersed in the different media, with the exception of aminated 50 nm particles which showed evidence of agglomeration. Plain 50, 200 nm and aminated 50 nm particles caused significant genotoxic effects in the presence of formamidopyrimidine-DNA glycosylase when dispersed in HBSS or LLF. These effects were reduced when the particles were dispersed in BSA and serum. There was no significant micronucleus formation produced by any of the particles when suspended in any of the dispersants. The data suggest that silica particles can produce a significant genotoxic effect according to the comet assay in A549 cells, possibly driven by an oxidative stress-dependent mechanism which may be modified depending on the choice of dispersant employed.

  14. Triptolide inhibits COX-2 expression by regulating mRNA stability in TNF-{alpha}-treated A549 cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Sun, Lixin; Zhang, Shuang; Jiang, Zhenzhou

    2011-12-09

    Highlights: Black-Right-Pointing-Pointer Triptolide inhibited COX-2 expression and the half-life of COX-2 mRNA is decreased. Black-Right-Pointing-Pointer The HuR protein shuttling from nucleus to cytoplasm is inhibited by triptolide. Black-Right-Pointing-Pointer Triptolide inhibited 3 Prime -UTR fluorescence reporter gene activity. Black-Right-Pointing-Pointer COX-2 mRNA binding to HuR is decreased by triptolide in pull-down experiments. -- Abstract: Cyclooxygenase-2 (COX-2) over-expression is frequently associated with human non-small-cell lung cancer (NSCLC) and involved in tumor proliferation, invasion, angiogenesis and resistance to apoptosis. In the present study, the effects of triptolide on COX-2 expression in A549 cells were investigated and triptolide was found to inhibit TNF-{alpha}-induced COX-2 expression.more » In our further studies, it was found that triptolide decreased the half-life of COX-2 mRNA dramatically and that it inhibited 3 Prime -untranslated region (3 Prime -UTR) fluorescence reporter gene activity. Meanwhile, triptolide inhibited the HuR shuttling from nucleus to cytoplasm. After triptolide treatment, decreased COX-2 mRNA in pull-down experiments with anti-HuR antibodies was observed, indicating that the decreased cytoplasmic HuR is responsible for the decreased COX-2 mRNA. Taken together, our results provided evidence for the first time that triptolide inhibited COX-2 expression by COX-2 mRNA stability modulation and post-transcriptional regulation. These results provide a novel mechanism of action for triptolide which may be important in the treatment of lung cancer.« less

  15. Combined treatment with apatinib and docetaxel in A549 xenograft mice and its cellular pharmacokinetic basis.

    PubMed

    Feng, Si-Qi; Wang, Guang-Ji; Zhang, Jing-Wei; Xie, Yuan; Sun, Run-Bin; Fei, Fei; Huang, Jing-Qiu; Wang, Ying; Aa, Ji-Ye; Zhou, Fang

    2018-05-17

    Apatinib, a small-molecule inhibitor of VEGFR-2, has attracted much attention due to its encouraging anticancer activity in third-line clinical treatment for many malignancies, including non-small cell lung cancer (NSCLC). Its usage in second-line therapy with chemotherapeutic drugs is still under exploration. In this study we investigated the antitumor effect of apatinib combined with docetaxel against NSCLC and its cellular pharmacokinetic basis. A549 xenograft nude mice were treated with apatinib (100 mg/kg every day for 20 days) combined with docetaxel (8 mg/kg, ip, every four days for 5 times). Apatinib significantly enhanced the antitumor effect of docetaxel and alleviated docetaxel-induced liver damage as well as decreased serum transaminases (ALT and AST). LC-MS/MS analysis revealed that apatinib treatment significantly increased the docetaxel concentration in tumors (up to 1.77 times) without enhancing the docetaxel concentration in the serum, heart, liver, lung and kidney. Furthermore, apatinib decreased docetaxel-induced upregulation of P-glycoprotein in tumors. The effects of apatinib on the uptake, efflux and subcellular distribution of docetaxel were investigated in A549 and A549/DTX (docetaxel-resistant) cells in vitro. A cellular pharmacokinetic study revealed that apatinib significantly increased cellular/subcellular accumulation (especially in the cytosol) and decreased the efflux of docetaxel in A549/DTX cells through P-gp, while apatinib exerted no significant effect on the cellular pharmacokinetics of docetaxel in A549 cells. Consequently, the IC 50 value of docetaxel in A549/DTX cells was more significantly decreased by apatinib than that in A549 cells. These results demonstrate that apatinib has potential for application in second-line therapy combined with docetaxel for NSCLC patients, especially for docetaxel-resistant or multidrug-resistant patients.

  16. The Pseudomonas aeruginosa exopolysaccharide Psl facilitates surface adherence and NF-kappaB activation in A549 cells.

    PubMed

    Byrd, Matthew S; Pang, Bing; Mishra, Meenu; Swords, W Edward; Wozniak, Daniel J

    2010-06-29

    In order for the opportunistic Gram-negative pathogen Pseudomonas aeruginosa to cause an airway infection, the pathogen interacts with epithelial cells and the overlying mucous layer. We examined the contribution of the biofilm polysaccharide Psl to epithelial cell adherence and the impact of Psl on proinflammatory signaling by flagellin. Psl has been implicated in the initial attachment of P. aeruginosa to biotic and abiotic surfaces, but its direct role in pathogenesis has not been evaluated (L. Ma, K. D. Jackson, R. M. Landry, M. R. Parsek, and D. J. Wozniak, J. Bacteriol. 188:8213-8221, 2006). Using an NF-kappaB luciferase reporter system in the human epithelial cell line A549, we show that both Psl and flagellin are necessary for full activation of NF-kappaB and production of the interleukin 8 (IL-8) chemokine. We demonstrate that Psl does not directly stimulate NF-kappaB activity, but indirectly as a result of increasing contact between bacterial cells and epithelial cells, it facilitates flagellin-mediated proinflammatory signaling. We confirm differential adherence of Psl and/or flagellin mutants by scanning electron microscopy and identify Psl-dependent membrane structures that may participate in adherence. Although we hypothesized that Psl would protect P. aeruginosa from recognition by the epithelial cell line A549, we instead observed a positive role for Psl in flagellin-mediated NF-kappaB activation, likely as a result of increasing contact between bacterial cells and epithelial cells.

  17. The Pseudomonas aeruginosa Exopolysaccharide Psl Facilitates Surface Adherence and NF-κB Activation in A549 Cells

    PubMed Central

    Byrd, Matthew S.; Pang, Bing; Mishra, Meenu; Swords, W. Edward; Wozniak, Daniel J.

    2010-01-01

    In order for the opportunistic Gram-negative pathogen Pseudomonas aeruginosa to cause an airway infection, the pathogen interacts with epithelial cells and the overlying mucous layer. We examined the contribution of the biofilm polysaccharide Psl to epithelial cell adherence and the impact of Psl on proinflammatory signaling by flagellin. Psl has been implicated in the initial attachment of P. aeruginosa to biotic and abiotic surfaces, but its direct role in pathogenesis has not been evaluated (L. Ma, K. D. Jackson, R. M. Landry, M. R. Parsek, and D. J. Wozniak, J. Bacteriol. 188:8213–8221, 2006). Using an NF-κB luciferase reporter system in the human epithelial cell line A549, we show that both Psl and flagellin are necessary for full activation of NF-κB and production of the interleukin 8 (IL-8) chemokine. We demonstrate that Psl does not directly stimulate NF-κB activity, but indirectly as a result of increasing contact between bacterial cells and epithelial cells, it facilitates flagellin-mediated proinflammatory signaling. We confirm differential adherence of Psl and/or flagellin mutants by scanning electron microscopy and identify Psl-dependent membrane structures that may participate in adherence. Although we hypothesized that Psl would protect P. aeruginosa from recognition by the epithelial cell line A549, we instead observed a positive role for Psl in flagellin-mediated NF-κB activation, likely as a result of increasing contact between bacterial cells and epithelial cells. PMID:20802825

  18. Low-dose carbon-based nanoparticle-induced effects in A549 lung cells determined by biospectroscopy are associated with increases in genomic methylation

    NASA Astrophysics Data System (ADS)

    Li, Junyi; Tian, Meiping; Cui, Li; Dwyer, John; Fullwood, Nigel J.; Shen, Heqing; Martin, Francis L.

    2016-02-01

    Nanotechnology has introduced many manufactured carbon-based nanoparticles (CNPs) into our environment, generating a debate into their risks and benefits. Numerous nanotoxicology investigations have been carried, and nanoparticle-induced toxic effects have been reported. However, there remain gaps in our knowledge, primarily regarding mechanism. Herein, we assessed the global alterations induced by CNPs in A549 lung cells using biospectroscopy techniques, including attenuated total reflection Fourier-transform infrared (ATR-FTIR) spectroscopy and surface-enhanced Raman spectroscopy (SERS). A549 cells were treated with fullerene (C60), long or short multi-walled carbon nanotubes, or single-walled carbon nanotubes at concentrations of 0.1 mg/L, 0.01 mg/L and 0.001 mg/L. Exposed cells were then analysed by ATR-FTIR spectroscopy and SERS. Spectra were pre-processed via computational analysis, and information on biochemical alterations in exposed cells were identified. Additionally, global DNA methylation levels in cells exposed to CNPs at 0.1 mg/L were determined using HPLC-MS and genetic regulators (for DNA methylation) were checked by quantitative real-time RT-PCR. It was found that CNPs exert marked effects in A549 cells and also contribute to increases in global DNA methylation. For the first time, this study highlights that real-world levels of nanoparticles can alter the methylome of exposed cells; this could have enormous implications for their regulatory assessment.

  19. Cyclin-dependent kinases regulate apoptosis of intestinal epithelial cells

    PubMed Central

    Bhattacharya, Sujoy; Ray, Ramesh M.; Johnson, Leonard R.

    2014-01-01

    Homeostasis of the gastrointestinal epithelium is dependent upon a balance between cell proliferation and apoptosis. Cyclin-dependent kinases (Cdks) are well known for their role in cell proliferation. Previous studies from our group have shown that polyamine-depletion of intestinal epithelial cells (IEC-6) decreases cyclin-dependent kinase 2 (Cdk2) activity, increases p53 and p21Cip1 protein levels, induces G1 arrest, and protects cells from camptothecin (CPT)-induced apoptosis. Although emerging evidence suggests that members of the Cdk family are involved in the regulation of apoptosis, their roles directing apoptosis of IEC-6 cells are not known. In this study, we report that inhibition of Cdk1, 2, and 9 (with the broad range Cdk inhibitor, AZD5438) in proliferating IEC-6 cells triggered DNA damage, activated p53 signaling, inhibited proliferation, and induced apoptosis. By contrast, inhibition of Cdk2 (with NU6140) increased p53 protein and activity, inhibited proliferation, but had no effect on apoptosis. Notably, AZD5438 sensitized, whereas, NU6140 rescued proliferating IEC-6 cells from CPT-induced apoptosis. However, in colon carcinoma (Caco2) cells with mutant p53, treatment with either AZD5438 or NU6140 blocked proliferation, albeit more robustly with AZD5438. Both Cdk inhibitors induced apoptosis in Caco2 cells in a p53-independent manner. In serum starved quiescent IEC-6 cells, both AZD5438 and NU6140 decreased TNF- /CPT-induced activation of p53 and, consequently, rescued cells from apoptosis, indicating that sustained Cdk activity is required for apoptosis of quiescent cells. Furthermore, AZD5438 partially reversed the protective effect of polyamine depletion whereas NU6140 had no effect. Together, these results demonstrate that Cdks possess opposing roles in the control of apoptosis in quiescent and proliferating cells. In addition, Cdk inhibitors uncouple proliferation from apoptosis in a p53-dependent manner. PMID:24242917

  20. Nrf2 mediates redox adaptation in NOX4-overexpressed non-small cell lung cancer cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Wu, Qipeng; Yao, Bei; Li, Ning

    The redox adaptation mechanisms in cancer cells are very complex and remain largely unclear. Our previous studies have confirmed that NADPH oxidase 4 (NOX4) is abundantly expressed in non-small cell lung cancer (NSCLC) and confers apoptosis resistance on NSCLC cells. However, the comprehensive mechanisms for NOX4-mediated oxidative resistance of cancer cells remain still undentified. The present study found that NOX4-derived H{sub 2}O{sub 2} enhanced the nuclear factor erythroid 2-related factor 2 (Nrf2) stability via disruption of redox-dependent proteasomal degradation and stimulated its activity through activation of PI3K signaling. Specifically, the results showed that ectopic NOX4 expression did not induce apoptosismore » of A549 cells; however, inhibition of Nrf2 resulted in obvious apoptotic death of NOX4-overexpressed A549 cells, accompanied by a significant increase in H{sub 2}O{sub 2} level and decrease in GSH content. Besides, inhibition of Nrf2 could suppress cell growth and efficiently reverse the enhancement effect of NOX4 on cell growth. The in vivo data confirmed that inhibition of Nrf2 could interfere apoptosis resistance in NOX4-overexpressed A549 tumors and led to cell growth inhibition. In conclusion, these results reveal that Nrf2 is critically involved in redox adaptation regulation in NOX4-overexpressed NSCLC cells. Therefore, NOX4 and Nrf2 may be promising combination targets against malignant progression of NSCLC. - Highlights: • NOX4-derived H{sub 2}O{sub 2} upregulates Nrf2 expression and activity in NSCLC. • Nrf2 confers apoptosis resistance in NOX4-overexpressed NSCLC cells. • Inhibition of Nrf2 reverses the enhancement effect of NOX4 on cell growth.« less

  1. MicroRNA-1 promotes apoptosis of hepatocarcinoma cells by targeting apoptosis inhibitor-5 (API-5).

    PubMed

    Li, Dong; Liu, Yu; Li, Hua; Peng, Jing-Jing; Tan, Yan; Zou, Qiang; Song, Xiao-Feng; Du, Min; Yang, Zheng-Hui; Tan, Yong; Zhou, Jin-Jun; Xu, Tao; Fu, Zeng-Qiang; Feng, Jian-Qiong; Cheng, Peng; chen, Tao; Wei, Dong; Su, Xiao-Mei; Liu, Huan-Yi; Qi, Zhong-Chun; Tang, Li-Jun; Wang, Tao; Guo, Xin; Hu, Yong-He; Zhang, Tao

    2015-01-02

    Although microRNA-1 (miR-1) is a known liver cancer suppressor, the role of miR-1 in apoptosis of hepatoma cells has remained largely unknown. Our study shows that ectopic miR-1 overexpression induced apoptosis of liver hepatocellular carcinoma (HepG2) cells. Apoptosis inhibitor 5 (API-5) was found to be a potential regulator of miR-1 induced apoptosis, using a bioinformatics approach. Furthermore, an inverse relationship between miR-1 and API-5 expression was observed in human liver cancer tissues and adjacent normal liver tissues. Negative regulation of API-5 expression by miR-1 was demonstrated to promote apoptosis of HepG2 cells. Our study provides a novel regulatory mechanism of miR-1 in the apoptosis of hepatoma cells. Copyright © 2014 Federation of European Biochemical Societies. Published by Elsevier B.V. All rights reserved.

  2. Apoptosis in capillary endothelial cells in ageing skeletal muscle

    PubMed Central

    Wang, Huijuan; Listrat, Anne; Meunier, Bruno; Gueugneau, Marine; Coudy-Gandilhon, Cécile; Combaret, Lydie; Taillandier, Daniel; Polge, Cécile; Attaix, Didier; Lethias, Claire; Lee, Kijoon; Goh, Kheng Lim; Béchet, Daniel

    2014-01-01

    The age-related loss of skeletal muscle mass and function (sarcopenia) is a consistent hallmark of ageing. Apoptosis plays an important role in muscle atrophy, and the intent of this study was to specify whether apoptosis is restricted to myofibre nuclei (myonuclei) or occurs in satellite cells or stromal cells of extracellular matrix (ECM). Sarcopenia in mouse gastrocnemius muscle was characterized by myofibre atrophy, oxidative type grouping, delocalization of myonuclei and ECM fibrosis. Terminal deoxynucleotidyl transferase-mediated dUTP nick end-labelling (TUNEL) indicated a sharp rise in apoptosis during ageing. TUNEL coupled with immunostaining for dystrophin, paired box protein-7 (Pax7) or laminin-2α, respectively, was used to identify apoptosis in myonuclei, satellite cells and stromal cells. In adult muscle, apoptosis was not detected in myofibres, but was restricted to stromal cells. Moreover, the age-related rise in apoptotic nuclei was essentially due to stromal cells. Myofibre-associated apoptosis nevertheless occurred in old muscle, but represented < 20% of the total muscle apoptosis. Specifically, apoptosis in old muscle affected a small proportion (0.8%) of the myonuclei, but a large part (46%) of the Pax7+ satellite cells. TUNEL coupled with CD31 immunostaining further attributed stromal apoptosis to capillary endothelial cells. Age-dependent rise in apoptotic capillary endothelial cells was concomitant with altered levels of key angiogenic regulators, perlecan and a perlecan domain V (endorepellin) proteolytic product. Collectively, our results indicate that sarcopenia is associated with apoptosis of satellite cells and impairment of capillary functions, which is likely to contribute to the decline in muscle mass and functionality during ageing. PMID:24245531

  3. Fisetin induces apoptosis through mitochondrial apoptosis pathway in human uveal melanoma cells.

    PubMed

    Wang, Kai; Hu, Dan-Ning; Lin, Hui-Wen; Yang, Wei-En; Hsieh, Yi-Hsien; Chien, Hsiang-Wen; Yang, Shun-Fa

    2018-05-01

    Fisetin, a diatery flavonoid, been reported that possess anticancer effects in various cancers. The purpose of the study was to investigate the antitumor effects of fisetin in cultured uveal melanoma cell lines and compared with normal retinal pigment epithelial (RPE) cells. MTT assay was used for evaluating cytotoxic effects of fisetin. Flow cytometry study was used for the determination of apoptosis. JC-1 fluorescent reader was used to determine mitochondrial transmembrane potential changes. The results shown that fisetin dose-dependently decreased the cell viability of uveal melanoma cells but not influenced the cell viability of RPE cells. Apoptosis of uveal melanoma cells was induced by fisetin efficiently. Fisetin inhibited antiapoptotic Bcl-2 family proteins and damaged the mitochondrial transmembrane potential. The levels of proapoptotic Bcl-2 proteins, cytochrome c, and various caspase activities were increased by fisetin. In conclusion, fisetin induces apoptosis of uveal melanoma cells selectively and may be a promising agent to be explored for the treatment of uveal melanoma. © 2018 Wiley Periodicals, Inc.

  4. Cytotoxic constituents from the mangrove endophytic Pestalotiopsis sp. induce G0/G1 cell cycle arrest and apoptosis in human cancer cells.

    PubMed

    Zhou, Jing; Li, Gang; Deng, Qin; Zheng, Dongyao; Yang, Xiaobo; Xu, Jing

    2017-10-31

    Chemical examination of Chinese mangrove Rhizophora mucronata endophytic Pestalotiopsis sp., yielded  11 known metabolites with various structure types, including demethylincisterol A 3 (1), dankasterone B (2), (22E, 24R)-ergosta-7,9(11), 22-triene-3β, 5α, 6α-triol (3), ergosta-5,7,22-trien-3-ol (4), 5, 8-epidioxy-5, 8-ergosta-6, 22E-dien-3-ol (5), stigmastan-3-one (6), stigmast-4-en-3-one (7), stigmast-4-en-6 -ol-3-one (8), flufuran (9), (2-cis, 4-trans)-abscisic acid (10), similanpyrone B (11). Their structures were unambiguously elucidated on the basis of extensive NMR spectroscopic and mass spectrometric analyses. Compounds 1, 4, 6-9 showed significant in vitro cytotoxicity against the human cancer cell lines Hela, A549 and HepG, of which compound 1 was the most potential with IC 50 values reaching nM degree ranging from 0.17 to 14.16 nM. Flow cytometric investigation demonstrated that compound 1 mainly inhibited cell cycle at G 0 /G 1 phase in a dose-dependent manner with a significant induction of apoptosis on the three tested cell lines. The involvement of the mitochondria in compound 1 induced apoptosis was investigated using MMP. We suggested that R. mucronata endophytic Pestalotiopsis sp. contained a potential anticancer compound demethylincisterol A 3 .

  5. 5-(Furan-2-yl)-4-(3,4,5-trimethoxyphenyl)-3H-1,2-dithiol-3-one oxime (6f), a new synthetic compound, causes human fibrosarcoma HT-1080 cell apoptosis by disrupting tubulin polymerisation and inducing G2/M arrest.

    PubMed

    Zuo, Daiying; Pang, Lili; Shen, Jiwei; Guan, Qi; Bai, Zhaoshi; Zhang, Huijuan; Li, Yao; Lu, Guodong; Zhang, Weige; Wu, Yingliang

    2017-06-01

    In the current study, we synthesized a series of new compounds targeting tubulin and tested their anti-proliferative activities. Among these new synthetic com-pounds, 5-(furan-2-yl)-4-(3,4,5-trimethoxyphenyl)-3H-1,2-dithiol-3-one oxime (6f) exhibited significant anti-proliferative activity against different human cancer cell lines including human gastric adenocarcinoma SGC-7901, human non-small cell lung cancer A549, and human fibrosarcoma HT-1080. As a result, 6f was selected to further test the sensitivity to different cancer cell lines including human cervical cancer cell line HeLa, human breast cancer cell line MCF-7, non-small cell lung cancer cell line A549, human liver carcinoma cell line HepG-2, human oral squamous cell carcinoma cell lines KB, SGC-7901 and HT-1080. Among these cell lines, HT-1080 and HeLa are the most sensitive. Therefore, HT-1080 was selected to further explore the properties of anti-proliferative activity and the underlying mechanisms. Our data proved that 6f exhibited strong anti-proliferative effects against HT-1080 cells in a time- and dose-dependent manner. We showed that the growth inhibitory effect of 6f in HT-1080 cells was related with microtubule depolymerisation. Molecular docking studies revealed that 6f interacted and bound efficiently with the colchicine-binding site of tubulin. In addition, 6f treatment induced G2/M cell cycle arrest dose-dependently and subsequently induced cell apoptosis. Western blot study indicated that upregulation of cyclin B1 and p-cdc2 was related with G2/M arrest. 6f-induced cell apoptosis was associated with both mitochondrial and death receptor pathway. In conclusion, our data showed that 6f, among the newly synthetic compounds, exhibited highest anti-proliferative activity by disrupting the microtubule polymerisation, causing G2/M arrest and subsequently inducing cell apoptosis in HT-1080 cells. Hence, 6f is a promising microtubule depolymerising agent for the treatment of various cancers

  6. A novel apoptosis correlated molecule: expression and characterization of protein Latcripin-1 from Lentinula edodes C(91-3).

    PubMed

    Liu, Ben; Zhong, Mintao; Lun, Yongzhi; Wang, Xiaoli; Sun, Wenchang; Li, Xingyun; Ning, Anhong; Cao, Jing; Zhang, Wei; Liu, Lei; Huang, Min

    2012-01-01

    An apoptosis correlated molecule-protein Latcripin-1 of Lentinula edodes C(91-3)-was expressed and characterized in Pichia pastoris GS115. The total RNA was obtained from Lentinula edodes C(91-3). According to the transcriptome, the full-length gene of Latcripin-1 was isolated with 3'-Full Rapid Amplification of cDNA Ends (RACE) and 5'-Full RACE methods. The full-length gene was inserted into the secretory expression vector pPIC9K. The protein Latcripin-1 was expressed in Pichia pastoris GS115 and analyzed by Sodium Dodecylsulfonate Polyacrylate Gel Electrophoresis (SDS-PAGE) and Western blot. The Western blot showed that the protein was expressed successfully. The biological function of protein Latcripin-1 on A549 cells was studied with flow cytometry and the 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyl-tetrazolium Bromide (MTT) method. The toxic effect of protein Latcripin-1 was detected with the MTT method by co-culturing the characterized protein with chick embryo fibroblasts. The MTT assay results showed that there was a great difference between protein Latcripin-1 groups and the control group (p < 0.05). There was no toxic effect of the characterized protein on chick embryo fibroblasts. The flow cytometry showed that there was a significant difference between the protein groups of interest and the control group according to apoptosis function (p < 0.05). At the same time, cell ultrastructure observed by transmission electron microscopy supported the results of flow cytometry. The work demonstrates that protein Latcripin-1 can induce apoptosis of human lung cancer cells A549 and brings new insights into and advantages to finding anti-tumor proteins.

  7. Ameliorative Effects of Dimetylthiourea and N-Acetylcysteine on Nanoparticles Induced Cyto-Genotoxicity in Human Lung Cancer Cells-A549

    PubMed Central

    Srivastava, Ritesh Kumar; Rahman, Qamar; Kashyap, Mahendra Pratap; Lohani, Mohtashim; Pant, Aditya Bhushan

    2011-01-01

    We study the ameliorative potential of dimetylthiourea (DMTU), an OH• radical trapper and N-acetylcysteine (NAC), a glutathione precursor/H2O2 scavenger against titanium dioxide nanoparticles (TiO2-NPs) and multi-walled carbon nanotubes (MWCNTs) induced cyto-genotoxicity in cultured human lung cancer cells-A549. Cytogenotoxicity was induced by exposing the cells to selected concentrations (10 and 50 µg/ml) of either of TiO2-NPs or MWCNTs for 24 h. Anti-cytogenotoxicity effects of DMTU and NAC were studied in two groups, i.e., treatment of 30 minutes prior to toxic insult (short term exposure), while the other group received DMTU and NAC treatment during nanoparticles exposure, i.e., 24 h (long term exposure). Investigations were carried out for cell viability, generation of reactive oxygen species (ROS), micronuclei (MN), and expression of markers of oxidative stress (HSP27, CYP2E1), genotoxicity (P53) and CYP2E1 dependent n- nitrosodimethylamine-demethylase (NDMA-d) activity. In general, the treatment of both DMTU and NAC was found to be effective significantly against TiO2-NPs and MWCNTs induced cytogenotoxicity in A549 cells. Long-term treatment of DMTU and NAC during toxic insults has shown better prevention than short-term pretreatment. Although, cells responded significantly to both DMTU and NAC, but responses were chemical specific. In part, TiO2-NPs induced toxic responses were mediated through OH• radicals generation and reduction in the antioxidant defense system. While in the case of MWCNTs, adverse effects were primarily due to altering/hampering the enzymatic antioxidant system. Data indicate the applicability of human lung cancer cells-A549 as a pre-screening tool to identify the target specific prophylactic and therapeutic potential of drugs candidate molecules against nanoparticles induced cellular damages. PMID:21980536

  8. Nanoparticles of Selaginella doederleinii leaf extract inhibit human lung cancer cells A549

    NASA Astrophysics Data System (ADS)

    Syaefudin; Juniarti, A.; Rosiyana, L.; Setyani, A.; Khodijah, S.

    2016-01-01

    The aim of the present study is to evaluate cytotoxicity effect of nanoparticles of Selaginella doederleinii (S. doederleinii) leaves extract. S. doederleinii was extracted by maceration method using 70%(v/v) ethanol as solvent. Phytochemical content was analyzed qualitatively by using Harborne and Thin Layer Chromatography (TLC) methods. Nanoparticle extract was prepared by ionic gelation using chitosan as encapsulant agent. Anticancer activity was performed by using 3-(4, 5-dimethylthiazol-2-yl)-2, 5-diphenyltetrazolium bromide (MTT) assay. The results showed that S. doederleinii contains of flavonoids. Nanoparticle of S. doederleinii leaves extract greatly inhibited A549 cells growth (cancer cells), with IC50 of 3% or 1020 μg/ml. These nanoparticles extract also inhibited the growth of Chang cells (normal cells), with IC50 of 4% or 1442 μg/ml. The effective concentration of nanoparticles extract which inhibits cancer cells without harming the normal cells is 0.5% or 167 μg/ml. Further studies are needed to obtain the concentration of nanoparticles extract which can selectively suppress cancer cells.

  9. Caspase dependent apoptotic inhibition of melanoma and lung cancer cells by tropical Rubus extracts.

    PubMed

    George, Blassan Plackal Adimuriyil; Abrahamse, Heidi; Hemmaragala, Nanjundaswamy M

    2016-05-01

    Rubus fairholmianus Gard. inhibits human melanoma (A375) and lung cancer (A549) cell growth by the caspase dependent apoptotic pathway. Herbal products have a long history of clinical use and acceptance. They are freely available natural compounds that can be safely used to prevent various ailments. The plants and plant derived products became the basis of traditional medicine system throughout the world for thousands of years. The effects of R. fairholmianus root acetone extract (RFRA) on the proliferation of A375 and A549 cells was examined in this study. RFRA led to a decrease in cell viability, proliferation and an increase in cytotoxicity in a dose dependent manner when compared with control and normal skin fibroblast cells (WS1). The morphology of treated cells supported apoptotic cell death. Annexin V/propidium iodide staining indicated that RFRA induced apoptosis in A375 and A549 cells and the percentages of early and late apoptotic populations significantly increased. Moreover, the apoptotic inducing ability of RFRA when analysing effector caspase 3/7 activity, indicated a marked increase in treated cells. In summary, we have shown the anticancer effects of RFRA in A375 and A549 cancer cells via induction of caspase dependent apoptosis in vitro. The extract is more effective against melanoma; which may suggest the usefulness of RFRA-based anticancer therapies. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  10. Identification of a Novel Pathway That Selectively Modulates Apoptosis of Breast Cancer Cells

    PubMed Central

    Tinnikov, Alexander A.; Yeung, Kay T.; Das, Sharmistha; Samuels, Herbert H.

    2014-01-01

    Expression of the nuclear receptor interacting factor 3 (NRIF3) coregulator in a wide variety of breast cancer cells selectively leads to rapid caspase-2–dependent apoptotic cell death. A novel death domain (DD1) was mapped to a 30– amino acid region of NRIF3. Because the cytotoxicity of NRIF3 and DD1 seems to be cell type–specific, these studies suggest that breast cancer cells contain a novel “death switch” that can be specifically modulated by NRIF3 or DD1. Using an MCF-7 cell cDNA library in a yeast two-hybrid screen, we cloned a factor that mediates apoptosis by DD1 and refer to this factor as DD1-interacting factor-1 (DIF-1). DIF-1 is a transcriptional repressor that mediates its effect through SirT1, and this repression is attenuated by the binding of NRIF3/DD1. DIF-1 expression rescues breast cancer cells from NRIF3/DD1-induced apoptosis. Small interfering RNA (siRNA) knockdown of DIF-1 selectively leads to apoptosis of breast cancer cells, further suggesting that DIF-1 plays a key role in NRIF3/DD1-mediated apoptosis. A protein kinase A inhibitor (H89) also elicits apoptosis of breast cancer cells but not of the other cell types examined, and DIF-1 also protects these cells from H89-mediated apoptosis. In addition, H89 incubation results in a rapid increase in NRIF3 levels and siRNA knockdown of NRIF3 protects breast cancer cells from H89-mediated apoptosis. Our results indicate that DIF-1 plays a key role in breast cancer cell survival and further characterizing this pathway may provide important insights into developing novel therapies to selec tively target breast cancer cells for apoptosis. PMID:19190336

  11. Cell renewal and apoptosis in macrostomum sp. [Lignano].

    PubMed

    Nimeth, K; Ladurner, P; Gschwentner, R; Salvenmoser, W; Rieger, R

    2002-01-01

    In platyhelminths, all cell renewal is accomplished by totipotent stem cells (neoblasts). Tissue maintenance is achieved in a balance between cell proliferation and apoptosis. It is known that in Macrostomum sp. the epidermis undergoes extensive cell renewal. Here we show that parenchymal cells also exhibit a high rate of cell turnover. We demonstrate cell renewal using continuous 5'bromo-2-deoxyuridine (BrdU) exposure. About one-third of all cells are replaced after 14 days. The high level of replacement requires an equivalent removal of cells by apoptosis. Cell death is characterized using a combination of three methods: (1). terminal deoxynucleotidyl transferase-mediated dUTP nick-end labelling (TUNEL), (2). specific binding of phosphatidyl-serine to fluorescent-labelled annexin V and (3). identification of apoptotic stages by ultrastructure. The number of cells observed in apoptosis is insufficient to explain the homeostasis of tissues in Macrostomum. Apoptosis-independent mechanisms may play an additional role in tissue dynamics.

  12. Up-Regulation of Pro-Inflammatory Cytokines and Chemokine Production in Avian Influenza H9N2 Virus-Infected Human Lung Epithelial Cell Line (A549).

    PubMed

    Farzin, Hamidreza; Toroghi, Reza; Haghparast, Alireza

    2016-01-01

    Influenza H9N2 virus mostly infects avian species but poses a potential health risk to humans. Little is known about the mammalian host immune responses to H9N2 virus. To obtain insight into the innate immune responses of human lung epithelial cells to the avian H9N2 virus, the expressions of pro-inflammatory cytokines and chemokine in the human airway epithelial cells infected with avian H9N2 virus were examined by real-time quantitative reverse transcription polymerase chain reaction (qRT-PCR) and enzyme-linked immunosorbent assay (ELISA). H9N2 virus was able to cultivate in the human lung epithelial cell line (A549) and stimulate production of pro-inflammatory cytokines (IL-1β, IL-6) and chemokine (IL-8). Expressions of cytokine genes were up-regulated to a significantly higher level for IL-1β (p < 0.01), IL-6 (p < 0.01 after 12 hours and p < 0.05 after 24 hours) and IL-8 (p < 0.01 after 12 hours and p < 0.001 after 24 hours) in virus-cultured A549 cells as compared with non-virus-cultured cells. The amount of IL-6 and IL-1β proteins secreted into the culture medium was also increased after virus culture infection of A549 cell line compared to non-virus-cultured A549 cells and were significant in both IL-1β (p < 0.05 in 18 hours and p < 0.001 in 24-48 hours harvested supernatant) and IL-6 (p < 0.001). Silencing the p65 component of NF-κB in A549 cells suppressed the stimulatory effects of influenza virus on secretion of pro-inflammatory cytokines and chemokine. The findings in this study will broaden our understanding of host innate immune mechanisms and the pathogenesis of H9N2 influenza viruses in human respiratory epithelium.

  13. Sulfamic acid promoted one-pot synthesis of phenanthrene fused-dihydrodibenzo-quinolinones: Anticancer activity, tubulin polymerization inhibition and apoptosis inducing studies.

    PubMed

    Kumar, Niggula Praveen; Thatikonda, Sowjanya; Tokala, Ramya; Kumari, S Sujana; Lakshmi, Uppu Jaya; Godugu, Chandraiah; Shankaraiah, Nagula; Kamal, Ahmed

    2018-05-01

    A facile one-pot method for the synthesis of new phenanthrene fused-dihydrodibenzo-quinolinone derivatives has been successfully accomplished by employing sulfamic acid as catalyst. These new compounds were evaluated for their in vitro cytotoxic potential against human lung (A549), prostate (PC-3 and DU145), breast (MCF-7) and colon (HT-29 and HCT-116) cancer cell lines. Among all the tested compounds, one of the derivatives 8p showed good anti-proliferative activity against A549 lung cancer cell line with an IC 50 of 3.17 ± 0.52 µM. Flow cytometric analyses revealed that compound 8p arrested both Sub G1 and G2/M phases of cell cycle in a dose dependent manner. The compound 8p also displayed significant inhibition of tubulin polymerization and disruption of microtubule network (IC 50 of 5.15 ± 0.15 µM). Molecular docking studies revealed that compound 8p efficiently interacted with critical amino acid Cys241 of the α/β-tubulin by a hydrogen bond (SH…O = 2.4 Å). Further, the effect of 8p on cell viability was also studied by AO/EB, DCFDA and DAPI staining. The apoptotic characteristic features revealed that 8p inhibited cell proliferation effectively through apoptosis by inducing the ROS generation. Analysis of mitochondrial membrane potential through JC-1 staining and annexin V binding assay indicated the extent of apoptosis in A549 cancer cells. Copyright © 2018 Elsevier Ltd. All rights reserved.

  14. The interaction between NOLC1 and IAV NS1 protein promotes host cell apoptosis and reduces virus replication.

    PubMed

    Zhu, Chunyu; Zheng, Fangliang; Zhu, Junfeng; Liu, Meichen; Liu, Na; Li, Xue; Zhang, Li; Deng, Zaidong; Zhao, Qi; Liu, Hongsheng

    2017-11-07

    NS1 of the influenza virus plays an important role in the infection ability of the influenza virus. Our previous research found that NS1 protein interacts with the NOLC1 protein of host cells, however, the function of the interaction is unknown. In the present study, the role of the interaction between the two proteins in infection was further studied. Several analyses, including the use of a pull-down assay, Co-IP, western blot analysis, overexpression, RNAi, flow cytometry, etc., were used to demonstrate that the NS1 protein of H3N2 influenza virus interacts with host protein NOLC1 and reduces the quantity of NOLC1. The interaction also promotes apoptosis in A549 host cells, while the suppression of NOLC1 protein reduces the proliferation of the H3N2 virus. Based on these data, it was concluded that during the process of infection, NS1 protein interacts with NOLC1 protein, reducing the level of NOLC1, and that the interaction between the two proteins promotes apoptosis of host cells, thus reducing the proliferation of the virus. These findings provide new information on the biological function of the interaction between NS1 and NOLC1.

  15. Curcumin sensitizes human lung cancer cells to apoptosis and metastasis synergistically combined with carboplatin.

    PubMed

    Kang, Ji Ho; Kang, Hye Seon; Kim, In Kyoung; Lee, Hwa Young; Ha, Jick Hwan; Yeo, Chang Dong; Kang, Hyun Hui; Moon, Hwa Sik; Lee, Sang Haak

    2015-11-01

    Although carboplatin is one of the standard chemotherapeutic agents for non-small cell lung cancer (NSCLC), it has limited therapeutic efficacy due to activation of a survival signaling pathway and the induction of multidrug resistance. Curcumin, a natural compound isolated from the plant Curcuma longa, is known to sensitize tumors to different chemotherapeutic agents. The aim of this study is to evaluate whether curcumin can chemosensitize lung cancer cells to carboplatin and to analyze the signaling pathway underlying this synergism. We investigated the synergistic effect of both agents on cell proliferation, apoptosis, invasion, migration, and expression of related signaling proteins using the human NSCLC cell line, A549. A549 cell was treated with different concentrations of curcumin and carboplatin alone and in combination. Combined treatment with curcumin and carboplatin inhibited tumor cell growth, migration, and invasion compared with either drug alone. Matrix metalloproteinase (MMP)-2 and MMP-9 were more efficiently downregulated by co-treatment than by each treatment alone. mRNA and protein expression of caspase-3 and caspase-9 and proapoptotic genes was increased in cells treated with a combination of curcumin and carboplatin, whereas expression of the antiapoptotic Bcl-2 gene was suppressed. Co-treatment of both agents substantially suppressed NF-κB activation and increased expression of p53. Phosphorylation of Akt, a protein upstream of NF-κB, was reduced, resulting in inhibition of the degradation of inhibitor of κB(IκBα), whereas the activity of extracellular signal-regulated kinase (ERK1/2) was enhanced. Our study demonstrated that the synergistic antitumor activity of curcumin combined with carboplatin is mediated by multiple mechanisms involving suppression of NF-κB via inhibition of the Akt/IKKα pathway and enhanced ERK1/2 activity. Based on this mechanism, curcumin has potential as a chemosensitizer for carboplatin in the treatment of

  16. Miniature Dielectric Barrier Discharge Nonthermal Plasma Induces Apoptosis in Lung Cancer Cells and Inhibits Cell Migration.

    PubMed

    Karki, Surya B; Yildirim-Ayan, Eda; Eisenmann, Kathryn M; Ayan, Halim

    2017-01-01

    Traditional cancer treatments like radiotherapy and chemotherapy have drawbacks and are not selective for killing only cancer cells. Nonthermal atmospheric pressure plasmas with dielectric barrier discharge (DBD) can be applied to living cells and tissues and have emerged as novel tools for localized cancer therapy. The purpose of this study was to investigate the different effects caused by miniature DBD (mDBD) plasma to A549 lung cancer cells. In this study, A549 lung cancer cells cultured in 12 well plates were treated with mDBD plasma for specified treatment times to assess the changes in the size of the area of cell detachment, the viability of attached or detached cells, and cell migration. Furthermore, we investigated an innovative mDBD plasma-based therapy for localized treatment of lung cancer cells through apoptotic induction. Our results indicate that plasma treatment for 120 sec causes apoptotic cell death in 35.8% of cells, while mDBD plasma treatment for 60 sec, 30 sec, or 15 sec causes apoptotic cell death in 20.5%, 14.1%, and 6.3% of the cell population, respectively. Additionally, we observed reduced A549 cell migration in response to mDBD plasma treatment. Thus, mDBD plasma system can be a viable platform for localized lung cancer therapy.

  17. Apoptosis-Dependent and Apoptosis-Independent Functions of Bim in Prostate Cancer Cells

    DTIC Science & Technology

    2005-03-01

    Independent Functions of Bim in Prostate Cancer Cells PRINCIPAL INVESTIGATOR: Dr. Dean Tang...SUBTITLE 5a. CONTRACT NUMBER Apoptosis-Dependent and Apoptosis-Independent Functions of Bim in Prostate Cancer Cells 5b. GRANT NUMBER DAMD17-03-1...Unlimited 13. SUPPLEMENTARY NOTES Original contains colored plates: ALL DTIC reproductions will be in black and white. 14. ABSTRACT

  18. Airborne particulate matter in vitro exposure induces cytoskeleton remodeling through activation of the ROCK-MYPT1-MLC pathway in A549 epithelial lung cells.

    PubMed

    Chirino, Yolanda I; García-Cuellar, Claudia María; García-García, Carlos; Soto-Reyes, Ernesto; Osornio-Vargas, Álvaro Román; Herrera, Luis A; López-Saavedra, Alejandro; Miranda, Javier; Quintana-Belmares, Raúl; Pérez, Irma Rosas; Sánchez-Pérez, Yesennia

    2017-04-15

    Airborne particulate matter with an aerodynamic diameter ≤10μm (PM 10 ) is considered a risk factor for the development of lung cancer. Little is known about the cellular mechanisms by which PM 10 is associated with cancer, but there is evidence that its exposure can lead to an acquired invasive phenotype, apoptosis evasion, inflammasome activation, and cytoskeleton remodeling in lung epithelial cells. Cytoskeleton remodeling occurs through actin stress fiber formation, which is partially regulated through ROCK kinase activation, we aimed to investigate if this protein was activated in response to PM 10 exposure in A549 lung epithelial cells. Results showed that 10μg/cm 2 of PM 10 had no influence on cell viability but increased actin stress fibers, cytoplasmic ROCK expression, and phosphorylation of myosin phosphatase-targeting 1 (MYPT1) and myosin light chain (MLC) proteins, which are targeted by ROCK. The inhibition of ROCK prevented actin stress fiber formation and the phosphorylation of MYPT1 and MLC, suggesting that PM 10 activated the ROCK-MYPT1-MLC pathway in lung epithelial cells. The activation of ROCK1 has been involved in the acquisition of malignant phenotypes, and its induction by PM 10 exposure could contribute to the understanding of PM 10 as a risk factor for cancer development through the mechanisms associated with invasive phenotype. Copyright © 2017 Elsevier B.V. All rights reserved.

  19. Up-regulation of 5-lipoxygenase by inhibition of cathepsin G enhances TRAIL-induced apoptosis through down-regulation of survivin

    PubMed Central

    Woo, Seon Min; Min, Kyoung-Jin; Seo, Seung Un; Kim, Shin; Park, Jong-Wook; Song, Dae Kyu; Lee, Hyun-Shik; Kim, Sang Hyun; Kwon, Taeg Kyu

    2017-01-01

    Cathepsin G is a serine protease secreted from activated neutrophils, it has important roles in inflammation and immune response. Moreover, cathepsin G promotes tumor cell-cell adhesion and migration in cancer cells. In this study, we investigated whether inhibition of cathepsin G could sensitize TRAIL-mediated apoptosis in cancer cells. An inhibitor of cathepsin G [Cathepsin G inhibitor I (Cat GI); CAS 429676-93-7] markedly induced TRAIL-mediated apoptosis in human renal carcinoma (Caki, ACHN, and A498), lung cancer (A549) and cervical cancer (Hela) cells. In contrast, combined treatment with Cat GI and TRAIL had no effect on apoptosis in normal cells [mesangial cell (MC) and human skin fibroblast (HSF)]. Cat GI induced down-regulation of survivin expression at the post-translational level, and overexpression of survivin markedly blocked apoptosis induced by combined treatment with Cat GI plus TRAIL. Interestingly, Cat GI induced down-regulation of survivin via 5-lipoxygenase (5-LOX)-mediated reactive oxygen species (ROS) production. Inhibition of 5-LOX by gene silencing (siRNA) or a pharmacological inhibitor of 5-LOX (zileuton) markedly attenuated combined treatment-induced apoptosis. Taken together, our results indicate that inhibition of cathepsin G sensitizes TRAIL-induced apoptosis through 5-LOX-mediated down-regulation of survivin expression. PMID:29290980

  20. Verocytotoxin-induced apoptosis of human microvascular endothelial cells.

    PubMed

    Pijpers, A H; van Setten, P A; van den Heuvel, L P; Assmann, K J; Dijkman, H B; Pennings, A H; Monnens, L A; van Hinsbergh, V W

    2001-04-01

    The pathogenesis of the epidemic form of hemolytic uremic syndrome is characterized by endothelial cell damage. In this study, the role of apoptosis in verocytotoxin (VT)-mediated endothelial cell death in human glomerular microvascular endothelial cells (GMVEC), human umbilical vein endothelial cells, and foreskin microvascular endothelial cells (FMVEC) was investigated. VT induced apoptosis in GMVEC and human umbilical vein endothelial cells when the cells were prestimulated with the inflammatory mediator tumor necrosis factor-alpha (TNF-alpha). FMVEC displayed strong binding of VT and high susceptibility to VT under basal conditions, which made them suitable for the study of VT-induced apoptosis without TNF-alpha interference. On the basis of functional (flow cytometry and immunofluorescence microscopy using FITC-conjugated annexin V and propidium iodide), morphologic (transmission electron microscopy), and molecular (agarose gel electrophoresis of cellular DNA fragments) criteria, it was documented that VT induced programmed cell death in microvascular endothelial cells in a dose- and time-dependent manner. Furthermore, whereas partial inhibition of protein synthesis by VT was associated with a considerable number of apoptotic cells, comparable inhibition of protein synthesis by cycloheximide was not. This suggests that additional pathways, independent of protein synthesis inhibition, may be involved in VT-mediated apoptosis in microvascular endothelial cells. Specific inhibition of caspases by Ac-Asp-Glu-Val-Asp-CHO, but not by Ac-Tyr-Val-Ala-Asp-CHO, was accompanied by inhibition of VT-induced apoptosis in FMVEC and TNF-alpha-treated GMVEC. These data indicate that VT can induce apoptosis in human microvascular endothelial cells.

  1. Inhibition of mitogen activated protein kinases increases the sensitivity of A549 lung cancer cells to the cytotoxicity induced by a kava chalcone analog

    PubMed Central

    Warmka, Janel K.; Solberg, Eric L.; Zeliadt, Nicholette A.; Srinivasan, Balasubramanian; Charlson, Aaron T.; Xing, Chengguo; Wattenberg, Elizabeth V.

    2012-01-01

    We are interested in investigating the biological activity of chalcones, a major class of compounds found in the beverage kava, in order to develop potent and selective chemopreventive candidates. Consumption of kava in the South Pacific Islands is inversely correlated with cancer incidence, even among smokers. Accordingly, chalcones have anti-cancer activities in animal and cell culture models. To investigate signaling pathways that affect chalcone action we studied a potent analog, (E)-3-(3-hydroxy-4-methoxyphenyl)-1-(3,4,5-trimethoxyphenyl)prop-2-en-1-one (chalcone-24). Chalcone-24 was selected from a series of chalcone analogs that were synthesized based on the structures derived from flavokawain compounds found in kava, and screened in A549 lung cancer cells for induction of cytotoxicity and inhibition of NF-κB, a transcription factor associated with cell survival. Incubation of A549 cells with chalcone-24 resulted in a dose-dependent inhibition of cell viability, inhibition of NF-κB, activation of caspases, and activation of extracellular signal regulated kinase 1/2 (ERK1/2) and c-Jun N-terminal kinase (JNK); ERK1/2 and JNK are mitogen activated protein kinases that play central roles in regulating cell fate. Pharmacological inhibitors of ERK1/2 or JNK increased the sensitivity of A549 cells to chalcone-24-induced cytotoxicity, without affecting NF-κB or caspase activity. These results will help refine the synthesis of chalcone analogs to maximize the combination of actions required to prevent and treat cancer. PMID:22771807

  2. Inhibition of mitogen activated protein kinases increases the sensitivity of A549 lung cancer cells to the cytotoxicity induced by a kava chalcone analog.

    PubMed

    Warmka, Janel K; Solberg, Eric L; Zeliadt, Nicholette A; Srinivasan, Balasubramanian; Charlson, Aaron T; Xing, Chengguo; Wattenberg, Elizabeth V

    2012-08-03

    We are interested in investigating the biological activity of chalcones, a major class of compounds found in the beverage kava, in order to develop potent and selective chemopreventive candidates. Consumption of kava in the South Pacific Islands is inversely correlated with cancer incidence, even among smokers. Accordingly, chalcones have anti-cancer activities in animal and cell culture models. To investigate signaling pathways that affect chalcone action we studied a potent analog, (E)-3-(3-hydroxy-4-methoxyphenyl)-1-(3,4,5-trimethoxyphenyl)prop-2-en-1-one (chalcone-24). Chalcone-24 was selected from a series of chalcone analogs that were synthesized based on the structures derived from flavokawain compounds found in kava, and screened in A549 lung cancer cells for induction of cytotoxicity and inhibition of NF-κB, a transcription factor associated with cell survival. Incubation of A549 cells with chalcone-24 resulted in a dose-dependent inhibition of cell viability, inhibition of NF-κB, activation of caspases, and activation of extracellular signal regulated kinase 1/2 (ERK1/2) and c-Jun N-terminal kinase (JNK); ERK1/2 and JNK are mitogen activated protein kinases that play central roles in regulating cell fate. Pharmacological inhibitors of ERK1/2 or JNK increased the sensitivity of A549 cells to chalcone-24-induced cytotoxicity, without affecting NF-κB or caspase activity. These results will help refine the synthesis of chalcone analogs to maximize the combination of actions required to prevent and treat cancer. Copyright © 2012 Elsevier Inc. All rights reserved.

  3. Quercetin Enhances the Antitumor Activity of Trichostatin A through Upregulation of p53 Protein Expression In Vitro and In Vivo

    PubMed Central

    Chan, Shu-Ting; Yang, Nae-Cherng; Huang, Chin-Shiu; Liao, Jiunn-Wang; Yeh, Shu-Lan

    2013-01-01

    This study investigated the effects of quercetin on the anti-tumor effect of trichostatin A (TSA), a novel anticancer drug, in vitro and in vivo and the possible mechanisms of these effects in human lung cancer cells. We first showed that quercetin (5 µM) significantly increased the growth arrest and apoptosis in A549 cells (expressing wild-type p53) induced by 25 ng/mL of (82.5 nM) TSA at 48 h by about 25% and 101%, respectively. However, such enhancing effects of quercetin (5 µM) were not significant in TSA-exposed H1299 cells (a p53 null mutant) or were much lower than in A549 cells. In addition, quercetin significantly increased TSA-induced p53 expression in A549 cells. Transfection of p53 siRNA into A549 cells significantly but not completely diminished the enhancing effects of quercetin on TSA-induced apoptosis. Furthermore, we demonstrated that quercetin enhanced TSA-induced apoptosis through the mitochondrial pathway. Transfection of p53 siRNA abolished such enhancing effects of quercetin. However, quercetin increased the acetylation of histones H3 and H4 induced by TSA in A549 cells, even with p53 siRNA transfection as well as in H1299 cells. In a xenograft mouse model of lung cancer, quercetin enhanced the antitumor effect of TSA. Tumors from mice treated with TSA in combination with quercetin had higher p53 and apoptosis levels than did those from control and TSA-treated mice. These data indicate that regulation of the expression of p53 by quercetin plays an important role in enhancing TSA-induced apoptosis in A549 cells. However, p53-independent mechanisms may also contribute to the enhancing effect of quercetin. PMID:23342112

  4. Curcumin-induced downregulation of Axl receptor tyrosine kinase inhibits cell proliferation and circumvents chemoresistance in non-small lung cancer cells.

    PubMed

    Kim, Kyung-Chan; Baek, Suk-Hwan; Lee, Chuhee

    2015-12-01

    Lung cancer is still in the first place in terms of both incidence and mortality. In the present study, we demonstrated the effect of curcumin, a phytochemical of the plant Curcuma longa, on expression and activation of Axl receptor tyrosine kinase (RTK) which plays an important role in cell survival, proliferation and anti-apoptosis. Curcumin treatment of non-small cell lung cancer (NSCLC) A549 and H460 cells, was found to decrease Axl protein as well as mRNA levels in a dose- and time-dependent manner. Axl promoter activity was also reduced by curcumin, indicating that curcumin downregulates Axl expression at the transcriptional level. Moreover, Axl phosphorylation in response to binding of its ligand, Gas6, was abrogated by curcumin, suggesting the inhibitory effect of curcumin on Gas6-induced Axl activation. We next found cytotoxic effect of cucumin on both the parental A549 and H460 cells, and their variants which are resistant to cisplatin (A549/CisR and H460/CisR) and paclitaxel (A549/TR and H460/TR). Exposure of these cells to curcumin resulted in dose-dependent decline of cell viability and clonogenic ability. It is further observed that the anti-proliferative effect of curcumin on A549 cells overexpressing Axl protein was reduced, while that on H460 cells transfected Axl specific siRNA was augmented, confirming that curcumin inhibits cell proliferation via downregulation of Axl expression. In addition, curcumin was found to cause the induction of p21, a cyclin-dependent kinase inhibitor, and reduction of X-linked inhibitor of apoptosis protein (XIAP), an anti-apoptotic molecule, in parental H460 cells as well as chemoresistant cells, H460/CisR and H460/TR. Taken together, our data imply that Axl RTK is a novel target of curcumin through which it exerts anti-proliferative effect in both parental and chemoresistant NSCLC cells.

  5. Effects of Human Parvovirus B19 and Bocavirus VP1 Unique Region on Tight Junction of Human Airway Epithelial A549 Cells

    PubMed Central

    Chiu, Chun-Ching; Shi, Ya-Fang; Yang, Jiann-Jou; Hsiao, Yuan-Chao; Tzang, Bor-Show; Hsu, Tsai-Ching

    2014-01-01

    As is widely recognized, human parvovirus B19 (B19) and human bocavirus (HBoV) are important human pathogens. Obviously, both VP1 unique region (VP1u) of B19 and HBoV exhibit the secreted phospholipase A2 (sPLA2)-like enzymatic activity and are recognized to participate in the pathogenesis of lower respiratory tract illnesses. However, exactly how, both VP1u from B19 and HBoV affect tight junction has seldom been addressed. Therefore, this study investigates how B19-VP1u and HBoV-VP1u may affect the tight junction of the airway epithelial A549 cells by examining phospholipase A2 activity and transepithelial electrical resistance (TEER) as well as performing immunoblotting analyses. Experimental results indicate that TEER is more significantly decreased in A549 cells by treatment with TNF-α (10 ng), two dosages of B19-VP1u and BoV-VP1u (400 ng and 4000 ng) or bee venom PLA2 (10 ng) than that of the control. Accordingly, more significantly increased claudin-1 and decreased occludin are detected in A549 cells by treatment with TNF-α or both dosages of HBoV-VP1u than that of the control. Additionally, more significantly decreased Na+/K+ ATPase is observed in A549 cells by treatment with TNF-α, high dosage of B19-VP1u or both dosages of BoV-VP1u than that of the control. Above findings suggest that HBoV-VP1u rather than B19 VP1u likely plays more important roles in the disruption of tight junction in the airway tract. Meanwhile, this discrepancy appears not to be associated with the secreted phospholipase A2 (sPLA2)-like enzymatic activity. PMID:25268969

  6. Transcriptome analysis of Spodoptera frugiperda Sf9 cells reveals putative apoptosis-related genes and a preliminary apoptosis mechanism induced by azadirachtin.

    PubMed

    Shu, Benshui; Zhang, Jingjing; Sethuraman, Veeran; Cui, Gaofeng; Yi, Xin; Zhong, Guohua

    2017-10-16

    As an important botanical pesticide, azadirachtin demonstrates broad insecticidal activity against many agricultural pests. The results of a previous study indicated the toxicity and apoptosis induction of azadirachtin in Spodoptera frugiperda Sf9 cells. However, the lack of genomic data has hindered a deeper investigation of apoptosis in Sf9 cells at a molecular level. In the present study, the complete transcriptome data for Sf9 cell line was accomplished using Illumina sequencing technology, and 97 putative apoptosis-related genes were identified through BLAST and KEGG orthologue annotations. Fragments of potential candidate apoptosis-related genes were cloned, and the mRNA expression patterns of ten identified genes regulated by azadirachtin were examined using qRT-PCR. Furthermore, Western blot analysis showed that six putative apoptosis-related proteins were upregulated after being treated with azadirachtin while the protein Bcl-2 were downregulated. These data suggested that both intrinsic and extrinsic apoptotic signal pathways comprising the identified potential apoptosis-related genes were potentially active in S. frugiperda. In addition, the preliminary results revealed that caspase-dependent or caspase-independent apoptotic pathways could function in azadirachtin-induced apoptosis in Sf9 cells.

  7. 5-Fluorouracil-induced apoptosis in cultured oral cancer cells.

    PubMed

    Tong, D; Poot, M; Hu, D; Oda, D

    2000-03-01

    Chemotherapy is commonly used to treat advanced oral squamous cell carcinoma (SCC) and is known to kill cancer cells through apoptosis. Our hypothesis states that 5-fluorouracil (5FU) also kills cultured oral epithelial cells through programmed cell death or apoptosis. Cultured oral cancer cells were exposed to an optimum dose of 20 mg/ml of 5FU. Cells were analyzed for changes in cell cycle distribution and induction of cell death including apoptosis. Normal control, human papilloma virus-immortalized (PP), ATCC SCC cell line (CA1) and two primary oral SCC cell lines (CA3 and -4) were studied. Inhibition of apoptosis by a pan-caspase inhibitor was used. SYTO 11 flow cytometry showed increased apoptosis in all 5FU-treated cell cultures compared to untreated controls. The results show biological variation in apoptotic response. CA1 had the lowest apoptotic rate of the cancer cell lines at 1.5%. Next lowest was CA3, followed by CA4 and PP. In addition, alteration in the G1 and S phase fractions were found. Untreated CA1 showed 28% G1, 53% S compared to 43% G1, and 40% S of treated. We investigated the pathway of apoptosis using the pan-caspase inhibitor IDN-1529 by methylthiazolyl diphenyl tetrazolium bromide (MTT) colorimetric analysis. Results showed mild inhibition of cell death when cells were incubated with 50 microM IDN-1529 for 24 h. This suggests a probable caspase-dependent apoptotic pathway. In conclusion, our data suggest that 5FU induces oral cancer cell death through apoptosis and that biological variation exists between normal and cancer cells and between different types of cancer cells themselves. Our data indicate that cultures of a useful in vitro model for chemosensitivity assays are possible. Our results also suggest a caspase-dependent pathway for chemocytotoxicity in oral SCC.

  8. The mitochondria-mediate apoptosis of Lepidopteran cells induced by azadirachtin.

    PubMed

    Huang, Jingfei; Lv, Chaojun; Hu, Meiying; Zhong, Guohua

    2013-01-01

    Mitochondria have been shown to play an important role in apoptosis using mammalian cell lines. However, this seems not to be the case in Drosophila, an insect model organism; thus more in-depth studies of insect cell apoptosis are necessary. In the present study, mitochondrial involvement during azadirachtin- and camptothecin-induced apoptosis in Spodoptera frugiperda Sf9 cells (isolated from Spodoptera frugiperda pupal ovarian tissue) was investigated. The results showed that both azadirachtin and camptothecin could induce apoptosis in Sf9 cells. Reactive oxygen species (ROS) generation, activation of mitochondrial permeability transition pores (MPTPs) and loss of mitochondrial membrane potential (MMP) were observed very early during apoptosis and were followed subsequently by the release of cytochrome-c from the mitochondria. Furthermore, the results also revealed that the opening of MPTPs and the loss of MMP induced by azadirachtin could be significantly inhibited by the permeability transition pore (PTP) inhibitor cyclosporin A (CsA), which was used to identify the key role of mitochondria in the apoptosis of Sf9 cells. However, in camptothecin-treated Sf9 cells, CsA could not suppress the opening of MPTPs and the loss of MMP when apoptosis was induced. The data from caspase-3 and caspase-9 activity assays and detection of apoptosis by morphological observation and flow cytometry also uncovered the different effect of CsA on the two botanical apoptosis inducers. Although different mechanisms of apoptosis induction exist, our study revealed that mitochondria play a crucial role in insect cell line apoptosis.

  9. The Mitochondria-Mediate Apoptosis of Lepidopteran Cells Induced by Azadirachtin

    PubMed Central

    Huang, Jingfei; Lv, Chaojun; Hu, Meiying; Zhong, Guohua

    2013-01-01

    Mitochondria have been shown to play an important role in apoptosis using mammalian cell lines. However, this seems not to be the case in Drosophila, an insect model organism; thus more in-depth studies of insect cell apoptosis are necessary. In the present study, mitochondrial involvement during azadirachtin- and camptothecin-induced apoptosis in Spodoptera frugiperda Sf9 cells (isolated from Spodoptera frugiperda pupal ovarian tissue) was investigated. The results showed that both azadirachtin and camptothecin could induce apoptosis in Sf9 cells. Reactive oxygen species (ROS) generation, activation of mitochondrial permeability transition pores (MPTPs) and loss of mitochondrial membrane potential (MMP) were observed very early during apoptosis and were followed subsequently by the release of cytochrome-c from the mitochondria. Furthermore, the results also revealed that the opening of MPTPs and the loss of MMP induced by azadirachtin could be significantly inhibited by the permeability transition pore (PTP) inhibitor cyclosporin A (CsA), which was used to identify the key role of mitochondria in the apoptosis of Sf9 cells. However, in camptothecin-treated Sf9 cells, CsA could not suppress the opening of MPTPs and the loss of MMP when apoptosis was induced. The data from caspase-3 and caspase-9 activity assays and detection of apoptosis by morphological observation and flow cytometry also uncovered the different effect of CsA on the two botanical apoptosis inducers. Although different mechanisms of apoptosis induction exist, our study revealed that mitochondria play a crucial role in insect cell line apoptosis. PMID:23516491

  10. Single-cell-precision microplasma-induced cancer cell apoptosis.

    PubMed

    Tan, Xiao; Zhao, Shasha; Lei, Qian; Lu, Xinpei; He, Guangyuan; Ostrikov, Kostya

    2014-01-01

    The issue of single-cell control has recently attracted enormous interest. However, in spite of the presently achievable intracellular-level physiological probing through bio-photonics, nano-probe-based, and some other techniques, the issue of inducing selective, single-cell-precision apoptosis, without affecting neighbouring cells remains essentially open. Here we resolve this issue and report on the effective single-cell-precision cancer cell treatment using the reactive chemistry of the localized corona-type plasma discharge around a needle-like electrode with the spot size ∼1 µm. When the electrode is positioned with the micrometer precision against a selected cell, a focused and highly-localized micro-plasma discharge induces apoptosis in the selected individual HepG2 and HeLa cancer cells only, without affecting any surrounding cells, even in small cell clusters. This is confirmed by the real-time monitoring of the morphological and structural changes at the cellular and cell nucleus levels after the plasma exposure.

  11. Apoptosis in HEp-2 cells infected with Ureaplasma diversum.

    PubMed

    Amorim, Aline Teixeira; Marques, Lucas Miranda; Santos, Angelita Maria Oliveira Gusmão; Martins, Hellen Braga; Barbosa, Maysa Santos; Rezende, Izadora Souza; Andrade, Ewerton Ferraz; Campos, Guilherme Barreto; Lobão, Tássia Neves; Cortez, Beatriz Araujo; Monezi, Telma Alvez; Machado-Santelli, Glaucia Maria; Timenetsky, Jorge

    2014-09-04

    Bacterial pathogens have many strategies for infecting and persisting in host cells. Adhesion, invasion and intracellular life are important features in the biology of mollicutes. The intracellular location of Ureaplasma diversum may trigger disturbances in the host cell. This includes activation or inhibition of pro and anti-apoptotic factors, which facilitate the development of host damage. The aim of the present study was to associate U. diversum infection in HEp-2 cells and apoptosis induction. Cells were infected for 72hs with four U. diversum clinical isolates and an ATCC strain. The U. diversum invasion was analyzed by Confocal Laser Scanning Microscopy and gentamicin invasion assay. The apoptosis was evaluated using pro-apoptotic and anti-apoptotic gene expression, and FITC Annexin V/Dead Cell Apoptosis Kit. The number of internalized ureaplasma in HEp-2 cells increased significantly throughout the infection. The flow cytometry analysis with fluorochromes to detect membrane depolarization and gene expression for caspase 2, 3 and 9 increased in infected cells after 24 hours. However, after 72 hours a considerable decrease of apoptotic cells was observed. The data suggests that apoptosis may be initially induced by some isolates in association with HEp-2 cells, but over time, there was no evidence of apoptosis in the presence of ureaplasma and HEp-2 cells. The initial increase and then decrease in apoptosis could be related to bacterial pathogen-associated molecular pattern (PAMPS). Moreover, the isolates of U. diversum presented differences in the studied parameters for apoptosis. It was also observed that the amount of microorganisms was not proportional to the induction of apoptosis in HEp-2 cells.

  12. Nur77 attenuates endothelin-1 expression via downregulation of NF-κB and p38 MAPK in A549 cells and in an ARDS rat model.

    PubMed

    Jiang, Yujie; Zeng, Yi; Huang, Xia; Qin, Yueqiu; Luo, Weigui; Xiang, Shulin; Sooranna, Suren R; Pinhu, Liao

    2016-12-01

    Acute respiratory distress syndrome (ARDS) is characterized by inflammatory injury to the alveolar and capillary barriers that results in impaired gas exchange and severe acute respiratory failure. Nuclear orphan receptor Nur77 has emerged as a regulator of gene expression in inflammation, and its role in the pathogenesis of ARDS is not clear. The objective of this study is to investigate the potential role of Nur77 and its underlying mechanism in the regulation of endothelin-1 (ET-1) expression in lipopolysaccharide (LPS)-induced A549 cells and an ARDS rat model. We demonstrate that LPS induced Nur77 expression and nuclear export in A549 cells. Overexpression of Nur77 markedly decreased basal and LPS-induced ET-1 expression in A549 cells, whereas knockdown of Nur77 increased the ET-1 expression. LPS-induced phosphorylation and nuclear translocation of NF-κB and p38 MAPK were blocked by Nur77 overexpression and augmented by Nur77 knockdown in A549 cells. In vivo, LPS induced Nur77 expression in lung in ARDS rats. Pharmacological activation of Nur77 by cytosporone B (CsnB) inhibited ET-1 expression in ARDS rats, decreased LPS-induced phosphorylation of NF-κB and p38 MAPK, and relieved lung, liver, and kidney injury. Pharmacological deactivation of Nur77 by 1,1-bis-(3'-indolyl)-1-(p-hydroxyphenyl)methane (DIM-C-pPhOH, C-DIM8) had no effect on ET-1 expression and lung injury. These results indicated that Nur77 decreases ET-1 expression by suppressing NF-κB and p38 MAPK in LPS-stimulated A549 cells in vitro, and, in an LPS-induced ARDS rat model, CsnB reduced ET-1 expression and lung injury in ARDS rats. Copyright © 2016 the American Physiological Society.

  13. Salinomycin overcomes ABC transporter-mediated multidrug and apoptosis resistance in human leukemia stem cell-like KG-1a cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Fuchs, Dominik; Institute of Immunology, University of Heidelberg, Im Neuenheimer Feld 305, D-69120 Heidelberg; Daniel, Volker

    2010-04-16

    Leukemia stem cells are known to exhibit multidrug resistance by expression of ATP-binding cassette (ABC) transporters which constitute transmembrane proteins capable of exporting a wide variety of chemotherapeutic drugs from the cytosol. We show here that human promyeloblastic leukemia KG-1a cells exposed to the histone deacetylase inhibitor phenylbutyrate resemble many characteristics of leukemia stem cells, including expression of functional ABC transporters such as P-glycoprotein, BCRP and MRP8. Consequently, KG-1a cells display resistance to the induction of apoptosis by various chemotherapeutic drugs. Resistance to apoptosis induction by chemotherapeutic drugs can be reversed by cyclosporine A, which effectively inhibits the activity ofmore » P-glycoprotein and BCRP, thus demonstrating ABC transporter-mediated drug resistance in KG-1a cells. However, KG-1a are highly sensitive to apoptosis induction by salinomycin, a polyether ionophore antibiotic that has recently been shown to kill human breast cancer stem cell-like cells and to induce apoptosis in human cancer cells displaying multiple mechanisms of drug and apoptosis resistance. Whereas KG-1a cells can be adapted to proliferate in the presence of apoptosis-inducing concentrations of bortezomib and doxorubicin, salinomycin does not permit long-term adaptation of the cells to apoptosis-inducing concentrations. Thus, salinomycin should be regarded as a novel and effective agent for the elimination of leukemia stem cells and other tumor cells exhibiting ABC transporter-mediated multidrug resistance.« less

  14. Loss of inhibitor of apoptosis proteins as a determinant of polyamine analog-induced apoptosis in human melanoma cells.

    PubMed

    Chen, Ying; Kramer, Debora L; Li, Fengzhi; Porter, Carl W

    2003-08-07

    We have previously shown that the clinically relevant polyamine analog N1,N11-diethylnorspermine (DENSPM) causes rapid apoptosis in human melanoma SK-MEL-28 cells via a series of events that include mitochondrial release of cytochrome c and activation of the caspase cascade. Upstream to these events, DENSPM downregulates polyamine biosynthesis and potently upregulates polyamine catabolism at the level of spermidine/spermine N1-acetyltransferase (SSAT). In searching for downstream effectors that either contribute to or abrogate the apoptotic response, we observed that DENSPM treatment of SK-MEL-28 cells for 30 h led to cytosolic release of Smac/Diablo, a mitochondrial protein known to bind and inhibit the function of inhibitor of apoptosis proteins (IAPs). Subsequently, we found that DENSPM markedly lowered survivin and ML-IAP protein (but not XIAP) levels by 18 h via an apparently Smac/Diablo-independent pathway. Proteasome inhibitors fully prevented survivin and ML-IAP protein loss as well as apoptosis, suggesting that the proteasome-mediated degradation of survivin and ML-IAP is causally linked to the cellular outcome. We also observed that structural analogs of DENSPM which differentially induced SSAT and apoptosis lowered survivin and ML-IAP levels in a manner that correlated with enzyme activity. The linkage between IAPs and SSAT was more directly established by the finding that selective prevention of SSAT induction by small interfering RNA prevented survivin and ML-IAP loss as well as apoptosis during DENSPM treatment. Among the melanoma cell lines (SK-MEL-28, MALME-3M, A375 and LOX), survivin degradation correlated temporally with the onset of DENSPM induced apoptosis or growth inhibition. By contrast, ML-IAP degradation occurred only during rapid apoptosis seen in SK-MEL-28 cells. These data suggest a sequence of events whereby DENSPM induction of SSAT leads to loss of IAP proteins and a more fulminate apoptotic response. The findings implicate survivin

  15. Peroxisome proliferators induce apoptosis in hepatoma cells.

    PubMed

    Canuto, R A; Muzio, G; Bonelli, G; Maggiora, M; Autelli, R; Barbiero, G; Costelli, P; Brossa, O; Baccino, F M

    1998-01-01

    In the AH-130 hepatoma, a poorly differentiated tumor, maintained by weekly transplantations in rats, a low percentage of cells spontaneously underwent apoptosis, mainly during the transition from logarithmic- to stationary-growth phase. It was possible to induce massive apoptosis of cells by treating them with clofibrate, a peroxisome proliferator and hypolipidemic drug. Similar results were obtained with HepG2 cells. With 1 mM clofibrate, apoptosis began to manifest itself after 1 h of treatment in vitro, and was assessed by morphological analysis, by DNA fragmentation carried out with agarose gel electrophoresis, and with flow cytometric determination of terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end-labeling. The mechanisms whereby clofibrate induces apoptosis are still unclear. Since the peroxisome proliferator-activated receptor was expressed at a very low level and was not stimulated by clofibrate in the AH-130 hepatoma cells, its involvement seems unlikely. Moreover, lipid peroxidation was not increased after clofibrate treatment. Phospholipids and cholesterol were significantly decreased. The decreased cholesterol content might suggest an inhibition of the mevalonate pathway and, therefore, of isoprenylation of proteins involved in cell proliferation.

  16. Novel anticancer agent, benzyldihydroxyoctenone, isolated from Streptomyces sp. causes G1 cell cycle arrest and induces apoptosis of HeLa cells.

    PubMed

    Lee, Chul-Hoon; Lim, Haeyoung; Moon, Sangik; Shin, Choonshik; Kim, Seunghyun; Kim, Bum-Joon; Lim, Yoongho

    2007-06-01

    In the course of screening for anticancer agents, a novel active compound, F3-2-5, was isolated from culture broth of Streptomyces sp., KACC91015. Its structure was identified using nuclear magnetic resonance, mass spectrometry, and molecular modeling experiments, and confirmed by total synthesis. The growth of various human cancer cell lines was inhibited in a dose-dependent manner by 0.06-0.48 mM F3-2-5 over 24 h. Its IC(50) values were estimated at 37 microM on HeLa, 72 microM on A549, and 190 microM on HT-29 cells. However, F3-2-5 had no antiproliferative effect on normal lymphocytes and normal fibroblasts used as controls. Moreover, it affected cell cycle regulation and caused apoptosis of the HeLa cells; chromatin condensation and DNA fragmentation were observed in cells exposed to 80 microM F3-2-5. Western blot analysis revealed that F3-2-5 inhibited phosphorylation of retinoblastoma protein (pRb) and reduced expression of cyclin-dependent kinase-4 and -6, and cyclin D1 and E, while levels of p53 and p21(WAF1/CIP1) increased. Taken together, these findings show that F3-2-5 inhibits proliferation of HeLa cells by inducing G(1) phase arrest as a consequence of inhibition of pRb phosphorylation following up-regulation of p21(WAF1/CIP1) and p53. Furthermore, apoptosis in HeLa cells treated with F3-2-5 was associated with an increase in Bax and p53, leading to release of cytochrome c, activation of caspase-3, and -8, and cleavage of poly (ADP-ribose) polymerase.

  17. Chalcones Enhance TRAIL-Induced Apoptosis in Prostate Cancer Cells

    PubMed Central

    Szliszka, Ewelina; Czuba, Zenon P; Mazur, Bogdan; Sedek, Lukasz; Paradysz, Andrzej; Krol, Wojciech

    2009-01-01

    Chalcones exhibit chemopreventive and antitumor effects. Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) is a naturally occurring anticancer agent that induces apoptosis in cancer cells and is not toxic to normal cells. We examined the cytotoxic and apoptotic effect of five chalcones in combination with TRAIL on prostate cancer cells. The cytotoxicity was evaluated by the MTT and LDH assays. The apoptosis was determined using flow cytometry with annexin V-FITC. Our study showed that all five tested chalcones: chalcone, licochalcone-A, isobavachalcone, xanthohumol, butein markedly augmented TRAIL-mediated apoptosis and cytotoxicity in prostate cancer cells and confirmed the significant role of chalcones in chemoprevention of prostate cancer. PMID:20161998

  18. Spontaneous apoptosis in a rat hepatoma cell line and its inhibition by dexamethasone.

    PubMed

    Chaya, D; Fougère-Deschatrette, C; Weiss, M C

    1996-01-01

    Cultures of dedifferentiated rat hepatoma Rab1-5-1 cells exhibit spontaneously a high level of mortality during the exponential growth phase. We demonstrate that these cells die by apoptosis, showing chromatin condensation and internucleosomal DNA fragmentation. Cells of the original H4II cell line and of its differentiated and dedifferentiated derivatives also die by apoptosis, but only in heavily confluent cultures. We evaluated mortality with time in Rab1-5-1 cultures by establishing growth curves, including quantification of floating cells, and conclude that up to half of the cells in a culture are lost to apoptosis. The production of apoptotic cells is abolished by the presence of 10(-6) M dexamethasone and this inhibition is reversible in 48 hours. Rab1-5-1 cells that spontaneously die by apoptosis with high frequency represent a novel model to investigate factors that regulate the spontaneous frequency of death, and to study the nature and the kinetics of commitment to the apoptotic pathway.

  19. Evaluation of Anti-Metastatic Potential of the Combination of Fisetin with Paclitaxel on A549 Non-Small Cell Lung Cancer Cells.

    PubMed

    Klimaszewska-Wiśniewska, Anna; Hałas-Wiśniewska, Marta; Grzanka, Alina; Grzanka, Dariusz

    2018-02-27

    The identification and development of new agents with a therapeutic potential as well as novel drug combinations are gaining the attention of scientists and clinicians as a plausible approach to improve therapeutic regimens for chemoresistant tumors. We have recently reported that the flavonoid fisetin (FIS), at physiologically attainable concentrations, acts synergistically with clinically achievable doses of paclitaxel (PTX) to produce growth inhibitory and pro-death effects on A549 human non-small cell lung cancer (NSCLC) cells. To further investigate a potential therapeutic efficacy of the combination of fisetin with paclitaxel, we decided to assess its impact on metastatic capability of A549 cells as well as its toxicity toward normal human lung fibroblast. Cell viability, cell migration, and invasion were measured by thiazolyl blue tetrazolium bromide (MTT) assay, wound healing assay, and Transwell chamber assay, respectively. The expression of metastasis-related genes was assessed with quantitative reverse transcriptase real-time polymerase chain reaction (qRT-PCR). Actin and vimentin filaments were examined under the fluorescence microscope. The combination of FIS and PTX significantly reduced cancer cell migration and invasion, at least partially, through a marked rearrangement of actin and vimentin cytoskeleton and the modulation of metastasis-related genes. Most of these effects of the combination treatment were significantly greater than those of individual agents. Paclitaxel alone was even more toxic to normal cells than the combination of this drug with the flavonoid, suggesting that FIS may provide some protection against PTX-mediated cytotoxicity. The combination of FIS and PTX is expected to have a synergistic anticancer efficacy and a significant potential for the treatment of NSCLC, however, further in vitro and in vivo studies are required to confirm this preliminary evidence.

  20. A Role for Serglycin Proteoglycan in Mast Cell Apoptosis Induced by a Secretory Granule-mediated Pathway*

    PubMed Central

    Melo, Fabio Rabelo; Waern, Ida; Rönnberg, Elin; Åbrink, Magnus; Lee, David M.; Schlenner, Susan M.; Feyerabend, Thorsten B.; Rodewald, Hans-Reimer; Turk, Boris; Wernersson, Sara; Pejler, Gunnar

    2011-01-01

    Mast cell secretory granules (secretory lysosomes) contain large amounts of fully active proteases bound to serglycin proteoglycan. Damage to the granule membrane will thus lead to the release of serglycin and serglycin-bound proteases into the cytosol, which potentially could lead to proteolytic activation of cytosolic pro-apoptotic compounds. We therefore hypothesized that mast cells are susceptible to apoptosis induced by permeabilization of the granule membrane and that this process is serglycin-dependent. Indeed, we show that wild-type mast cells are highly sensitive to apoptosis induced by granule permeabilization, whereas serglycin-deficient cells are largely resistant. The reduced sensitivity of serglycin−/− cells to apoptosis was accompanied by reduced granule damage, reduced release of proteases into the cytosol, and defective caspase-3 activation. Mechanistically, the apoptosis-promoting effect of serglycin involved serglycin-dependent proteases, as indicated by reduced sensitivity to apoptosis and reduced caspase-3 activation in cells lacking individual mast cell-specific proteases. Together, these findings implicate serglycin proteoglycan as a novel player in mast cell apoptosis. PMID:21123167

  1. A Novel Apoptosis Correlated Molecule: Expression and Characterization of Protein Latcripin-1 from Lentinula edodes C91–3

    PubMed Central

    Liu, Ben; Zhong, Mintao; Lun, Yongzhi; Wang, Xiaoli; Sun, Wenchang; Li, Xingyun; Ning, Anhong; Cao, Jing; Zhang, Wei; Liu, Lei; Huang, Min

    2012-01-01

    An apoptosis correlated molecule—protein Latcripin-1 of Lentinula edodes C91–3—was expressed and characterized in Pichia pastoris GS115. The total RNA was obtained from Lentinula edodes C91–3. According to the transcriptome, the full-length gene of Latcripin-1 was isolated with 3′-Full Rapid Amplification of cDNA Ends (RACE) and 5′-Full RACE methods. The full-length gene was inserted into the secretory expression vector pPIC9K. The protein Latcripin-1 was expressed in Pichia pastoris GS115 and analyzed by Sodium Dodecylsulfonate Polyacrylate Gel Electrophoresis (SDS-PAGE) and Western blot. The Western blot showed that the protein was expressed successfully. The biological function of protein Latcripin-1 on A549 cells was studied with flow cytometry and the 3-(4,5-Dimethylthiazol-2-yl)-2,5-Diphenyl-tetrazolium Bromide (MTT) method. The toxic effect of protein Latcripin-1 was detected with the MTT method by co-culturing the characterized protein with chick embryo fibroblasts. The MTT assay results showed that there was a great difference between protein Latcripin-1 groups and the control group (p < 0.05). There was no toxic effect of the characterized protein on chick embryo fibroblasts. The flow cytometry showed that there was a significant difference between the protein groups of interest and the control group according to apoptosis function (p < 0.05). At the same time, cell ultrastructure observed by transmission electron microscopy supported the results of flow cytometry. The work demonstrates that protein Latcripin-1 can induce apoptosis of human lung cancer cells A549 and brings new insights into and advantages to finding anti-tumor proteins. PMID:22754362

  2. Oxidative Stress Facilitates IFN-γ-Induced Mimic Extracellular Trap Cell Death in A549 Lung Epithelial Cancer Cells.

    PubMed

    Lin, Chiou-Feng; Chen, Chia-Ling; Chien, Shun-Yi; Tseng, Po-Chun; Wang, Yu-Chih; Tsai, Tsung-Ting

    2016-01-01

    We previously demonstrated that IFN-γ induces an autophagy-regulated mimic extracellular trap cell death (ETosis) in A549 human lung cancer cells. Regarding reactive oxygen species (ROS) are involved in ETosis, this study investigated the role of oxidative stress. After IFN-γ stimulation, a necrosis-like cell death mimic ETosis occurred accompanied by the inhibition of cell growth, aberrant nuclear staining, and nucleosome release. ROS were generated in a time-dependent manner with an increase in NADPH oxidase component protein expression. STAT1-mediated IFN regulatory factor-1 activation was essential for upregulating ROS production. By genetically silencing p47phox, IFN-γ-induced ROS and mimic ETosis were significantly attenuated. This mechanistic study indicated that ROS may mediate DNA damage followed by histone H3 citrullination. Furthermore, ROS promoted IFN-γ-induced mimic ETosis in cooperation with autophagy. These findings further demonstrate that ROS regulates IFN-γ-induced mimic ETosis in lung epithelial malignancy.

  3. Oxidative Stress Facilitates IFN-γ-Induced Mimic Extracellular Trap Cell Death in A549 Lung Epithelial Cancer Cells

    PubMed Central

    Lin, Chiou-Feng; Chen, Chia-Ling; Chien, Shun-Yi; Tseng, Po-Chun; Wang, Yu-Chih; Tsai, Tsung-Ting

    2016-01-01

    We previously demonstrated that IFN-γ induces an autophagy-regulated mimic extracellular trap cell death (ETosis) in A549 human lung cancer cells. Regarding reactive oxygen species (ROS) are involved in ETosis, this study investigated the role of oxidative stress. After IFN-γ stimulation, a necrosis-like cell death mimic ETosis occurred accompanied by the inhibition of cell growth, aberrant nuclear staining, and nucleosome release. ROS were generated in a time-dependent manner with an increase in NADPH oxidase component protein expression. STAT1-mediated IFN regulatory factor-1 activation was essential for upregulating ROS production. By genetically silencing p47phox, IFN-γ-induced ROS and mimic ETosis were significantly attenuated. This mechanistic study indicated that ROS may mediate DNA damage followed by histone H3 citrullination. Furthermore, ROS promoted IFN-γ-induced mimic ETosis in cooperation with autophagy. These findings further demonstrate that ROS regulates IFN-γ-induced mimic ETosis in lung epithelial malignancy. PMID:27575372

  4. Nrf2 but not autophagy inhibition is associated with the survival of wild-type epidermal growth factor receptor non-small cell lung cancer cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zhou, Yan

    Non-small cell lung cancer (NSCLC) is one of the most common malignancies in the world. Icotinib and Gefitinib are two epidermal growth factor receptor (EGFR) tyrosine kinase inhibitors (TKIs) that have been used to treat NSCLC. While it is well known that mutations of EGFR can affect the sensitivity of NSCLC to the EGFR-TKI, other mechanisms may also be adopted by lung cancer cells to develop resistance to EGFR-TKI treatment. Cancer cells can use multiple adaptive mechanisms such as activation of autophagy and Nrf2 to protect against various stresses and chemotherapeutic drugs. Whether autophagy or Nrf2 activation contributes to themore » resistance of NSCLC to EGFR-TKI treatment in wild-type EGFR NSCLC cells remains elusive. In the present study, we confirmed that Icotinib and Gefitinib induced apoptosis in EGFR mutant HCC827 but not in EGFR wild-type A549 NSCLC cells. Icotinib and Gefitinib did not induce autophagic flux or inhibit mTOR in A549 cells. Moreover, suppression of autophagy by chloroquine, a lysosomal inhibitor, did not affect Icotinib- or Gefitinib-induced cell death in A549 cells. In contrast, Brusatol, an Nrf2 inhibitor, significantly suppressed the cell survival of A549 cells. However, Brusatol did not further sensitize A549 cells to EGFR TKI-induced cell death. Results from this study suggest that inhibition of Nrf2 can decrease cell vitality of EGFR wild-type A549 cells independent of autophagy. - Highlights: • Cancer cells use adaptive mechanisms against chemotherapy. • Autophagy is not essential for the drug resistance of lung cancer A549 cells. • Inhibition of Nrf2 decreases cell survival of lung cancer A549 cells.« less

  5. Evaluation of cytotoxicity of new trans-palladium(II) complex in human cells in vitro.

    PubMed

    Kontek, Renata; Matławska-Wasowska, Ksenia; Kalinowska-Lis, Urszula; Kontek, Bogdan; Ochocki, Justyn

    2011-01-01

    Studies of cytotoxicity allow to elucidate the mechanisms by which chemical compounds influence cells and tissues. On the basis of the structural analogy between platinum(II) and palladium(II) complexes, a variety of studies on palladium(II) compounds as potential anticancer drugs have been carried out (1, 2). The cytotoxicity was evaluated by MTT assay. Abilities of trans-palladium(II) complex containing diethyl (pyridin-2-ylmethyl)phosphates as non-leaving ligands (trans-[PdCl2(2-pmOpe 2)]) to induce apoptosis and necrosis in normal lymphocytes, A549 cells and HT29 cell lines were performed by use of fluorochrome staining. The obtained results revealed, that the new trans-palladium(II) complex was more cytotoxic against A549 and HT29 tumor cells than on the normal lymphocytes in vitro. The novel complex induces apoptosis in all tested cells, but in lymphocytes to a lesser degree. The compound tested also induced significant amounts of necrotic cells, which exceeded the level of apoptotic cell fractions. The results demonstrate that the trans-Pd(II) complex showed substantial cytotoxic activity against A549 and HT29 tumor cells and indicate that the new trans-palladium(II) complex effectively inhibited cancer cells growth.

  6. 5-alpha-reductase type I (SRD5A1) is up-regulated in non-small cell lung cancer but does not impact proliferation, cell cycle distribution or apoptosis

    PubMed Central

    2012-01-01

    Background Non-small cell lung cancer (NSCLC) is one of the most frequent malignancies and has a high mortality rate due to late detection and lack of efficient treatments. Identifying novel drug targets for this indication may open the way for new treatment strategies. Comparison of gene expression profiles of NSCLC and normal adjacent tissue (NAT) allowed to determine that 5-alpha-reductase type I (SRD5A1) was up-regulated in NSCLC compared to NAT. This raised the question whether SRD5A1 was involved in sustained proliferation and survival of NSCLC. Methods siRNA-mediated silencing of SRD5A1 was performed in A549 and NCI-H460 lung cancer cell lines in order to determine the impact on proliferation, on distribution during the different phases of the cell cycle, and on apoptosis/necrosis. In addition, lung cancer cell lines were treated with 4-azasteroids, which specifically inhibit SRD5A1 activity, and the effects on proliferation were measured. Statistical analyses using ANOVA and post-hoc Tamhane-T2-test were performed. In the case of non-parametric data, the Kruskal-Wallis test and the post-hoc Mann-Whitney-U-test were used. Results The knock-down of SRDA51 expression was very efficient with the SRD5A1 transcripts being reduced to 10% of control levels. Knock-down efficiency was furthermore confirmed at the protein level. However, no effect of SRD5A1 silencing was observed in the proliferation assay, the cell cycle analysis, and the apoptosis/necrosis assay. Treatment of lung cancer cell lines with 4-azasteroids did not significantly inhibit proliferation. Conclusions In summary, the results suggest that SRD5A1 is not a crucial enzyme for the sustained proliferation of NSCLC cell lines. PMID:22257483

  7. 5-alpha-reductase type I (SRD5A1) is up-regulated in non-small cell lung cancer but does not impact proliferation, cell cycle distribution or apoptosis.

    PubMed

    Kapp, Friedrich G; Sommer, Anette; Kiefer, Thomas; Dölken, Gottfried; Haendler, Bernard

    2012-01-18

    Non-small cell lung cancer (NSCLC) is one of the most frequent malignancies and has a high mortality rate due to late detection and lack of efficient treatments. Identifying novel drug targets for this indication may open the way for new treatment strategies. Comparison of gene expression profiles of NSCLC and normal adjacent tissue (NAT) allowed to determine that 5-alpha-reductase type I (SRD5A1) was up-regulated in NSCLC compared to NAT. This raised the question whether SRD5A1 was involved in sustained proliferation and survival of NSCLC. siRNA-mediated silencing of SRD5A1 was performed in A549 and NCI-H460 lung cancer cell lines in order to determine the impact on proliferation, on distribution during the different phases of the cell cycle, and on apoptosis/necrosis. In addition, lung cancer cell lines were treated with 4-azasteroids, which specifically inhibit SRD5A1 activity, and the effects on proliferation were measured. Statistical analyses using ANOVA and post-hoc Tamhane-T2-test were performed. In the case of non-parametric data, the Kruskal-Wallis test and the post-hoc Mann-Whitney-U-test were used. The knock-down of SRDA51 expression was very efficient with the SRD5A1 transcripts being reduced to 10% of control levels. Knock-down efficiency was furthermore confirmed at the protein level. However, no effect of SRD5A1 silencing was observed in the proliferation assay, the cell cycle analysis, and the apoptosis/necrosis assay. Treatment of lung cancer cell lines with 4-azasteroids did not significantly inhibit proliferation. In summary, the results suggest that SRD5A1 is not a crucial enzyme for the sustained proliferation of NSCLC cell lines.

  8. Long non‑coding RNA AK001796 contributes to cisplatin resistance of non‑small cell lung cancer.

    PubMed

    Liu, Bin; Pan, Chun-Feng; Ma, Teng; Wang, Jun; Yao, Guo-Liang; Wei, Ke; Chen, Yi-Jiang

    2017-10-01

    Cisplatin (DDP)‑based chemotherapy is the most widely used therapy for non‑small cell lung cancer (NSCLC). However, the existence of chemoresistance has become a major limitation in its efficacy. Long non‑coding RNAs (lncRNAs) have been shown to be involved in chemotherapy drug resistance. The aim of the present study was to investigate the biological role of lncRNA AK001796 in cisplatin‑resistant NSCLC A549/DDP cells. Reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) analysis was performed to monitor the differences in the expression of AK001796 in cisplatin-resistant (A549/DDP) cells and parental A549 cells. Cellular sensitivity to cisplatin and cell viability were examined using an MTT assay. Cell apoptosis and cell cycle distribution were measured using flow cytometry. The expression levels of cell cycle proteins cyclin C (CCNC), baculoviral IAP repeat containing 5 (BIRC5), cyclin‑dependent kinase 1 (CDK1) and G2 and S phase‑expressed 1 (GTSE1) were assessed using RT‑qPCR and western blot analyses. It was found that the expression of AK001796 was increased in A549/DDP cells, compared with that in A549 cells. The knockdown of AK001796 by small interfering RNA reduced cellular cisplatin resistance and cell viability, and resulted in cell‑cycle arrest, with a marked increase in the proportion of A549/DDP cells in the G0/G1 phase. By contrast, the knockdown of AK001796 increased the number of apoptotic cancer cells during cisplatin treatment. It was also shown that the knockdown of AK001796 positively induced the expression of cell apoptosis‑associated factors, CCNC and BIRC5, and suppressed the expression of cell cycle‑associated factors, CDK1 and GTSE5. Taken together, these findings indicated that lncRNA AK001796 increased the resistance of NSCLC cells to cisplatin through regulating cell apoptosis and cell proliferation, and thus provides an attractive therapeutic target for NSCLC.

  9. [Progress on mechanism of cell apoptosis induced by rubella virus].

    PubMed

    Li, Zhen-mei; Chu, Fu-lu; Liu, Ying; Wang, Zhi-yu

    2013-09-01

    Rubella virus (RV), a member of the family Togaviridae, can induce apoptosis of host cells in vitro. Protein kinases of the Ras-Raf-MEK-ERK pathway and PI3K-Akt pathway play essential roles in virus multiplication, cell survival and apoptosis. Proteins p53 and TAp63 that bind to specific DNA sequences stimulate Bax in a manner to produce functional pores that facilitate release of mitochondrial cytochrome c and downstream caspase activation. In this review, the molecular mechanisms of RV-induced cell apoptosis, including RV-infected cell lines, pathological changes in cell components and apoptosis signaling pathways are summarized.

  10. Mitochondria are the primary target in the induction of apoptosis by chiral ruthenium(II) polypyridyl complexes in cancer cells.

    PubMed

    Wang, Jin-Quan; Zhang, Ping-Yu; Qian, Chen; Hou, Xiao-Juan; Ji, Liang-Nian; Chao, Hui

    2014-03-01

    A series of novel chiral ruthenium(II) polypyridyl complexes (Δ-Ru1, Λ-Ru1, Δ-Ru2, Λ-Ru2, Δ-Ru3, Λ-Ru3) were synthesized and evaluated to determine their antiproliferative activities. Colocalization, inductively coupled plasma mass spectrometry, and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay studies showed that these ruthenium(II) complexes accumulated preferentially in the mitochondria and exhibited cytotoxicity against various cancer cells in vitro. The complex Δ-Ru1 is of particular interest because it was found to have half-maximal inhibitory concentrations comparable to those of cisplatin and better activity than cisplatin against a cisplatin-resistant cell line, A549-CP/R. Δ-Ru1 induced alterations in the mitochondrial membrane potential and triggered intrinsic mitochondria-mediated apoptosis in HeLa cells, which involved the regulation of Bcl-2 family members and the activation of caspases. Taken together, these data suggest that Δ-Ru1 may be a novel mitochondria-targeting anticancer agent.

  11. Regulation of programmed cell death or apoptosis in atherosclerosis.

    PubMed

    Geng, Y J

    1997-01-01

    Intimal thickening caused by accumulation of cells, lipids, and connective tissue characterizes atherosclerosis, an arterial disease that leads to cardiac and cerebral infarction. Apoptosis, or genetically programmed cell death, is important for the development and morphogenesis of organs and tissues. As in other tissues, cells of cardiovascular tissues can undergo apoptosis. Increased apoptosis has been found in both human and animal atherosclerotic lesions, mediating tissue turnover and lesion development. In addition to vascular cells, many activated immune cells, mainly macrophages and T cells, are present in atherosclerotic lesions, where these cells produce biologically active substances such as the proinflammatory cytokines tumor necrosis factor, interleukin-1 (IL-1), and interferon-gamma. Simultaneous exposure to these cytokines may trigger apoptosis of vascular smooth muscle cells. The products of death-regulating genes including Fas/Fas ligand, members of IL-1 beta cysteinyl protease (caspase) family, the tumor suppressive gene p53, and the protooncogene c-myc have been found in vascular cells and may participate in the regulation of vascular apoptosis during the development of atherosclerosis. Abnormal occurrence of apoptosis may take place in atherosclerotic lesions, including attenuation or acceleration of the apoptotic death process. The former may cause an increase in the cellularity of the lesions, and the latter can reduce cellular components important for maintaining the integrity and stability of the plaques. Clarification of the molecular mechanism that regulates apoptosis may help design a new strategy for treatment of patients with atherosclerosis and its major complications, heart attack and stroke.

  12. Low-power laser irradiation inhibits amyloid beta-induced cell apoptosis

    NASA Astrophysics Data System (ADS)

    Zhang, Heng; Wu, Shengnan

    2011-03-01

    The deposition and accumulation of amyloid-β-peptide (Aβ) in the brain are considered a pathological hallmark of Alzheimer's disease(AD). Apoptosis is a contributing pathophysiological mechanism of AD. Low-power laser irradiation (LPLI), a non-damage physical therapy, which has been used clinically for decades of years, is shown to promote cell proliferation and prevent apoptosis. Recently, low-power laser irradiation (LPLI) has been applied to moderate AD. In this study, Rat pheochromocytoma (PC12) cells were treated with amyloid beta 25-35 (Aβ25-35) for induction of apoptosis before LPLI treatment. We measured cell viability with CCK-8 according to the manufacture's protocol, the cell viability assays show that low fluence of LPLI (2 J/cm2 ) could inhibit the cells apoptosis. Then using statistical analysis of proportion of apoptotic cells by flow cytometry based on Annexin V-FITC/PI, the assays also reveal that low fluence of LPLI (2 J/cm2 ) could inhibit the Aβ-induced cell apoptosis. Taken together, we demonstrated that low fluence of LPLI (2 J/cm2 ) could inhibit the Aβ-induced cell apoptosis, these results directly point to a therapeutic strategy for the treatment of AD through LPLI.

  13. Activities of ten essential oils towards Propionibacterium acnes and PC-3, A-549 and MCF-7 cancer cells.

    PubMed

    Zu, Yuangang; Yu, Huimin; Liang, Lu; Fu, Yujie; Efferth, Thomas; Liu, Xia; Wu, Nan

    2010-04-30

    Ten essential oils, namely, mint (Mentha spicata L., Lamiaceae), ginger (Zingiber officinale Rosc., Zingiberaceae), lemon (Citrus limon Burm.f., Rutaceae), grapefruit (Citrus paradisi Macf., Rutaceae), jasmine (Jasminum grandiflora L., Oleaceae), lavender (Mill., Lamiaceae), chamomile (Matricaria chamomilla L., Compositae), thyme (Thymus vulgaris L., Lamiaceae), rose (Rosa damascena Mill., Rosaceae) and cinnamon (Cinnamomum zeylanicum N. Lauraceae) were tested for their antibacterial activities towards Propionibacterium acnes and in vitro toxicology against three human cancer cell lines. Thyme, cinnamon and rose essential oils exhibited the best antibacterial activities towards P. acnes, with inhibition diameters of 40 +/- 1.2 mm, 33.5 +/- 1.5 mm and 16.5 +/- 0.7 mm, and minimal inhibitory concentrations of 0.016% (v/v), 0.016% (v/v) and 0.031% (v/v), respectively. Time-kill dynamic procedures showed that thyme, cinnamon, rose, and lavender essential oils exhibited the strongest bactericidal activities at a concentration of 0.25% (v/v), and P. acnes was completely killed after 5 min. The thyme essential oil exhibited the strongest cytotoxicity towards three human cancer cells. Its inhibition concentration 50% (IC(50)) values on PC-3, A549 and MCF-7 tumor cell lines were 0.010% (v/v), 0.011% (v/v) and 0.030% (v/v), respectively. The cytotoxicity of 10 essential oils on human prostate carcinoma cell (PC-3) was significantly stronger than on human lung carcinoma (A549) and human breast cancer (MCF-7) cell lines.

  14. [Effects of methyl tertiary butyl ether on cell cycle and cell apoptosis].

    PubMed

    Zhou, W; Huang, G; Zhang, H; Ye, S

    2000-07-01

    To explore the effects of the new gasoline additive, methyl tertiary butyl ether (MTBE) on cell cycle and cell apoptosis. Flow cytometry was used to evaluate the effect of MTBE (1, 2, 4 microl/ml, 24 h) on NIH/3T3 cell cycles; and the effect of MTBE on Hela cell apoptosis was evaluated by detecting cell survival using crystal violet staining. Flow cytometry showed that MTBE could change NIH/3T3 cell cycles, decrease the number of cells in S stage, and arrest cells at G(2) + M stage. The results suggested that MTBE could affect NIH/3T3 cell cycles and induce cell proliferation. This situation existed 48 hours after the treatment, and cell cycles came back normal 96 hours after the treatment. By detecting cell survival using crystal violet staining, we found that MTBE could inhibit the apoptosis of Hela cells which was induced by tumor necrosis factor (TNF)alpha and cycloheximide. MTBE's carcinogenicity to animals may relate to induction of cell proliferation and inhibition of cell apoptosis.

  15. Nonylphenol diethoxylate inhibits apoptosis induced in PC12 cells.

    PubMed

    Liu, Chuang; Sun, Yongkun; Song, Yutong; Saito, Takeshi; Kurasaki, Masaaki

    2016-11-01

    Nonylphenol and short-chain nonylphenol ethoxylates such as NP 2 EO are present in aquatic environment as wastewater contaminants, and their toxic effects on aquatic species have been reported. Apoptosis has been shown to be induced by serum deprivation or copper treatment. To understand the toxicity of nonylphenol diethoxylate, we investigated the effects of NP 2 EO on apoptosis induced by serum deprivation and copper by using PC12 cell system. Nonylphenol diethoxylate itself showed no toxicity and recovered cell viability from apoptosis. In addition, nonylphenol diethoxylate decreased DNA fragmentation caused by apoptosis in PC12 cells. This phenomenon was confirmed after treating apoptotic PC12 cells with nonylphenol diethoxylate, whereas the cytochrome c release into the cytosol decreased as compared to that in apoptotic cells not treated with nonylphenol diethoxylates. Furthermore, Bax contents in apoptotic cells were reduced after exposure to nonylphenol diethoxylate. Thus, nonylphenol diethoxylate has the opposite effect on apoptosis in PC12 cells compared to nonylphenol, which enhances apoptosis induced by serum deprivation. The difference in structure of the two compounds is hypothesized to be responsible for this phenomenon. These results indicated that nonylphenol diethoxylate has capability to affect cell differentiation and development and has potentially harmful effect on organisms because of its unexpected impact on apoptosis. © 2015 Wiley Periodicals, Inc. Environ Toxicol 31: 1389-1398, 2016. © 2015 Wiley Periodicals, Inc.

  16. Osteoblasts Protect AML Cells from SDF-1-Induced Apoptosis

    PubMed Central

    Kremer, Kimberly N.; Dudakovic, Amel; McGee-Lawrence, Meghan E.; Philips, Rachael L.; Hess, Allan D.; Smith, B. Douglas; van Wijnen, Andre J.; Karp, Judith E.; Kaufmann, Scott H.; Westendorf, Jennifer J.; Hedin, Karen E.

    2014-01-01

    The bone marrow provides a protective environment for acute myeloid leukemia (AML) cells that often allows leukemic stem cells to survive standard chemotherapeutic regimens. Targeting these leukemic stem cells within the bone marrow is critical for preventing relapse. We recently demonstrated that SDF-1, a chemokine abundant in the bone marrow, induces apoptosis in AML cell lines and in patient samples expressing high levels of its receptor, CXCR4. Here we show that a subset of osteoblast lineage cells within the bone marrow can protect AML cells from undergoing apoptosis in response to the SDF-1 naturally present in that location. In co-culture systems, osteoblasts at various stages of differentiation protected AML cell lines and patient isolates from SDF-1-induced apoptosis. The differentiation of the osteoblast cell lines, MC3T3 and W-20-17, mediated this protection via a cell contact-independent mechanism. In contrast, bone marrow-derived mesenchymal cells, the precursors of osteoblasts, induced apoptosis in AML cells via a CXCR4-dependent mechanism and failed to protect AML cells from exogenously added SDF-1. These results indicate that osteoblasts in the process of differentiation potently inhibit the SDF-1-driven apoptotic pathway of CXCR4-expressing AML cells residing in the bone marrow. Drugs targeting this protective mechanism could potentially provide a new approach to treating AML by enhancing the SDF-1-induced apoptosis of AML cells residing within the bone marrow microenvironment. PMID:24851270

  17. INDUCTION OF CELL CYCLE ARREST AND APOPTOSIS BY ORMENIS ERIOLEPIS A MORROCAN ENDEMIC PLANT IN VARIOUS HUMAN CANCER CELL LINES.

    PubMed

    Belayachi, Lamiae; Aceves-Luquero, Clara; Merghoub, Nawel; de Mattos, Silvia Fernández; Amzazi, Saaîd; Villalonga, Priam; Bakri, Youssef

    2017-01-01

    Ormenis eriolepis Coss (Asteraceae) is an endemic Moroccan subspecies, traditionally named "Hellala" or "Fergoga". It's usually used for its hypoglycemic effect as well as for the treatment of stomacal pain. As far as we know, there is no scientific exploration of anti tumoral activity of Ormenis eriolepis extracts. In this regard, we performed a screening of organic extracts and fractions in a panel of both hematological and solid cancer cell lines, to evaluate the potential in vitro anti tumoral activity and to elucidate the respective mechanisms that may be responsible for growth arrest and cell death induction. The plant was extracted using organic solvents, and four different extracts were screened on Jurkat, Jeko-1, TK-6, LN229, SW620, U2OS, PC-3 and NIH3T3 cells. Cell viability assays revealed that, the IC50 values were (11,63±5,37μg/ml) for Jurkat, (13,33±1,67μg/ml) for Jeko-1, (41,67±1,98μg/ml) for LN229 and (19,31±4,88μg/ml) for PC-3 cells upon treatment with Oe-DF and Oe-HE respectively. Both the fraction and extract exhibited no effects on TK6 and NIH3T3. Cytometry analysis accompanied by DNA damage signaling protein levels monitoring (p-H2A.X), showed that both the Dichloromethane Fraction and Hexanic extract induce DNA double stranded breaks (DSBs) accompanied by cell cycle arrest in G1 (Jurkat, Jeko -1 and LN22) and G2/M (PC-3) phases which is agreed with the caspase activity observed. Additional experiments with selective inhibitors of stress and survival pathways (JNK, MAPK, Rho, p53, and JAK3) indicated that none of these pathways was significantly involved in apoptosis induction. The bioactive compound analysis by CG/MS indicated that the major compounds in Oe-DF were: Linoleic Acid (15,89%), Podophyllotoxin (17,89%) and Quercetin (22,95%). For Oe-HE the major molecules were: Linoleic Acid (9,76%), α-curcumene (7,07%), α-bisabolol (5,49%), Campesterol (4,41%), Stigmasterol (14,08%) and β-sitosterol (7,49%). Our data suggest that

  18. INDUCTION OF CELL CYCLE ARREST AND APOPTOSIS BY ORMENIS ERIOLEPIS A MORROCAN ENDEMIC PLANT IN VARIOUS HUMAN CANCER CELL LINES

    PubMed Central

    Belayachi, Lamiae; Aceves-Luquero, Clara; Merghoub, Nawel; de Mattos, Silvia Fernández; Amzazi, Saaîd; Villalonga, Priam; Bakri, Youssef

    2017-01-01

    Background: Ormenis eriolepis Coss (Asteraceae) is an endemic Moroccan subspecies, traditionally named “Hellala” or “Fergoga”. It’s usually used for its hypoglycemic effect as well as for the treatment of stomacal pain. As far as we know, there is no scientific exploration of anti tumoral activity of Ormenis eriolepis extracts. Materials and Methods: In this regard, we performed a screening of organic extracts and fractions in a panel of both hematological and solid cancer cell lines, to evaluate the potential in vitro anti tumoral activity and to elucidate the respective mechanisms that may be responsible for growth arrest and cell death induction. The plant was extracted using organic solvents, and four different extracts were screened on Jurkat, Jeko-1, TK-6, LN229, SW620, U2OS, PC-3 and NIH3T3 cells. Results: Cell viability assays revealed that, the IC50 values were (11,63±5,37μg/ml) for Jurkat, (13,33±1,67μg/ml) for Jeko-1, (41,67±1,98μg/ml) for LN229 and (19,31±4,88μg/ml) for PC-3 cells upon treatment with Oe-DF and Oe-HE respectively. Both the fraction and extract exhibited no effects on TK6 and NIH3T3. Cytometry analysis accompanied by DNA damage signaling protein levels monitoring (p-H2A.X), showed that both the Dichloromethane Fraction and Hexanic extract induce DNA double stranded breaks (DSBs) accompanied by cell cycle arrest in G1 (Jurkat, Jeko -1 and LN22) and G2/M (PC-3) phases which is agreed with the caspase activity observed. Additional experiments with selective inhibitors of stress and survival pathways (JNK, MAPK, Rho, p53, and JAK3) indicated that none of these pathways was significantly involved in apoptosis induction. The bioactive compound analysis by CG/MS indicated that the major compounds in Oe-DF were: Linoleic Acid (15,89%), Podophyllotoxin (17,89%) and Quercetin (22,95%). For Oe-HE the major molecules were: Linoleic Acid (9,76%), α-curcumene (7,07%), α-bisabolol (5,49%), Campesterol (4,41%), Stigmasterol (14

  19. Acetyl-CoA carboxylase-alpha inhibitor TOFA induces human cancer cell apoptosis.

    PubMed

    Wang, Chun; Xu, Canxin; Sun, Mingwei; Luo, Dixian; Liao, Duan-Fang; Cao, Deliang

    2009-07-31

    Acetyl-CoA carboxylase-alpha (ACCA) is a rate-limiting enzyme in long chain fatty acid synthesis, playing a critical role in cellular energy storage and lipid synthesis. ACCA is upregulated in multiple types of human cancers and small interfering RNA-mediated ACCA silencing in human breast and prostate cancer cells results in oxidative stress and apoptosis. This study reports for the first time that TOFA (5-tetradecyloxy-2-furoic acid), an allosteric inhibitor of ACCA, is cytotoxic to lung cancer cells NCI-H460 and colon carcinoma cells HCT-8 and HCT-15, with an IC(50) at approximately 5.0, 5.0, and 4.5 microg/ml, respectively. TOFA at 1.0-20.0 microg/ml effectively blocked fatty acid synthesis and induced cell death in a dose-dependent manner. The cell death was characterized with PARP cleavage, DNA fragmentation, and annexin-V staining, all of which are the features of the apoptosis. Supplementing simultaneously the cells with palmitic acids (100 microM), the end-products of the fatty acid synthesis pathway, prevented the apoptosis induced by TOFA. Taken together, these data suggest that TOFA is a potent cytotoxic agent to lung and colon cancer cells, inducing apoptosis through disturbing their fatty acid synthesis.

  20. Acetyl-CoA Carboxylase-α Inhibitor TOFA Induces Human Cancer Cell Apoptosis

    PubMed Central

    Wang, Chun; Xu, Canxin; Sun, Mingwei; Luo, Dixian; Liao, Duan-fang; Cao, Deliang

    2009-01-01

    Acetyl-CoA carboxylase-α (ACCA) is a rate-limiting enzyme in long chain fatty acid synthesis, playing a critical role in cellular energy storage and lipid synthesis. ACCA is upregulated in multiple types of human cancers and small interfering RNA-mediated ACCA silencing in human breast and prostate cancer cells results in oxidative stress and apoptosis. This study reports for the first time that TOFA (5-tetradecyloxy-2-furoic acid), an allosteric inhibitor of ACCA, is cytotoxic to lung cancer cells NCI-H460 and colon carcinoma cells HCT-8 and HCT-15, with an IC50 at approximately 5.0, 5.0, and 4.5 μg/ml, respectively. TOFA at 1.0–20.0 μg/ml effectively blocked fatty acid synthesis and induced cell death in a dose-dependent manner. The cell death was characterized with PARP cleavage, DNA fragmentation, and annexin-V staining, all of which are the features of the apoptosis. Supplementing simultaneously the cells with palmitic acids (100 μM), the end-products of the fatty acid synthesis pathway, prevented the apoptosis induced by TOFA. Taken together, these data suggest that TOFA is a potent cytotoxic agent to lung and colon cancer cells, inducing apoptosis through disturbing their fatty acid synthesis. PMID:19450551

  1. Asiatic Acid (AA) Sensitizes Multidrug-Resistant Human Lung Adenocarcinoma A549/DDP Cells to Cisplatin (DDP) via Downregulation of P-Glycoprotein (MDR1) and Its Targets.

    PubMed

    Cheng, Qilai; Liao, Meixiang; Hu, Haibo; Li, Hongliang; Wu, Longhuo

    2018-01-01

    P-glycoprotein (P-gp, i.e., MDR1) is associated with the phenotype of multidrug resistance (MDR) and causes chemotherapy failure in the management of cancers. Searching for effective MDR modulators and combining them with anticancer drugs is a promising strategy against MDR. Asiatic acid (AA), a natural triterpene isolated from the plant Centella asiatica, may have an antitumor activity. The present study assessed the reversing effect of AA on MDR and possible molecular mechanisms of AA action in MDR1-overexpressing cisplatin (DDP)-resistant lung cancer cells, A549/DDP. Human lung adenocarcinoma A549/DDP cells were either exposed to different concentrations of AA or treated with DDP, and their viability was measured by the MTT assay. A Rhodamine 123 efflux assay, immunofluorescent staining, ATPase assay, reverse-transcription PCR (RT-PCR), and western blot analysis were conducted to elucidate the mechanisms of action of AA on MDR. Our results showed that AA significantly enhanced the cytotoxicity of DDP toward A549/DDP cells but not its parental A549 cells. Furthermore, AA strongly inhibited P-gp expression by blocking MDR1 gene transcription and increased the intracellular accumulation of the P-gp substrate Rhodamine 123 in A549/DDP cells. Nuclear factor (NF)-kB (p65) activity, IkB degradation, and NF-kB/p65 nuclear translocation were markedly inhibited by pretreatment with AA. Additionally, AA inhibited the MAPK-ERK pathway, as indicated by decreased phosphorylation of ERK1 and -2, AKT, p38, and JNK, thus resulting in reduced activity of the Y-box binding protein 1 (YB1) via blockage of its nuclear translocation. AA reversed P-gp-mediated MDR by inhibition of P-gp expression. This effect was likely related to downregulation of YB1, and this effect was mediated by the NF-kB and MAPK-ERK pathways. AA may be useful as an MDR reversal agent for combination therapy in clinical trials. © 2018 The Author(s). Published by S. Karger AG, Basel.

  2. Hyperthermia: an effective strategy to induce apoptosis in cancer cells.

    PubMed

    Ahmed, Kanwal; Tabuchi, Yoshiaki; Kondo, Takashi

    2015-11-01

    Heat has been used as a medicinal and healing modality throughout human history. The combination of hyperthermia (HT) with radiation and anticancer agents has been used clinically and has shown positive results to a certain extent. However, the clinical results of HT treatment alone have been only partially satisfactory. Cell death following HT treatment is a function of both temperature and treatment duration. HT induces cancer cell death through apoptosis; the degree of apoptosis and the apoptotic pathway vary in different cancer cell types. HT-induced reactive oxygen species production are responsible for apoptosis in various cell types. However, the underlying mechanism of signal transduction and the genes related to this process still need to be elucidated. In this review, we summarize the molecular mechanism of apoptosis induced by HT, enhancement of heat-induced apoptosis, and the genetic network involved in HT-induced apoptosis.

  3. Graphene as a nanocarrier for tamoxifen induces apoptosis in transformed cancer cell lines of different origins.

    PubMed

    Misra, Santosh K; Kondaiah, Paturu; Bhattacharya, Santanu; Rao, C N R

    2012-01-09

    A cationic amphiphile, cholest-5en-3β-oxyethyl pyridinium bromide (PY(+) -Chol), is able to efficiently disperse exfoliated graphene (GR) in water by the physical adsorption of PY(+) -Chol on the surface of GR to form stable, dark aqueous suspensions at room temperature. The GR-PY(+) -Chol suspension can then be used to solubilize Tamoxifen Citrate (TmC), a breast cancer drug, in water. The resulting TmC-GR-PY(+) -Chol is stable for a long time without any precipitation. Fluorescence emission and UV absorption spectra indicate the existence of noncovalent interactions between TmC, GR, and PY(+) -Chol in these suspensions. Electron microscopy shows the existence of segregated GR sheets and TmC 'ribbons' in the composite suspensions. Atomic force microscopy indicates the presence of 'extended' structures of GR-PY(+) -Chol, which grows wider in the presence of TmC. The slow time-dependent release of TmC is noticed in a reconstituted cell culture medium, a property useful as a drug carrier. TmC-GR-PY(+) -Chol selectively enhanced the cell death (apoptosis) of the transformed cancer cells compared to normal cells. This potency is found to be true for a wide range of transformed cancer cells viz. HeLa, A549, ras oncogene-transformed NIH3T3, HepG2, MDA-MB231, MCF-7, and HEK293T compared to the normal cell HEK293 in vitro. Confocal microscopy confirmed the high efficiency of TmC-GR-PY(+) -Chol in delivering the drug to the cells, compared to the suspensions devoid of GR. Copyright © 2012 WILEY-VCH Verlag GmbH & Co. KGaA, Weinheim.

  4. Folic acid inhibits homocysteine-induced cell apoptosis in human umbilical vein endothelial cells.

    PubMed

    Cui, Shanshan; Li, Wen; Wang, Pengyan; Lv, Xin; Gao, Yuxia; Huang, Guowei

    2017-12-18

    Homocysteine may be responsible for vascular endothelial cell injury, which occurs early in the pathology of cardiovascular disease. Homocysteine metabolism requires enzymatic interaction with vitamins such as folic acid, vitamin B12, and vitamin B6. We hypothesized that folic acid alleviated homocysteine-induced vascular injury by regulating the metabolic pathway of apoptosis. Human umbilical vein endothelial cells were incubated for 48 h with folic acid at the concentrations of 0-1000 nmol/L, in combination with either 1000 μmol/L homocysteine or vehicle for the first 24 h. We then assessed cell viability and apoptosis by methyl thiazolyl tetrazolium assay and flow cytometry, respectively. To further investigate how folic acid influenced cell apoptosis, we also analyzed the activities of caspase-3/7 and the mRNA and protein expressions of BCL2, BAX, TP53, CASP3, and CASP8 in human umbilical vein endothelial cells. We showed that folic acid increased cell viability and decreased apoptosis in a dose-dependent manner, and that this effect was mediated by decreased caspase-3/7 activity, upregulated BCL2/BAX ratio, and downregulated TP53, CASP3, and CASP8 expressions. Thus, we conclude that folic acid inhibits cell apoptosis and ameliorates homocysteine toxicity by regulating the expression of apoptosis-related genes in human umbilical vein endothelial cells.

  5. Small Molecular TRAIL Inducer ONC201 Induces Death in Lung Cancer Cells: A Preclinical Study

    PubMed Central

    Feng, Yuan; Zhou, Jihong; Li, Zhanhua; Jiang, Ying; Zhou, Ying

    2016-01-01

    Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) selectively targets cancer cells. The present preclinical study investigated the anti-cancer efficiency of ONC201, a first-in-class small molecule TRAIL inducer, in lung cancer cells. We showed that ONC201 was cytotoxic and anti-proliferative in both established (A549 and H460 lines) and primary human lung cancer cells. It was yet non-cytotoxic to normal lung epithelial cells. Further, ONC201 induced exogenous apoptosis activation in lung cancer cells, which was evidenced by TRAIL/death receptor-5 (DR5) induction and caspase-8 activation. The caspase-8 inhibitor or TRAIL/DR5 siRNA knockdown alleviated ONC201’s cytotoxicity against lung cancer cells. Molecularly, ONC201 in-activated Akt-S6K1 and Erk signalings in lung cancer cells, causing Foxo3a nuclear translocation. For the in vivo studies, intraperitoneal injection of ONC201 at well-tolerated doses significantly inhibited xenografted A549 tumor growth in severe combined immunodeficient (SCID) mice. Further, ONC201 administration induced TRAIL/DR5 expression, yet inactivated Akt-S6K1 and Erk in tumor tissues. These results of the study demonstrates the potent anti-lung cancer activity by ONC201. PMID:27626799

  6. Small Molecular TRAIL Inducer ONC201 Induces Death in Lung Cancer Cells: A Preclinical Study.

    PubMed

    Feng, Yuan; Zhou, Jihong; Li, Zhanhua; Jiang, Ying; Zhou, Ying

    2016-01-01

    Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) selectively targets cancer cells. The present preclinical study investigated the anti-cancer efficiency of ONC201, a first-in-class small molecule TRAIL inducer, in lung cancer cells. We showed that ONC201 was cytotoxic and anti-proliferative in both established (A549 and H460 lines) and primary human lung cancer cells. It was yet non-cytotoxic to normal lung epithelial cells. Further, ONC201 induced exogenous apoptosis activation in lung cancer cells, which was evidenced by TRAIL/death receptor-5 (DR5) induction and caspase-8 activation. The caspase-8 inhibitor or TRAIL/DR5 siRNA knockdown alleviated ONC201's cytotoxicity against lung cancer cells. Molecularly, ONC201 in-activated Akt-S6K1 and Erk signalings in lung cancer cells, causing Foxo3a nuclear translocation. For the in vivo studies, intraperitoneal injection of ONC201 at well-tolerated doses significantly inhibited xenografted A549 tumor growth in severe combined immunodeficient (SCID) mice. Further, ONC201 administration induced TRAIL/DR5 expression, yet inactivated Akt-S6K1 and Erk in tumor tissues. These results of the study demonstrates the potent anti-lung cancer activity by ONC201.

  7. c-Myc plays a key role in TADs-induced apoptosis and cell cycle arrest in human hepatocellular carcinoma cells

    PubMed Central

    Zhang, Dongdong; Qi, Junpeng; Liu, Rui; Dai, Bingling; Ma, Weina; Zhan, Yingzhuan; Zhang, Yanmin

    2015-01-01

    Cancer cell growth is complicated progression which is regulated and controlled by multiple factors including cell cycle, migration and apoptosis. In present study, we report that TADs, a novel derivative of taspine, has an essential role in resisting hepatocellular carcinoma growth (including arrest cell cycle) and migration, and inducing cell apoptosis. Our findings demonstrated that the TADs showed good inhibition on the hepatoma cell growth and migration, and good action on apoptosis induction. Using genome-wide microarray analysis, we found the down-regulated growth and apoptosis factors, and selected down-regulated genes were confirmed by Western blot. Knockdown of a checkpoint c-Myc by siRNA significantly attenuated tumor inhibition and apoptosis effects of TADs. Moreover, our results indicated TADs could simultaneously increase cyclin D1 protein levels and decrease amount of cyclin E, cyclin B1 and cdc2 of the cycle proteins, and also TADs reduced Bcl-2 expression, and upregulated Bad, Bak and Bax activities. In conclusion, these results illustrated that TADs is a key factor in growth and apoptosis signaling inhibitor, has potential in cancer therapy. PMID:26045987

  8. c-Myc plays a key role in TADs-induced apoptosis and cell cycle arrest in human hepatocellular carcinoma cells.

    PubMed

    Zhang, Dongdong; Qi, Junpeng; Liu, Rui; Dai, Bingling; Ma, Weina; Zhan, Yingzhuan; Zhang, Yanmin

    2015-01-01

    Cancer cell growth is complicated progression which is regulated and controlled by multiple factors including cell cycle, migration and apoptosis. In present study, we report that TADs, a novel derivative of taspine, has an essential role in resisting hepatocellular carcinoma growth (including arrest cell cycle) and migration, and inducing cell apoptosis. Our findings demonstrated that the TADs showed good inhibition on the hepatoma cell growth and migration, and good action on apoptosis induction. Using genome-wide microarray analysis, we found the down-regulated growth and apoptosis factors, and selected down-regulated genes were confirmed by Western blot. Knockdown of a checkpoint c-Myc by siRNA significantly attenuated tumor inhibition and apoptosis effects of TADs. Moreover, our results indicated TADs could simultaneously increase cyclin D1 protein levels and decrease amount of cyclin E, cyclin B1 and cdc2 of the cycle proteins, and also TADs reduced Bcl-2 expression, and upregulated Bad, Bak and Bax activities. In conclusion, these results illustrated that TADs is a key factor in growth and apoptosis signaling inhibitor, has potential in cancer therapy.

  9. Centchroman induces redox-dependent apoptosis and cell-cycle arrest in human endometrial cancer cells.

    PubMed

    Shyam, Hari; Singh, Neetu; Kaushik, Shweta; Sharma, Ramesh; Balapure, Anil K

    2017-04-01

    Centchroman (CC) or Ormeloxifene has been shown to induce apoptosis and cell cycle arrest in various types of cancer cells. This has, however, not been addressed for endometrial cancer cells where its (CC) mechanism of action remains unclear. This study focuses on the basis of antineoplasticity of CC by blocking the targets involved in the cell cycle, survival and apoptosis in endometrial cancer cells. Ishikawa Human Endometrial Cancer Cells were cultured under estrogen deprived medium, exposed to CC and analyzed for proliferation and apoptosis. Additionally, we also analyzed oxidative stress induced by CC. Cell viability studies confirmed the IC 50 of CC in Ishikawa cells to be 20 µM after 48 h treatment. CC arrests the cells in G0/G1 phase through cyclin D1 and cyclin E mediated pathways. Phosphatidylserine externalization, nuclear morphology changes, DNA fragmentation, PARP cleavage, and alteration of Bcl-2 family protein expression clearly suggest ongoing apoptosis in the CC treated cells. Activation of caspase 3 & 9, up-regulation of AIF and inhibition of apoptosis by z-VAD-fmk clearly explains the participation of the intrinsic pathway of programmed cell death. Further, the increase of ROS, loss of MMP, inhibition of antioxidant (MnSOD, Cu/Zn-SOD and GST) and inhibition of apoptosis with L-NAC suggests CC induced oxidative stress leading to apoptosis via mitochondria mediated pathway. Therefore, CC could be a potential therapeutic agent for the treatment of Endometrial Cancer adjunct to its utility as a contraceptive and an anti-breast cancer agent.

  10. Exposure to diethylhexyl phthalate (DEHP) and monoethylhexyl phthalate (MEHP) promotes the loss of alveolar epithelial phenotype of A549 cells.

    PubMed

    Rafael-Vázquez, L; García-Trejo, Semiramis; Aztatzi-Aguilar, O G; Bazán-Perkins, B; Quintanilla-Vega, B

    2018-05-17

    Di(2-ethylhexyl) phthalate (DEHP) is a widely used plasticizer that is metabolized to mono(2-ethylhexyl) phthalate (MEHP). Inhalation is an important exposure route for both phthalates, and their effects on lungs include inflammation, alteration of postnatal maturation (alveolarization), enlarged airspaces and cell differentiation changes, suggesting that alveolar epithelial cells-2 (AEC) are targets of phthalates. This study evaluated the cell progression, epithelial and mesenchymal markers, including surfactant secretion in A549 cells (AEC) that were exposed to DEHP (1-100 μM) or MEHP (1-50 μM) for 24-72 h. The results showed an increased cell proliferation at all concentrations of each phthalate at 24 and 48 h. Cell migration showed a concentration-dependent increase at 24 and 48 h of exposure to either phthalate and enlarged structures were seen. Decreased levels of both surfactants (SP-B/SP-C) were observed after the exposure to either phthalate at 48 h, and of SP-C positive cells exposed to MEHP, suggesting a loss of the epithelial phenotype. While a decrease in the epithelial marker E-cadherin and an increase in the mesenchymal marker fibronectin were observed following exposure to either phthalate. Our results showed that DEHP and MEHP altered the structure and migration of A549 cells and promoted the loss of the epithelial phenotype. Copyright © 2018 Elsevier B.V. All rights reserved.

  11. TU-H-CAMPUS-TeP3-01: Gold Nanoparticle-Enhanced Radiation Therapy in In Vitro A549 Lung Carcinoma: Studies in Both Traditional Monolayer and Three Dimensional Cell Culture Models

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Oumano, M; University of Massachusetts Lowell, Lowell, MA; Ngwa, W

    Purpose: To measure the increase in in vitro radiosensitivity for A549 lung carcinoma cells due to gold nanoparticle (GNP) radiation dose enhancement in both traditional monolayer and three dimensional (3D) cell culture models. Methods: A γH2AX immunofluorescence assay is performed on monolayer A549 cell culture and quantitatively analyzed to measure the increase in double strand breaks (DSBs) resulting from GNP dose enhancement. A clonogenic survival assay (CSA) is then performed on monolayer A549 cell culture to assess true viability after treatment. And lastly, another γH2AX assay is performed on 3D A549 multicellular nodules overlaid on a bed of growth factormore » reduced matrigel to measure dose response in a model that better recapitulates treatment response to actual tumors in vivo. Results: The first γH2AX assay performed on the monolayer cell culture shows a significant increase in DSBs due to GNP dose enhancement. The maximum average observed increase in normalized fluorescent intensity for monolayer cell culture is 171% for the 6Gy-treatment groups incubated in 0.556 mg Au/ml solution. The CSA performed on monolayer cell culture also shows considerable GNP dose enhancement. The maximum decrease in the normalized surviving fraction is 12% for the 4Gy-treatment group incubated in 0.556 mg Au/ml. And lastly, the GNP dose enhancement is confirmed to be mitigated in three dimensional cell culture models as compared to the traditional monolayer model. The maximum average observed dose enhancement for 3D cell culture is 19% for the 6Gy-treatment groups and incubated in 0.556 mg Au/ml. Conclusion: A marked increase in radiosensitivity is observed for A549 lung carcinoma cells when treated with GNPs plus radiation as opposed to radiation alone. Traditional monolayer cell culture also shows a much more pronounced radiation dose enhancement than 3D cell culture.« less

  12. Bupivacaine-induced apoptosis independently of WDR35 expression in mouse neuroblastoma Neuro2a cells

    PubMed Central

    2012-01-01

    Background Bupivacaine-induced neurotoxicity has been shown to occur through apoptosis. Recently, bupivacaine was shown to elicit reactive oxygen species (ROS) production and induce apoptosis accompanied by activation of p38 mitogen-activated protein kinase (MAPK) in a human neuroblastoma cell line. We have reported that WDR35, a WD40-repeat protein, may mediate apoptosis through caspase-3 activation. The present study was undertaken to test whether bupivacaine induces apoptosis in mouse neuroblastoma Neuro2a cells and to determine whether ROS, p38 MAPK, and WDR35 are involved. Results Our results showed that bupivacaine induced ROS generation and p38 MAPK activation in Neuro2a cells, resulting in apoptosis. Bupivacaine also increased WDR35 expression in a dose- and time-dependent manner. Hydrogen peroxide (H2O2) also increased WDR35 expression in Neuro2a cells. Antioxidant (EUK-8) and p38 MAPK inhibitor (SB202190) treatment attenuated the increase in caspase-3 activity, cell death and WDR35 expression induced by bupivacaine or H2O2. Although transfection of Neuro2a cells with WDR35 siRNA attenuated the bupivacaine- or H2O2-induced increase in expression of WDR35 mRNA and protein, in contrast to our previous studies, it did not inhibit the increase in caspase-3 activity in bupivacaine- or H2O2-treated cells. Conclusions In summary, our results indicated that bupivacaine induced apoptosis in Neuro2a cells. Bupivacaine induced ROS generation and p38 MAPK activation, resulting in an increase in WDR35 expression, in these cells. However, the increase in WDR35 expression may not be essential for the bupivacaine-induced apoptosis in Neuro2a cells. These results may suggest the existence of another mechanism of bupivacaine-induced apoptosis independent from WDR35 expression in Neuro2a cells. PMID:23227925

  13. Tumor-targeting magnetic lipoplex delivery of short hairpin RNA suppresses IGF-1R overexpression of lung adenocarcinoma A549 cells in vitro and in vivo

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Wang, Chunmao; Ding, Chao; Kong, Minjian

    2011-07-08

    Highlights: {yields} We compared lipofection with magnetofection about difference of transfection efficiency on delivery a therapeutic gene in vitro and in vivo. {yields} We investigated the difference of shRNA induced by magnetofection and lipofection into A549 cell and subcutaneous tumor to knockdown IGF-1R overexpressed in A549 cell and A549 tumor. {yields} We investigated in vivo shRNA silenced IGF-1R overexpression 24, 48, and 72 h after shRNA intravenous injection into tumor-bearing mice by way of magnetofection and lipofection. {yields} Our results showed that magnetofection could achieve therapeutic gene targeted delivery into special site, which contributed to targeted gene therapy of lungmore » cancers. -- Abstract: Liposomal magnetofection potentiates gene transfection by applying a magnetic field to concentrate magnetic lipoplexes onto target cells. Magnetic lipoplexes are self-assembling ternary complexes of cationic lipids with plasmid DNA associated with superparamagnetic iron oxide nanoparticles (SPIONs). Type1insulin-like growth factor receptor (IGF-1R), an important oncogene, is frequently overexpressed in lung cancer and mediates cancer cell proliferation and tumor growth. In this study, we evaluated the transfection efficiency (percentage of transfected cells) and therapeutic potential (potency of IGF-1R knockdown) of liposomal magnetofection of plasmids expressing GFP and shRNAs targeting IGF-1R (pGFPshIGF-1Rs) in A549 cells and in tumor-bearing mice as compared to lipofection using Lipofectamine 2000. Liposomal magnetofection provided a threefold improvement in transgene expression over lipofection and transfected up to 64.1% of A549 cells in vitro. In vitro, IGF-1R specific-shRNA transfected by lipofection inhibited IGF-1R protein by 56.1 {+-} 6% and by liposomal magnetofection by 85.1 {+-} 3%. In vivo delivery efficiency of the pGFPshIGF-1R plasmid into the tumor was significantly higher in the liposomal magnetofection group than in the

  14. Label-free measurements on cell apoptosis using a terahertz metamaterial-based biosensor

    NASA Astrophysics Data System (ADS)

    Zhang, Caihong; Liang, Lanju; Ding, Liang; Jin, Biaobing; Hou, Yayi; Li, Chun; Jiang, Ling; Liu, Weiwei; Hu, Wei; Lu, Yanqing; Kang, Lin; Xu, Weiwei; Chen, Jian; Wu, Peiheng

    2016-06-01

    Label-free, real-time, and in-situ measurement on cell apoptosis is highly desirable in cell biology. We propose here a design of terahertz (THz) metamaterial-based biosensor for meeting this requirement. This metamaterial consists of a planar array of five concentric subwavelength gold ring resonators on a 10 μm-thick polyimide substrate, which can sense the change of dielectric environment above the metamaterial. We employ this sensor to an oral cancer cell (SCC4) with and without cisplatin, a chemotherapy drug for cancer treatment, and find a linear relation between cell apoptosis measured by Flow Cytometry and the relative change of resonant frequencies of the metamaterial measured by THz time-domain spectroscopy. This implies that we can determine the cell apoptosis in a label-free manner. We believe that this metamaterial-based biosensor can be developed into a cheap, label-free, real-time, and in-situ detection tool, which is of significant impact on the study of cell biology.

  15. MiR-509-3-5p causes aberrant mitosis and anti-proliferative effect by suppression of PLK1 in human lung cancer A549 cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Wang, Xian-Hui; Lu, Yao; Liang, Jing-Jing

    MicroRNAs (miRNAs) are potent post-transcriptional regulators of gene expression and play roles in DNA damage response (DDR). PLK1 is identified as a modulator of DNA damage checkpoint. Although down-regulation of PLK1 by certain microRNAs has been reported, little is known about the interplay between PLK1 and miR-509-3-5p in DDR. Here we have demonstrated that miR-509-3-5p repressed PLK1 expression by targeting PLK1 3′-UTR, thereby causing mitotic aberration and growth arrest of human lung cancer A549 cells. Repression of PLK1 by miR-509-3-5p was further evidenced by over-expression of miR-509-3-5p in A549, HepG2 and HCT116p53{sup −/−} cancer cells, in which PLK1 protein wasmore » suppressed. Consistently, miR-509-3-5p was stimulated, while PLK1 protein was down-regulated in A549 cells exposed to CIS and ADR, suggesting that suppression of PLK1 by miR-509-3-5p is a component of CIS/ADR-induced DDR pathway. Flow cytometry and immunofluorescence labeling showed that over-expression of miR-509-3-5p in A549 induced G2/M arrest and aberrant mitosis characterized by abnormal bipolar mitotic spindles, condensed chromosomes, lagging DNA and chromosome bridges. In addition, over-expression of miR-509-3-5p markedly blocked A549 cell proliferation and sensitized the cells to CIS and ADR treatment. Taken together, miR-509-3-5p is a feasible suppressor for cancer by targeting PLK1. Our data may provide aid in potential design of combined chemotherapy and in our better understanding of the roles of microRNAs in response to DNA damage. - Highlights: • MiR-509-3-5p represses PLK1 expression by targeting PLK1 3ГЉВ№-UTR. • Expression of miR-509-3-5p is induced and PLK1 repressed upon DNA damage. • Overexpression of miR-509-3-5p induces G2/M arrest and aberrant mitosis. • MiR-509-3-5p inhibits cell proliferation and sensitizes cells to DNA damage agents.« less

  16. The role of ARK in stress-induced apoptosis in Drosophila cells

    PubMed Central

    Zimmermann, Katja C.; Ricci, Jean-Ehrland; Droin, Nathalie M.; Green, Douglas R.

    2002-01-01

    The molecular mechanisms of apoptosis are highly conserved throughout evolution. The homologs of genes essential for apoptosis in Caenorhabditis elegans and Drosophila melanogaster have been shown to be important for apoptosis in mammalian systems. Although a homologue for CED-4/apoptotic protease-activating factor (Apaf)-1 has been described in Drosophila, its exact function and the role of the mitochondrial pathway in its activation remain unclear. Here, we used the technique of RNA interference to dissect apoptotic signaling pathways in Drosophila cells. Inhibition of the Drosophila CED-4/Apaf-1–related killer (ARK) homologue resulted in pronounced inhibition of stress-induced apoptosis, whereas loss of ARK did not protect the cells from Reaper- or Grim-induced cell death. Reduction of DIAP1 induced rapid apoptosis in these cells, whereas the inhibition of DIAP2 expression did not but resulted in increased sensitivity to stress-induced apoptosis; apoptosis in both cases was prevented by inhibition of ARK expression. Cells in which cytochrome c expression was decreased underwent apoptosis induced by stress stimuli, Reaper or Grim. These results demonstrate the central role of ARK in stress-induced apoptosis, which appears to act independently of cytochrome c. Apoptosis induced by Reaper or Grim can proceed via a distinct pathway, independent of ARK. PMID:11901172

  17. Glutamine drives glutathione synthesis and contributes to radiation sensitivity of A549 and H460 lung cancer cell lines

    PubMed Central

    Sappington, Daniel R.; Siegel, Eric R.; Hiatt, Gloria; Desai, Abhishek; Penney, Rosalind B.; Jamshidi-Parsian, Azemat; Griffin, Robert J.; Boysen, Gunnar

    2016-01-01

    Background Increased glutamine uptake is known to drive cancer cell proliferation, making tumor cells glutamine-dependent. Glutamine provides additional carbon and nitrogen sources for cell growth. The first step in glutamine utilization is its conversion to glutamate by glutaminase (GLS). Glutamate is a precursor for glutathione synthesis, and we investigated the hypothesis that glutamine drives glutathione synthesis and thereby contributes to cellular defense systems. Methods The importance of glutamine for glutathione synthesis was studied in H460 and A549 lung cancer cell lines using glutamine-free medium and Bis-2-(5-phenyl-acetamido-1,3,4-thiadiazol-2-yl)ethyl sulfide (BPTES) a GLS inhibitor. Metabolic activities were determined by targeted mass spectrometry. Results A significant correlation between glutamine consumption and glutathione excretion was demonstrated in H460 and A549 tumor cells. Culturing in the presence of [13C5]glutamine demonstrated that by 12 hrs >50% of excreted glutathione is derived from glutamine. Culturing in glutamine-free medium or treatment with BPTES, a glutaminase (GLS)-specific inhibitor, reduced cell proliferation and viability, and abolished glutathione excretion. Treatment with glutathione-ester prevented BPTES induced cytotoxicity. Inhibition of GLS markedly radiosensitized the lung tumor cell lines, suggesting an important role of glutamine-derived glutathione in determining radiation sensitivity. Conclusions We demonstrate here for the first time that a significant amount of extracellular glutathione is directly derived from glutamine. This finding adds yet another important function to the already known glutamine dependence of tumor cells and probably tumors as well. General significance Glutamine is essential for synthesis and excretion of glutathione to promote cell growth and viability. PMID:26825773

  18. Apoptosis inhibitor of macrophage (AIM) reduces cell number in canine histiocytic sarcoma cell lines.

    PubMed

    Uchida, Mona; Saeki, Kohei; Maeda, Shingo; Tamahara, Satoshi; Yonezawa, Tomohiro; Matsuki, Naoaki

    2016-10-01

    Apoptosis inhibitor of macrophage (AIM) is initially reported to protect macrophages from apoptosis. In this study, we determined the effect of AIM on the macrophage-derived tumor, histiocytic sarcoma cell lines (HS) of dogs. Five HS and five other tumor cell lines were used. When recombinant canine AIM was applied to non-serum culture media, cell numbers of all the HS and two of other tumor cell lines decreased dose-dependently. The DNA fragmentation, TUNEL staining and flow cytometry tests revealed that AIM induced both of apoptosis and cell cycle arrest in the HS. Although AIM is known as an apoptosis inhibitor, these results suggest that a high dose of AIM could have an opposite function in HS and some tumor cell lines.

  19. Tumor Necrosis Factor-related Apoptosis-inducing Ligand (TRAIL)-Troglitazone-induced Apoptosis in Prostate Cancer Cells Involve AMP-activated Protein Kinase*

    PubMed Central

    Santha, Sreevidya; Viswakarma, Navin; Das, Subhasis; Rana, Ajay; Rana, Basabi

    2015-01-01

    Prostate cancer (PCa) is one of the most frequently diagnosed cancers in men with limited treatment options for the hormone-resistant forms. Development of novel therapeutic options is critically needed to target advanced forms. Here we demonstrate that combinatorial treatment with the thiazolidinedione troglitazone (TZD) and TNF-related apoptosis-inducing ligand (TRAIL) can induce significant apoptosis in various PCa cells independent of androgen receptor status. Because TZD is known to activate AMP-activated protein kinase (AMPK), we determined whether AMPK is a molecular target mediating this apoptotic cascade by utilizing PCa cell lines stably overexpressing AMPKα1 dominant negative (C4-2-DN) or empty vector (C4-2-EV). Our results indicated a significantly higher degree of apoptosis with TRAIL-TZD combination in C4-2-EV cells compared with C4-2-DN cells. Similarly, results from a 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay showed a larger reduction of viability of C4-2-EV cells compared with C4-2-DN cells when treated with TRAIL-TZD, thus suggesting that C4-2-DN cells were more apoptosis-resistant. Additionally, siRNA-mediated knockdown of endogenous AMPKα1 expression showed a reduction of TRAIL-TZD-induced apoptosis, further confirming the participation of AMPK in mediating this apoptosis. Apoptosis induction by this combinatorial treatment was also associated with a cleavage of β-catenin that was inhibited in both C4-2-DN cells and those cells in which AMPKα1 was knocked down. In addition, time course studies showed an increase in pACCS79 (AMPK target) levels coinciding with the time of apoptosis. These studies indicate the involvement of AMPK in TRAIL-TZD-mediated apoptosis and β-catenin cleavage and suggest the possibility of utilizing AMPK as a therapeutic target in apoptosis-resistant prostate cancer. PMID:26198640

  20. JAK/STAT autocontrol of ligand-producing cell number through apoptosis.

    PubMed

    Borensztejn, Antoine; Boissoneau, Elisabeth; Fernandez, Guillaume; Agnès, François; Pret, Anne-Marie

    2013-01-01

    During development, specific cells are eliminated by apoptosis to ensure that the correct number of cells is integrated in a given tissue or structure. How the apoptosis machinery is activated selectively in vivo in the context of a developing tissue is still poorly understood. In the Drosophila ovary, specialised follicle cells [polar cells (PCs)] are produced in excess during early oogenesis and reduced by apoptosis to exactly two cells per follicle extremity. PCs act as an organising centre during follicle maturation as they are the only source of the JAK/STAT pathway ligand Unpaired (Upd), the morphogen activity of which instructs distinct follicle cell fates. Here we show that reduction of Upd levels leads to prolonged survival of supernumerary PCs, downregulation of the pro-apoptotic factor Hid, upregulation of the anti-apoptotic factor Diap1 and inhibition of caspase activity. Upd-mediated activation of the JAK/STAT pathway occurs in PCs themselves, as well as in adjacent terminal follicle and interfollicular stalk cells, and inhibition of JAK/STAT signalling in any one of these cell populations protects PCs from apoptosis. Thus, a Stat-dependent unidentified relay signal is necessary for inducing supernumerary PC death. Finally, blocking apoptosis of PCs leads to specification of excess adjacent border cells via excessive Upd signalling. Our results therefore show that Upd and JAK/STAT signalling induce apoptosis of supernumerary PCs to control the size of the PC organising centre and thereby produce appropriate levels of Upd. This is the first example linking this highly conserved signalling pathway with developmental apoptosis in Drosophila.

  1. GSK-3β mediates dexamethasone-induced pancreatic β cell apoptosis

    PubMed Central

    Guo, Bin; Zhang, Wenjian; Xu, Shiqing; Lou, Jinning; Wang, Shuxia; Men, Xiuli

    2015-01-01

    Aims Glucocorticoids, such as dexamethasone, are widely used anti-inflammatory drugs. Their use is frequently associated with the development of steroid- associated diabetes. Pancreatic β-cell dysfunction has been suggested to be one of the main causes of steroid-associated diabetes. However, the mechanism is not fully understood. Glycogen synthase kinase-3β (GSK-3β) is a multifunctional serine/threonine kinase and plays an important role in energy metabolism, cell growth and apoptosis. Therefore, the contribution of GSK-3β in dexamethasone-induced pancreatic β-cell apoptosis was determined in the present study. Main Methods The effect of dexamethasone treatment on rat pancreatic β-cell line (INS-1) apoptosis (determined by TUNEL and Flow Cytometry), generation of reactive oxidative stress (ROS), and the phosphorylation status of GSK-3β was determined. The inhibitory effect of GSK-3β inhibitor-lithium chloride (LiCl) on dexamethasone-induced β-cell apoptosis was also evaluated. Key Findings Dexamethasone (0.1 μM) treatment induced INS-1 apoptosis, which was associated with increased GSK-3β activation and increased NOX4-derived ROS generation. Pretreatment of INS-1 with LiCl inhibited dexamethasone induced ROS generation and INS-1 apoptosis. Significance This study provides a new mechanism of Dex induced pancreatic β cell apoptosis and may serve as a new therapeutic option for treating GCs induced diabetes. PMID:26606859

  2. Pellino-1 confers chemoresistance in lung cancer cells by upregulating cIAP2 through Lys63-mediated polyubiquitination

    PubMed Central

    Koh, Jaemoon; Chung, Doo Hyun

    2016-01-01

    Pellino-1 is an E3 ubiquitin ligase that mediates immune receptor signaling pathways. The role of Pellino-1 in oncogenesis of lung cancer was investigated in this study. Pellino-1 expression was increased in human lung cancer cell lines compared with non-neoplastic lung cell lines. Pellino-1 overexpression in human lung cancer cells, A549 and H1299 cells, increased the survival and colony forming ability. Pellino-1 overexpression in these cells also conferred resistance to cisplatin- or paclitaxel-induced apoptosis. In contrast, depletion of Pellino-1 decreased the survival of A549 and H1299 cells and sensitized these cells to cisplatin- and paclitaxel-induced apoptosis. Pellino-1 overexpression in A549 and H1299 cells upregulated the expression of inhibitor of apoptosis (IAP) proteins, including cIAP1 and cIAP2, while Pellino-1 depletion downregulated these molecules. Notably, Pellino-1 directly interacted with cIAP2 and stabilized cIAP2 through lysine63-mediated polyubiquitination via its E3 ligase activity. Pellino-1-mediated chemoresistance in lung cancer cells was dependent on the induction of cIAP2. Moreover, a strong positive correlation between Pellino-1 and the cIAP2 expression was observed in human lung adenocarcinoma tissues. Taken together, these results demonstrate that Pellino-1 contributes to lung oncogenesis through the overexpression of cIAP2 and promotion of cell survival and chemoresistance. Pellino-1 might be a novel oncogene and potential therapeutic target in lung cancer. PMID:27248820

  3. EVA1A inhibits GBM cell proliferation by inducing autophagy and apoptosis

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Shen, Xue; Kan, Shifeng; Liu, Zhen

    Eva-1 homolog A (EVA1A) is a novel lysosome and endoplasmic reticulum-associated protein involved in autophagy and apoptosis. In this study, we constructed a recombinant adenovirus 5-EVA1A vector (Ad5-EVA1A) to overexpress EVA1A in glioblastoma (GBM) cell lines and evaluated its anti-tumor activities in vitro and in vivo. We found that overexpression of EVA1A in three GBM cell lines (U251, U87 and SHG44) resulted in a suppression of tumor cell growth via activation of autophagy and induction of cell apoptosis in a dose- and time-dependent manner. EVA1A-mediated autophagy was associated with inactivation of the mTOR/RPS6KB1 signaling pathway. Furthermore in vivo, overexpression ofmore » EVA1A successfully inhibited tumor growth in NOD/SCID mice. Our data suggest that EVA1A-induced autophagy and apoptosis play a role in suppressing the development of GBM and their up-regulation may be an effective method for treating this form of cancer. - Highlights: • Overexpression of EVA1A suppresses GBM cell growth. • EVA1A induces autophagy through the mTOR/RPS6KB1 pathway. • EVA1A induces GBM cell apoptosis. • EVA1A inhibits the development of GBM in vivo.« less

  4. Pleuropterus multiflorus (Hasuo) mediated straightforward eco-friendly synthesis of silver, gold nanoparticles and evaluation of their anti-cancer activity on A549 lung cancer cell line.

    PubMed

    Castro-Aceituno, Verónica; Abbai, Ragavendran; Moon, Seong Soo; Ahn, Sungeun; Mathiyalagan, Ramya; Kim, Yu-Jin; Kim, Yeon-Ju; Yang, Deok Chun

    2017-09-01

    Pleuropterus multiflorus (Hasuo) is a widely used medicinal plant in Korea and China for treating amnesia, isnomia, heart throbbing etc. With the constructive idea of promoting the wide-spread usage of P. multiflorus, we propose its indirect usage in the form of biologically active silver (Pm-AgNPs) and gold nanoparticles (Pm-AuNPs). The synthesized nanoparticles were predominantly spherical, crystalline with the Z-average hydrodynamic diameter of 274.8nm and 104.8nm respectively. Also, proteins and phenols were identified as the major players involved in their synthesis and stability. Further, Pm-AgNPs at 25μg/mL were significantly cytotoxic to lung cancer cells, whereas, Pm-AuNPs were not cytotoxic to both normal keratinocyte and lung cancer cells even at 100μg/mL. In addition, further evaluation of the anti-cancer activity of these new nanoparticles, such as migration and apoptosis, shown that Pm-AgNPs have a potential therapeutic effect on A549 lung cancer cell treatment. To the best of our knowledge, this is the first report dissecting out the ability of the endemic P. multiflorus for the synthesis of bioactive silver and gold nanoparticle which would open up doors for its extensive usage in medicinal field. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  5. Salidroside induces cell-cycle arrest and apoptosis in human breast cancer cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Hu, Xiaolan, E-mail: huxiaolan1998@yahoo.com.cn; Zhang, Xianqi; Qiu, Shuifeng

    2010-07-16

    Research highlights: {yields} Salidroside inhibits the growth of human breast cancer cells. {yields} Salidroside induces cell-cycle arrest of human breast cancer cells. {yields} Salidroside induces apoptosis of human breast cancer cell lines. -- Abstract: Recently, salidroside (p-hydroxyphenethyl-{beta}-D-glucoside) has been identified as one of the most potent compounds isolated from plants of the Rhodiola genus used widely in traditional Chinese medicine, but pharmacokinetic data on the compound are unavailable. We were the first to report the cytotoxic effects of salidroside on cancer cell lines derived from different tissues, and we found that human breast cancer MDA-MB-231 cells (estrogen receptor negative) weremore » sensitive to the inhibitory action of low-concentration salidroside. To further investigate the cytotoxic effects of salidroside on breast cancer cells and reveal possible ER-related differences in response to salidroside, we used MDA-MB-231 cells and MCF-7 cells (estrogen receptor-positive) as models to study possible molecular mechanisms; we evaluated the effects of salidroside on cell growth characteristics, such as proliferation, cell cycle duration, and apoptosis, and on the expression of apoptosis-related molecules. Our results demonstrated for the first time that salidroside induces cell-cycle arrest and apoptosis in human breast cancer cells and may be a promising candidate for breast cancer treatment.« less

  6. Sulfamic and succinic acid derivatives of 25-OH-PPD and their activities to MCF-7, A-549, HCT-116, and BGC-823 cell lines.

    PubMed

    Zhou, Wu-Xi; Cao, Jia-Qing; Wang, Xu-De; Guo, Jun-Hui; Zhao, Yu-Qing

    2017-02-15

    In the search for new anti-tumor agents with higher potency than our previously identified compound 1 (25-OH-PPD, 25-hydroxyprotopanaxadiol), 12 novel sulfamic and succinic acid derivatives that could improve water solubility and contribute to good drug potency and pharmacokinetic profiles were designed and synthesized. Their in vitro anti-tumor activities in MCF-7, A-549, HCT-116, and BGC-823 cell lines and one normal cell line were tested by standard MTT assay. Results showed that compared with compound 1, compounds 2, 3, and 7 exhibited higher cytotoxic activity on A-549 and BGC-823 cell lines, together with lower toxicity in the normal cell. In particular, compound 2 exhibited the best anti-tumor activity in the in vitro assays, which may provide valuable data for the research and development of new anti-tumor agents. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. Apoptotic cells can induce non-autonomous apoptosis through the TNF pathway

    PubMed Central

    Pérez-Garijo, Ainhoa; Fuchs, Yaron; Steller, Hermann

    2013-01-01

    Apoptotic cells can produce signals to instruct cells in their local environment, including ones that stimulate engulfment and proliferation. We identified a novel mode of communication by which apoptotic cells induce additional apoptosis in the same tissue. Strong induction of apoptosis in one compartment of the Drosophila wing disc causes apoptosis of cells in the other compartment, indicating that dying cells can release long-range death factors. We identified Eiger, the Drosophila tumor necrosis factor (TNF) homolog, as the signal responsible for apoptosis-induced apoptosis (AiA). Eiger is produced in apoptotic cells and, through activation of the c-Jun N-terminal kinase (JNK) pathway, is able to propagate the initial apoptotic stimulus. We also show that during coordinated cell death of hair follicle cells in mice, TNF-α is expressed in apoptotic cells and is required for normal cell death. AiA provides a mechanism to explain cohort behavior of dying cells that is seen both in normal development and under pathological conditions. DOI: http://dx.doi.org/10.7554/eLife.01004.001 PMID:24066226

  8. Microenvironment mesenchymal cells protect ovarian cancer cell lines from apoptosis by inhibiting XIAP inactivation

    PubMed Central

    Castells, M; Milhas, D; Gandy, C; Thibault, B; Rafii, A; Delord, J-P; Couderc, B

    2013-01-01

    Epithelial ovarian carcinoma is characterized by high frequency of recurrence (70% of patients) and carboplatin resistance acquisition. Carcinoma-associated mesenchymal stem cells (CA-MSC) have been shown to induce ovarian cancer chemoresistance through trogocytosis. Here we examined CA-MSC properties to protect ovarian cancer cells from carboplatin-induced apoptosis. Apoptosis was determined by Propidium Iodide and Annexin-V-FITC labelling and poly-ADP-ribose polymerase cleavage analysis. We showed a significant increase of inhibitory concentration 50 and a 30% decrease of carboplatin-induced apoptosis in ovarian cancer cells incubated in the presence of CA-MSC-conditioned medium (CM). A molecular analysis of apoptosis signalling pathway in response to carboplatin revealed that the presence of CA-MSC CM induced a 30% decrease of effector caspases-3 and -7 activation and proteolysis activity. CA-MSC secretions promoted Akt and X-linked inhibitor of apoptosis protein (XIAP; caspase inhibitor from inhibitor of apoptosis protein (IAP) family) phosphorylation. XIAP depletion by siRNA strategy permitted to restore apoptosis in ovarian cancer cells stimulated by CA-MSC CM. The factors secreted by CA-MSC are able to confer chemoresistance to carboplatin in ovarian cancer cells through the inhibition of effector caspases activation and apoptosis blockade. Activation of the phosphatidylinositol 3-kinase (PI3K)/Akt signalling pathway and the phosphorylation of its downstream target XIAP underlined the implication of this signalling pathway in ovarian cancer chemoresistance. This study reveals the potentialities of targeting XIAP in ovarian cancer therapy. PMID:24176845

  9. Titanium dioxide nanoparticles-mediated in vitro cytotoxicity does not induce Hsp70 and Grp78 expression in human bronchial epithelial A549 cells.

    PubMed

    Aueviriyavit, Sasitorn; Phummiratch, Duangkamol; Kulthong, Kornphimol; Maniratanachote, Rawiwan

    2012-10-01

    Titanium dioxide nanoparticles (TiO(2)NPs) are increasingly being used in various industrial applications including the production of paper, plastics, cosmetics and paints. With the increasing number of nano-related products, the concern of governments and the general public about the health and environmental risks, especially with regard to occupational and other environmental exposure, are gradually increasing. However, there is insufficient knowledge about the actual affects upon human health and the environment, as well as a lack of suitable biomarkers for assessing TiO(2)NP-induced cytotoxicity. Since the respiratory tract is likely to be the main exposure route of industrial workers to TiO(2)NPs, we investigated the cytotoxicity of the anatase and rutile crystalline forms of TiO(2)NPs in A549 cells, a human alveolar type II-like epithelial cell line. In addition, we evaluated the transcript and protein expression levels of two heat shock protein (HSP) members, Grp78 and Hsp70, to ascertain their suitability as biomarkers of TiO(2)NP-induced toxicity in the respiratory system. Ultrastructural observations confirmed the presence of TiO(2)NPs inside cells. In vitro exposure of A549 cells to the anatase or rutile forms of TiO(2)NPs led to cell death and induced intracellular ROS generation in a dose-dependent manner, as determined by the MTS and dichlorofluorescein (DCF) assays, respectively. In contrast, the transcript and protein expression levels of Hsp70 and Grp78 did not change within the same TiO(2)NPs dose range (25-500 μg/ml). Thus, whilst TiO(2)NPs can cause cytotoxicity in A549 cells, and thus potentially in respiratory cells, Hsp70 and Grp78 are not suitable biomarkers for evaluating the acute toxicological effects of TiO(2)NPs in the respiratory system.

  10. Evodiamine Induces Cell Growth Arrest, Apoptosis and Suppresses Tumorigenesis in Human Urothelial Cell Carcinoma Cells.

    PubMed

    Shi, Chung-Sheng; Li, Jhy-Ming; Chin, Chih-Chien; Kuo, Yi-Hung; Lee, Ying-Ray; Huang, Yun-Ching

    2017-03-01

    Evodiamine, an indole alkaloid derived from Evodia rutaecarpa, exhibits pharmacological activities including vasodilatation, analgesia, anti-cardiovascular disease, anti-Alzheimer's disease, anti-inflammation, and anti-tumor activity. This study analyzes the anti-tumor effects of evodiamine on cellular growth, tumorigenesis, cell cycle and apoptosis induction of human urothelial cell carcinoma (UCC) cells. The present study showed that evodiamine significantly inhibited the proliferation of UCC cells in a dose- and time-dependent manner. Also, evodiamine suppressed the tumorigenesis of UCC cells in vitro. Moreover, evodiamine caused G 2 /M cell-cycle arrest and induced caspase-dependent apoptosis in UCC cells. Finally, we demonstrated that evodiamine exhibits better cytotoxic than 5-fluorouracil, a clinical chemotherapeutic drug, for UCC cells. Evodiamine induces growth inhibition, tumorigenesis suppression, cell-cycle arrest, and apoptosis induction in human UCC cells. Therefore, this agent displays a therapeutic potential for treating human UCC cells and is worthy for further investigation. Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved.

  11. Identification of a Raloxifene Analog That Promotes AhR-Mediated Apoptosis in Cancer Cells.

    PubMed

    Jang, Hyo Sang; Pearce, Martin; O'Donnell, Edmond F; Nguyen, Bach Duc; Truong, Lisa; Mueller, Monica J; Bisson, William H; Kerkvliet, Nancy I; Tanguay, Robert L; Kolluri, Siva Kumar

    2017-12-01

    We previously reported that raloxifene, an estrogen receptor modulator, is also a ligand for the aryl hydrocarbon receptor (AhR). Raloxifene induces apoptosis in estrogen receptor-negative human cancer cells through the AhR. We performed structure-activity studies with seven raloxifene analogs to better understand the structural requirements of raloxifene for induction of AhR-mediated transcriptional activity and apoptosis. We identified Y134 as a raloxifene analog that activates AhR-mediated transcriptional activity and induces apoptosis in MDA-MB-231 human triple negative breast cancer cells. Suppression of AhR expression strongly reduced apoptosis induced by Y134, indicating the requirement of AhR for Y134-induced apoptosis. Y134 also induced apoptosis in hepatoma cells without having an effect on cell cycle regulation. Toxicity testing on zebrafish embryos revealed that Y134 has a significantly better safety profile than raloxifene. Our studies also identified an analog of raloxifene that acts as a partial antagonist of the AhR, and is capable of inhibiting AhR agonist-induced transcriptional activity. We conclude that Y134 is a promising raloxifene analog for further optimization as an anti-cancer agent targeting the AhR.

  12. CDB-4124 does not cause apoptosis in cultured fibroid cells.

    PubMed

    Roeder, Hilary; Jayes, Friederike; Feng, Liping; Leppert, Phyllis C

    2011-09-01

    Selective progesterone receptor modulators (SPRMs), such as asoprisnil (J867) and ulipristal (CDB-2914), have been shown to reduce fibroid volume in vivo and to induce apoptosis in vitro. CDB-4124 (telapristone), a SPRM with different side groups, also reduced fibroid volume in vivo, and we hypothesized that this SPRM would also cause apoptosis in cultured fibroid cells. Immortalized, progesterone receptor-positive fibroid cells, known to be capable of apoptosis, were grown to 80% confluence in serum-containing media. Cells were then treated for 48 hours in serum-free media with 0, 10, 100, or 1000 nmol/L CDB-4124. Actinomycin-D and staurosporine were used as positive controls to induce apoptosis. Apoptosis was quantified using a TUNEL-fluorescein kit. Images were captured with a widefield-fluorescence microscope and analyzed using MetaMorph image analysis software. To validate results, Western blots of total cell lysates were probed for cleaved caspase-3 (c-CASP3). Experiments were repeated 3 times using independent cell batches. Analysis of 19 712 nuclei indicated 14.8% ± 10.9% (mean ± SEM), 8.4% ± 4.6%, 8.2% ± 4.7%, and 9.3% ± 6.3% apoptosis in 0, 10, 100, and 1000 nmol/L CDB-4124-treated cells, respectively. There was no evidence of elevated c-CASP3 over vehicle control after treatment with CDB-4124. CDB-4124 did not significantly induce apoptosis in cultured fibroid cells under the conditions described suggesting apoptosis may not be the main pathway responsible for CDB-4124-induced fibroid shrinkage. Variations in SPRM biological effects may be due to differences in fibroid source cells, binding kinetics, or extracellular matrix characteristics, and can be exploited in further investigations of the mechanisms of action of SPRMs in fibroid biology.

  13. [Enhanced growth inhibition by combined two pathway inhibitors on K-ras mutated non-small cell lung cancer cells].

    PubMed

    Yang, Zhenli; Li, Zhanwen; Feng, Hailiang; Bian, Xiaocui; Liu, Yanyan; Liu, Yuqin

    2014-09-01

    To evaluate the effect of combined targeting of MEK and PI3K signaling pathways on K-ras mutated non-small cell lung cancer cell line A549 cells and the relevant mechanisms. A549 cells were treated with different concentrations of two inhibitors. Growth inhibition was determined by MTT assay. According to the results of MTT test, the cells were divided into four groups: the control group, PI3K inhibitor group (GDC-0941,0.5 and 5.0 µmol/L), combination group I (0.5 µmol/L AZD6244+0.5 µmol/L GDC-0941) and combination group II (5.0 µmol/L AZD6244+5.0 µmol/L GDC-0941). The cell cycle and apoptosis were analyzed by flow cytometry. The expression of proteins related to apoptosis was tested with Western blot. Both GDC-0941 and AZD6244 inhibited the cell proliferation. The combination group II led to a stronger growth inhibition. The combination group I showed an antagonistic effect and combination group II showed an additive or synergistic effect. Compared with the control group, the combination group I led to reduced apoptotic rate [(20.70 ± 0.99)% vs. (18.65 ± 0.92 )%, P > 0.05]; Combination group II exhibited enhanced apoptotic rate [(37.85 ± 3.18)% vs. (52.27 ± 4.36)%, P < 0.01]. In addition, in the combination group II, more A549 cells were arrested in G0/G1 phase and decreased S phase (P < 0.01), due to the reduced expressions of CyclinD1 and Cyclin B1, the increased cleaved PARP and the diminished ratio of Bcl-2/Bax. For single K-ras mutated NSCLC cell line A549 cells, combination of RAS/MEK/ERK and PI3K/AKT/mTOR inhibition showed synergistic effects depending on the drug doses. Double pathways targeted therapy may be beneficial for these patients.

  14. Diabetes and apoptosis: neural crest cells and neural tube.

    PubMed

    Chappell, James H; Wang, Xiao Dan; Loeken, Mary R

    2009-12-01

    Birth defects resulting from diabetic pregnancy are associated with apoptosis of a critical mass of progenitor cells early during the formation of the affected organ(s). Insufficient expression of genes that regulate viability of the progenitor cells is responsible for the apoptosis. In particular, maternal diabetes inhibits expression of a gene, Pax3, that encodes a transcription factor which is expressed in neural crest and neuroepithelial cells. As a result of insufficient Pax3, cardiac neural crest and neuroepithelial cells undergo apoptosis by a process dependent on the p53 tumor suppressor protein. This, then provides a cellular explanation for the cardiac outflow tract and neural tube and defects induced by diabetic pregnancy.

  15. Diabetes and apoptosis: neural crest cells and neural tube

    PubMed Central

    Chappell, James H.; Dan Wang, Xiao

    2016-01-01

    Birth defects resulting from diabetic pregnancy are associated with apoptosis of a critical mass of progenitor cells early during the formation of the affected organ(s). Insufficient expression of genes that regulate viability of the progenitor cells is responsible for the apoptosis. In particular, maternal diabetes inhibits expression of a gene, Pax3, that encodes a transcription factor which is expressed in neural crest and neuroepithelial cells. As a result of insufficient Pax3, cardiac neural crest and neuroepithelial cells undergo apoptosis by a process dependent on the p53 tumor suppressor protein. This, then provides a cellular explanation for the cardiac outflow tract and neural tube and defects induced by diabetic pregnancy. PMID:19333760

  16. Overexpression of RBM5 induces autophagy in human lung adenocarcinoma cells.

    PubMed

    Su, Zhenzhong; Wang, Ke; Li, Ranwei; Yin, Jinzhi; Hao, Yuqiu; Lv, Xuejiao; Li, Junyao; Zhao, Lijing; Du, Yanwei; Li, Ping; Zhang, Jie

    2016-02-29

    Dysfunctions in autophagy and apoptosis are closely interacted and play an important role in cancer development. RNA binding motif 5 (RBM5) is a tumor suppressor gene, which inhibits tumor cells' growth and enhances chemosensitivity through inducing apoptosis in our previous studies. In this study, we investigated the relationship between RBM5 overexpression and autophagy in human lung adenocarcinoma cells. Human lung adenocarcinoma cancer (A549) cells were cultured in vitro and were transiently transfected with a RBM5 expressing plasmid (GV287-RBM5) or plasmid with scrambled control sequence. RBM5 expression was determined by semi-quantitative reverse transcription polymerase chain reaction (RT-PCR) and Western blot. Intracellular LC-3 I/II, Beclin-1, lysosome associated membrane protein-1 (LAMP1), Bcl-2, and NF-κB/p65 protein levels were detected by Western blot. Chemical staining with monodansylcadaverine (MDC) and acridine orange (AO) was applied to detect acidic vesicular organelles (AVOs). The ultrastructure changes were observed under transmission electron microscope (TEM). Then, transplanted tumor models of A549 cells on BALB/c nude mice were established and treated with the recombinant plasmids carried by attenuated Salmonella to induce RBM5 overexpression in tumor tissues. RBM5, LC-3, LAMP1, and Beclin1 expression was determined by immunohistochemistry staining in plasmids-treated A549 xenografts. Our study demonstrated that overexpression of RBM5 caused an increase in the autophagy-related proteins including LC3-I, LC3-II, LC3-II/LC3-I ratio, Beclin1, and LAMP1 in A549 cells. A large number of autophagosomes with double-membrane structure and AVOs were detected in the cytoplasm of A549 cells transfected with GV287-RBM5 at 24 h. We observed that the protein level of NF-κB/P65 was increased and the protein level of Bcl-2 decreased by RBM5 overexpression. Furthermore, treatment with an autophagy inhibitor, 3-MA, enhanced RBM5-induced cell death and

  17. Physiology and pathophysiology of apoptosis in epithelial cells of the liver, pancreas, and intestine.

    PubMed

    Jones, B A; Gores, G J

    1997-12-01

    Cell death of gastrointestinal epithelial cells occurs by a process referred to as apoptosis. In this review, we succinctly define apoptosis and summarize the role of apoptosis in the physiology and pathophysiology of epithelial cells in the liver, pancreas, and small and large intestine. The physiological mediators regulating apoptosis in gastrointestinal epithelial cells, when known, are discussed. Selected pathophysiological consequences of excessive apoptosis and inhibition of apoptosis are used to illustrate the significance of apoptosis in disease processes. These examples demonstrate that excessive apoptosis may result in epithelial cell atrophy, injury, and dysfunction, whereas inhibition of apoptosis results in hyperplasia and promotes malignant transformation. The specific cellular mechanisms responsible for dysregulation of epithelial cell apoptosis during pathophysiological disturbances are emphasized. Potential future areas of physiological research regarding apoptosis in gastrointestinal epithelia are highlighted when appropriate.

  18. Effects of celecoxib on cell apoptosis and Fas, FasL and Bcl-2 expression in a BGC-823 human gastric cancer cell line.

    PubMed

    Li, Qian; Peng, Jie; Liu, Ting; Zhang, Guiying

    2017-09-01

    Fas, which is an apoptotic-related protein, has an important role in cell apoptosis. Fas ligand (FasL) binds to Fas and activates apoptosis signal transduction. We previously demonstrated that the efficiency of celecoxib inhibited the proliferation and apoptosis of HT-29 colon cancer cell line. The BGC823 cell line was used as an experimental model to evaluate the potential role of celecoxib on gastric cancer cell apoptosis. Inhibitory effects of celecoxib on cell viability were determined by MTT assay. Cell apoptosis was evaluated by flow cytometric analysis and laser confocal microscopy. The results of the present study demonstrated that celecoxib inhibited the viability of BGC823 cells in a concentration- and time-dependent manner. Furthermore, the effect of BGC823 cells apoptosis was increased in a concentration-dependent manner. Western blotting was used to determine the protein expression levels of Fas, FasL, and B-cell lymphoma-2 (Bcl-2). During the celecoxib-induced apoptosis of BGC823 cells, celecoxib upregulated Fas expression and downregulated FasL and Bcl-2 expression in a concentration-dependent manner. These results suggest that celecoxib inhibited the growth and induced apoptosis of BGC823 gastric cancer cells by regulating the protein expression of Fas, FasL and Bcl-2.

  19. Nuclear events of apoptosis in vitro in cell-free mitotic extracts: a model system for analysis of the active phase of apoptosis

    PubMed Central

    1993-01-01

    We have developed a cell-free system that induces the morphological transformations characteristic of apoptosis in isolated nuclei. The system uses extracts prepared from mitotic chicken hepatoma cells following a sequential S phase/M phase synchronization. When nuclei are added to these extracts, the chromatin becomes highly condensed into spherical domains that ultimately extrude through the nuclear envelope, forming apoptotic bodies. The process is highly synchronous, and the structural changes are completed within 60 min. Coincident with these morphological changes, the nuclear DNA is cleaved into a nucleosomal ladder. Both processes are inhibited by Zn2+, an inhibitor of apoptosis in intact cells. Nuclear lamina disassembly accompanies these structural changes in added nuclei, and we show that lamina disassembly is a characteristic feature of apoptosis in intact cells of mouse, human and chicken. This system may provide a powerful means of dissecting the biochemical mechanisms underlying the final stages of apoptosis. PMID:8408207

  20. N-Desmethyldauricine Induces Autophagic Cell Death in Apoptosis-Defective Cells via Ca2+ Mobilization.

    PubMed

    Law, Betty Y K; Mok, Simon W F; Chen, Juan; Michelangeli, Francesco; Jiang, Zhi-Hong; Han, Yu; Qu, Yuan Q; Qiu, Alena C L; Xu, Su-Wei; Xue, Wei-Wei; Yao, Xiao-Jun; Gao, Jia Y; Javed, Masood-Ul-Hassan; Coghi, Paolo; Liu, Liang; Wong, Vincent K W

    2017-01-01

    Resistance of cancer cells to chemotherapy remains a significant problem in oncology. Mechanisms regulating programmed cell death, including apoptosis, autophagy or necrosis, in the treatment of cancers have been extensively investigated over the last few decades. Autophagy is now emerging as an important pathway in regulating cell death or survival in cancer therapy. Recent studies demonstrated variety of natural small-molecules could induce autophagic cell death in apoptosis-resistant cancer cells, therefore, discovery of novel autophagic enhancers from natural products could be a promising strategy for treatment of chemotherapy-resistant cancer. By computational virtual docking analysis, biochemical assays, and advanced live-cell imaging techniques, we have identified N -desmethyldauricine (LP-4), isolated from rhizoma of Menispermum dauricum DC as a novel inducer of autophagy. LP-4 was shown to induce autophagy via the Ulk-1-PERK and Ca 2+ /Calmodulin-dependent protein kinase kinase β (CaMKKβ)-AMPK-mTOR signaling cascades, via mobilizing calcium release through inhibition of SERCA, and importantly, lead to autophagic cell death in a panel of cancer cells, apoptosis-defective and apoptosis-resistant cells. Taken together, this study provides detailed insights into the cytotoxic mechanism of a novel autophagic compound that targeting the apoptosis resistant cancer cells, and new implication on drug discovery from natural products for drug resistant cancer therapy.

  1. A Vitex agnus-castus extract inhibits cell growth and induces apoptosis in prostate epithelial cell lines.

    PubMed

    Weisskopf, M; Schaffner, W; Jundt, G; Sulser, T; Wyler, S; Tullberg-Reinert, H

    2005-10-01

    Extracts of Vitex agnus-castus fruits (VACF) are described to have beneficial effects on disorders related to hyperprolactinemia (cycle disorders, premenstrual syndrome). A VACF extract has recently been shown to exhibit antitumor activities in different human cancer cell lines. In the present study, we explored the antiproliferative effects of a VACF extract with a particular focus on apoptosis-inducing and potential cytotoxic effects. Three different human prostate epithelial cell lines (BPH-1, LNCaP, PC-3) representing different disease stages and androgen responsiveness were chosen. The action of VACF on cell viability was assessed using the WST-8-tetrazolium assay. Cell proliferation in cells receiving VACF alone or in combination with a pan-caspase inhibitor (Z-VAD-fmk) was quantified using a Crystal Violet assay. Flow cytometric cell cycle analysis and measurement of DNA fragmentation using an ELISA method were used for studying the induction of apoptosis. Lactate dehydrogenase (LDH) activity was determined as a marker of cytotoxicity. The extract inhibited proliferation of all three cell lines in a concentration-dependent manner with IC (50) values below 10 microg/mL after treatment for 48 h. Cell cycle analysis and DNA fragmentation assays suggest that part of the cells were undergoing apoptosis. The VACF-induced decrease in cell number was partially inhibited by Z-VAD-fmk, indicating a caspase-dependent apoptotic cell death. However, the concentration-dependent LDH activity of VACF treated cells indicated cytotoxic effects as well. These data suggest that VACF contains components that inhibit proliferation and induce apoptosis in human prostate epithelial cell lines. The extract may be useful for the prevention and/or treatment not only of benign prostatic hyperplasia but also of human prostate cancer.

  2. Cytotoxic and apoptosis-inducing activities against human lung cancer cell lines of cassaine diterpenoids from the bark of Erythrophleum fordii.

    PubMed

    Ha, Manh Tuan; Tran, Manh Hung; Phuong, Thien Thuong; Kim, Jeong Ah; Woo, Mi Hee; Choi, Jae Sue; Lee, Suhyun; Lee, Jeong Hyung; Lee, Hyeong Kyu; Min, Byung Sun

    2017-07-01

    A phytochemical investigation into the bark of Erythrophleum fordii yielded four new compounds, two new cassaine diterpenoids (erythrofordin T and U, 1 and 2) and two new cassaine diterpenoid amines (erythroformine A and B, 6 and 7), as well as nine known compounds. We report for the first time the isolation of erythrofordin V (3) from a natural source and that of the remaining eight known diterpenoids (4-5, 8-13) from E. fordii. All structures were elucidated using spectroscopic analysis. Cytotoxic activity of the isolated compounds (1-13) was examined in vitro against three non-small cell lung cancer cell lines (A549, NCI-H1975, and NCI-H1229) using the MTT assay. Cassaine diterpene amines (6-10, 12, 13) exhibited potent cytotoxic activity against all three cell lines with IC 50 values between 0.4μM and 5.9μM. Erythroformine B (7) significantly induced apoptosis in all three cancer cells in a concentration-dependent manner. Copyright © 2017 Elsevier Ltd. All rights reserved.

  3. Effects of Notch2 and Notch3 on Cell Proliferation and Apoptosis of Trophoblast Cell Lines.

    PubMed

    Zhao, Wei-Xiu; Zhuang, Xu; Huang, Tao-Tao; Feng, Ran; Lin, Jian-Hua

    2015-01-01

    To investigate the effect of Notch2 and Notch3 on cell proliferation and apoptosis of two trophoblast cell lines, BeWo and JAR. Notch2 and Notch3 expression in BeWo and JAR cells was upregulated or downregulated using lentivirus-mediated overexpression or RNA interference. The effect of Notch2 and Notch3 on cell proliferation was assessed by the CCK-8 assay. The effect of Notch2 and Notch3 on the apoptosis of BeWo and JAR cells was evaluated by flow cytometry using the Annexin V-PE Apoptosis kit. Lentivirus-based overexpression vectors were constructed by cloning the full-length coding sequences of human Notch2 and Notch3 C-terminally tagged with GFP or GFP alone (control) into a lentivirus-based expression vector. Lentivirus-based gene silencing vectors were prepared by cloning small interfering sequences targeting human Notch2 and Notch3 and scrambled control RNA sequence into a lentivirus-based gene knockdown vector. The effect of Notch2 and Notch3 on cell proliferation was assessed by the CCK-8 assay. And the effect of Notch2 and Notch3 on the apoptosis of BeWo and JAR cells was evaluated by flow cytometry using the Annexin V PE Apoptosis kit. We found that the downregulation of Notch2 and Notch3 gene expression in BeWo and JAR cells resulted in an increase in cell proliferation, while upregulation of Notch3 and Notch2 expression led to a decrease in cell proliferation. Moreover, the overexpression of Notch3 and Notch2 in BeWo and JAR cells reduced apoptosis in these trophoblast cell lines, whereas apoptosis was increased in the cells in which the expression of Notch3 and Notch2 was downregulated. Notch2 and Notch3 inhibited both cell proliferation and cell apoptosis in BeWo and JAR trophoblast cell lines.

  4. Analysis of A549 cell proteome alteration in response to recombinant influenza A virus nucleoprotein and its interaction with cellular proteins, a preliminary study.

    PubMed

    Kumar, D; Tiwari, K; Rajala, M S

    Influenza A virus undergoes frequent changes of antigenicity and contributes to seasonal epidemics or unpredictable pandemics. Nucleoprotein, encoded by gene segment 5, is an internal protein of the virus and is conserved among strains of different host origins. In the current study, we analyzed the differentially expressed proteins in A549 cells transiently transfected with the recombinant nucleoprotein of influenza A virus by 2D gel electrophoresis. The resolved protein spots on gel were identified by MALDI-TOF/Mass spectrometry analysis. The majority of the host proteins detected to be differentially abundant in recombinant nucleoprotein-expressing cells as compared to vector-transfected cells are the proteins of metabolic pathways, glycolytic enzymes, molecular chaperones and cytoskeletal proteins. We further demonstrated the interaction of virus nucleoprotein with some of the identified host cellular proteins. In vitro binding assay carried out using the purified recombinant nucleoprotein (pET29a+NP-His) and A549 cell lysate confirmed the interaction between nucleoprotein and host proteins, such as alpha enolase 1, pyruvate kinase and β-actin. The preliminary data of our study provides the information on virus nucleoprotein interaction with proteins involved in glycolysis. However, studies are ongoing to understand the significance of these interactions in modulating the host factors during virus replication.

  5. Ruthenium(II) Complexes with 2-Phenylimidazo[4,5-f][1,10]phenanthroline Derivatives that Strongly Combat Cisplatin-Resistant Tumor Cells

    NASA Astrophysics Data System (ADS)

    Zeng, Leli; Chen, Yu; Liu, Jiangping; Huang, Huaiyi; Guan, Ruilin; Ji, Liangnian; Chao, Hui

    2016-01-01

    Cisplatin was the first metal-based therapeutic agent approved for the treatment of human cancers, but its clinical activity is greatly limited by tumor drug resistance. This work utilized the parent complex [Ru(phen)2(PIP)]2+ (1) to develop three Ru(II) complexes (2-4) with different positional modifications. These compounds exhibited similar or superior cytotoxicities compared to cisplatin in HeLa, A549 and multidrug-resistant (A549R) tumor cell lines. Complex 4, the most potent member of the series, was highly active against A549R cancer cells (IC50 = 0.8 μM). This complex exhibited 178-fold better activity than cisplatin (IC50 = 142.5 μM) in A549R cells. 3D multicellular A549R tumor spheroids were also used to confirm the high proliferative and cytotoxic activity of complex 4. Complex 4 had the greatest cellular uptake and had a tendency to accumulate in the mitochondria of A549R cells. Further mechanistic studies showed that complex 4 induced A549R cell apoptosis via inhibition of thioredoxin reductase (TrxR), elevated intracellular ROS levels, mitochondrial dysfunction and cell cycle arrest, making it an outstanding candidate for overcoming cisplatin resistance.

  6. Lipopolysaccharide Stimulates Butyric Acid-Induced Apoptosis in Human Peripheral Blood Mononuclear Cells

    PubMed Central

    Kurita-Ochiai, Tomoko; Fukushima, Kazuo; Ochiai, Kuniyasu

    1999-01-01

    We previously reported that butyric acid, an extracellular metabolite from periodontopathic bacteria, induced apoptosis in murine thymocytes, splenic T cells, and human Jurkat T cells. In this study, we examined the ability of butyric acid to induce apoptosis in peripheral blood mononuclear cells (PBMC) and the effect of bacterial lipopolysaccharide (LPS) on this apoptosis. Butyric acid significantly inhibited the anti-CD3 monoclonal antibody- and concanavalin A-induced proliferative responses in a dose-dependent fashion. This inhibition of PBMC growth by butyric acid depended on apoptosis in vitro. It was characterized by internucleosomal DNA digestion and revealed by gel electrophoresis followed by a colorimetric DNA fragmentation assay to occur in a concentration-dependent fashion. Butyric acid-induced PBMC apoptosis was accompanied by caspase-3 protease activity but not by caspase-1 protease activity. LPS potentiated butyric acid-induced PBMC apoptosis in a dose-dependent manner. Flow-cytometric analysis revealed that LPS increased the proportion of sub-G1 cells and the number of late-stage apoptotic cells induced by butyric acid. Annexin V binding experiments with fractionated subpopulations of PBMC in flow cytometory revealed that LPS accelerated the butyric acid-induced CD3+-T-cell apoptosis followed by similar levels of both CD4+- and CD8+-T-cell apoptosis. The addition of LPS to PBMC cultures did not cause DNA fragmentation, suggesting that LPS was unable to induce PBMC apoptosis directly. These data suggest that LPS, in combination with butyric acid, potentiates CD3+ PBMC T-cell apoptosis and plays a role in the apoptotic depletion of CD4+ and CD8+ cells. PMID:9864191

  7. DNA fragmentation and cell cycle arrest: a hallmark of apoptosis induced by crocin from kashmiri saffron in a human pancreatic cancer cell line.

    PubMed

    Bakshi, Hamid; Sam, Smitha; Rozati, Roya; Sultan, Phalisteen; Islam, Tajamul; Rathore, Babita; Lone, Zahoor; Sharma, Manik; Triphati, Jagrati; Saxena, Ramesh Chand

    2010-01-01

    Apoptosis, a widely important mechanism that contributes to cell growth reduction, is reported to be induced by Crocus sativus in different cancer types. The present study was designed to elucidate apoptosis induction by crocin, a main component of Crocus sativus in a human pancreatic cancer cell line (BxPC-3). Cell viability was measured by MTT assay, Hoechest33258 staining was used to detect the chromatin condensation characteristic of apoptosis, and DNA fragmentation was assessed by gel electrophoresis and cell cycle analysis by flow cytometry. Crocin induced apoptosis and G1-phase cell cycle arrest of BxPC-3 cells, while decreasing cell viability in a dose dependent and time dependent manner. Cells treated with 10μg/L crocin exhibited apoptotic morphology (brightly blue-fluorescent condensed nuclei on Hoechst 33258 staining) and reduction of volume. DNA analysis revealed typical ladders as early as 12 hours after treatment indicative of apoptosis. Our preclinical study demonstrated a pancreatic cancer cell line to be highly sensitive to crocin-mediated growth inhibition and apoptotic cell death. Although the molecular mechanisms of crocin action are not yet clearly understood, it appears to have potential as a therapeutic agent.

  8. A chimeric antigen receptor for TRAIL-receptor 1 induces apoptosis in various types of tumor cells.

    PubMed

    Kobayashi, Eiji; Kishi, Hiroyuki; Ozawa, Tatsuhiko; Hamana, Hiroshi; Nakagawa, Hidetoshi; Jin, Aishun; Lin, Zhezhu; Muraguchi, Atsushi

    2014-10-31

    Tumor necrosis factor (TNF)-related apoptosis-inducing ligand (TRAIL) and its associated receptors (TRAIL-R/TR) are attractive targets for cancer therapy because TRAIL induces apoptosis in tumor cells through TR while having little cytotoxicity on normal cells. Therefore, many agonistic monoclonal antibodies (mAbs) specific for TR have been produced, and these induce apoptosis in multiple tumor cell types. However, some TR-expressing tumor cells are resistant to TR-specific mAb-induced apoptosis. In this study, we constructed a chimeric antigen receptor (CAR) of a TRAIL-receptor 1 (TR1)-specific single chain variable fragment (scFv) antibody (TR1-scFv-CAR) and expressed it on a Jurkat T cell line, the KHYG-1 NK cell line, and human peripheral blood lymphocytes (PBLs). We found that the TR1-scFv-CAR-expressing Jurkat cells killed target cells via TR1-mediated apoptosis, whereas TR1-scFv-CAR-expressing KHYG-1 cells and PBLs killed target cells not only via TR1-mediated apoptosis but also via CAR signal-induced cytolysis, resulting in cytotoxicity on a broader range if target cells than with TR1-scFv-CAR-expressing Jurkat cells. The results suggest that TR1-scFv-CAR could be a new candidate for cancer gene therapy. Copyright © 2014 Elsevier Inc. All rights reserved.

  9. Osthole induces apoptosis, suppresses cell-cycle progression and proliferation of cancer cells.

    PubMed

    Jarząb, Agata; Grabarska, Aneta; Kiełbus, Michał; Jeleniewicz, Witold; Dmoszyńska-Graniczka, Magdalena; Skalicka-Woźniak, Krystyna; Sieniawska, Elwira; Polberg, Krzysztof; Stepulak, Andrzej

    2014-11-01

    The aim of the present study was to determine the effects of osthole on cell proliferation and viability, cell-cycle progression and induction of apoptosis in human laryngeal cancer RK33 and human medulloblastoma TE671 cell lines. Cell viability was measured by means of the MTT method and cell proliferation by the 5-bromo-2-deoxyuridine (BrdU) incorporation assay. Cell-cycle progression was determined by flow cytometry, and induction of apoptosis by release of oligonucleosomes to the cytosol. The gene expression was estimated by a quantitative polymerase chain reaction (qPCR) method. High-performance counter-current chromatography (HPCCC) was applied for isolation of osthole from fruits of Mutellina purpurea. Osthole decreased proliferation and cell viability of cancer cells in a dose-dependent manner. The tested compound induced apoptosis, increased the cell numbers in G1 and decreased cell number in S/G2 phases of the cell cycle, differentially regulating CDKN1A and TP53 gene expression depending on cancer cell type. Osthole could be considered as a potential compound for cancer therapy and chemoprevention. Copyright© 2014 International Institute of Anticancer Research (Dr. John G. Delinassios), All rights reserved.

  10. Opium induces apoptosis in jurkat cells.

    PubMed

    Igder, Somayeh; Asadikaram, Gholam Reza; Sheykholeslam, Farzaneh; Sayadi, Ahmad Reza; Mahmoodi, Mehdi; Kazemi Arababadi, Mohammad; Rasaee, Mohammad Javad

    2013-01-01

    The direct effect of some opioids on immune cells has been demonstrated. The aim of this study was to assess the apoptotic effect of opium on Jurkat T lymphocyte cells. Different concentrations of opium (2.86 × 10-3 to 2.86 × 10-11 g/ml) were added to 24-well plates containing 5 × 105 Jurkat cells. Apoptotic events were assessed after 6, 24, and 72 hours by flow-cytometric detection of surface phosphatidylserine. Significant differences in apoptosis of Jurkat cells were seen at 24 and 72 hours in different concentrations of opium (P < 0.05). After 72 hours, significant increase in necrosis of Jurkat cells was seen in opium concentration of 2.85 × 10-3 g/ml compared to cells without opium (control) (P < 0.05). These results showed that opium directly increases apoptosis and necrosis of T lymphocytes. This effect may play a role in immune dysfunction in opium addicts.

  11. Opium Induces Apoptosis in Jurkat Cells

    PubMed Central

    Igder, Somayeh; Asadikaram, Gholam Reza; Sheykholeslam, Farzaneh; Sayadi, Ahmad Reza; Mahmoodi, Mehdi; Kazemi Arababadi, Mohammad; Rasaee, Mohammad Javad

    2013-01-01

    Background The direct effect of some opioids on immune cells has been demonstrated. The aim of this study was to assess the apoptotic effect of opium on Jurkat T lymphocyte cells. Methods Different concentrations of opium (2.86 × 10-3 to 2.86 × 10-11 g/ml) were added to 24-well plates containing 5 × 105 Jurkat cells. Apoptotic events were assessed after 6, 24, and 72 hours by flow-cytometric detection of surface phosphatidylserine. Findings Significant differences in apoptosis of Jurkat cells were seen at 24 and 72 hours in different concentrations of opium (P < 0.05). After 72 hours, significant increase in necrosis of Jurkat cells was seen in opium concentration of 2.85 × 10-3 g/ml compared to cells without opium (control) (P < 0.05). Conclusion These results showed that opium directly increases apoptosis and necrosis of T lymphocytes. This effect may play a role in immune dysfunction in opium addicts. PMID:24494155

  12. Adipose-derived stromal cells inhibit prostate cancer cell proliferation inducing apoptosis

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Takahara, Kiyoshi; Ii, Masaaki, E-mail: masaii@art.osaka-med.ac.jp; Inamoto, Teruo

    2014-04-18

    Highlights: • AdSC transplantation exhibits inhibitory effect on tumor progressions of PCa cells. • AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway. • High expression of the TGF-β1 gene in AdSCs. - Abstract: Mesenchymal stem cells (MSCs) have generated a great deal of interest in the field of regenerative medicine. Adipose-derived stromal cells (AdSCs) are known to exhibit extensive proliferation potential and can undergo multilineage differentiation, sharing similar characteristics to bone marrow-derived MSCs. However, as the effect of AdSCs on tumor growth has not been studied sufficiently, we assessed the degree to which AdSCs affect the proliferationmore » of prostate cancer (PCa) cell. Human AdSCs exerted an inhibitory effect on the proliferation of androgen-responsive (LNCaP) and androgen-nonresponsive (PC3) human PCa cells, while normal human dermal fibroblasts (NHDFs) did not, and in fact promoted PCa cell proliferation to a degree. Moreover, AdSCs induced apoptosis of LNCaP cells and PC3 cells, activating the caspase3/7 signaling pathway. cDNA microarray analysis suggested that AdSC-induced apoptosis in both LNCaP and PC3 cells was related to the TGF-β signaling pathway. Consistent with our in vitro observations, local transplantation of AdSCs delayed the growth of tumors derived from both LNCaP- and PC3-xenografts in immunodeficient mice. This is the first preclinical study to have directly demonstrated that AdSC-induced PCa cell apoptosis may occur via the TGF-β signaling pathway, irrespective of androgen-responsiveness. Since autologous AdSCs can be easily isolated from adipose tissue without any ethical concerns, we suggest that therapy with these cells could be a novel approach for patients with PCa.« less

  13. [Suppression of COX-2 protein to cell apoptosis in non-small cell lung cancer].

    PubMed

    Sun, Limei; Zhao, Yue; Wang, Lujian; Song, Min; Song, Jiye

    2007-06-20

    One of mechanisms of carcinogenesis is suppression of cell apoptosis which leads to accumulation of aberrant cells. The aim of this study is to investigate cell apoptosis and COX-2 protein expression in non-small cell lung cancer (NSCLC). Cell apoptosis, expression of COX-2 and microvessel density (MVD) were detcted in 111 NSCLC samples by TdT-mediated dUTP nick end labeling (TUNEL) technique and immunohistochemical staining. The positive rate of COX-2 protein expression was 67.6% (75/111), and there were 53 patients with high level cell apoptosis (47.7%). Expression of COX-2 protien was significantly related to TNM stages (P=0.025) and lymph node metastasis (P=0.018). The MVD in NSCLC tissues with positive COX-2 expression was significantly higher than that in negative expression ones (P=0.000). COX model showed that lymph node metastasis (P=0.006) and positive expression of COX-2 protein (P=0.000) were independent prognostic factors of NSCLC. The expression of COX-2 protein may suppress cell apoptosis of tumor, and it may serve as a potential marker of prognosis for NSCLC.

  14. Cystic fibrosis epithelial cells are primed for apoptosis as a result of increased Fas (CD95).

    PubMed

    Chen, Qiwei; Pandi, Sudha Priya Soundara; Kerrigan, Lauren; McElvaney, Noel G; Greene, Catherine M; Elborn, J Stuart; Taggart, Clifford C; Weldon, Sinéad

    2018-02-24

    Previous work suggests that apoptosis is dysfunctional in cystic fibrosis (CF) airways with conflicting results. We evaluated the relationship between dysfunctional cystic fibrosis transmembrane conductance regulator (CFTR) and apoptosis in CF airway epithelial cells. Apoptosis and associated caspase activity were analysed in non-CF and CF tracheal and bronchial epithelial cell lines. Basal levels of apoptosis and activity of caspase-3 and caspase-8 were significantly increased in CF epithelial cells compared to controls, suggesting involvement of extrinsic apoptosis signalling, which is mediated by the activation of death receptors, such as Fas (CD95). Increased levels of Fas were observed in CF epithelial cells and bronchial brushings from CF patients compared to non-CF controls. Neutralisation of Fas significantly inhibited caspase-3 activity in CF epithelial cells compared to untreated cells. In addition, activation of Fas significantly increased caspase-3 activity and apoptosis in CF epithelial cells compared to control cells. Overall, these results suggest that CF airway epithelial cells are more sensitive to apoptosis via increased levels of Fas and subsequent activation of the Fas death receptor pathway, which may be associated with dysfunctional CFTR. Copyright © 2018 European Cystic Fibrosis Society. All rights reserved.

  15. The omega-3 polyunsaturated fatty acid docosahexaenoic acid inhibits proliferation and progression of non-small cell lung cancer cells through the reactive oxygen species-mediated inactivation of the PI3K /Akt pathway.

    PubMed

    Yin, Yuanqin; Sui, Chengguang; Meng, Fandong; Ma, Ping; Jiang, Youhong

    2017-05-03

    Docosahexaenoic acid(DHA) inhibits tumor growth and progression in various cancers, including lung cancer. However, the mechanisms involved remain unclear. The aim of this study was to identify the mechanism of DHA in inhibiting progression of non-small cell lung cancer (NSCLC) in vitro. The proliferation of A549 was tested by MTT, and cell apoptosis was analysed using flow cytometer. The migration and invasion were examined respectively by wound healing assay and Transwell invasion assay. The level of ROS (reactive oxygen species, ROS) was checked by DCF (dichlorodihydrofluorescein, DCF) production in cells. The apoptosis associated protein (caspase-3, PARP,Bax,Bcl-2 and survivin) and metastases associated proteins including HEF1, MMP9 and VEGF were detected by Western blot, and the same method was used in the expression of PI3K and Akt. DHA inhibited proliferation and induced apoptosis of A549 cells. Moreover, it suppressed the invasion and metastasis of A549 cells, while downregulating the levels of metastasis-associated proteins, including HEF1, matrix metallopeptidase (MMP9), and vascular endothelial growth factor (VEGF), in a dose -dependent manner. In addition, DHA inactivated Akt phosphorylation. All of these responses were associated with the accumulation of intracellular ROS. DHA downregulated the level of antioxidant enzymes such as catalase, while the antioxidant N-acetyl-cysteine (NAC) reversed the effect of DHA, which further validated our findings. The present study demonstrates that DHA inhibits the development of non-small lung tumors through an ROS-mediated inactivation of the PI3K/Akt signaling pathway.

  16. Deficiency in methionine, tryptophan, isoleucine, or choline induces apoptosis in cultured cells.

    PubMed

    Yen, Chi-Liang E; Mar, Mei-Heng; Craciunescu, Corneliu N; Edwards, Lloyd J; Zeisel, Steven H

    2002-07-01

    Cells in culture die by apoptosis when deprived of the essential nutrient choline. We now report that cells (both proliferating PC12 cells and postmitotic neurons isolated from fetal rat brains) undergo apoptosis when deprived of other individual essential nutrients (methionine, tryptophan or isoleucine). In PC12 cells, deficiencies of each nutrient independently led to ceramide accumulation and to caspase activation, both recognized signals of several apoptotic pathways. A similar profile of caspases was activated in PC12 cells deprived of choline, methionine, tryptophan or isoleucine. More than one caspase was involved and these caspases appeared to transmit parallel signals for apoptosis induction because only broad-spectrum caspase inhibitors, but not inhibitors for specific individual caspases inhibited apoptosis in choline- or methionine-deprived cells. The induction of these caspase-dependent apoptosis pathways likely did not involve the same upstream signals. Choline deficiency perturbed choline metabolism but did not affect protein synthesis, whereas amino acid deficiencies inhibited protein synthesis but did not perturb choline metabolism. In addition, a subclone of PC12 cells that was resistant to choline deficiency-induced apoptosis was not resistant to tryptophan deficiency-induced apoptosis. These observations suggest that deficiency of each studied nutrient activates different pathways for signaling apoptosis that ultimately converge on a common execution pathway.

  17. A mononuclear Cu(II) complex with 5,6-diphenyl-3-(2-pyridyl)-1,2,4-triazine: Synthesis, crystal structure, DNA- and BSA-binding, molecular modeling, and anticancer activity against MCF-7, A-549, and HT-29 cell lines.

    PubMed

    Anjomshoa, Marzieh; Hadadzadeh, Hassan; Torkzadeh-Mahani, Masoud; Fatemi, Seyed Jamilaldin; Adeli-Sardou, Mahboubeh; Rudbari, Hadi Amiri; Nardo, Viviana Mollica

    2015-01-01

    The copper(II) complex of 1,2,4-triazine derivatives, [Cu(dppt)2(H2O)](PF6)2(dppt is 5,6-diphenyl-3-(2-pyridyl)-1,2,4-triazine), has been synthesized and fully characterized by spectroscopic methods and single crystal X-ray diffraction. The in vitro DNA-binding studies of the complex have been investigated by several methods. The results showed that the complex intercalates into the base pairs of DNA. The complex also indicated good binding propensity to BSA. The results of molecular docking and molecular dynamic simulation methods confirm the experimental results. Finally, the in vitro cytotoxicity indicate that the complex has excellent anticancer activity against the three human carcinoma cell lines, MCF-7, A-549, and HT-29, with IC50 values of 9.8, 7.80, and 4.50 μM, respectively. The microscopic analyses of the cancer cells demonstrate that the Cu(II) complex apparently induced apoptosis. Copyright © 2015 Elsevier Masson SAS. All rights reserved.

  18. A microtubule inhibitor, ABT-751, induces autophagy and delays apoptosis in Huh-7 cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Wei, Ren-Jie

    The objective was to investigate the upstream mechanisms of apoptosis which were triggered by a novel anti-microtubule drug, ABT-751, in hepatocellular carcinoma-derived Huh-7 cells. Effects of ABT-751 were evaluated by immunocytochemistry, flow cytometric, alkaline comet, soft agar, immunoblotting, CytoID, green fluorescent protein-microtubule associated protein 1 light chain 3 beta detection, plasmid transfection, nuclear/cytosol fractionation, coimmunoprecipitation, quantitative reverse transcription-polymerase chain reaction, small-hairpin RNA interference and mitochondria/cytosol fractionation assays. Results showed that ABT-751 caused dysregulation of microtubule, collapse of mitochondrial membrane potential, generation of reactive oxygen species (ROS), DNA damage, G{sub 2}/M cell cycle arrest, inhibition of anchorage-independent cell growth and apoptosismore » in Huh-7 cells. ABT-751 also induced early autophagy via upregulation of nuclear TP53 and downregulation of the AKT serine/threonine kinase (AKT)/mechanistic target of rapamycin (MTOR) pathway. Through modulation of the expression levels of DNA damage checkpoint proteins and G{sub 2}/M cell cycle regulators, ABT-751 induced G{sub 2}/M cell cycle arrest. Subsequently, ABT-751 triggered apoptosis with marked downregulation of B-cell CLL/lymphoma 2, upregulation of mitochondrial BCL2 antagonist/killer 1 and BCL2 like 11 protein levels, and cleavages of caspase 8 (CASP8), CASP9, CASP3 and DNA fragmentation factor subunit alpha proteins. Suppression of ROS significantly decreased ABT-751-induced autophagic and apoptotic cells. Pharmacological inhibition of autophagy significantly increased the percentages of ABT-751-induced apoptotic cells. The autophagy induced by ABT-751 plays a protective role to postpone apoptosis by exerting adaptive responses following microtubule damage, ROS and/or impaired mitochondria. - Highlights: • An anti-microtubule agent, ABT-751, induces autophagy and apoptosis in Huh-7

  19. Anticancer activity of silver nanoparticles from Panax ginseng fresh leaves in human cancer cells.

    PubMed

    Castro-Aceituno, Verónica; Ahn, Sungeun; Simu, Shakina Yesmin; Singh, Priyanka; Mathiyalagan, Ramya; Lee, Hyun A; Yang, Deok Chun

    2016-12-01

    The pharmaceutical role of silver nanoparticles has been increased over the last decades, especially those synthesized through herbal medicinal plants, due to their variety of pharmacological importance. Panax ginseng Meyer (P. ginseng) has been widely used as a therapeutic herbal medicine for a long time in cancer treatment. In this study, the cytotoxic and oxidative effect of a novel silver nanoparticles synthesized from P. ginseng fresh leaves (P.g AgNPs) were evaluated in different human cancer cell lines. In addition, the effect of P.g AgNPs on cell migration, apoptosis and the determination of the mechanism involve was determinate by the use of A549 lung cancer cell line. It was found that P.g AgNPs treatment inhibited cell viability and induced oxidative stress in A549, MCF7 and HepG2 cancer cell lines. Likewise, P.g AgNPs treatment inhibited the epidermal growth factor (EGF)-enhanced migration, as well as decreased the mRNA levels and phosphorylation of EGF receptors in A549 cells. Moreover, P.g AgNPs modified the morphology of the cell nucleus and increase apoptosis percentage; this effect was linked to the stimulation of p38 MAPK/p53 pathway. Taken together, our results showed that P.g AgNPs exhibited anti-cancer activity in A549 and the regulation of EGFR/p38 MAPK/p53 pathway might be the possible mechanism of its anti-activity. Further experiments are suggested to determinate the mechanism by which P.g AgNPs induce cytotoxicity and ROS generation in MCF-7 and HepG2 cells. Copyright © 2016 Elsevier Masson SAS. All rights reserved.

  20. Identification of a Short Cell-Penetrating Peptide from Bovine Lactoferricin for Intracellular Delivery of DNA in Human A549 Cells.

    PubMed

    Liu, Betty R; Huang, Yue-Wern; Aronstam, Robert S; Lee, Han-Jung

    2016-01-01

    Cell-penetrating peptides (CPPs) have been shown to deliver cargos, including protein, DNA, RNA, and nanomaterials, in fully active forms into live cells. Most of the CPP sequences in use today are based on non-native proteins that may be immunogenic. Here we demonstrate that the L5a CPP (RRWQW) from bovine lactoferricin (LFcin), stably and noncovalently complexed with plasmid DNA and prepared at an optimal nitrogen/phosphate ratio of 12, is able to efficiently enter into human lung cancer A549 cells. The L5a CPP delivered a plasmid containing the enhanced green fluorescent protein (EGFP) coding sequence that was subsequently expressed in cells, as revealed by real-time PCR and fluorescent microscopy at the mRNA and protein levels, respectively. Treatment with calcium chloride increased the level of gene expression, without affecting CPP-mediated transfection efficiency. Zeta-potential analysis revealed that positively electrostatic interactions of CPP/DNA complexes correlated with CPP-mediated transport. The L5a and L5a/DNA complexes were not cytotoxic. This biomimetic LFcin L5a represents one of the shortest effective CPPs and could be a promising lead peptide with less immunogenic for DNA delivery in gene therapy.

  1. 31 CFR 549.303 - Entity.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 31 Money and Finance:Treasury 3 2011-07-01 2011-07-01 false Entity. 549.303 Section 549.303 Money... CONTROL, DEPARTMENT OF THE TREASURY LEBANON SANCTIONS REGULATIONS General Definitions § 549.303 Entity. The term entity means a partnership, association, trust, joint venture, corporation, group, subgroup...

  2. Sodium ascorbate inhibits growth via the induction of cell cycle arrest and apoptosis in human malignant melanoma A375.S2 cells.

    PubMed

    Lin, Shuw-Yuan; Lai, Wan-Wen; Chou, Chi-Chung; Kuo, Hsiu-Maan; Li, Te-Mao; Chung, Jing-Gung; Yang, Jen-Hung

    2006-12-01

    Vitamin C has been reported to be useful in the treatment and prevention of cancer. Inconsistent effects from growth stimulation to induction of apoptosis of malignant tumor cells, however, have been reported. Melanoma is an increasingly common and potentially lethal malignancy. It was reported that melanoma cells were more susceptible to ascorbate toxicity than any other tumor cells. The mechanisms accounting for ascorbate-induced apoptosis in human melanoma cells, however, have remained unclear. This study was undertaken to investigate the effect of sodium ascorbate on cytotoxicity and apoptosis in human malignant melanoma A375.S2 cells. A375.S2 cells were incubated with a certain range of concentrations of sodium ascorbate for various time periods. In order to examine the effects of sodium ascorbate on cell proliferation, cell cycle, apoptosis and necrosis, we performed 4,6-diamidino-2-phenylindole dihydrochloride assays and flow cytometry analysis. Polymerase chain reaction was used to examine the mRNA levels of p53, p21, p27, cyclin A, cyclin E, CDK2 and CDK4, which are associated with cell cycle S-phase arrest and apoptosis. Flow cytometric analysis showed that sodium ascorbate significantly induced cell cycle arrest and apoptosis in the A375.S2 cell line in a dose-dependent manner. The increased expressions of p53 and p21, and the decreased expressions of cyclin A, cyclin E, CDK2 and CDK4, indicated the cell cycle arrest at G1/S phase after the cells had been treated with sodium ascorbate. Induction of apoptosis involved an increase in the levels of p53, p21 and cellular Ca, and a decrease in mitochondrial membrane potential and activation of caspase 3 before culminating in apoptosis in sodium ascorbate-treated A375.S2 cells.

  3. Different effects of a novel CaO-MgO-SiO₂-based multiphase glass-ceramic on cell behaviors of normal and cancer cells in vitro.

    PubMed

    Zhang, Mengjiao; Chen, Xianchun; Pu, Ximing; Liao, Xiaoming; Huang, Zhongbing; Yin, Guangfu

    2014-04-01

    The effects in vitro of a novel multiphase glass-ceramic (with nominal composition of 43.19% CaO, 7.68% MgO, and 49.13% SiO2 in weight percent) on cell adhesion, proliferation, differentiation and ultrastructure of human osteosarcoma cell line MG63, mouse fibroblasts L929, and human lung adenocarcinoma epithelial cell line A549 were investigated in this research. Scanning electron microscopy (SEM) micrographs revealed that the surface morphology of this glass-ceramic was beneficial to cell adhesion. The glass-ceramic extracts at certain concentrations could stimulate the proliferation and differentiation of MG63 and L929 cells, whereas inhibit A549 proliferation, which might be resulted from the released Si ions. In addition, when cultured with 0.1mg/mL glass-ceramic powder suspension, the cell ultrastructure of MG63 showed abundant organelles and L929 displayed the phenomena of cellular stress response. While more interestingly, A549 exhibited chromatin condensation, mitochondria swell and RER expansion, which was presumed to be early signs of apoptosis. These results suggest that this novel CaO-MgO-SiO2-based multiphase glass-ceramic has potential for bone regeneration and tissue engineering applications. Copyright © 2013 Elsevier B.V. All rights reserved.

  4. Over-expression of angiotensin II type 2 receptor gene induces cell death in lung adenocarcinoma cells.

    PubMed

    Pickel, Lara; Matsuzuka, Takaya; Doi, Chiyo; Ayuzawa, Rie; Maurya, Dharmendra Kumar; Xie, Sheng-Xue; Berkland, Cory; Tamura, Masaaki

    2010-02-01

    The endogenous angiotensin II (Ang II) type 2 receptor (AT 2) has been shown to mediate apoptosis in cardiovascular tissues. Thus, the aim of this study was to explore the anti-cancer effect of AT 2 over-expression on lung adenocarcinoma cells in vitro using adenoviral (Ad), FuGENE, and nanoparticle vectors. All three gene transfection methods efficiently transfected AT 2 cDNA into lung cancer cells but caused minimal gene transfection in normal lung epithelial cells. Ad-AT 2 significantly attenuated multiple human lung cancer cell growth (A549 and H358) as compared to the control viral vector, Ad-LacZ, when cell viability was examined by direct cell count. Examination of annexin V by flow cytometry revealed the activation of the apoptotic pathway via AT 2 over-expression. Western Blot analysis confirmed the activation of caspase-3. Similarly, poly (lactide-co-glycolic acid) (PLGA) biodegradable nanoparticles encapsulated AT 2 plasmid DNA were shown to be effectively taken up into the lung cancer cell. Nanoparticle-based AT 2 gene transfection markedly increased AT 2 expression and resultant cell death in A549 cells. These results indicate that AT 2 over-expression effectively attenuates growth of lung adenocarcinoma cells through intrinsic apoptosis. Our results also suggest that PLGA nanoparticles can be used as an efficient gene delivery vector for lung adenocarcinoma targeted therapy.

  5. [Construction of lentiviral mediated CyPA siRNA and its functions in non-small cell lung cancer].

    PubMed

    FENG, Yan-ming; WU, Yi-ming; TU, Xin-ming; XU, Zheng-shun; WU, Wei-dong

    2010-02-01

    To construct a lentiviral-vector-mediated CyPA small interference RNA (siRNA) and study its function in non-small cell lung cancer. First, four target sequences were selected according to CyPA mRNA sequence, the complementary DNA contained both sense and antisense oligonucleotides were designed, synthesized and cloned into the pGCL-GFP vector, which contained U6 promoter and green fluorescent protein (GFP). The resulting lentiviral vector containing CyPA shRNA was named Lv-shCyPA, and it was confirmed by PCR and sequencing. Next, it was cotransfected by Lipofectamine 2000 along with pHelper1.0 and pHelper 2.0 into 293T cells to package lentivirus particles. At the same time, the packed virus infected non-small cell lung cancer cell (A549), the level of CyPA protein at 5 d after infection was detected by Western Blot to screen the target of CyPA. A549 were infected with Lv-shCyPA and grown as xenografts in severe combined immunodeficient mice. Cell cycle and apoptosis were measured by FCM. It was confirmed by PCR and DNA sequencing that lentiviral-vector-mediated CyPA siRNA (Lv-shCyPA) producing CyPA shRNA was constructed successfully. The titer of concentrated virus were 1 x 10(7) TU/ml. Flow cytometric analysis demonstrated G2-M phase (11.40% +/- 0.68%) was decreased relatively in A549/LvshCyPA compared with control groups (14.52% +/- 1.19%) (P<0.05). The apoptosis rate of A549/Lv-shCyPA (5.01% +/- 0.5%) was higher than control groups (0.35% +/- 0.17%) (P<0.05). Visible tumors were only detectable at 6th day after inoculated by A549/Lv-shCyPA. The xenograft tumors of A549/Lv-shCyPA remarkably delayed tumor growth and remained at a similarly small average size at 38th days after inoculation compared with the control group (P < 0.05). Lentiviral-vector-mediated siRNA technique effectively inhibits the expression of CyPA, induces the NSCLC cell apoptosis, inhibits the tumor growth. Elucidation of the precise role of CypA in these pathways may lead to new targeted

  6. The fruit juice of Morinda citrifolia (noni) downregulates HIF-1α protein expression through inhibition of PKB, ERK-1/2, JNK-1 and S6 in manganese-stimulated A549 human lung cancer cells.

    PubMed

    Jang, Byeong-Churl

    2012-03-01

    High exposure of manganese is suggested to be a risk factor for many lung diseases. Evidence suggests anticancerous and antiangiogenic effects by products derived from Morinda citrifolia (noni) fruit. In this study, we investigated the effect of noni fruit juice (NFJ) on the expression of HIF-1α, a tumor angiogenic transcription factor in manganese-chloride (manganese)-stimulated A549 human lung carcinoma cells. Treatment with manganese largely induced expression of HIF-1α protein but did not affect HIF-1α mRNA expression in A549 cells, suggesting the metal-mediated co- and/or post-translational HIF-1α upregulation. Manganese treatment also led to increased phosphorylation of extracellular-regulated protein kinase-1/2 (ERK-1/2), c-Jun N-terminal kinase-1 (JNK-1), protein kinase B (PKB), S6 and eukaryotic translation initiation factor-2α (eIF-2α) in A549 cells. Of note, the exposure of NFJ inhibited the manganese-induced HIF-1α protein upregulation in a concentration-dependent manner. Importantly, as assessed by results of pharmacological inhibition and siRNA transfection studies, the effect of NFJ on HIF-1α protein downregulation seemed to be largely associated with the ability of NFJ to interfere with the metal's signaling to activate PKB, ERK-1/2, JNK-1 and S6 in A549 cells. It was further shown that NFJ could repress the induction of HIF-1α protein by desferoxamine or interleukin-1β (IL-1β), another HIF-1α inducer in A549 cells. Thus, the present study provides the first evidence that NFJ has the ability to strongly downregulate manganese-induced HIF-1α protein expression in A549 human lung cancer cells, which may suggest the NFJ-mediated beneficial effects on lung pathologies in which manganese and HIF-1α overexpression play pathogenic roles.

  7. Role of TGF-β signaling in curcumin-mediated inhibition of tumorigenicity of human lung cancer cells

    PubMed Central

    Datta, Raktima; Halder, Sunil K.

    2014-01-01

    Purpose Curcumin has been shown to have potent anti-cancer activities like inhibition of cell proliferation, induction of apoptosis, and suppression of angiogenesis. Transforming growth factor-β (TGF-β) signaling plays a complex role in tumor suppression and promotion depending on the tumor type and stage. However, the effect of curcumin on TGF-β signaling in cancer cells and the role of TGF-β signaling in curcumin-induced anticancer activities have not been determined. Here, we investigate the role of curcumin on TGF-β signaling, and whether TGF-β signaling is involved in the antitumor activities of curcumin. Methods Human non-small cell lung cancer (NSCLC) cell lines, ACC-LC-176 (without TGF-β signaling), H358, and A549 (with TGF-β signaling) were treated with curcumin to determine cell growth, apoptosis, and tumorigenicity. Antitumor activities of curcumin were determined using these cell lines and an in vivo mouse model. We also tested the effect of curcumin on TGF-β/Smad signaling by western blotting and by luciferase assays. Results Curcumin inhibited cell growth and induced apoptosis of all three NSCLC cell lines in vitro and in vivo. It significantly reduced subcutaneous tumor growth by these three cell lines irrespective of TGF-β signaling status. Curcumin inhibited TGF-β-induced Smad2/3 phosphorylation and transcription in H358 and A549 cells, but not in ACC-LC-176 cells. Conclusions Curcumin reduces tumorigenicity of human lung cancer cells in vitro and in vivo by inhibiting cell proliferation and promoting apoptosis. These results suggest that TGF-β signaling is not directly involved in curcumin-mediated growth inhibition, induction of apoptosis, and inhibition of tumorigenicity. PMID:23224523

  8. Impaired lysosomal activity mediated autophagic flux disruption by graphite carbon nanofibers induce apoptosis in human lung epithelial cells through oxidative stress and energetic impairment.

    PubMed

    Mittal, Sandeep; Sharma, Pradeep Kumar; Tiwari, Ratnakar; Rayavarapu, Raja Gopal; Shankar, Jai; Chauhan, Lalit Kumar Singh; Pandey, Alok Kumar

    2017-04-28

    Graphite carbon nanofibers (GCNF) have emerged as a potential alternative of carbon nanotubes (CNT) for various biomedical applications due to their superior physico-chemical properties. Therefore in-depth understanding of the GCNF induced toxic effects and underlying mechanisms in biological systems is of great interest. Currently, autophagy activation by nanomaterials is recognized as an emerging toxicity mechanism. However, the association of GCNF induced toxicity with this form of cell death is largely unknown. In this study, we have assessed the possible mechanism; especially the role of autophagy, underlying the GCNF induced toxicity. Human lung adenocarcinoma (A549) cells were exposed to a range of GCNF concentrations and various cellular parameters were analyzed (up to 48 h). Transmission electron microscopy, immunofluorescent staining, western blot and quantitative real time PCR were performed to detect apoptosis, autophagy induction, lysosomal destabilization and cytoskeleton disruption in GCNF exposed cells. DCFDA assay was used to evaluate the reactive oxygen species (ROS) production. Experiments with N-acetyl-L-cysteine (NAC), 3-methyladenine (3-MA) and LC3 siRNA was carried out to confirm the involvement of oxidative stress and autophagy in GCNF induced cell death. Comet assay and micronucleus (MN) assay was performed to assess the genotoxicity potential. In the present study, GCNF was found to induce nanotoxicity in human lung cells through autophagosomes accumulation followed by apoptosis via intracellular ROS generation. Mechanistically, impaired lysosomal function and cytoskeleton disruption mediated autophagic flux blockade was found to be the major cause of accumulation rather than autophagy induction which further activates apoptosis. The whole process was in line with the increased ROS level and their pharmacological inhibition leads to mitigation of GCNF induced cell death. Moreover the inhibition of autophagy attenuates apoptosis indicating

  9. Bupivacaine induces apoptosis via ROS in the Schwann cell line.

    PubMed

    Park, C J; Park, S A; Yoon, T G; Lee, S J; Yum, K W; Kim, H J

    2005-09-01

    Local anesthetics have been generally accepted as being safe. However, recent clinical trials and basic studies have provided strong evidence for the neurotoxicity of local anesthetics, especially through apoptosis. We hypothesized that local anesthetics cause neural complications through Schwann cell apoptosis. Among local anesthetics tested on the Schwann cell line, RT4-D6P2T, bupivacaine significantly induced cell death, measured by the methyl tetrazolium (MTT) assay, in a dose- (LD50 = 476 microM) and time-dependent manner. The bupivacaine-induced generation of reactive oxygen species (ROS), which was initiated within 5 hrs and preceded the activation of caspase-3 and poly ADP-ribose polymerase (PARP) degradation, was suggested to trigger apoptosis, exhibited by Hoechst 33258 nuclear staining and DNA fragmentation. Furthermore, concomitant block of ROS by anti-oxidants significantly inhibited bupivacaine-induced apoptosis. Among the local anesthetics for peripheral neural blocks, bupivacaine induced apoptosis in the Schwann cell line, which may be associated with ROS production.

  10. A reactive oxygen species activation mechanism contributes to JS-K-induced apoptosis in human bladder cancer cells.

    PubMed

    Qiu, Mingning; Chen, Lieqian; Tan, Guobin; Ke, Longzhi; Zhang, Sai; Chen, Hege; Liu, Jianjun

    2015-10-13

    Reactive oxygen species (ROS) and cellular oxidant stress are regulators of cancer cells. The alteration of redox status, which is induced by increased generation of ROS, results in increased vulnerability to oxidative stress. The aim of this study is to investigate the influence of O2-(2,4-dinitrophenyl) 1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate (JS-K, C13H16N6O8) on proliferation and apoptosis in bladder cancer cells and explored possible ROS-related mechanisms. Our results indicated that JS-K could suppress bladder cancer cell proliferation in a concentration- and time-dependent manner and induce apoptosis and ROS accumulation in a concentration-dependent manner. With increasing concentrations of JS-K, expression of proteins that are involved in cell apoptosis increased in a concentration-dependent manner. Additionally, the antioxidant N-acetylcysteine (NAC) reversed JS-K-induced cell apoptosis; conversely, the prooxidant oxidized glutathione (GSSG) exacerbated JS-K-induced cell apoptosis. Furthermore, we found that nitrites, which were generated from the oxidation of JS-K-released NO, induced apoptosis in bladder cancer cells to a lower extent through the ROS-related pathway. In addition, JS-K was shown to enhance the chemo-sensitivity of doxorubicin in bladder cancer cells. Taken together, the data suggest that JS-K-released NO induces bladder cancer cell apoptosis by increasing ROS levels, and nitrites resulting from oxidation of NO have a continuous apoptosis-inducing effect.

  11. A reactive oxygen species activation mechanism contributes to JS-K-induced apoptosis in human bladder cancer cells

    PubMed Central

    Qiu, Mingning; Chen, Lieqian; Tan, Guobin; Ke, Longzhi; Zhang, Sai; Chen, Hege; Liu, Jianjun

    2015-01-01

    Reactive oxygen species (ROS) and cellular oxidant stress are regulators of cancer cells. The alteration of redox status, which is induced by increased generation of ROS, results in increased vulnerability to oxidative stress. The aim of this study is to investigate the influence of O2-(2,4-dinitrophenyl) 1-[(4-ethoxycarbonyl)piperazin-1-yl]diazen-1-ium-1,2-diolate (JS-K, C13H16N6O8) on proliferation and apoptosis in bladder cancer cells and explored possible ROS-related mechanisms. Our results indicated that JS-K could suppress bladder cancer cell proliferation in a concentration- and time-dependent manner and induce apoptosis and ROS accumulation in a concentration-dependent manner. With increasing concentrations of JS-K, expression of proteins that are involved in cell apoptosis increased in a concentration-dependent manner. Additionally, the antioxidant N-acetylcysteine (NAC) reversed JS-K-induced cell apoptosis; conversely, the prooxidant oxidized glutathione (GSSG) exacerbated JS-K-induced cell apoptosis. Furthermore, we found that nitrites, which were generated from the oxidation of JS-K-released NO, induced apoptosis in bladder cancer cells to a lower extent through the ROS-related pathway. In addition, JS-K was shown to enhance the chemo-sensitivity of doxorubicin in bladder cancer cells. Taken together, the data suggest that JS-K-released NO induces bladder cancer cell apoptosis by increasing ROS levels, and nitrites resulting from oxidation of NO have a continuous apoptosis-inducing effect. PMID:26458509

  12. Apoptosis in fish: environmental factors and programmed cell death.

    PubMed

    AnvariFar, Hossein; Amirkolaie, Abdolsamad Keramat; Miandare, Hamed Kolangi; Ouraji, Hossein; Jalali, M Ali; Üçüncü, Sema İşisağ

    2017-06-01

    Apoptosis, a form of programmed cell death, is a critical component in maintaining homeostasis and growth in all tissues and plays a significant role in immunity and cytotoxicity. In contrast to necrosis or traumatic cell death, apoptosis is a well-controlled and vital process characterized mainly by cytoplasmic shrinkage, chromatin condensation, DNA fragmentation, membrane blebbing and apoptotic bodies. Our understanding of apoptosis is partly based on observations in invertebrates but mainly in mammals. Despite the great advantages of fish models in studying vertebrate development and diseases and the tremendous interest observed in recent years, reports on apoptosis in fish are still limited. Although apoptotic machinery is well conserved between aquatic and terrestrial organisms throughout the history of evolution, some differences exist in key components of apoptotic pathways. Core parts of apoptotic machinery in fish are virtually expressed as equivalent to the mammalian models. Some differences are, however, evident, such as the extrinsic and intrinsic pathways of apoptosis including lack of a C-terminal region in the Fas-associated protein with a death domain in fish. Aquatic species inhabit a complex and highly fluctuating environment, making these species good examples to reveal features of apoptosis that may not be easily investigated in mammals. Therefore, in order to gain a wider view on programmed cell death in fish, interactions between the main environmental factors, chemicals and apoptosis are discussed in this review. It is indicated that apoptosis can be induced in fish by exposure to environmental stressors during different stages of the fish life cycle.

  13. miR-320a regulates cell proliferation and apoptosis in multiple myeloma by targeting pre-B-cell leukemia transcription factor 3

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Lu, Yinghao; Department of Hematology, Affiliated Hospital of Guizhou Medical University, The Hematopoietic Stem Cell Transplant Center of Guizhou Province, Blood Diseases Diagnosis and Treatment Center of Guizhou Province, Guiyang, 550004, Guizhou Province; Wu, Depei, E-mail: wudepei@medmail.com.cn

    2016-05-13

    Aberrant expression of microRNAs (miRNAs) is implicated in cancer development and progression. While miR-320a is reported to be deregulated in many malignancy types, its biological role in multiple myeloma (MM) remains unclear. Here, we observed reduced expression of miR-320a in MM samples and cell lines. Ectopic expression of miR-320a dramatically suppressed cell viability and clonogenicity and induced apoptosis in vitro. Mechanistic investigation led to the identification of Pre-B-cellleukemia transcription factor 3 (PBX3) as a novel and direct downstream target of miR-320a. Interestingly, reintroduction of PBX3 abrogated miR-320a-induced MM cell growth inhibition and apoptosis. In a mouse xenograft model, miR-320a overexpression inhibitedmore » tumorigenicity and promoted apoptosis. Our findings collectively indicate that miR-320a inhibits cell proliferation and induces apoptosis in MM cells by directly targeting PBX3, supporting its utility as a novel and potential therapeutic agent for miRNA-based MM therapy. -- Highlights: •Expression of miR-320a in MM cell induces apoptosis in vitro. •miR-320a represses PBX3 via targeting specific sequences in the 3′UTR region. •Exogenous expression of PBX3 reverses the effects of miR-320a in inhibiting MM cell growth and promoting apoptosis. •Overexpression of miR-320a inhibits tumor growth and increases apoptosis in vivo.« less

  14. Isogambogenic acid induces apoptosis-independent autophagic cell death in human non-small-cell lung carcinoma cells.

    PubMed

    Yang, Jianhong; Zhou, Yongzhao; Cheng, Xia; Fan, Yi; He, Shichao; Li, Shucai; Ye, Haoyu; Xie, Caifeng; Wu, Wenshuang; Li, Chunyan; Pei, Heying; Li, Luyuan; Wei, Zhe; Peng, Aihua; Wei, Yuquan; Li, Weimin; Chen, Lijuan

    2015-01-09

    To overcome drug resistance caused by apoptosis deficiency in patients with non-small cell lung carcinoma (NSCLC), there is a need to identify other means of triggering apoptosis-independent cancer cell death. We are the first to report that isogambogenic acid (iso-GNA) can induce apoptosis-independent autophagic cell death in human NSCLC cells. Several features of the iso-GNA-treated NSCLC cells indicated that iso-GNA induced autophagic cell death. First, there was no evidence of apoptosis or cleaved caspase 3 accumulation and activation. Second, iso-GNA treatment induced the formation of autophagic vacuoles, increased LC3 conversion, caused the appearance of autophagosomes and increased the expression of autophagy-related proteins. These findings provide evidence that iso-GNA induces autophagy in NSCLC cells. Third, iso-GNA-induced cell death was inhibited by autophagic inhibitors or by selective ablation of Atg7 and Beclin 1 genes. Furthermore, the mTOR inhibitor rapamycin increased iso-GNA-induced cell death by enhancing autophagy. Finally, a xenograft model provided additional evidence that iso-GNA exhibited anticancer effect through inducing autophagy-dependent cell death in NSCLC cells. Taken together, our results demonstrated that iso-GNA exhibited an anticancer effect by inducing autophagy-dependent cell death in NSCLC cells, which may be an effective chemotherapeutic agent that can be used against NSCLC in a clinical setting.

  15. Isogambogenic acid induces apoptosis-independent autophagic cell death in human non-small-cell lung carcinoma cells

    PubMed Central

    Yang, Jianhong; Zhou, Yongzhao; Cheng, Xia; Fan, Yi; He, Shichao; Li, Shucai; Ye, Haoyu; Xie, Caifeng; Wu, Wenshuang; Li, Chunyan; Pei, Heying; Li, Luyuan; Wei, Zhe; Peng, Aihua; Wei, Yuquan; Li, Weimin; Chen, Lijuan

    2015-01-01

    To overcome drug resistance caused by apoptosis deficiency in patients with non-small cell lung carcinoma (NSCLC), there is a need to identify other means of triggering apoptosis-independent cancer cell death. We are the first to report that isogambogenic acid (iso-GNA) can induce apoptosis-independent autophagic cell death in human NSCLC cells. Several features of the iso-GNA-treated NSCLC cells indicated that iso-GNA induced autophagic cell death. First, there was no evidence of apoptosis or cleaved caspase 3 accumulation and activation. Second, iso-GNA treatment induced the formation of autophagic vacuoles, increased LC3 conversion, caused the appearance of autophagosomes and increased the expression of autophagy-related proteins. These findings provide evidence that iso-GNA induces autophagy in NSCLC cells. Third, iso-GNA-induced cell death was inhibited by autophagic inhibitors or by selective ablation of Atg7 and Beclin 1 genes. Furthermore, the mTOR inhibitor rapamycin increased iso-GNA-induced cell death by enhancing autophagy. Finally, a xenograft model provided additional evidence that iso-GNA exhibited anticancer effect through inducing autophagy-dependent cell death in NSCLC cells. Taken together, our results demonstrated that iso-GNA exhibited an anticancer effect by inducing autophagy-dependent cell death in NSCLC cells, which may be an effective chemotherapeutic agent that can be used against NSCLC in a clinical setting. PMID:25571970

  16. Leptospermum flavescens Constituent-LF1 Causes Cell Death through the Induction of Cell Cycle Arrest and Apoptosis in Human Lung Carcinoma Cells

    PubMed Central

    Navanesan, Suerialoasan; Abdul Wahab, Norhanom; Manickam, Sugumaran; Sim, Kae Shin

    2015-01-01

    Leptospermum flavescens Sm. (Myrtaceae), locally known as ‘Senna makki’ is a smallish tree that is widespread and recorded to naturally occur in the montane regions above 900 m a.s.l from Burma to Australia. Although the species is recorded to be used traditionally to treat various ailments, there is limited data on biological and chemical investigations of L. flavescens. The aim of the present study was to investigate and understand the ability of L. flavescens in inducing cell death in lung cancer cells. The cytotoxic potentials of the extraction yields (methanol, hexane, ethyl acetate and water extracts as wells as a semi pure fraction, LF1) were evaluated against two human non-small cell lung carcinoma cell lines (A549 and NCI-H1299) using the MTT assay. LF1 showed the greatest cytotoxic effect against both cell lines with IC50 values of 7.12 ± 0.07 and 9.62 ± 0.50 μg/ml respectively. LF1 treated cells showed a sub-G1 region in the cell cycle analysis and also caused the presence of apoptotic morphologies in cells stained with acridine orange and ethidium bromide. Treatment with LF1 manifested an apoptotic population in cells that were evaluated using the Annexin V/ propidium iodide assay. Increasing dosage of LF1 caused a rise in the presence of activated caspase-3 enzymes in treated cells. Blockage of cell cycle progression was also observed in LF1-treated cells. These findings suggest that LF1 induces apoptosis and cell cycle arrest in treated lung cancer cells. Further studies are being conducted to isolate and identify the active compound as well to better understand the mechanism involved in inducing cell death. PMID:26287817

  17. [Phloretin induces apoptosis of BEL-7402 cells in vitro].

    PubMed

    Luo, Hui; Wang, Ya-jun; Chen, Jie; Liu, Jiang-qin; Zhang, Hai-tao

    2008-07-01

    To examine the effect of phloretin on apoptosis of BEL-7402 cells. The viability changes of BEL- 7402 cells as a result of phloretin-induced toxicity were analyzed using MTT assay, and the cell morphology changes were observed with fluorescence microscope. Flow cytometry was used to analyze the cell cycle and mitochondrial membrane potential changes, and chromogenic substrate assay performed to detect caspase activity. Phloretin induced obvious cytotoxicity against BEL-7402 cells with IC50 of 89.23 microg/mL. The growth curve demonstrated decreased growth of the cells as phloretin concentration increased. Cell apoptosis occurred 24 h after treatment with 40-160 microg/mL phloretin. Morphological, the cells exposed to phloretin exhibited nuclear chromatin condensation and increased fluorescence intensity. The activity of caspase-9 reached the peak level 12 h after phloretin exposure, and leak levels of caspase-6 and caspase-3 activities occurred 18 and 24 h after the exposure, respectively. Phloretin can induce BEL-7402 cell apoptosis though the mitochondrial pathway.

  18. Nucleoprotein of influenza A virus negatively impacts antiapoptotic protein API5 to enhance E2F1-dependent apoptosis and virus replication.

    PubMed

    Mayank, A K; Sharma, S; Nailwal, H; Lal, S K

    2015-12-17

    Apoptosis of host cells profoundly influences virus propagation and dissemination, events that are integral to influenza A virus (IAV) pathogenesis. The trigger for activation of apoptosis is regulated by an intricate interplay between cellular and viral proteins, with a strong bearing on IAV replication. Though the knowledge of viral proteins and mechanisms employed by IAV to induce apoptosis has advanced considerably of late, we know relatively little about the repertoire of host factors targeted by viral proteins. Thus, identification of cellular proteins that are hijacked by the virus will help us not only to understand the molecular underpinnings of IAV-induced apoptosis, but also to design future antiviral therapies. Here we show that the nucleoprotein (NP) of IAV directly interacts with and suppresses the expression of API5, a host antiapoptotic protein that antagonizes E2F1-dependent apoptosis. siRNA-mediated depletion of API5, in NP-overexpressed as well as IAV-infected cells, leads to upregulation of apoptotic protease activating factor 1 (APAF1), a downstream modulator of E2F1-mediated apoptosis, and cleavage of caspases 9 and 3, although a reciprocal pattern of these events was observed on ectopic overexpression of API5. In concordance with these observations, annexin V and 7AAD staining assays exhibit downregulation of early and late apoptosis in IAV-infected or NP-transfected cells on overexpression of API5. Most significantly, while overexpression of API5 decreases viral titers, cellular NP protein as well as mRNA levels in IAV-infected A549 cells, silencing of API5 expression causes a steep rise in the same parameters. From the data reported in this manuscript, we propose a proapoptotic role for NP in IAV pathogenesis, whereby it suppresses expression of antiapoptotic factor API5, thus potentiating the E2F1-dependent apoptotic pathway and ensuring viral replication.

  19. Undecylprodigiosin conjugated monodisperse gold nanoparticles efficiently cause apoptosis in colon cancer cells in vitro

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Nikodinovic-Runic, Jasmina; Mojic, Marija; Kang, Yijin

    2014-01-01

    Bacterial pigment undecylprodigiosin (UP) was produced using Streptomyces sp. JS520 and conjugated to monodisperse gold nanoparticles (UP-Au). Both UP and UP-Au showed cytocidal activity towards melanoma (A375), lung carcinoma (A549), breast cancer (MCF-7) and colon cancer (HCT-116) cells, inducing apoptosis with IC50 values ranging from 0.4 to 4 mu g ml(-1). Unconjugated UP had a tendency to lose its activity over time and to change biophysical characteristics over pH. The loss of the pigment potency was overcome by conjugation with gold nanoparticles. UP-Au exhibited high stability over pH 3.8 to 7.4 and its activity remained unaffected in time. Nano-packing changedmore » the mechanism of UP toxicity by converting the intracellular signals from a mitochondrial dependent to a mitochondrial independent apoptotic process. The availability of nonpyrogenic UP in high amounts, together with specific anticancer activity and improved stability in the complex with gold nanoparticles, presents a novel platform for further development of UP-Au complexes as an anticancer drug suitable for clinical applications.« less

  20. Molecular Cell Biology of Apoptosis and Necroptosis in Cancer.

    PubMed

    Dillon, Christopher P; Green, Douglas R

    Cell death is a major mechanism to eliminate cells in which DNA is damaged, organelles are stressed, or oncogenes are overexpressed, all events that would otherwise predispose cells to oncogenic transformation. The pathways that initiate and execute cell death are complex, genetically encoded, and subject to significant regulation. Consequently, while these pathways are often mutated in malignancy, there is considerable interest in inducing cell death in tumor cells as therapy. This chapter addresses our current understanding of molecular mechanisms contributing to two cell death pathways, apoptotic cell death and necroptosis, a regulated form of necrotic cell death. Apoptosis can be induced by a wide variety of signals, leading to protease activation that dismantles the cell. We discuss the physiological importance of each apoptosis pathway and summarize their known roles in cancer suppression and the current efforts at targeting each pathway therapeutically. The intricate mechanistic link between death receptor-mediated apoptosis and necroptosis is described, as well as the potential opportunities for utilizing necroptosis in the treatment of malignancy.

  1. Marine Bacterial Polysaccharide EPS11 Inhibits Cancer Cell Growth via Blocking Cell Adhesion and Stimulating Anoikis

    PubMed Central

    Cao, Ruobing; Jin, Weihua; Shan, Yeqi; Wang, Ju; Liu, Ge; Kuang, Shan

    2018-01-01

    Tumor cells that acquire metastatic potential have developed resistance to anoikis, a cell death process, after detachment from their primary site to the second organ. In this study, we investigated the molecular mechanisms of a novel marine bacterial polysaccharide EPS11 which exerts its cytotoxic effects through affecting cancer cell adhesion and anoikis. Firstly, we found that EPS11 could significantly affect cell proliferation and block cell adhesion in A549 cells. We further demonstrated that the expression of several cell adhesion associated proteins is downregulated and the filiform structures of cancer cells are destroyed after EPS11 treatment. Interestingly, the destruction of filiform structures in A549 cells by EPS11 is in a dose-dependent manner, and the inhibitory tendency is very consistent with that observed in the cell adhesion assay, which confirms that filiform structures play important roles in modulating cell adhesion. Moreover, we showed that EPS11 induces apoptosis of A549 cells through stimulating βIII-tubulin associated anoikis: (i) EPS11 inhibits the expression of βIII-tubulin in both transcription and translation levels; and (ii) EPS11 treatment dramatically decreases the phosphorylation of protein kinase B (PKB or AKT), a critical downstream effector of βIII-tubulin. Importantly, EPS11 evidently inhibits the growth of A549-derived tumor xenografts in vivo. Thus, our results suggest that EPS11 may be a potential candidate for human non-small cell lung carcinoma treatment via blocking filiform structure mediated adhesion and stimulating βIII-tubulin associated anoikis. PMID:29518055

  2. A superoxide anion generator, pyrogallol, inhibits the growth of HeLa cells via cell cycle arrest and apoptosis.

    PubMed

    Kim, Sang Wook; Han, Yong Whan; Lee, Soo Teik; Jeong, Hey Jin; Kim, Seong Hun; Kim, In Hee; Lee, Seung Ok; Kim, Dae Ghon; Kim, Suhn Hee; Kim, Sung Zoo; Park, Woo Hyun

    2008-02-01

    We investigated the in vitro effects of pyrogallol on cell growth, cell cycle regulation, and apoptosis in HeLa cells. Pyrogallol inhibited the growth of HeLa cells with an IC(50) of approximately 45 microM. Pyrogallol induced arrest during all phases of the cell cycle and also very efficiently resulted in apoptosis in HeLa cells, as evidenced by flow cytometric detection of sub-G1 DNA content, annexin V binding assay, and DAPI staining. This apoptotic process was accompanied by the loss of mitochondrial transmembrane potential (DeltaPsi(m)), Bcl-2 decrease, caspase-3 activation, and PARP cleavage. Pan-caspase inhibitor (Z-VAD) could rescue some HeLa cells from pyrogallol-induced cell death, while caspase-8 and -9 inhibitors unexpectedly enhanced the apoptosis. When we examined the changes of the ROS, H(2)O(2) or O(2)(*-) in pyrogallol-treated cells, H(2)O(2) was slightly increased and O(2)(*-) significantly was increased. In addition, we detected a decreased GSH content in pyrogallol-treated cells. Only pan-caspase inhibitor showing recovery of GSH depletion and reduced intracellular O(2)(*-) level decreased PI staining in pyrogallol-treated HeLa cells, which indicates dead cells. In summary, we have demonstrated that pyrogallol as a generator of ROS, especially O(2) (*-), potently inhibited the growth of HeLa cells through arrests during all phases of the cell cycle and apoptosis. (c) 2007 Wiley-Liss, Inc.

  3. Apoptosis transcriptional mechanism of feline infectious peritonitis virus infected cells.

    PubMed

    Shuid, Ahmad Naqib; Safi, Nikoo; Haghani, Amin; Mehrbod, Parvaneh; Haron, Mohd Syamsul Reza; Tan, Sheau Wei; Omar, Abdul Rahman

    2015-11-01

    Apoptosis has been postulated to play an important role during feline infectious peritonitis virus (FIPV) infection; however, its mechanism is not well characterized. This study is focused on apoptosis and transcriptional profiling of FIPV-infected cells following in vitro infection of CRFK cells with FIPV 79-1146 WSU. Flow cytometry was used to determine mode of cell death in first 42 h post infection (hpi). FIPV infected cells underwent early apoptosis at 9 hpi (p < 0.05) followed by late apoptosis at 12 hpi (p < 0.05) and necrosis from 24 hpi (p < 0.05). Then, next generation sequencing was performed on 9 hpi and control uninfected cells by Illumina analyzer. An aggregate of 4546 genes (2229 down-regulated and 2317 up-regulated) from 17 cellular process, 11 molecular functions and 130 possible biological pathways were affected by FIPV. 131 genes from apoptosis cluster (80 down-regulated and 51 up-regulated) along with increase of apoptosis, p53, p38 MAPK, VEGF and chemokines/cytokines signaling pathways were probably involved in apoptosis process. Six of the de-regulated genes expression (RASSF1, BATF2, MAGEB16, PDCD5, TNFα and TRAF2) and TNFα protein concentration were analyzed by RT-qPCR and ELISA, respectively, at different time-points. Up-regulations of both pro-apoptotic (i.e. PDCD5) and anti-apoptotic (i.e. TRAF2) were detected from first hpi and continuing to deregulate during apoptosis process in the infected cells.

  4. Cancerous inhibitor of protein phosphatase 2A determines bortezomib-induced apoptosis in leukemia cells

    PubMed Central

    Liu, Chun-Yu; Shiau, Chung-Wai; Kuo, Hsin-Yu; Huang, Hsiang-Po; Chen, Ming-Huang; Tzeng, Cheng-Hwai; Chen, Kuen-Feng

    2013-01-01

    The multiple cellular targets affected by proteasome inhibition implicate a potential role for bortezomib, a first-in-class proteasome inhibitor, in enhancing antitumor activities in hematologic malignancies. Here, we examined the antitumor activity and drug targets of bortezomib in leukemia cells. Human leukemia cell lines were used for in vitro studies. Drug efficacy was evaluated by apoptosis assays and associated molecular events assessed by Western Blot. Gene silencing was performed by small interference RNA. Drug was tested in vivo in xenograft models of human leukemia cell lines and in primary leukemia cells. Clinical samples were assessed by immunohistochemical staining. Bortezomib differentially induced apoptosis in leukemia cells that was independent of its proteasome inhibition. Cancerous inhibitor of protein phosphatase 2A, a cellular inhibitor of protein phosphatase 2A, mediated the apoptotic effect of bortezomib. Bortezomib increased protein phosphatase 2A activity in sensitive leukemia cells (HL-60 and KG-1), but not in resistant cells (MOLT-3 and K562). Bortezomib’s downregulation of cancerous inhibitor of protein phosphatase 2A and phospho-Akt correlated with its drug sensitivity. Furthermore, cancerous inhibitor of protein phosphatase 2A negatively regulated protein phosphatase 2A activity. Ectopic expression of CIP2A up-regulated phospho-Akt and protected HL-60 cells from bortezomib-induced apoptosis, whereas silencing CIP2A overcame the resistance to bortezomib-induced apoptosis in MOLT3 and K562 cells. Importantly, bortezomib exerted in vivo antitumor activity in HL-60 xenografted tumors and induced cell death in some primary leukemic cells. Cancerous inhibitor of protein phosphatase 2A was expressed in leukemic blasts from bone marrow samples. Cancerous inhibitor of protein phosphatase 2A plays a major role in mediating bortezomib-induced apoptosis in leukemia cells. PMID:22983581

  5. Ursodeoxycholic Acid Induces Death Receptor-mediated Apoptosis in Prostate Cancer Cells

    PubMed Central

    Lee, Won Sup; Jung, Ji Hyun; Panchanathan, Radha; Yun, Jeong Won; Kim, Dong Hoon; Kim, Hye Jung; Kim, Gon Sup; Ryu, Chung Ho; Shin, Sung Chul; Hong, Soon Chan; Choi, Yung Hyun; Jung, Jin-Myung

    2017-01-01

    Background Bile acids have anti-cancer properties in a certain types of cancers. We determined anticancer activity and its underlying molecular mechanism of ursodeoxycholic acid (UDCA) in human DU145 prostate cancer cells. Methods Cell viability was measured with an MTT assay. UDCA-induced apoptosis was determined with flow cytometric analysis. The expression levels of apoptosis-related signaling proteins were examined with Western blotting. Results UDCA treatment significantly inhibited cell growth of DU145 in a dose-dependent manner. It induced cellular shrinkage and cytoplasmic blebs and accumulated the cells with sub-G1 DNA contents. Moreover, UDCA activated caspase 8, suggesting that UDCA-induced apoptosis is associated with extrinsic pathway. Consistent to this finding, UDCA increased the expressions of tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) receptor, death receptor 4 (DR4) and death receptor 5 (DR5), and TRAIL augmented the UDCA-induced cell death in DU145 cells. In addition, UDCA also increased the expressions of Bax and cytochrome c and decreased the expression of Bcl-xL in DU145 cells. This finding suggests that UDCA-induced apoptosis may be involved in intrinsic pathway. Conclusions UDCA induces apoptosis via extrinsic pathway as well as intrinsic pathway in DU145 prostate cancer cells. UDCA may be a promising anti-cancer agent against prostate cancer. PMID:28382282

  6. Effect of caffeine on radiation-induced apoptosis in TK6 cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Zhen, W.; Vaughan, A.T.M.

    1995-02-01

    Apoptosis has been measured in cells of the human TK6 lymphoblastoid cell line by recording the release of endonuclease-digested DNA from affected cells using flow cytometry. In asynchronously dividing cells, DNA degradation characteristic of apoptosis was first seen 12 h after irradiation as a defined DNA fluorescent peak of sub-G{sub 1}-phase content, reaching a maximum of 30-50% of the population by 24-72 h. Treating cells with 2 mM caffeine either before or up to 3 h after irradiation eliminated the degradation of DNA entirely. In addition, the percentage of cells in which apoptosis could be detected microscopically decreased from 62.4more » {+-} 0.95% to 16.7 {+-} 1.5% 72 h after caffeine treatment. Delaying caffeine treatment for 12 h after irradiation reduced DNA degradation by approximately 50% compared to cells receiving radiation alone. DNA degradation induced by serum deprivation was unaffected by caffeine treatment. These data support the contention that irradiation of TK6 cells produces a long-lived cellular signal which triggers apoptosis. Apoptosis produced by serum deprivation does not operate through the same pathway. 36 refs., 5 figs.« less

  7. Knockdown of hepatoma-derived growth factor-related protein-3 induces apoptosis of H1299 cells via ROS-dependent and p53-independent NF-κB activation

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Yun, Hong Shik; Department of Life Science, College of Natural Sciences, Hanyang University, Seoul 133-791; Baek, Jeong-Hwa

    2014-07-11

    Highlights: • HRP-3 is a radiation- and anticancer drug-responsive protein in H1299 cells. • Depletion of HRP-3 induces apoptosis of radio- and chemoresistant H1299 cells. • Depletion of HRP-3 promotes ROS generation via inhibition of the Nrf2/HO-1 pathway. • ROS generation enhances NF-κB activity, which acts as an upstream signal in the c-Myc/Noxa apoptotic pathway. - Abstract: We previously identified hepatoma-derived growth factor-related protein-3 (HRP-3) as a radioresistant biomarker in p53 wild-type A549 cells and found that p53-dependent induction of the PUMA pathway was a critical event in regulating the radioresistant phenotype. Here, we found that HRP-3 knockdown regulates themore » radioresistance of p53-null H1299 cells through a distinctly different molecular mechanism. HRP-3 depletion was sufficient to cause apoptosis of H1299 cells by generating substantial levels of reactive oxygen species (ROS) through inhibition of the Nrf2/HO-1 antioxidant pathway. Subsequent, ROS-dependent and p53-independent NF-κB activation stimulated expression of c-Myc and Noxa proteins, thereby inducing the apoptotic machinery. Our results thus extend the range of targets for the development of new drugs to treat both p53 wild-type or p53-null radioresistant lung cancer cells.« less

  8. [The role of apoptosis of granulosa cells in follicular atresia].

    PubMed

    Grotowski, W; Lecybył, R; Warenik-Szymankiewicz, A; Trzeciak, W H

    1997-07-01

    Apoptosis plays an important role in the process of morphogenesis and embryogenesis. Its increase or inhibition is an etiopathological factor in many different diseases. It has recently been shown that apoptosis of granulosa cells is one of the main mechanisms responsible for follicular atresia. There are many other factors influencing the process of granulosa cells apoptosis, among them the most important are: RnGH, FSH, LH, sex hormones (estrogens and androgens), growth factors and their receptors (EGF/TGF-alpha, FGF, IGF-1) and cytokines (e.g. TNF-alpha). The article presents data concerning the regulatory mechanisms of granulosa cells apoptosis in the ovary.

  9. 3,3'-diindolylmethane potentiates tumor necrosis factor-related apoptosis-inducing ligand-induced apoptosis of gastric cancer cells.

    PubMed

    Ye, Yang; Miao, Shuhan; Wang, Yan; Zhou, Jianwei; Lu, Rongzhu

    2015-05-01

    Tumor necrosis factor-related apoptosis-inducing ligand (TRAIL) specifically kills cancer cells without destroying the majority of healthy cells. However, numerous types of cancer cell, including gastric cancer cells, tend to be resistant to TRAIL. The bioactive product 3,3'-diindolylmethane (DIM), which is derived from cruciferous vegetables, is also currently recognized as a candidate anticancer agent. In the present study, a Cell Counting Kit 8 cell growth assay and an Annexin V-fluorescein isothiocyanate apoptosis assay were performed to investigate the potentiating effect of DIM on TRAIL-induced apoptosis in gastric cancer cells, and the possible mechanisms of this potentiation. The results obtained demonstrated that, compared with TRAIL or DIM treatment alone, co-treatment with TRAIL (25 or 50 ng/ml) and DIM (10 µmol/l) induced cytotoxic and apoptotic effects in BGC-823 and SGC-7901 gastric cancer cells. Furthermore, western blot analysis revealed that the protein expression levels of death receptor 5 (DR5), CCAAT/enhancer binding protein homologous protein (CHOP) and glucose-regulated protein 78 (GRP78) were upregulated in the co-treated gastric cancer cells. To the best of our knowledge, the present study is the first to provide evidence that DIM sensitizes TRAIL-induced inhibition of proliferation and apoptosis in gastric cancer cells, accompanied by the upregulated expression of DR5, CHOP and GRP78 proteins, which may be involved in endoplasmic reticulum stress mechanisms.

  10. EVA1A inhibits GBM cell proliferation by inducing autophagy and apoptosis.

    PubMed

    Shen, Xue; Kan, Shifeng; Liu, Zhen; Lu, Guang; Zhang, Xiaoyan; Chen, Yingyu; Bai, Yun

    2017-03-01

    Eva-1 homolog A (EVA1A) is a novel lysosome and endoplasmic reticulum-associated protein involved in autophagy and apoptosis. In this study, we constructed a recombinant adenovirus 5-EVA1A vector (Ad5-EVA1A) to overexpress EVA1A in glioblastoma (GBM) cell lines and evaluated its anti-tumor activities in vitro and in vivo. We found that overexpression of EVA1A in three GBM cell lines (U251, U87 and SHG44) resulted in a suppression of tumor cell growth via activation of autophagy and induction of cell apoptosis in a dose- and time-dependent manner. EVA1A-mediated autophagy was associated with inactivation of the mTOR/RPS6KB1 signaling pathway. Furthermore in vivo, overexpression of EVA1A successfully inhibited tumor growth in NOD/SCID mice. Our data suggest that EVA1A-induced autophagy and apoptosis play a role in suppressing the development of GBM and their up-regulation may be an effective method for treating this form of cancer. Copyright © 2017 Elsevier Inc. All rights reserved.

  11. Release behavior and toxicity profiles towards A549 cell lines of ciprofloxacin from its layered zinc hydroxide intercalation compound.

    PubMed

    Abdul Latip, Ahmad Faiz; Hussein, Mohd Zobir; Stanslas, Johnson; Wong, Charng Choon; Adnan, Rohana

    2013-01-01

    Layered hydroxides salts (LHS), a layered inorganic compound is gaining attention in a wide range of applications, particularly due to its unique anion exchange properties. In this work, layered zinc hydroxide nitrate (LZH), a family member of LHS was intercalated with anionic ciprofloxacin (CFX), a broad spectrum antibiotic via ion exchange in a mixture solution of water:ethanol. Powder x-ray diffraction (XRD), Fourier transform infrared (FTIR) and thermogravimetric analysis (TGA) confirmed the drug anions were successfully intercalated in the interlayer space of LZH. Specific surface area of the obtained compound was increased compared to that of the host due to the different pore textures between the two materials. CFX anions were slowly released over 80 hours in phosphate-buffered saline (PBS) solution due to strong interactions that occurred between the intercalated anions and the host lattices. The intercalation compound demonstrated enhanced antiproliferative effects towards A549 cancer cells compared to the toxicity of CFX alone. Strong host-guest interactions between the LZH lattice and the CFX anion give rise to a new intercalation compound that demonstrates sustained release mode and enhanced toxicity effects towards A549 cell lines. These findings should serve as foundations towards further developments of the brucite-like host material in drug delivery systems.

  12. Release behavior and toxicity profiles towards A549 cell lines of ciprofloxacin from its layered zinc hydroxide intercalation compound

    PubMed Central

    2013-01-01

    Background Layered hydroxides salts (LHS), a layered inorganic compound is gaining attention in a wide range of applications, particularly due to its unique anion exchange properties. In this work, layered zinc hydroxide nitrate (LZH), a family member of LHS was intercalated with anionic ciprofloxacin (CFX), a broad spectrum antibiotic via ion exchange in a mixture solution of water:ethanol. Results Powder x-ray diffraction (XRD), Fourier transform infrared (FTIR) and thermogravimetric analysis (TGA) confirmed the drug anions were successfully intercalated in the interlayer space of LZH. Specific surface area of the obtained compound was increased compared to that of the host due to the different pore textures between the two materials. CFX anions were slowly released over 80 hours in phosphate-buffered saline (PBS) solution due to strong interactions that occurred between the intercalated anions and the host lattices. The intercalation compound demonstrated enhanced antiproliferative effects towards A549 cancer cells compared to the toxicity of CFX alone. Conclusions Strong host-guest interactions between the LZH lattice and the CFX anion give rise to a new intercalation compound that demonstrates sustained release mode and enhanced toxicity effects towards A549 cell lines. These findings should serve as foundations towards further developments of the brucite-like host material in drug delivery systems. PMID:23849189

  13. Platinum Nanoparticles Induce Apoptosis on Raw 264.7 Macrophage Cells.

    PubMed

    Loan, Ta Thi; Do, Le Thanh; Yoo, Hoon

    2018-02-01

    The cellular effects of platinum nanoparticles (PNP05, average size of 5 nm, and PNP30, average size of 30 nm) were investigated on murine leukemia Raw 264.7 cells. Cells treated with various concentrations of PNPs showed size-dependent cytotoxicity in an MTT assay with PNP5 of smaller nanoparticles higher toxicity than PNP30. Investigations on cell morphology, Annexin V assay, DNA fragmentation and the activity of caspase-3/-7 showed that PNPs induced apoptosis on Raw 264.7 cells by changing cell morphology and density, increasing cell population in apoptosis and causing nucleus fragmentation. Further study on caspase activity by Western blotting revealed that the apoptosis was induced by the activation of caspase-3 and -7. In addition, PNPs inactivated DNA repair system, generating dose-dependent DNA ladder bands on agarose gel electrophoresis. Taken together, PNPs triggered cytotoxicity on Raw 264.7 cells by suppressing cell growth/survival and inducing apoptosis.

  14. Is Osteopontin a Friend or Foe of Cell Apoptosis in Inflammatory Gastrointestinal and Liver Diseases?

    PubMed

    Iida, Tomoya; Wagatsuma, Kohei; Hirayama, Daisuke; Nakase, Hiroshi

    2017-12-21

    Osteopontin (OPN) is involved in a variety of biological processes, including bone remodeling, innate immunity, acute and chronic inflammation, and cancer. The expression of OPN occurs in various tissues and cells, including intestinal epithelial cells and immune cells such as macrophages, dendritic cells, and T lymphocytes. OPN plays an important role in the efficient development of T helper 1 immune responses and cell survival by inhibiting apoptosis. The association of OPN with apoptosis has been investigated. In this review, we described the role of OPN in inflammatory gastrointestinal and liver diseases, focusing on the association of OPN with apoptosis. OPN changes its association with apoptosis depending on the type of disease and the phase of disease activity, acting as a promoter or a suppressor of inflammation and inflammatory carcinogenesis. It is essential that the roles of OPN in those diseases are elucidated, and treatments based on its mechanism are developed.

  15. Upregulation of erythropoietin receptor in UT-7/EPO cells inhibits simulated microgravity-induced cell apoptosis

    NASA Astrophysics Data System (ADS)

    Zou, Li-xue; Cui, Shao-yan; Zhong, Jian; Yi, Zong-chun; Sun, Yan; Fan, Yu-bo; Zhuang, Feng-yuan

    2011-07-01

    Hematopoietic progenitor cell proliferation can be altered in either spaceflight or under simulated microgravity experiments on the ground, however, the underlying mechanism remains unknown. Our previous study showed that exposure of the human erythropoietin (EPO)-dependent leukemia cell line UT-7/EPO to conditions of simulated microgravity significantly inhibited the cellular proliferation rate and induced cell apoptosis. We postulated that the downregulation of the erythropoietin receptor (EPOR) expression in UT-7/EPO cells under simulated microgravity may be a possible reason for microgravity triggered apoptosis. In this paper, a human EPOR gene was transferred into UT-7/EPO cells and the resulting expression of EPOR on the surface of UT-7/EPO cells increased approximately 61% ( p < 0.05) as selected by the antibiotic G418. It was also shown through cytometry assays and morphological observations that microgravity-induced apoptosis markedly decreased in these UT-7/EPO-EPOR cells. Thus, we concluded that upregulation of EPOR in UT-7/EPO cells could inhibit the simulated microgravity-induced cell apoptosis in this EPO dependent cell line.

  16. 6-shogaol induces autophagic cell death then triggered apoptosis in colorectal adenocarcinoma HT-29 cells.

    PubMed

    Li, Ting-Yi; Chiang, Been-Huang

    2017-09-01

    6-shogaol is a phytochemical of dietary ginger, we found that 6-shogaol could induced both autophagic and apoptotic death in human colon adenocarcinoma (HT-29) cells. Results of this study showed that 6-shogal induced cell cycle arrest, autophagy, and apoptosis in HT-29 cells in a time sequence. After 6h, 6-shogal induced apparent G2/M arrest, then the HT-29 cells formed numerous autophagosomes in each phase of the cell cycle. After 18h, increases in acidic vesicles and LAMP-1 (Lysosome-associated membrane proteins 1) showed that 6-shogaol had caused autophagic cell death. After 24h, cell shrinkage and Caspase-3/7 activities rising, suggesting that apoptotic cell death had increased. And after 48h, the result of TUNEL assay indicated the highest occurrence of apoptosis upon 6-shogaol treatment. It appeared that apoptosis is triggered by autophagy in 6-shogaol treated HT-29 cells, the damage of autophagic cell death initiated apoptosis program. Copyright © 2017 Elsevier Masson SAS. All rights reserved.

  17. Dihydroartemisinin-induced apoptosis in human acute monocytic leukemia cells

    PubMed Central

    Cao, Jia-Tian; Mo, Hui-Min; Wang, Yue; Zhao, Kai; Zhang, Tian-Tian; Wang, Chang-Qian; Xu, Kai-Lin; Han, Zhi-Hua

    2018-01-01

    Dihydroartemisinin (DHA) is a derivative of artemisinin. The present study aimed to investigate whether DHA induces apoptosis in the THP-1 human acute monocytic leukemia cell line (AMoL), and to identify the relative molecular mechanisms. The results of the present study demonstrated that the viability of THP-1 cells were inhibited by DHA in a dose- and time-dependent manner, which was accompanied by morphological characteristics associated with apoptosis. After 24 h of 200 µM DHA treatment, the proportion of apoptotic cells was significantly increased compared with the untreated controls (P<0.01). In addition, DHA downregulated the levels of B-cell lymphoma (Bcl)-2, protein kinase B (Akt)1, Akt2 and Akt3 gene expression, and increased the expression of the Bcl-2-associated X protein apoptosis regulator. The protein expression of phospho-Akt and phospho-extracellular signal-regulated kinase (ERK) was also decreased, and the protein expression level of cleaved caspase-3 was increased following treatment with DHA. Therefore, DHA may induce apoptosis in the AMoL THP-1 cell line via currently unknown underlying molecular mechanisms, including the downregulation of ERK and Akt, and the activation of caspase-3. PMID:29435054

  18. Knockdown of HIF-1α and IL-8 induced apoptosis of hepatocellular carcinoma triggers apoptosis of vascular endothelial cells.

    PubMed

    Choi, Sung Hoon; Park, Jun Yong; Kang, Wonseok; Kim, Seung Up; Kim, Do Young; Ahn, Sang Hoon; Ro, Simon Wonsang; Han, Kwang-Hyub

    2016-01-01

    A local hypoxic microenvironment is one of the most important characteristics of solid tumors. Hypoxia inducible factor-1α (HIF-1α) and Interleukin-8 (IL-8) activate tumor survival from hypoxic-induced apoptosis in each pathway. This study aimed to evaluate whether knockdown of HIF-1α and IL-8 induced apoptosis of the hepatocellular carcinoma (HCC) and endothelial cell lines. HCC cell lines were infected with adenovirus-expressing shRNA for HIF-1α and IL-8 and maintained under hypoxic conditions (1% O2, 24 h). The expression levels of HIF-1α and both apoptotic and growth factors were examined by real-time quantitative PCR and western blot. We also investigated apoptosis by TUNEL assay (FACS and Immunofluorescence) and measured the concentration of cytochrome C. Inhibition of HIF-1α and IL-8 up-regulated the expression of apoptotic factors while downregulating anti-apoptotic factors simultaneously. Knockdown of HIF-1α and IL-8 increased the concentration of cytochrome C and enhanced DNA fragmentation in HCC cell lines. Moreover, culture supernatant collected from the knockdown of HIF-1α and IL-8 in HCC cell lines induced apoptosis in human umbilical vein endothelial cells under hypoxia, and the expression of variable apoptotic ligand increased from HCC cell lines, time-dependently. These data suggest that adenovirus-mediated knockdown of HIF-1α and IL-8 induced apoptosis in HCC cells and triggered apoptosis of vascular endothelial cells.

  19. Lentiviral Delivery of HIV-1 Vpr Protein Induces Apoptosis in Transformed Cells

    NASA Astrophysics Data System (ADS)

    Stewart, Sheila A.; Poon, Betty; Jowett, Jeremy B. M.; Xie, Yiming; Chen, Irvin S. Y.

    1999-10-01

    Most current anticancer therapies act by inducing tumor cell stasis followed by apoptosis. HIV-1 Vpr effectively induces apoptosis of T cells after arrest of cells at a G2/M checkpoint. Here, we investigated whether this property of Vpr could be exploited for use as a potential anticancer agent. As a potentially safer alternative to transfer of genes encoding Vpr, we developed a method to efficiently introduce Vpr protein directly into cells. Vpr packaged into HIV-1 virions lacking a genome induced efficient cell cycle arrest and apoptosis. Introduction of Vpr into tumor cell lines of various tissue origin, including those bearing predisposing mutations in p53, XPA, and hMLH1, induced cell cycle arrest and apoptosis with high efficiency. Significantly, apoptosis mediated by virion-associated Vpr was more effective on rapidly dividing cells compared with slow-growing cells, thus, in concept, providing a potential differential effect between some types of tumor cells and surrounding normal cells. This model system provides a rationale and proof of concept for the development of potential cancer therapeutic agents based on the growth-arresting and apoptotic properties of Vpr.

  20. Ultra-sensitive assay for paclitaxel in intracellular compartments of A549 cells using liquid chromatography-tandem mass spectrometry.

    PubMed

    Wang, Tingting; Ma, Wenxiao; Sun, Yantong; Yang, Yan; Zhang, Weiping; Fawcett, J Paul; Du, Hongwei; Gu, Jingkai

    2013-01-01

    A high-performance liquid chromatography-tandem mass spectrometric (LC-MS/MS) method for the determination of paclitaxel in intracellular compartments using docetaxel as internal standard (IS) has been developed and validated. A549 cancer cells (10(6)) were incubated with paclitaxel (2ng/mL) for up to 4h and then subjected to sequential extraction of cytosolic, membrane/organelle, nuclear and cytoskeleton soluble protein. Fractions were ultrasonicated to release protein bound paclitaxel after which drug was extracted using liquid-liquid extraction with diethyl ether:dichloromethane (2:1, v/v). Chromatographic separation was then carried out on an Ascentis Express C18 column (50mm×4.6mm, 2.7μm) with a mobile phase of acetonitrile:0.1% formic acid in water (50:50, v/v). Detection involved electrospray positive ionization followed by multiple reactions monitoring of the precursor-to-product ion transitions of paclitaxel at m/z 854.4→286.3 and docetaxel at m/z 808.6→226.1. Assay validation based on samples of total cell extract in the same buffer as protein fractions showed the assay was linear over the range 2-600pg/mL with intra- and inter-day precision (as relative standard deviation) and accuracy (as relative error) of <7% and <±12%, respectively. Recovery was approximately 70% and matrix effects were minimal. The distribution of paclitaxel in subcellular components of A549 cancer cells was mainly into the cytoskeletal compartment. Copyright © 2012 Elsevier B.V. All rights reserved.

  1. [Knock-down of BCL11A expression in breast cancer cells promotes MDA-MB-231 cell apoptosis].

    PubMed

    Li, Hongli; Gui, Chen; Yan, Lijun

    2016-11-01

    Objective To detect the expression and pathological significance of B-cell CLL/lymphoma 11A (BCL11A) in breast cancer and investigate the effect of its silencing on the apoptosis of human MDA-MB-231 breast cancer cells. MethodsImmunohistochemistry was used to detect the expression of BCL11A in 62 cases of human breast cancer tissues and 8 cases of normal tissues. We synthesized siRNA targeting BCL11A, and then siRNA was transfected into MDA-MB-231 cells. Forty-eight hours later, the suppression effect of siRNA on BCL11A was determined by quantitative real-time PCR and Western blotting. The apoptosis of MDA-MB-231 cells was detected by flow cytometry. Results The BCL11A protein was mainly expressed in cytoplasm. The expression level of BCL11A in breast cancer tissues was higher than that in paracancerous tissues. The expression had correlations with tumor grade, tumor stage, while it had no correlations with the patients' age and tumor size. BCL11A-siRNA significantly suppressed the expression of BCL11A mRNA and protein as compared with the control group. MDA-MB-231 cells transfected with BCL11A-siRNA had higher apoptosis rate compared with the control group. Conclusion The BCL11A protein is highly expressed in breast cancer and knock-down of BCL11A promotes the apoptosis of MDA-MB-231 cells.

  2. Huperzine A protects neural stem cells against Aβ-induced apoptosis in a neural stem cells and microglia co-culture system

    PubMed Central

    Zhu, Ning; Lin, Jizong; Wang, Kewan; Wei, Meidan; Chen, Qingzhuang; Wang, Yong

    2015-01-01

    Objectives: This study aims to explore whether Huperzine A (HupA) could protect neural stem cells against amyloid beta-peptide Aβ induced apoptosis in a neural stem cells (NSCs) and microglia co-culture system. Methods: Rat NSCs and microglial cells were isolated, cultured and identified with immunofluorescence Assays (IFA). Co-culture systems of NSCs and microglial cells were employed using Transwell Permeable Supports. The effects of Aβ1-42 on NSCs were studied in 4 groups using co-culture systems: NSCs, Aβ+NSCs, co-culture and Aβ+co-culture groups. Bromodeoxyuridine (BrdU) incorporation and flow cytometry were utilized to assess the differences of proliferation, differentiation and apoptosis of NSCs between the groups. LQ test was performed to assess the amounts of IL-6, TNF-α and MIP-α secreted, and flow cytometry and Western blotting were used to assess apoptosis of NSCs and the expressions of Bcl-2 and Bax in each group. Results: IFA results showed that isolated rat NSCs were nestin-positive and microglial cells were CD11b/c-positive. Among all the groups, the Aβ+co-culture group has the lowest BrdU expression level, the lowest MAP2-positive, ChAT-positive cell counts and the highest NSC apoptosis rate. Smaller amounts of IL-6, TNF-α and MIP-α were being secreted by microglial cells in the HupA+Aβ+co-culture group compared with those in the Aβ+ co-culture group. Also the Bcl-2: Bax ratio was much higher in the HupA+Aβ+co-culture group than in the Aβ+co-culture group. Conclusions: HupA inhibits cell apoptosis through restraining microglia’s inflammatory response induced by Aβ1-42. PMID:26261518

  3. Mechanism of colon cancer cell apoptosis mediated by pyropheophorbide-a methylester photosensitization.

    PubMed

    Matroule, J Y; Carthy, C M; Granville, D J; Jolois, O; Hunt, D W; Piette, J

    2001-07-05

    Pyropheophorbide-a methylester (PPME) is a second generation of photosensitizers used in photodynamic therapy (PDT). We demonstrated that PPME photosensitization triggered apoptosis of colon cancer cells as measured by using several classical parameters such as DNA laddering, PARP cleavage, caspase activation and mitochondrial release of cytochrome c. Preincubation of cells with N-acetyl cysteine (NAC) or pyrolidine dithiocarbamate (PDTC) protected against apoptosis mediated by PPME photosensitization showing that reactive oxygen species (ROS) are involved as second messengers. On the other hand, photosensitization carried out in the presence of deuterium oxide (D2O) which enhances singlet oxygen (1O2) lifetime only increases necrosis without affecting apoptosis. Since PPME was localized in the endoplasmic reticulum (ER)/Golgi system and lysosomes, other messengers than ROS were tested such as calcium, Bid, Bap31, phosphorylated Bcl-2 and caspase-12 but none was clearly identified as being involved in triggering cytochrome c release from mitochondria. On the other hand, we demonstrated that the transduction pathways leading to NF-kappaB activation and apoptosis were clearly independent although NF-kappaB was shown to counteract apoptosis mediated by PPME photosensitization.

  4. Nuclear orphan receptor TLX affects gene expression, proliferation and cell apoptosis in beta cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Shi, Xiaoli; Xiong, Xiaokan; Dai, Zhe

    2015-12-04

    Nuclear orphan receptor TLX is an essential regulator of the growth of neural stem cells. However, its exact function in pancreatic islet cells is still unknown. In the present study, gene expression profiling analysis revealed that overexpression of TLX in beta cell line MIN6 causes suppression of 176 genes and upregulation of 49 genes, including a cadre of cell cycle, cell proliferation and cell death control genes, such as Btg2, Ddit3 and Gadd45a. We next examined the effects of TLX overexpression on proliferation, apoptosis and insulin secretion in MIN6 cells. Proliferation analysis using EdU assay showed that overexpression of TLXmore » increased percentage of EdU-positive cells. Cell cycle and apoptosis analysis revealed that overexpression of TLX in MIN6 cells resulted in higher percentage of cells exiting G1 into S-phase, and a 58.8% decrease of cell apoptosis induced by 0.5 mM palmitate. Moreover, TLX overexpression did not cause impairment of insulin secretion. Together, we conclude that TLX is among factors capable of controlling beta cell proliferation and survival, which may serve as a target for the development of novel therapies for diabetes. - Highlights: • TLX overexpression in MIN6 cell causes significant expression changes of 225 genes. • TLX overexpression promotes MIN6 cell proliferation and decreases cell apoptosis. • TLX overexpression does not cause impairment of insulin secretion.« less

  5. Apoptosis induced by cold shock in vitro is dependent on cell growth phase.

    PubMed

    Soloff, B L; Nagle, W A; Moss, A J; Henle, K J; Crawford, J T

    1987-06-15

    Chinese hamster V79 fibroblast cells were exposed to brief periods of cold but non-freezing temperatures at different points on the population growth curve. Upon rewarming, cells at the transition from logarithmic to stationary growth exhibited apoptosis (programmed cell death). Cells in other stages of growth, or after reentry into logarithmic growth by refeeding, did not exhibit apoptosis. Apoptosis was expressed by marked cytoplasmic blebbing, by a characteristic non-random fragmentation of DNA into nucleosomal-sized pieces, and by loss of colony-forming ability. The data suggest that cold shock served as a stimulus for susceptible cells to undergo apoptosis. Thus, the experiments describe a new in vitro system for studying the mechanisms of apoptosis.

  6. Zinc finger protein 598 inhibits cell survival by promoting UV-induced apoptosis.

    PubMed

    Yang, Qiaohong; Gupta, Romi

    2018-01-19

    UV is one of the major causes of DNA damage induced apoptosis. However, cancer cells adopt alternative mechanisms to evade UV-induced apoptosis. To identify factors that protect cancer cells from UV-induced apoptosis, we performed a genome wide short-hairpin RNA (shRNA) screen, which identified Zinc finger protein 598 (ZNF598) as a key regulator of UV-induced apoptosis. Here, we show that UV irradiation transcriptionally upregulates ZNF598 expression. Additionally, ZNF598 knockdown in cancer cells inhibited UV-induced apoptosis. In our study, we observe that ELK1 mRNA level as well as phosphorylated ELK1 levels was up regulated upon UV irradiation, which was necessary for UV irradiation induced upregulation of ZNF598. Cells expressing ELK1 shRNA were also resistant to UV-induced apoptosis, and phenocopy ZNF598 knockdown. Upon further investigation, we found that ZNF598 knockdown inhibits UV-induced apoptotic gene expression, which matches with decrease in percentage of annexin V positive cell. Similarly, ectopic expression of ZNF598 promoted apoptotic gene expression and also increased annexin V positive cells. Collectively, these results demonstrate that ZNF598 is a UV irradiation regulated gene and its loss results in resistance to UV-induced apoptosis.

  7. A nitric oxide burst precedes apoptosis in angiosperm and gymnosperm callus cells and foliar tissues.

    PubMed

    Pedroso, M C; Magalhaes, J R; Durzan, D

    2000-06-01

    Leaves and callus of Kalanchoë daigremontiana and Taxus brevifolia were used to investigate nitric oxide-induced apoptosis in plant cells. The effect of nitric oxide (NO) was studied by using a NO donor, sodium nitroprusside (SNP), a nitric oxide-synthase (NOS) inhibitor, N:(G)-monomethyl-L-arginine (NMMA), and centrifugation (an apoptosis-inducing treatment in these species). NO production was visualized in cells and tissues with a specific probe, diaminofluorescein diacetate (DAF-2 DA). DNA fragmentation was detected in situ by the terminal deoxynucleotidyl transferase-mediated dUTP nick end labelling (TUNEL) method. In both species, NO was detected diffused in the cytosol of epidermal cells and in chloroplasts of guard cells and leaf parenchyma cells. Centrifugation increased NO production, DNA fragmentation and subsequent cell death by apoptosis. SNP mimicked centrifugation results. NMMA significantly decreased NO production and apoptosis in both species. The inhibitory effect of NMMA on NO production suggests that a putative NOS is present in Kalanchoë and Taxus cells. The present results demonstrated the involvement of NO on DNA damage leading to cell death, and point to a potential role of NO as a signal molecule in these plants.

  8. Autophagy regulates chlorpyrifos-induced apoptosis in SH-SY5Y cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Park, Jae Hyeon; Hanyang Biomedical Research Institute, Seoul; Graduate School of Biomedical Science and Engineering, Hanyang University, Seoul

    Recent studies have shown that up-regulation of autophagy may be a tractable therapeutic intervention for clearing disease-causing proteins, including α-synuclein, ubiquitin, and other misfolded or aggregated proteins in pesticide-induced neurodegeneration. In a previous study, we reported that chlorpyrifos (CPF)-induced mitochondria-dependent apoptosis is mediated through reactive oxygen species in SH-SY5Y cells. In this study, we explored a novel pharmacotherapeutic approach to prevent CPF neurotoxicity involving the regulation of autophagy. We investigated the modulation of CPF-induced apoptosis according to autophagy regulation. We found that CPF induced apoptosis in SH-SY5Y cells, as demonstrated by the activation of caspase-3 and nuclear condensation. In addition,more » we observed that cells treated with CPF underwent autophagic cell death by monitoring the expression of LC3-II and p62. Pretreatment with the autophagy inducer rapamycin significantly enhanced the cell viability of CPF-exposed cells, and the enhancement of cell viability was partially due to alleviation of CPF-induced apoptosis via a decrease in levels of cleaved caspase-3. Specifically, rapamycin pretreatment decreased Bax and increased Bcl-2 expression in mitochondria. In addition, rapamycin significantly decreased cytochrome c release in from mitochondria into the cytosol. However, pretreatment of cells with the autophagy inhibitor, 3-methyladenine (3MA), remarkably increased CPF toxicity in these cells; this with correlated with increased expression of Bax and decreased expression of Bcl-2 in mitochondria. Our results suggest that CPF-induced cytotoxicity is modified by autophagy regulation and that rapamycin protects against CPF-induced apoptosis by enhancing autophagy. Pharmacologic induction of autophagy by rapamycin may be a useful treatment strategy in neurodegenerative disorders. - Highlights: ► Chlorpyrifos (CPF) is cytotoxic to SH-SY5Y cells ► CPF-induced cytotoxicity is

  9. LED-activated pheophorbide a in ovarian cancer cells: Cytotoxicity and apoptosis induction

    NASA Astrophysics Data System (ADS)

    Liu, L.; Xu, C. S.; Xia, X. S.; Leung, A. W. N.

    2011-02-01

    Pheophorbide a (Pa) from Chinese herbal medicine Scutellaria Barbata and Silkworm excreta has been proved to be potential photosensitizer. The present study investigated the cytotoxicity of ovarian cancer cells induced by LED-activated Pa using light microscopy with the SRB staining. We further investigated the apoptosis of the cells 6 h after LED-activated Pa using of the flow cytometer with PI staining and nuclear staining. The results showed that LED-activated Pa remarkably caused cell death of ovarian cancer cells. The condensation of chromatin, nuclear fragmentations, and 12.3% of cells containing subdiploid levels of DNA were found in the ovarian cancer cells after the treatment of LED-activated Pa. These data demonstrated that LED-activated Pa could cause significant cytotoxicity and apoptosis of ovarian cancer cells.

  10. Disruption of Redox Homeostasis in Tumor Necrosis Factor-Induced Apoptosis in a Murine Hepatocyte Cell Line

    PubMed Central

    Pierce, Robert H.; Campbell, Jean S.; Stephenson, Alyssa B.; Franklin, Christopher C.; Chaisson, Michelle; Poot, Martin; Kavanagh, Terrance J.; Rabinovitch, Peter S.; Fausto, Nelson

    2000-01-01

    Tumor necrosis factor (TNF) is a mediator of the acute phase response in the liver and can initiate proliferation and cause cell death in hepatocytes. We investigated the mechanisms by which TNF causes apoptosis in hepatocytes focusing on the role of oxidative stress, antioxidant defenses, and mitochondrial damage. The studies were conducted in cultured AML12 cells, a line of differentiated murine hepatocytes. As is the case for hepatocytes in vivo, AML12 cells were not sensitive to cell death by TNF alone, but died by apoptosis when exposed to TNF and a small dose of actinomycin D (Act D). Morphological signs of apoptosis were not detected until 6 hours after the treatment and by 18 hours ∼50% of the cells had died. Exposure of the cells to TNF+Act D did not block NFκB nuclear translocation, DNA binding, or its overall transactivation capacity. Induction of apoptosis was characterized by oxidative stress indicated by the loss of NAD(P)H and glutathione followed by mitochondrial damage that included loss of mitochondrial membrane potential, inner membrane structural damage, and mitochondrial condensation. These changes coincided with cytochrome C release and the activation of caspases-8, -9, and -3. TNF-induced apoptosis was dependent on glutathione levels. In cells with decreased levels of glutathione, TNF by itself in the absence of transcriptional blocking acted as an apoptotic agent. Conversely, the antioxidant α-lipoic acid, that protected against the loss of glutathione in cells exposed to TNF+Act D completely prevented mitochondrial damage, caspase activation, cytochrome C release, and apoptosis. The results demonstrate that apoptosis induced by TNF+Act D in AML12 cells involves oxidative injury and mitochondrial damage. As injury was regulated to a larger extent by the glutathione content of the cells, we suggest that the combination of TNF+Act D causes apoptosis because Act D blocks the transcription of genes required for antioxidant defenses. PMID

  11. Identification of a Short Cell-Penetrating Peptide from Bovine Lactoferricin for Intracellular Delivery of DNA in Human A549 Cells

    PubMed Central

    Liu, Betty R.; Huang, Yue-Wern; Aronstam, Robert S.; Lee, Han-Jung

    2016-01-01

    Cell-penetrating peptides (CPPs) have been shown to deliver cargos, including protein, DNA, RNA, and nanomaterials, in fully active forms into live cells. Most of the CPP sequences in use today are based on non-native proteins that may be immunogenic. Here we demonstrate that the L5a CPP (RRWQW) from bovine lactoferricin (LFcin), stably and noncovalently complexed with plasmid DNA and prepared at an optimal nitrogen/phosphate ratio of 12, is able to efficiently enter into human lung cancer A549 cells. The L5a CPP delivered a plasmid containing the enhanced green fluorescent protein (EGFP) coding sequence that was subsequently expressed in cells, as revealed by real-time PCR and fluorescent microscopy at the mRNA and protein levels, respectively. Treatment with calcium chloride increased the level of gene expression, without affecting CPP-mediated transfection efficiency. Zeta-potential analysis revealed that positively electrostatic interactions of CPP/DNA complexes correlated with CPP-mediated transport. The L5a and L5a/DNA complexes were not cytotoxic. This biomimetic LFcin L5a represents one of the shortest effective CPPs and could be a promising lead peptide with less immunogenic for DNA delivery in gene therapy. PMID:26942714

  12. Induction of apoptosis of liver cancer cells by nanosecond pulsed electric fields (nsPEFs).

    PubMed

    He, Ling; Xiao, Deyou; Feng, Jianguo; Yao, Chenguo; Tang, Liling

    2017-02-01

    The application of nanosecond pulsed electric fields (nsPEFs) is a novel method to induce the death of cancer cells. NsPEFs could directly function on the cell membrane and activate the apoptosis pathways, then induce apoptosis in various cell lines. However, the nsPEFs-inducing-apoptosis action sites and the exact pathways are not clear now. In this study, nsPEFs were applied to the human liver cancer cells HepG2 with different parameters. By apoptosis assay, morphological observation, detecting the mitochondrial membrane potential (ΔΨ m ), intracellular calcium ion concentration ([Ca 2+ ]i) and the expressions of key apoptosis factors, we demonstrated that nsPEFs could induce the morphology of cell apoptosis, the change in ΔΨ m , [Ca 2+ ]i and the upregulation of some key apoptosis factors, which revealed the responses of liver cancer cells and indicated that cells may undergo apoptosis through the mitochondria-dependent pathway after nsPEFs were applied.

  13. A COMPREHENSIVE STUDY ON APOPTOSIS INDUCTION BY AZADIRACHTIN IN Spodoptera frugiperda CULTURED CELL LINE Sf9.

    PubMed

    Shu, Benshui; Wang, Wenxiang; Hu, Qingbo; Huang, Jingfei; Hu, Meiying; Zhong, Guohua

    2015-07-01

    The induction of apoptosis by azadirachtin, a well-known botanical tetranortriterpenoid isolated from the neem tree (Azadirachta indica A. Juss) and other members of the Meliaceae, was investigated in Spodoptera frugiperda cultured cell line (Sf9). Morphological changes in Sf9 cells treated by various concentrations of azadirachtin were observed at different times under light microscopy. Morphological and biochemical analysis indicated that Sf9 cells treated by 1.5 μg/mL azadirachtin showed typical morphological changes, which were indicative of apoptosis and a clear DNA ladder. The flow cytometry analysis showed the apoptosis rate reached a maximum value of 32.66% at 24 h with 1.5 μg/mL azadirachtin in Sf9 cells. The inhibition of Sf9 cell proliferation suggested that the effect of azadirachtin was dose dependent and the EC50 at 48 and 72 h was 2.727 × 10(-6) and 6.348 × 10(-9) μg/mL, respectively. The treatment of azadirachtin in Sf9 cells could significantly increase the activity of Sf caspase-1, but showed no effect on the activity of Topo I, suggesting that the apoptosis induced by azadirachtinin Sf9 cells is through caspase-dependent pathway. These results provided not only a series of morphological, biochemical, and toxicological comprehensive evidences for induction of apoptosis by azadirachtin, but also a reference model for screening insect cell apoptosis inducers from natural compounds. © 2015 Wiley Periodicals, Inc.

  14. Aminomethylphosphonic Acid and Methoxyacetic Acid Induce Apoptosis in Prostate Cancer Cells

    PubMed Central

    Parajuli, Keshab R.; Zhang, Qiuyang; Liu, Sen; You, Zongbing

    2015-01-01

    Aminomethylphosphonic acid (AMPA) and its parent compound herbicide glyphosate are analogs to glycine, which have been reported to inhibit proliferation and promote apoptosis of cancer cells, but not normal cells. Methoxyacetic acid (MAA) is the active metabolite of ester phthalates widely used in industry as gelling, viscosity and stabilizer; its exposure is associated with developmental and reproductive toxicities in both rodents and humans. MAA has been reported to suppress prostate cancer cell growth by inducing growth arrest and apoptosis. However, it is unknown whether AMPA and MAA can inhibit cancer cell growth. In this study, we found that AMPA and MAA inhibited cell growth in prostate cancer cell lines (LNCaP, C4-2B, PC-3 and DU-145) through induction of apoptosis and cell cycle arrest at the G1 phase. Importantly, the AMPA-induced apoptosis was potentiated with the addition of MAA, which was due to downregulation of the anti-apoptotic gene baculoviral inhibitor of apoptosis protein repeat containing 2 (BIRC2), leading to activation of caspases 7 and 3. These results demonstrate that the combination of AMPA and MAA can promote the apoptosis of prostate cancer cells, suggesting that they can be used as potential therapeutic drugs in the treatment of prostate cancer. PMID:26006246

  15. Aminomethylphosphonic acid and methoxyacetic acid induce apoptosis in prostate cancer cells.

    PubMed

    Parajuli, Keshab R; Zhang, Qiuyang; Liu, Sen; You, Zongbing

    2015-05-22

    Aminomethylphosphonic acid (AMPA) and its parent compound herbicide glyphosate are analogs to glycine, which have been reported to inhibit proliferation and promote apoptosis of cancer cells, but not normal cells. Methoxyacetic acid (MAA) is the active metabolite of ester phthalates widely used in industry as gelling, viscosity and stabilizer; its exposure is associated with developmental and reproductive toxicities in both rodents and humans. MAA has been reported to suppress prostate cancer cell growth by inducing growth arrest and apoptosis. However, it is unknown whether AMPA and MAA can inhibit cancer cell growth. In this study, we found that AMPA and MAA inhibited cell growth in prostate cancer cell lines (LNCaP, C4-2B, PC-3 and DU-145) through induction of apoptosis and cell cycle arrest at the G1 phase. Importantly, the AMPA-induced apoptosis was potentiated with the addition of MAA, which was due to downregulation of the anti-apoptotic gene baculoviral inhibitor of apoptosis protein repeat containing 2 (BIRC2), leading to activation of caspases 7 and 3. These results demonstrate that the combination of AMPA and MAA can promote the apoptosis of prostate cancer cells, suggesting that they can be used as potential therapeutic drugs in the treatment of prostate cancer.

  16. Hsp27 Inhibition with OGX-427 Sensitizes Non-Small Cell Lung Cancer Cells to Erlotinib and Chemotherapy.

    PubMed

    Lelj-Garolla, Barbara; Kumano, Masafumi; Beraldi, Eliana; Nappi, Lucia; Rocchi, Palma; Ionescu, Diana N; Fazli, Ladan; Zoubeidi, Amina; Gleave, Martin E

    2015-05-01

    Non-small cell lung cancer (NSCLC) is the most frequent cause of death from cancer worldwide. Despite the availability of active chemotherapy regimens and EGFR tyrosine kinase inhibitors, all advanced patients develop recurrent disease after first-line therapy. Although Hsp27 is a stress-induced chaperone that promotes acquired resistance in several cancers, its relationship to treatment resistance in NSCLC has not been defined. Understanding adaptive responses of acquired resistance will help guide new strategies to control NSCLC. Hsp27 levels were evaluated in an HCC827 erlotinib-resistant-derived cell line (HCC-827Resistant), and sensitivity to erlotinib was examined in Hsp27-overexpressing A549 cells. The role of Hsp27 in both erlotinib and cytotoxic treatment resistance was evaluated in HCC-827 and A549 NSCLC cells using the Hsp27 antisense drug OGX-427. The effect of OGX-427 in combination with erlotinib was also assessed in mice bearing A549 xenografts. Hsp27 is induced by erlotinib and protects NSCLC cells from treatment-induced apoptosis, whereas OGX-427 sensitizes NSCLC cells to erlotinib. Interestingly, increased resistance to erlotinib was observed when Hsp27 was increased either in HCC827 erlotinib-resistant or overexpressing A549 cells. Combining OGX-427 with erlotinib significantly enhanced antitumor effects in vitro and delayed A549 xenograft growth in vivo. OGX-427 also significantly enhanced the activity of cytotoxic drugs used for NSCLC. These data indicate that treatment-induced Hsp27 contributes to the development of resistance, and provides preclinical proof-of-principle that inhibition of stress adaptive pathways mediated by Hsp27 enhances the activity of erlotinib and chemotherapeutics. ©2015 American Association for Cancer Research.

  17. EF24, a novel synthetic curcumin analog, induces apoptosis in cancer cells via a redox-dependent mechanism.

    PubMed

    Adams, Brian K; Cai, Jiyang; Armstrong, Jeff; Herold, Marike; Lu, Yang J; Sun, Aiming; Snyder, James P; Liotta, Dennis C; Jones, Dean P; Shoji, Mamoru

    2005-03-01

    In this study, we show that the novel synthetic curcumin analog, EF24, induces cell cycle arrest and apoptosis by means of a redox-dependent mechanism in MDA-MB-231 human breast cancer cells and DU-145 human prostate cancer cells. Cell cycle analysis demonstrated that EF24 causes a G2/M arrest in both cell lines, and that this cell cycle arrest is followed by the induction of apoptosis as evidenced by caspase-3 activation, phosphatidylserine externalization and an increased number of cells with a sub-G1 DNA fraction. In addition, we demonstrate that EF24 induces a depolarization of the mitochondrial membrane potential, suggesting that the compound may also induce apoptosis by altering mitochondrial function. EF24, like curcumin, serves as a Michael acceptor reacting with glutathione (GSH) and thioredoxin 1. Reaction of EF24 with these agents in vivo significantly reduced intracellular GSH as well as oxidized GSH in both the wild-type and Bcl-xL overexpressing HT29 human colon cancer cells. We therefore propose that the anticancer effect of a novel curcumin analog, EF24, is mediated in part by redox-mediated induction of apoptosis.

  18. [Effects of cucurmosin on the cell proliferation and apoptosis in human pancreatic PANC-1 cells].

    PubMed

    Xu, Chun-Sen; Huang, He-Guang; Chen, Ming-Huang

    2012-02-01

    To observe the effects of cucurmosin (CUS) on the cell proliferation and apoptosis in pancreatic PANC-1 cells. The inhibition of CUS on the PANC-1 cell growth was observed using MTT assay. The inhibition ratio of CUS on the pancreatic orthotopic transplantation was in vivo observed in the NOD/SCID mouse model. The changes of microstructure of the apoptosis-inducing effect of CUS on PANC-1 was observed under electron microscope. The cell cycle and apoptosis after CUS intervention was detected using flow cytometry. The Caspase-3 activity after CUS treatment was detected using enzyme linked immunospecific assay (ELISA). Treatment with CUS at the dose of 0.125, 0.25, and 0.5 mg/kg inhibited the growth of pancreatic carcinoma PANC-1 xenografs with the ratio of 45.2%, 50.0%, and 59.7%, respectively (P < 0.05). After exposure to 10 microg/mL CUS for 24 h, most cells presented typical morphologic changes of apoptosis such as chromatin condensation and shrunken nucleus. The apoptotic cells increased. Some nuclear shrinkage and fragmentation, as well as the apoptotic body were observed when cells were exposed to CUS for 72 h. Being exposed to 0, 2.5, 10.0, and 40.0 microg/mL of the CUS for 72 h, the percentage of G0/G1 phase cells was 46.56% +/- 5.08%, 53.33% +/- 5.05%, 67.50% +/- 6.50%, and 77.00% +/- 6.73%, respectively (P < 0.05). The apoptosis ratio was 2.50% +/- 0.13%, 8.30% +/- 1.23%, 23.40% +/- 2.45%, and 48.50% +/- 3.65% shown by Annexin V/PI (P < 0.05). The Caspase-3 activity (unit) was 0.009 +/- 0.002, 0.011 +/- 0.003, 0.035 +/- 0.009, and 0.065 +/- 0.009, respectively (P < 0.05). These data showed that CUS induced the apoptosis of PANC-1 cells in a dose-dependent maner. Being exposed to 40.0 microg/mL of the CUS for 24, 48, and 72 h, the percentage of G0/ G1 phase cells was 56.60% +/- 6.65%, 67.83% +/- 6.76%, and 77.00% +/- 6.73%, respectively (P < 0.05), the apoptosis ratio was 16.51% +/- 2.97%, 38.51% +/- 2.38%, and 48.50% +/- 3.65% shown by Annexin V/PI (P < 0

  19. Oxidative Stress-Responsive Apoptosis Inducing Protein (ORAIP) Plays a Critical Role in High Glucose-Induced Apoptosis in Rat Cardiac Myocytes and Murine Pancreatic β-Cells.

    PubMed

    Yao, Takako; Fujimura, Tsutomu; Murayama, Kimie; Okumura, Ko; Seko, Yoshinori

    2017-10-18

    We previously identified a novel apoptosis-inducing humoral factor in the conditioned medium of hypoxic/reoxygenated-cardiac myocytes. We named this novel post-translationally-modified secreted-form of eukaryotic translation initiation factor 5A Oxidative stress-Responsive Apoptosis-Inducing Protein (ORAIP). We confirmed that myocardial ischemia/reperfusion markedly increased plasma ORAIP levels and rat myocardial ischemia/reperfusion injury was clearly suppressed by neutralizing anti-ORAIP monoclonal antibodies (mAbs) in vivo. In this study, to investigate the mechanism of cell injury of cardiac myocytes and pancreatic β-cells involved in diabetes mellitus (DM), we analyzed plasma ORAIP levels in DM model rats and the role of ORAIP in high glucose-induced apoptosis of cardiac myocytes in vitro. We also examined whether recombinant-ORAIP induces apoptosis in pancreatic β-cells. Plasma ORAIP levels in DM rats during diabetic phase were about 18 times elevated as compared with non-diabetic phase. High glucose induced massive apoptosis in cardiac myocytes (66.2 ± 2.2%), which was 78% suppressed by neutralizing anti-ORAIP mAb in vitro. Furthermore, recombinant-ORAIP clearly induced apoptosis in pancreatic β-cells in vitro. These findings strongly suggested that ORAIP plays a pivotal role in hyperglycemia-induced myocardial injury and pancreatic β-cell injury in DM. ORAIP will be a biomarker and a critical therapeutic target for cardiac injury and progression of DM itself.

  20. Clinostat rotation induces apoptosis in luteal cells of the pregnant rat

    NASA Technical Reports Server (NTRS)

    Yang, Hyunwon; Bhat, Ganapathy K.; Sridaran, Rajagopala

    2002-01-01

    Recent studies have shown that microgravity induces changes at the cellular level, including apoptosis. However, it is unknown whether microgravity affects luteal cell function. This study was performed to assess whether microgravity conditions generated by clinostat rotation induce apoptosis and affect steroidogenesis by luteal cells. Luteal cells isolated from the corpora lutea of Day 8 pregnant rats were placed in equal numbers in slide flasks (chamber slides). One slide flask was placed in the clinostat and the other served as a stationary control. At 48 h in the clinostat, whereas the levels of progesterone and total cellular protein decreased, the number of shrunken cells increased. To determine whether apoptosis occurred in shrunken cells, Comet and TUNEL assays were performed. At 48 h, the percentage of apoptotic cells in the clinostat increased compared with that in the control. To investigate how the microgravity conditions induce apoptosis, the active mitochondria in luteal cells were detected with JC-1 dye. Cells in the control consisted of many active mitochondria, which were evenly distributed throughout the cell. In contrast, cells in the clinostat displayed fewer active mitochondria, which were distributed either to the outer edge of the cell or around the nucleus. These results suggest that mitochondrial dysfunction induced by clinostat rotation could lead to apoptosis in luteal cells and suppression of progesterone production.

  1. Huperzine A derivative M3 protects PC12 cells against sodium nitroprusside-induced apoptosis

    PubMed Central

    Ning, Na; Hu, Jin-feng; Yuan, Yu-he; Zhang, Xin-yuan; Dai, Jun-gui; Chen, Nai-hong

    2012-01-01

    Aim: To investigate the effects of M3, a derivative of huperzine A, on the apoptosis induced by sodium nitroprusside (SNP) in PC12 cells. Methods: Cell viability was detected using MTT method. Apoptosis was examined with annexin V/prodium iodide (PI) stain. The levels of reactive oxygen species (ROS) were measured using fluorophotometric quantitation. The amount of malonaldehyde (MDA) was determined with MDA detection kits. The expression of caspase-3 and Hsp70 were analyzed using Western blotting. Results: Exposure of PC12 cells to SNP (200 μmol/L) for 24 h decreased the cell viability to 69.0% of that in the control group. Pretreatment with M3 (10 μmol/L) or huperzine A (10 μmol/L) significantly protected the cells against SNP-induced injury and apoptosis; the ratio of apoptotic bodies in PC12 cells was decreased from 27.3% to 15.0%. Pretreatment with M3 (10 μmol/L) significantly decreased ROS and MDA levels, and increased the expression of Hsp70 in the cells. Quercetin (10 μmol/L) blocked the protective effect of M3, while did not influence on that of huperzine A. Conclusion: M3 protects PC12 cells against SNP-induced apoptosis, possible due to ROS scavenging and Hsp70 induction. PMID:22120967

  2. Dihydroartemisinin induces apoptosis preferentially via a Bim-mediated intrinsic pathway in hepatocarcinoma cells.

    PubMed

    Qin, Guiqi; Zhao, ChuBiao; Zhang, Lili; Liu, Hongyu; Quan, Yingyao; Chai, Liuying; Wu, Shengnan; Wang, Xiaoping; Chen, Tongsheng

    2015-08-01

    This report is designed to dissect the detail molecular mechanism by which dihydroartemisinin (DHA), a derivative of artemisinin, induces apoptosis in human hepatocellular carcinoma (HCC) cells. DHA induced a loss of the mitochondrial transmemberane potential (ΔΨm), release of cytochrome c, activation of caspases, and externalization of phosphatidylserine indicative of apoptosis induction. Compared with the modest inhibitory effects of silencing Bax, silencing Bak largely prevented DHA-induced ΔΨm collapse and apoptosis though DHA induced a commensurable activation of Bax and Bak, demonstrating a key role of the Bak-mediated intrinsic apoptosis pathway. DHA did not induce Bid cleavage and translocation from cytoplasm to mitochondria and had little effects on the expressions of Puma and Noxa, but did increase Bim and Bak expressions and decrease Mcl-1 expression. Furthermore, the cytotoxicity of DHA was remarkably reduced by silencing Bim, and modestly but significantly reduced by silencing Puma or Noxa. Silencing Bim or Noxa preferentially reduced DHA-induced Bak activation, while silencing Puma preferentially reduced DHA-induced Bax activation, demonstrating that Bim and to a lesser extent Noxa act as upstream mediators to trigger the Bak-mediated intrinsic apoptosis pathway. In addition, silencing Mcl-1 enhanced DHA-induced Bak activation and apoptosis. Taken together, our data demonstrate a crucial role of Bim in preferentially regulating the Bak/Mcl-1 rheostat to mediate DHA-induced apoptosis in HCC cells.

  3. Osthole inhibits proliferation and induces apoptosis in human osteosarcoma cells.

    PubMed

    Ding, Yong; Lu, Xiongwei; Hu, Xiaopeng; Ma, Jie; Ding, Huan

    2014-02-01

    The purpose of this study was to investigate the effect of osthole on osteosarcoma cell proliferation and apoptosis. Cell counting Kit-8 assay was performed to establish the effects of osthole on osteosarcoma MG-63 cell proliferation. Annexin V-FITC/PI was performed to analyze the apoptotic rate of the cells. The inhibitory effects of osthole on the expression of BCL-2, BAX, and caspase-3 were detected by Western blotting. Osthole inhibited the growth of human osteosarcoma MG-63 cells by inhibiting cell proliferation and induced cell apoptosis. Western blotting demonstrated that osthole downregulated the expressions of BCL-2 and caspase-3 and upregulated the expression of BAX in human osteosarcoma cells. Osthole can inhibit osteosarcoma cell proliferation and induced apoptosis effectively in a dose-dependent manner through downregulating the expression of BCL-2 and caspase-3 proteins levels and upregulating the expression of BAX proteins levels.

  4. Propolis Augments Apoptosis Induced by Butyrate via Targeting Cell Survival Pathways

    PubMed Central

    Drago, Eric; Bordonaro, Michael; Lee, Seon; Atamna, Wafa; Lazarova, Darina L.

    2013-01-01

    Diet is one of the major lifestyle factors affecting incidence of colorectal cancer (CC), and despite accumulating evidence that numerous diet-derived compounds modulate CC incidence, definitive dietary recommendations are not available. We propose a strategy that could facilitate the design of dietary supplements with CC-preventive properties. Thus, nutrient combinations that are a source of apoptosis-inducers and inhibitors of compensatory cell proliferation pathways (e.g., AKT signaling) may produce high levels of programmed death in CC cells. Here we report the combined effect of butyrate, an apoptosis inducer that is produced through fermentation of fiber in the colon, and propolis, a honeybee product, on CC cells. We established that propolis increases the apoptosis of CC cells exposed to butyrate through suppression of cell survival pathways such as the AKT signaling. The programmed death of CC cells by combined exposure to butyrate and propolis is further augmented by inhibition of the JNK signaling pathway. Analyses on the contribution of the downstream targets of JNK signaling, c-JUN and JAK/STAT, to the apoptosis of butyrate/propolis-treated CC cells ascertained that JAK/STAT signaling has an anti-apoptotic role; whereas, the role of cJUN might be dependent upon regulatory cell factors. Thus, our studies ascertained that propolis augments apoptosis of butyrate-sensitive CC cells and re-sensitizes butyrate-resistant CC cells to apoptosis by suppressing AKT signaling and downregulating the JAK/STAT pathway. Future in vivo studies should evaluate the CC-preventive potential of a dietary supplement that produces high levels of colonic butyrate, propolis, and diet-derived JAK/STAT inhibitors. PMID:24023824

  5. [Influence of human gastrointestinal tract bacterial pathogens on host cell apoptosis].

    PubMed

    Wronowska, Weronika; Godlewska, Renata; Jagusztyn-Krynicka, Elzbieta Katarzyna

    2005-01-01

    Several pathogenic bacteria are able to trigger apoptosis in the host cell, but the mechanisms by which it occurs differ, and the resulting pathology can take different courses. Induction and/or blockage of programmed cell death upon infection is a result of complex interaction of bacterial proteins with cellular proteins involved in signal transduction and apoptosis. In this review we focus on pro/anti-apoptotic activities exhibited by two enteric pathogens Salmonella enterica, Yersinia spp. and gastric pathogen Helicobacter pylori. We present current knowledge on how interaction between mammalian and bacterial cell relates to the molecular pathways of apoptosis, and what is the role of apoptosis in pathogenesis.

  6. Balance between senescence and apoptosis is regulated by telomere damage-induced association between p16 and caspase-3.

    PubMed

    Panneer Selvam, Shanmugam; Roth, Braden M; Nganga, Rose; Kim, Jisun; Cooley, Marion A; Helke, Kristi L; Smith, Charles D; Ogretmen, Besim

    2018-05-10

    Telomerase activation protects cells from telomere damage by delaying senescence and inducing cell immortalization, whereas telomerase inhibition mediates rapid senescence or apoptosis. However, the cellular mechanisms that determine telomere damage-dependent senescence versus apoptosis induction are largely unknown. Here, we demonstrate that telomerase instability mediated by silencing of sphingosine kinase 2 (SPHK2) and sphingosine 1-phosphate (S1P), which binds and stabilizes telomerase, induces telomere damage-dependent caspase-3 activation and apoptosis, but not senescence, in p16-deficient lung cancer cells or tumors. These outcomes were prevented by knockdown of a tumor-suppressor protein, transcription factor 21 (TCF21), or by ectopic expression of WT human telomerase reverse transcriptase (hTERT), but not mutant hTERT with altered S1P binding. Interestingly, SphK2-deficient mice exhibited accelerated aging and telomerase instability that increased telomere damage and senescence via p16 activation especially in testes tissues, but not in apoptosis. Moreover, p16 silencing in SphK2-/- mouse embryonic fibroblasts activated caspase-3 and apoptosis without inducing senescence. Further, ectopic WT p16 expression in p16-deficient A549 lung cancer cells prevented TCF21 and caspase-3 activation, and resulted in senescence in response to SphK2/S1P inhibition and telomere damage. Mechanistically, a p16 mutant with impaired [MS2] caspase-3 association did not prevent telomere damage-induced apoptosis, indicating that an association between p16 and caspase-3 proteins forces senescence induction by inhibiting caspase-3 activation and apoptosis.[MS3]  These results suggest that p16 plays a direct role in telomere damage-dependent senescence by limiting apoptosis via binding to caspase-3, revealing a direct link between telomere damage-dependent senescence and apoptosis with regards to aging and cancer. Published under license by The American Society for Biochemistry

  7. Restraining reactive oxygen species in Listeria monocytogenes promotes the apoptosis of glial cells.

    PubMed

    Li, Sen; Li, Yixuan; Chen, Guowei; Zhang, Jingchen; Xu, Fei; Wu, Man

    2017-07-01

    Listeria monocytogenes is a facultative anaerobic foodborne pathogen that can traverse the blood-brain barrier and cause brain infection. L. monocytogenes infection induces host cell apoptosis in several cell types. In this study, we investigated the apoptosis of human glioma cell line U251 invaded by L. monocytogenes and evaluated the function of bacterial reactive oxygen species (ROS) during infection. Bacterial ROS level was reduced by carrying out treatment with N-acetyl cysteine (NAC) and diphenyleneiodonium chloride (DPI). After infection, the apoptosis of U251 cells was examined by flow cytometry assay and propidium iodide staining. DPI and NAC efficiently decreased ROS level in L. monocytogenes without affecting bacterial growth. Moreover, the apoptosis of glial cells was enhanced upon invasion of DPI- and NAC-pretreated L. monocytogenes. Results indicate that the apoptosis of glial cells can be induced by L. monocytogenes, and that the inhibition of bacterial ROS increases the apoptosis of host cells.

  8. BIGH3 protein and macrophages in retinal endothelial cell apoptosis.

    PubMed

    Mondragon, Albert A; Betts-Obregon, Brandi S; Moritz, Robert J; Parvathaneni, Kalpana; Navarro, Mary M; Kim, Hong Seok; Lee, Chi Fung; LeBaron, Richard G; Asmis, Reto; Tsin, Andrew T

    2015-01-01

    Diabetes is a pandemic disease with a higher occurrence in minority populations. The molecular mechanism to initiate diabetes-associated retinal angiogenesis remains largely unknown. We propose an inflammatory pathway of diabetic retinopathy in which macrophages in the diabetic eye provide TGFβ to retinal endothelial cells (REC) in the retinal microvasculature. In response to TGFβ, REC synthesize and secrete a pro-apoptotic BIGH3 (TGFβ-Induced Gene Human Clone 3) protein, which acts in an autocrine loop to induce REC apoptosis. Rhesus monkey retinal endothelial cells (RhREC) were treated with dMCM (cell media of macrophages treated with high glucose and LDL) and assayed for apoptosis (TUNEL), BIGH3 mRNA (qPCR), and protein (Western blots) expressions. Cells were also treated with ΤGFβ1 and 2 for BIGH3 mRNA and protein expression. Inhibition assays were carried out using antibodies for TGFβ1 and for BIGH3 to block apoptosis and mRNA expression. BIGH3 in cultured RhREC cells were identified by immunohistochemistry (IHC). Distribution of BIGH3 and macrophages in the diabetic mouse retina was examined with IHC. RhRECs treated with dMCM or TGFβ showed a significant increase in apoptosis and BIGH3 protein expression. Recombinant BIGH3 added to RhREC culture medium led to a dose-dependent increase in apoptosis. Antibodies (Ab) directed against BIGH3 and TGFβ, as well as TGFβ receptor blocker resulted in a significant reduction in apoptosis induced by either dMCM, TGFβ or BIGH3. IHC showed that cultured RhREC constitutively expressed BIGH3. Macrophage and BIGH3 protein were co-localized to the inner retina of the diabetic mouse eye. Our results support a novel inflammatory pathway for diabetic retinopathy. This pathway is initiated by TGFβ released from macrophages, which promotes synthesis and release of BIGH3 protein by REC and REC apoptosis.

  9. Induction of apoptosis in HT-29 colon cancer cells by phloretin.

    PubMed

    Park, So Young; Kim, Eun Ji; Shin, Hyun-Kyung; Kwon, Dae Young; Kim, Myung Sunny; Surh, Young-Joon; Park, Jung Han Yoon

    2007-12-01

    Phloretin, which is present in apples and pears, has been found to inhibit the growth of several cancer cells and induce apoptosis of B16 melanoma and HL60 human leukemia cells. The present study examined whether and how phloretin induces apoptosis of HT-29 human colon cancer cells. Phloretin (0-100 micromol/L) substantially decreased viable cell number and induced apoptosis of HT-29 cells in a dose-dependent manner. Western blot analysis of total cell lysates revealed that phloretin increased the protein levels of Bax but had no effect on Bcl-2. In addition, phloretin induced cleavage of caspase-8, -9, -7, and -3 and poly(ADP-ribose) polymerase. Furthermore, phloretin increased the levels of cytochrome c and Smac/Diablo in the cytosol. The present results indicate that phloretin inhibits HT-29 cell growth by inducing apoptosis, which may be mediated through changes in mitochondrial membrane permeability and activation of the caspase pathways.

  10. Arecoline decreases interleukin-6 production and induces apoptosis and cell cycle arrest in human basal cell carcinoma cells

    DOE Office of Scientific and Technical Information (OSTI.GOV)

    Huang, Li-Wen; Hsieh, Bau-Shan; Cheng, Hsiao-Ling

    2012-01-15

    Arecoline, the most abundant areca alkaloid, has been reported to decrease interleukin-6 (IL-6) levels in epithelial cancer cells. Since IL-6 overexpression contributes to the tumorigenic potency of basal cell carcinoma (BCC), this study was designed to investigate whether arecoline altered IL-6 expression and its downstream regulation of apoptosis and the cell cycle in cultured BCC-1/KMC cells. BCC-1/KMC cells and a human keratinocyte cell line, HaCaT, were treated with arecoline at concentrations ranging from 10 to 100 μg/ml, then IL-6 production and expression of apoptosis- and cell cycle progress-related factors were examined. After 24 h exposure, arecoline inhibited BCC-1/KMC cell growthmore » and decreased IL-6 production in terms of mRNA expression and protein secretion, but had no effect on HaCaT cells. Analysis of DNA fragmentation and chromatin condensation showed that arecoline induced apoptosis of BCC-1/KMC cells in a dose-dependent manner, activated caspase-3, and decreased expression of the anti-apoptotic protein Bcl-2. In addition, arecoline induced progressive and sustained accumulation of BCC-1/KMC cells in G2/M phase as a result of reducing checkpoint Cdc2 activity by decreasing Cdc25C phosphatase levels and increasing p53 levels. Furthermore, subcutaneous injection of arecoline led to decreased BCC-1/KMC tumor growth in BALB/c mice by inducing apoptosis. This study demonstrates that arecoline has potential for preventing BCC tumorigenesis by reducing levels of the tumor cell survival factor IL-6, increasing levels of the tumor suppressor factor p53, and eliciting cell cycle arrest, followed by apoptosis. Highlights: ► Arecoline has potential to prevent against basal cell carcinoma tumorigenesis. ► It has more effectiveness on BCC as compared with a human keratinocyte cell line. ► Mechanisms involved including reducing tumor cells’ survival factor IL-6, ► Decreasing Cdc25C phosphatase, enhancing tumor suppressor factor p53,

  11. Inflammatory effects induced by selected limonene oxidation products: 4-OPA, IPOH, 4-AMCH in human bronchial (16HBE14o-) and alveolar (A549) epithelial cell lines.

    PubMed

    Lipsa, Dorelia; Leva, Paolo; Barrero-Moreno, Josefa; Coelhan, Mehmet

    2016-11-16

    Limonene, a monoterpene abundantly present in most of the consumer products (due to its pleasant citrus smell), easily undergoes ozonolysis leading to several limonene oxidation products (LOPs) such as 4-acetyl-1-methylcyclohexene (4-AMCH), 4-oxopentanal (4-OPA) and 3-isopropenyl-6-oxoheptanal (IPOH). Toxicological studies have indicated that human exposure to limonene and ozone can cause adverse airway effects. However, little attention has been paid to the potential health impact of specific LOPs, in particular of IPOH, 4-OPA and 4-AMCH. This study evaluates the cytotoxic effects of the selected LOPs on human bronchial epithelial (16HBE14o-) and alveolar epithelial (A549) cell lines by generating concentration-response curves using the neutral red uptake assay and analyzing the inflammatory response with a series of cytokines/chemokines. The cellular viability was mostly reduced by 4-OPA [IC 50 =1.6mM (A549) and 1.45mM (16HBE14o-)] when compared to IPOH [IC 50 =3.5mM (A549) and 3.4mM (16HBE14o-)] and 4-AMCH [IC 50 could not be calculated]. As a result from the inflammatory response, IPOH [50μM] induced an increase of both IL-6 and IL-8 secretion in A549 (1.5-fold change) and in 16HBE14o- (2.8- and 7-fold change respectively). 4-OPA [50μM] treatment of A549 increased IL-6 (1.4-times) and IL-8 (1.3-times) levels, while in 16HBE14o- had an opposite effect. A549 treated with 4-AMCH [50μM] elevate both IL-6 and IL-8 levels by 1.2-times, while in 16HBE14o- had an opposite effect. Based on our results, lung cellular injury characterized by inflammatory cytokine release was observed for both cell lines treated with the selected chemicals at concentrations that did not affect their cellular viability. Copyright © 2016 The Authors. Published by Elsevier Ireland Ltd.. All rights reserved.

  12. GNB2 is a mediator of lidocaine-induced apoptosis in rat pheochromocytoma PC12 cells.

    PubMed

    Tan, Yonghong; Wang, Qiong; Zhao, Baisong; She, Yingjun; Bi, Xiaobao

    2016-05-01

    Lidocaine has been recognized to induce neurotoxicity. However, the molecular mechanism underlying this effect, especially the critical molecules in cells that mediated the lidocaine-induced apoptosis were unclear. In the present study, PC12 cells were administrated with lidocaine for 48h. Using MTT assay and flow cytometry, we found lidocaine significantly decreased the cell proliferation and S phases in PC12 cells with treatment concentrations, and significantly enhanced cell apoptosis with treatment concentrations. Two-dimensional gel electrophoresis (2-DE) analysis and LC-MS/MS were used to identification of protein biomarkers. Six proteins were identified. Among them, three were up-expressed including ANXA6, GNB2 and STMN1, other three were down-expressed including ubiquitin-linke protein 7 (UBL7), DDAH2 and BLVRB. Using qRT-PCR, we confirmed that lidocaine up-regulated the mRNA expression of STMN1, GNB2, ANXA6 and DDAH2, and found that the GNB2 had the largest change (about increased by 6.4 folds). The up-regulation of GNB2 by lidocaine was also validated by western blot. After transfected with 100μM GNB2-Rat-453 siRNA, the expression of GNB2 in PC12 cells was almost completely inhibited; and the cell proliferation and cells in S phases were significantly enhanced, cell apoptosis including both early apoptosis and later apoptosis were significantly reduced in the presence of 0.5mM lidocaine for 48h. Therefore, neuronal apoptosis was induced by lidocaine and this effect was mediated by GNB2. Further research is needed to assess the clinical relevance and exact mechanism of neuronal apoptosis caused by lidocaine. Copyright © 2016 Elsevier Inc. All rights reserved.

  13. Development of apoptosis-resistant dihydrofolate reductase-deficient Chinese hamster ovary cell line.

    PubMed

    Lee, Suk Kyoo; Lee, Gyun Min

    2003-06-30

    Apoptosis-resistant dihydrofolate reductase-deficient CHO cell line (dhfr(-) CHO-bcl2) was developed by introduction of the bcl-2 gene into the dhfr(-) CHO cell line (DUKX-B11, ATCC CRL-9096) and subsequent selection of clones stably overexpressing Bcl-2 in the absence of selection pressure. When the dhfr(-) CHO-bcl2 cell line was used as a host cell line for development of a recombinant CHO (rCHO) cell line expressing a humanized antibody, it displayed stable expression of the bcl-2 gene during rCHO cell line development and no detrimental effect of Bcl-2 overexpression on specific antibody productivity. Taken together, the results obtained demonstrate that the use of an apoptosis-resistant dhfr(-) CHO cell line as the host cell line saves the effort of establishing an apoptosis-resistant rCHO cell line and expedites the development process of apoptosis-resistant rCHO cells producing therapeutic proteins. Copyright 2003 Wiley Periodicals, Inc. Biotechnol Bioeng 82: 872-876, 2003.

  14. Selective induction of apoptosis in human gastric cancer cells by Lactobacillus kefiri (PFT), a novel kefir product

    PubMed Central

    GHONEUM, MAMDOOH; FELO, NOURAN

    2015-01-01

    The present study was undertaken to evaluate the effect of Lactobacillus kefiri (PFT), a novel kefir product, on apoptosis of gastric cancer cells (AGS), breast cancer cells (4T1), and human peripheral blood mononuclear cells (PBMCs). Cells were cultured with PFT and apoptosis was determined by flow cytometry using 7-AAD dye and cytospin preparation. Mitochondrial dysfunction and expression of Bcl2 were monitored by flow cytometry. Results showed that PFT induced apoptosis in AGS gastric cancer cells in a dose-dependent manner. Apoptosis was detected at a concentration of 0.3 mg/ml (20.8%), increased to 25.8% at 0.6 mg/ml, 37% at 1.2 mg/ml, 53.1% at 2.5 mg/ml, and peaked at 66.3% at 5.0 mg/ml. Apoptosis is associated with the decreased polarization of mitochondrial membrane potential (MMP) and decreased Bcl2 expression. PFT-treated AGS cells manifested membrane blebbing, nuclear condensation, and fragmentation as identified in cytospin cytocentrifuge Giemsa stained preparations. On the other hand, flow cytometry analysis showed that PFT did not induce apoptosis in 4T1 breast cancer cells nor in PBMCs. These results suggest that PFT is safe for white blood cells and selectively induces apoptotic effects in gastric cancer cells. Hence, it may have potential as a therapeutic agent for the treatment of gastric cancers. PMID:26251956

  15. Selective induction of apoptosis in human gastric cancer cells by Lactobacillus kefiri (PFT), a novel kefir product.

    PubMed

    Ghoneum, Mamdooh; Felo, Nouran

    2015-10-01

    The present study was undertaken to evaluate the effect of Lactobacillus kefiri (PFT), a novel kefir product, on apoptosis of gastric cancer cells (AGS), breast cancer cells (4T1), and human peripheral blood mononuclear cells (PBMCs). Cells were cultured with PFT and apoptosis was determined by flow cytometry using 7-AAD dye and cytospin preparation. Mitochondrial dysfunction and expression of Bcl2 were monitored by flow cytometry. Results showed that PFT induced apoptosis in AGS gastric cancer cells in a dose-dependent manner. Apoptosis was detected at a concentration of 0.3 mg/ml (20.8%), increased to 25.8% at 0.6 mg/ml, 37% at 1.2 mg/ml, 53.1% at 2.5 mg/ml, and peaked at 66.3% at 5.0 mg/ml. Apoptosis is associated with the decreased polarization of mitochondrial membrane potential (MMP) and decreased Bcl2 expression. PFT-treated AGS cells manifested membrane blebbing, nuclear condensation, and fragmentation as identified in cytospin cytocentrifuge Giemsa stained preparations. On the other hand, flow cytometry analysis showed that PFT did not induce apoptosis in 4T1 breast cancer cells nor in PBMCs. These results suggest that PFT is safe for white blood cells and selectively induces apoptotic effects in gastric cancer cells. Hence, it may have potential as a therapeutic agent for the treatment of gastric cancers.

  16. IFN-gamma Impairs Release of IL-8 by IL-1beta-stimulated A549 Lung Carcinoma Cells

    PubMed Central

    Boost, Kim A; Sadik, Christian D; Bachmann, Malte; Zwissler, Bernhard; Pfeilschifter, Josef; Mühl, Heiko

    2008-01-01

    Background Production of interferon (IFN)-γ is key to efficient anti-tumor immunity. The present study was set out to investigate effects of IFNγ on the release of the potent pro-angiogenic mediator IL-8 by human A549 lung carcinoma cells. Methods A549 cells were cultured and stimulated with interleukin (IL)-1β alone or in combination with IFNγ. IL-8 production by these cells was analyzed with enzyme linked immuno sorbent assay (ELISA). mRNA-expression was analyzed by real-time PCR and RNase protection assay (RPA), respectively. Expression of inhibitor-κ Bα, cellular IL-8, and cyclooxygenase-2 was analyzed by Western blot analysis. Results Here we demonstrate that IFNγ efficiently reduced IL-8 secretion under the influence of IL-1β. Surprisingly, real-time PCR analysis and RPA revealed that the inhibitory effect of IFNγ on IL-8 was not associated with significant changes in mRNA levels. These observations concurred with lack of a modulatory activity of IFNγ on IL-1β-induced NF-κB activation as assessed by cellular IκB levels. Moreover, analysis of intracellular IL-8 suggests that IFNγ modulated IL-8 secretion by action on the posttranslational level. In contrast to IL-8, IL-1β-induced cyclooxygenase-2 expression and release of IL-6 were not affected by IFNγ indicating that modulation of IL-1β action by this cytokine displays specificity. Conclusion Data presented herein agree with an angiostatic role of IFNγ as seen in rodent models of solid tumors and suggest that increasing T helper type 1 (Th1)-like functions in lung cancer patients e.g. by local delivery of IFNγ may mediate therapeutic benefit via mechanisms that potentially include modulation of pro-angiogenic IL-8. PMID:18801189

  17. Upregulation of miR-98 Inhibits Apoptosis in Cartilage Cells in Osteoarthritis.

    PubMed

    Wang, Gui-Long; Wu, Yu-Bo; Liu, Jia-Tian; Li, Cui-Yun

    2016-11-01

    We aimed to investigate the effects of microRNA-98 (miR-98) on apoptosis in cartilage cells of osteoarthritis (OA) patients. Knee cartilage tissue samples were collected from 31 OA patients, 21 autopsies, and 26 amputation patients due to trauma. The clinicopathological data were recorded. Quantitative real-time polymerase chain reaction was performed to compare the miR-98 expression levels from cartilage cells obtained from the OA and non-OA patients. Clinicopathological characteristics of the patients were also analyzed. Primary chondrocytes were separated from cartilage tissues and transfected with plasmids or siRNA to overexpress or inhibit miR-98. Annexin V-PI double staining and TUNEL assays were used to examine apoptosis in the primary chondrocytes after transfection. Finally, a rat OA model was used to confirm the effects of miR-98 on apoptosis in cartilage cells in vivo. Compared with the normal cartilage tissues, miR-98 expression was reduced in the OA cartilage tissues (p < 0.01). The miR-98 expression levels were also significantly correlated with the OA stage (p < 0.05). In vitro, transfection with the miR-98 inhibitor increased apoptosis in the cartilage cells (p < 0.05), and transfection with a miR-98 mimic inhibited apoptosis in cartilage cells (p < 0.05). In the OA rat model, exogenous injection of the miR-98 mimic inhibited apoptosis in the rat cartilage cells thus alleviating OA. MiR-98 expression is reduced in the cartilage cells of OA patients and the overexpression of miR-98 inhibits cartilage cell apoptosis, while inhibition of microRNA-98 leads to cartilage cell apoptosis. These findings provide a theoretical basis for the development of novel targeted therapies for OA.

  18. IL-3 induces apoptosis in a ras-transformed myeloid cell line.

    PubMed

    Ahmed, N; Anderson, S M; Berridge, M V

    1999-04-01

    Growth factors promote cell survival and proliferation. Homeostasis is maintained by programmed cell death which occurs when the growth stimulus is withdrawn, in response to negative growth regulators such as interferons, TNF-alpha and CD95 ligand, or following differentiation. Although acutely-transforming oncogenes often overcome the need for growth factors, growth regulatory cytokines can influence proliferative responses of transformed cells. In this study we investigated the effects of IL-3 on the proliferative responses of parental bone marrow-derived 32D cells and cells transformed with ras and abl oncogenes. We show that treatment of ras-transformed 32D cells with IL-3 reduced proliferative responses and decreased colony-forming ability. These effects were exacerbated in the absence of serum and associated with inhibition of tyrosine kinase activity, down-regulation of RAS and MYC expression, and induction of apoptosis as indicated by DNA fragmentation. In contrast, treatment of parental 32D cells with IL-3, which is obligatory for cell survival and proliferation, increased tyrosine kinase activity, upregulated MYC and RAS expression and maintained DNA integrity. With abl-transformed cells, proliferation and colony-forming ability were also inhibited by IL-3. Tyrosine kinase activity and MYC expression were reduced, but early apoptosis was not evident. Calcium uptake however, was stimulated by IL-3 in both parental and oncogene-transformed cells. These results suggest that threshold levels of tyrosine kinase activity are necessary for cell survival and proliferation and that with ras-transformed cells, IL-3 treatment may result in this threshold being breached. We conclude that in some situations, growth-promoting cytokines can inhibit proliferation of transformed cells and induce cell death by apoptosis.

  19. The Marine Fungal Metabolite, Dicitrinone B, Induces A375 Cell Apoptosis through the ROS-Related Caspase Pathway

    PubMed Central

    Chen, Li; Gong, Mei-Wei; Peng, Zhen-Fei; Zhou, Tong; Ying, Min-Gang; Zheng, Qiu-Hong; Liu, Qin-Ying; Zhang, Qi-Qing

    2014-01-01

    Dicitrinone B, a rare carbon-bridged citrinin dimer, was isolated from the marine-derived fungus, Penicillium citrinum. It was reported to have antitumor effects on tumor cells previously; however, the details of the mechanism remain unclear. In this study, we found that dicitrinone B inhibited the proliferation of multiple tumor types. Among them, the human malignant melanoma cell, A375, was confirmed to be the most sensitive. Morphologic evaluation, cell cycle arrest and apoptosis rate analysis results showed that dicitrinone B significantly induced A375 cell apoptosis. Subsequent observation of reactive oxygen species (ROS) accumulation and mitochondrial membrane potential (MMP) reduction revealed that the apoptosis induced by dicitrinone B may be triggered by over-producing ROS. Further studies indicated that the apoptosis was associated with both intrinsic and extrinsic apoptosis pathways under the regulation of Bcl-2 family proteins. Caspase-9, caspase-8 and caspase-3 were activated during the process, leading to PARP cleavage. The pan-caspase inhibitor, Z-VAD-FMK, could reverse dicitrinone B-induced apoptosis, suggesting that it is a caspase-dependent pathway. Our data for the first time showed that dicitrinone B inhibits the proliferation of tumor cells by inducing cell apoptosis. Moreover, compared with the first-line chemotherapy drug, 5-fluorouracil (5-Fu), dicitrinone B showed much more potent anticancer efficacy, suggesting that it might serve as a potential antitumor agent. PMID:24699111

  20. High glucose induces apoptosis via upregulation of Bim expression in proximal tubule epithelial cells.

    PubMed

    Zhang, Xiao-Qian; Dong, Jian-Jun; Cai, Tian; Shen, Xue; Zhou, Xiao-Jun; Liao, Lin

    2017-04-11

    Diabetic nephropathy is the primary cause of end-stage renal disease. Apoptosis of tubule epithelial cells is a major feature of diabetic nephropathy. The mechanisms of high glucose (HG) induced apoptosis are not fully understood. Here we demonstrated that, HG induced apoptosis via upregulating the expression of proapoptotic Bcl-2 homology domain 3 (BH3)-only protein Bim protein, but not bring a significant change in the baseline level of autophagy in HK2 cells. The increase of Bim expression was caused by the ugregulation of transcription factors, FOXO1 and FOXO3a. Bim expression initiates BAX/BAK-mediated mitochondria-dependent apoptosis. Silence of Bim by siRNA in HK2 cells prevented HG-induced apoptosis and also sensitized HK2 cells to autophagy during HG treatment. The autophagy inhibitor 3-MA increased the injury in Bim knockdown HK2 cells by retriggering apoptosis. The above results suggest a Bim-independent apoptosis pathway in HK2 cells, which normally could be inhibited by autophagy. Overall, our results indicate that HG induces apoptosis via up-regulation of Bim expression in proximal tubule epithelial cells.

  1. Tumor Response to Radiotherapy Regulated by Endothelial Cell Apoptosis

    NASA Astrophysics Data System (ADS)

    Garcia-Barros, Monica; Paris, Francois; Cordon-Cardo, Carlos; Lyden, David; Rafii, Shahin; Haimovitz-Friedman, Adriana; Fuks, Zvi; Kolesnick, Richard

    2003-05-01

    About 50% of cancer patients receive radiation therapy. Here we investigated the hypothesis that tumor response to radiation is determined not only by tumor cell phenotype but also by microvascular sensitivity. MCA/129 fibrosarcomas and B16F1 melanomas grown in apoptosis-resistant acid sphingomyelinase (asmase)-deficient or Bax-deficient mice displayed markedly reduced baseline microvascular endothelial apoptosis and grew 200 to 400% faster than tumors on wild-type microvasculature. Thus, endothelial apoptosis is a homeostatic factor regulating angiogenesis-dependent tumor growth. Moreover, these tumors exhibited reduced endothelial apoptosis upon irradiation and, unlike tumors in wild-type mice, they were resistant to single-dose radiation up to 20 grays (Gy). These studies indicate that microvascular damage regulates tumor cell response to radiation at the clinically relevant dose range.

  2. Protective role of Osthole on myocardial cell apoptosis induced by doxorubicin in rats.

    PubMed

    Xu, Hongdang; Han, Yu; Zhang, Mengwei; Yan, Min; Gao, Chuanyu

    2015-01-01

    To explore the effect of Osthole on protecting myocardial cell apoptosis induced by doxorubicin during cardiac failure in rats. Myocardial cells isolated from the newborn SD rats were separated into three groups: cells treated with 1 μmol doxorubicin, cells treated with Osthole at three concentrations of 10, 20, and 40 μmol, cells treated neither with Osthole nor with doxorubicin were the control groups. Consequently, cell apoptosis of myocardial cells in each group was analyzed using TUNEL assay. Also, expressions of oxidase, NADPH, and ROS in myocardial cells were analyzed using different biological methods. Moreover, expressions of cell apoptosis associated proteins were analyzed using Western blotting. Compared with the controls, the results showed that cells received Osthole and doxorubicin treatments performed high percentage of cell apoptosis, suggesting that Osthole could anesis myocardial cell apoptosis induced by doxorubicin (P<0.05). Osthole of 10 μmol depressed the expressions of cell apoptosis associated proteins including Caspase-3 and Cytc, and enhancing expression of Bcl-XL expression (P<0.05). Osthole of 20 μmol significantly decreased the generation of intracellar superoxidase, NADPH, and NADPH activity in myocardial cells treated with doxorubicin (P<0.05). Moreover, Osthole of 20 μmol could significantly increase phosphorylated elF2α level in cells. Our study suggested that Osthole may play a protective role in suppressing myocardial apoptosis induced by doxorubicin through inhibiting NADPH and superoxidase production and downstream phosphorylated elF2α.

  3. Protective role of Osthole on myocardial cell apoptosis induced by doxorubicin in rats

    PubMed Central

    Xu, Hongdang; Han, Yu; Zhang, Mengwei; Yan, Min; Gao, Chuanyu

    2015-01-01

    Objective: To explore the effect of Osthole on protecting myocardial cell apoptosis induced by doxorubicin during cardiac failure in rats. Methods: Myocardial cells isolated from the newborn SD rats were separated into three groups: cells treated with 1 μmol doxorubicin, cells treated with Osthole at three concentrations of 10, 20, and 40 μmol, cells treated neither with Osthole nor with doxorubicin were the control groups. Consequently, cell apoptosis of myocardial cells in each group was analyzed using TUNEL assay. Also, expressions of oxidase, NADPH, and ROS in myocardial cells were analyzed using different biological methods. Moreover, expressions of cell apoptosis associated proteins were analyzed using Western blotting. Results: Compared with the controls, the results showed that cells received Osthole and doxorubicin treatments performed high percentage of cell apoptosis, suggesting that Osthole could anesis myocardial cell apoptosis induced by doxorubicin (P<0.05). Osthole of 10 μmol depressed the expressions of cell apoptosis associated proteins including Caspase-3 and Cytc, and enhancing expression of Bcl-XL expression (P<0.05). Osthole of 20 μmol significantly decreased the generation of intracellar superoxidase, NADPH, and NADPH activity in myocardial cells treated with doxorubicin (P<0.05). Moreover, Osthole of 20 μmol could significantly increase phosphorylated elF2α level in cells. Conclusion: Our study suggested that Osthole may play a protective role in suppressing myocardial apoptosis induced by doxorubicin through inhibiting NADPH and superoxidase production and downstream phosphorylated elF2α. PMID:26617794

  4. P53 protein in proliferation, repair and apoptosis of cells.

    PubMed

    Wawryk-Gawda, Ewelina; Chylińska-Wrzos, Patrycja; Lis-Sochocka, Marta; Chłapek, Katarzyna; Bulak, Kamila; Jędrych, Marian; Jodłowska-Jędrych, Barbara

    2014-05-01

    The p53 protein is an important factor of many intra- and extracellular processes. This protein regulates the repair of cellular DNA and induces apoptosis. It is also responsible for the regulation of the senescence and the cell entering the subsequent stages of the cellular cycle. The protein p53 is also involved in inhibiting angiogenesis and the induction of oxidative shock. In our study, we examined the activity of p53 protein in the uterine epithelial cells in rats treated with cladribine. Its action is mainly based on apoptosis induction. We compared the activity of p53 protein in cells with a high apoptosis index and in cells with active repair mechanisms and high proliferation index. We observed stronger p53 protein expression in the epithelial cells of the materials taken 24 h after the last dose of 2-CdA associated with the active process of apoptosis and inhibition of proliferation. After 4 weeks from the last dose of cladribine, the stronger expression of p53 protein was associated with both the existing changes in the cell's genome, the effects of the ongoing repair mechanisms, as well as the high proliferation activity.

  5. Aggregation of lipid rafts activates c-met and c-Src in non-small cell lung cancer cells.

    PubMed

    Zeng, Juan; Zhang, Heying; Tan, Yonggang; Sun, Cheng; Liang, Yusi; Yu, Jinyang; Zou, Huawei

    2018-05-30

    Activation of c-Met, a receptor tyrosine kinase, induces radiation therapy resistance in non-small cell lung cancer (NSCLC). The activated residual of c-Met is located in lipid rafts (Duhon et al. Mol Carcinog 49:739-49, 2010). Therefore, we hypothesized that disturbing the integrity of lipid rafts would restrain the activation of the c-Met protein and reverse radiation resistance in NSCLC. In this study, a series of experiments was performed to test this hypothesis. NSCLC A549 and H1993 cells were incubated with methyl-β-cyclodextrin (MβCD), a lipid raft inhibitor, at different concentrations for 1 h before the cells were X-ray irradiated. The following methods were used: clonogenic (colony-forming) survival assays, flow cytometry (for cell cycle and apoptosis analyses), immunofluorescence microscopy (to show the distribution of proteins in lipid rafts), Western blotting, and biochemical lipid raft isolation (purifying lipid rafts to show the distribution of proteins in lipid rafts). Our results showed that X-ray irradiation induced the aggregation of lipid rafts in A549 cells, activated c-Met and c-Src, and induced c-Met and c-Src clustering to lipid rafts. More importantly, MβCD suppressed the proliferation of A549 and H1993 cells, and the combination of MβCD and radiation resulted in additive increases in A549 and H1993 cell apoptosis. Destroying the integrity of lipid rafts inhibited the aggregation of c-Met and c-Src to lipid rafts and reduced the expression of phosphorylated c-Met and phosphorylated c-Src in lipid rafts. X-ray irradiation induced the aggregation of lipid rafts and the clustering of c-Met and c-Src to lipid rafts through both lipid raft-dependent and lipid raft-independent mechanisms. The lipid raft-dependent activation of c-Met and its downstream pathways played an important role in the development of radiation resistance in NSCLC cells mediated by c-Met. Further studies are still required to explore the molecular mechanisms of the

  6. Mitochondria-derived reactive oxygen species drive GANT61-induced mesothelioma cell apoptosis.

    PubMed

    Lim, Chuan Bian; Prêle, Cecilia M; Baltic, Svetlana; Arthur, Peter G; Creaney, Jenette; Watkins, D Neil; Thompson, Philip J; Mutsaers, Steven E

    2015-01-30

    Gli transcription factors of the Hedgehog (Hh) pathway have been reported to be drivers of malignant mesothelioma (MMe) cell survival. The Gli inhibitor GANT61 induces apoptosis in various cancer cell models, and has been associated directly with Gli inhibition. However various chemotherapeutics can induce cell death through generation of reactive oxygen species (ROS) but whether ROS mediates GANT61-induced apoptosis is unknown. In this study human MMe cells were treated with GANT61 and the mechanisms regulating cell death investigated. Exposure of MMe cells to GANT61 led to G1 phase arrest and apoptosis, which involved ROS but not its purported targets, GLI1 or GLI2. GANT61 triggered ROS generation and quenching of ROS protected MMe cells from GANT61-induced apoptosis. Furthermore, we demonstrated that mitochondria are important in mediating GANT61 effects: (1) ROS production and apoptosis were blocked by mitochondrial inhibitor rotenone; (2) GANT61 promoted superoxide formation in mitochondria; and (3) mitochondrial DNA-deficient LO68 cells failed to induce superoxide, and were more resistant to apoptosis induced by GANT61 than wild-type cells. Our data demonstrate for the first time that GANT61 induces apoptosis by promoting mitochondrial superoxide generation independent of Gli inhibition, and highlights the therapeutic potential of mitochondrial ROS-mediated anticancer drugs in MMe.

  7. 28 CFR 549.71 - Inmates affected.

    Code of Federal Regulations, 2011 CFR

    2011-07-01

    ... 28 Judicial Administration 2 2011-07-01 2011-07-01 false Inmates affected. 549.71 Section 549.71 Judicial Administration BUREAU OF PRISONS, DEPARTMENT OF JUSTICE INSTITUTIONAL MANAGEMENT MEDICAL SERVICES Fees for Health Care Services § 549.71 Inmates affected. This subpart applies to: (a) Any individual...

  8. 28 CFR 549.71 - Inmates affected.

    Code of Federal Regulations, 2014 CFR

    2014-07-01

    ... 28 Judicial Administration 2 2014-07-01 2014-07-01 false Inmates affected. 549.71 Section 549.71 Judicial Administration BUREAU OF PRISONS, DEPARTMENT OF JUSTICE INSTITUTIONAL MANAGEMENT MEDICAL SERVICES Fees for Health Care Services § 549.71 Inmates affected. This subpart applies to: (a) Any individual...

  9. 28 CFR 549.71 - Inmates affected.

    Code of Federal Regulations, 2012 CFR

    2012-07-01

    ... 28 Judicial Administration 2 2012-07-01 2012-07-01 false Inmates affected. 549.71 Section 549.71 Judicial Administration BUREAU OF PRISONS, DEPARTMENT OF JUSTICE INSTITUTIONAL MANAGEMENT MEDICAL SERVICES Fees for Health Care Services § 549.71 Inmates affected. This subpart applies to: (a) Any individual...

  10. 28 CFR 549.71 - Inmates affected.

    Code of Federal Regulations, 2010 CFR

    2010-07-01

    ... 28 Judicial Administration 2 2010-07-01 2010-07-01 false Inmates affected. 549.71 Section 549.71 Judicial Administration BUREAU OF PRISONS, DEPARTMENT OF JUSTICE INSTITUTIONAL MANAGEMENT MEDICAL SERVICES Fees for Health Care Services § 549.71 Inmates affected. This subpart applies to: (a) Any individual...

  11. 28 CFR 549.71 - Inmates affected.

    Code of Federal Regulations, 2013 CFR

    2013-07-01

    ... 28 Judicial Administration 2 2013-07-01 2013-07-01 false Inmates affected. 549.71 Section 549.71 Judicial Administration BUREAU OF PRISONS, DEPARTMENT OF JUSTICE INSTITUTIONAL MANAGEMENT MEDICAL SERVICES Fees for Health Care Services § 549.71 Inmates affected. This subpart applies to: (a) Any individual...

  12. Nano Copper Induces Apoptosis in PK-15 Cells via a Mitochondria-Mediated Pathway.

    PubMed

    Zhang, Hui; Chang, Zhenyu; Mehmood, Khalid; Abbas, Rao Zahid; Nabi, Fazul; Rehman, Mujeeb Ur; Wu, Xiaoxing; Tian, Xinxin; Yuan, Xiaodan; Li, Zhaoyang; Zhou, Donghai

    2018-01-01

    Nano-sized copper particles are widely used in various chemical, physical, and biological fields. However, earlier studies have shown that nano copper particles (40-100 μg/mL) can induce cell toxicity and apoptosis. Therefore, this study was conducted to investigate the role of nano copper in mitochondrion-mediated apoptosis in PK-15 cells. The cells were treated with different doses of nano copper (20, 40, 60, and 80 μg/mL) to determine the effects of apoptosis using acridine orange/ethidium bromide (AO/EB) fluorescence staining and a flow cytometry assay. The levels of malondialdehyde (MDA) and superoxide dismutase (SOD) in the PK-15 cells were examined using commercially available kits. Moreover, the mRNA levels of the Bax, Bid, Caspase-3, and CYCS genes were assessed by real-time PCR. The results revealed that nano copper exposure induced apoptosis and changed the mitochondrial membrane potential. In addition, nano copper significantly altered the levels of the Bax, Bid, Caspase-3, and CYCS genes at a concentration of 40 μg/mL. To summarize, nano copper significantly (P < 0.05) decreased the level of SOD and increased the level of MDA in PK-15 cells. Altogether, these results suggest that nano copper can play an important role in inducing the apoptotic pathway in PK-15 cells, which may be the mechanism by which nano copper induces nephrotoxicity.

  13. A Novel Bcl-x Isoform Connected to the T Cell Receptor Regulates Apoptosis in T Cells

    PubMed Central

    Yang, Xiao-Feng; Weber, Georg F.

    2014-01-01

    Summary We define a novel Bcl-x isoform, Bcl-xγ, that is generated by alternative splicing and characterized by a unique 47 amino acid C-terminus. Bcl-xγ is expressed primarily in thymocytes, where it may depend on an interaction between the TCR and host MHC products, and in mature T cells, where its expression is associated with ligation of the T cell receptor. Overexpression of Bcl-xγ in T cells inhibits activation-induced apoptosis; inhibition of Bcl-xγ, after stable expression of Bcl-xγ antisense cDNA, enhances activation-induced apoptosis. In contrast to other Bcl-x isoforms, cells that fail to express Bcl-xγ after CD3 ligation undergo programmed cell death, while activated T cells that express Bcl-xγ are spared. Identification of Bcl-xγ helps provide amolecular explanation of T cell activation and death after antigen engagement. PMID:9390687

  14. The impact of anticancer activity upon Beta vulgaris extract mediated biosynthesized silver nanoparticles (ag-NPs) against human breast (MCF-7), lung (A549) and pharynx (Hep-2) cancer cell lines.

    PubMed

    Venugopal, K; Ahmad, H; Manikandan, E; Thanigai Arul, K; Kavitha, K; Moodley, M K; Rajagopal, K; Balabhaskar, R; Bhaskar, M

    2017-08-01

    The present study tried for a phyto-synthetic method of producing silver nanoparticles (Ag-NPs) with size controlled as and eco-friendly route that can lead to their advanced production with decorative tranquil morphology. By inducing temperature fluctuation of the reaction mixture from 25 to 80°C the plasmon resonance band raised slowly which had an ultimate effect on size and shape of Ag-NPs as shown by UV-visible spectroscopy and TEM results. The biosynthesized nanoparticles showed good cytotoxic impact against MCF-7, A549 and Hep2 cells compared to normal cell lines. Compared to control plates, the percentage of cell growth inhibition was found to be high with as concentrations of Ag-NPs becomes more as determined by MTT assay. The AO/EtBr staining observations demonstrated that the mechanism of cell death induced by Ag-NPs was due to apoptosis in cancer cells. These present results propose that the silver nanoparticles (Ag-NPs) may be utilized as anticancer agents for the treatment of various cancer types. However, there is a need for study of in vivo examination of these nanoparticles to find their role and mechanism inside human body. Further, studies we plan to do biomarker fabrication from the green synthesized plant extract nanoparticles like silver, gold and copper nanoparticles with optimized shape and sizes and their enhancement of these noble nanoparticles. Copyright © 2017. Published by Elsevier B.V.

  15. Raman spectrum reveals the cell cycle arrest of Triptolide-induced leukemic T-lymphocytes apoptosis

    NASA Astrophysics Data System (ADS)

    Zhang, Daosen; Feng, Yanyan; Zhang, Qinnan; Su, Xin; Lu, Xiaoxu; Liu, Shengde; Zhong, Liyun

    2015-04-01

    Triptolide (TPL), a traditional Chinese medicine extract, possesses anti-inflammatory and anti-tumor properties. Though some research results have implicated that Triptolide (TPL) can be utilized in the treatment of leukemia, it remains controversial about the mechanism of TPL-induced leukemic T-lymphocytes apoptosis. In this study, combining Raman spectroscopic data, principal component analysis (PCA) and atomic force microscopy (AFM) imaging, both the biochemical changes and morphological changes during TPL-induced cell apoptosis were presented. In contrast, the corresponding data during Daunorubicin (DNR)-induced cell apoptosis was also exhibited. The obtained results showed that Raman spectral changes during TPL-induced cell apoptosis were greatly different from DNR-induced cell apoptosis in the early stage of apoptosis but revealed the high similarity in the late stage of apoptosis. Moreover, above Raman spectral changes were respectively consistent with the morphological changes of different stages during TPL-induced apoptosis or DNR-induced apoptosis, including membrane shrinkage and blebbing, chromatin condensation and the formation of apoptotic bodies. Importantly, it was found that Raman spectral changes with TPL-induced apoptosis or DNR-induced apoptosis were respectively related with the cell cycle G1 phase arrest or G1 and S phase arrest.

  16. Apoptosis and Vocal Fold Disease: Clinically Relevant Implications of Epithelial Cell Death

    ERIC Educational Resources Information Center

    Novaleski, Carolyn K.; Carter, Bruce D.; Sivasankar, M. Preeti; Ridner, Sheila H.; Dietrich, Mary S.; Rousseau, Bernard

    2017-01-01

    Purpose: Vocal fold diseases affecting the epithelium have a detrimental impact on vocal function. This review article provides an overview of apoptosis, the most commonly studied type of programmed cell death. Because apoptosis can damage epithelial cells, this article examines the implications of apoptosis on diseases affecting the vocal fold…

  17. Apoptosis and accidental cell death in cultured human keratinocytes after thermal injury.

    PubMed

    Matylevitch, N P; Schuschereba, S T; Mata, J R; Gilligan, G R; Lawlor, D F; Goodwin, C W; Bowman, P D

    1998-08-01

    The respective roles of apoptosis and accidental cell death after thermal injury were evaluated in normal human epidermal keratinocytes. By coupling the LIVE/DEAD fluorescence viability assay with the terminal deoxynucleotidyl transferase-mediated dUTP nick end-labeling (TUNEL) method and ultrastructural morphology, these two processes could be distinguished. Cells were grown on glass coverslips with a microgrid pattern so that the results of several staining procedures performed sequentially could be visualized in the same cells after heating at temperatures of up to 72 degrees C for 1 second. After exposure to temperatures of 58 to 59 degrees C, cells died predominantly by apoptosis; viable cells became TUNEL positive, indicating degradation of DNA. After exposure to temperatures of 60 to 66 degrees C, both TUNEL-positive viable cells and TUNEL-positive nonviable cells were observed, indicating that apoptosis and accidental cell death were occurring simultaneously. Cells died almost immediately after exposure to temperatures above 72 degrees C, presumably from heat fixation. The fluorescent mitochondrial probe MitoTracker Orange indicated that cells undergoing apoptosis became TUNEL positive before loss of mitochondrial function. Nucleosomal fragmentation of DNA analyzed by enzyme-linked immunosorbent assay and gel electrophoresis occurred after exposure to temperatures of 58 to 59 degrees C. The characteristic morphological findings of cells undergoing apoptosis, by transmission electron microscopy, included cellular shrinkage, cytoplasmic budding, and relatively intact mitochondria. Depending on temperature and time of exposure, normal human epidermal keratinocytes may die by apoptosis, accidental cell death, or heat fixation.

  18. Chrysin and its phosphate ester inhibit cell proliferation and induce apoptosis in Hela cells.

    PubMed

    Zhang, Ting; Chen, Xiaolan; Qu, Lingbo; Wu, Jinglan; Cui, Ran; Zhao, Yufen

    2004-12-01

    To improve the biological activities of chrysin (CR), we synthesize Diethyl Chysin-7-yl phosphate (CPE: C(19)H(19)O(7)P) and tetraethyl bis-phosphoric ester of chrysin (CP: C(23)H(28)O(10)P(2)) through a simplified Atheron-Todd reaction. The interactions of the CR and CPE with lysozyme were explored by electrospray ionization mass spectrometry (ESI) and fluorescence spectrometry method. Experimental results indicate that CPE could form the noncovalent compound with lysozyme, while the interaction of the CR with lysozyme was not detected. In addition, whether and how the compounds CPE and CP affect proliferation and apoptosis in human cervical cancer Hela cells were investigated. Moreover, the effects of CPE and CP in Hela cells were compared with that of the nonmodified CR compound. The Hela cells were co-cultured with CR, CP, and CPE as experimental groups, respectively, and corresponding control groups treated without CR, CP, and CPE. The proliferation and apoptosis were detected using MTT assay, HCl denatured-methyl green-pyronin staining, PCNA immunohistochemistry and TUNEL techniques. The cell growth IC(50), relative absorbance (RA), proliferating index (PI), PCNA-IR (immunoreactivity IR) integration value (IV), and apoptosis index (AI) were calculated and their correlation was analyzed in each group. The results show that all CR, CP, and CPE could inhibit proliferation and induce apoptosis in Hela cells. Moreover, the effects of CP and CPE were more potent than that of CR. The CP and CPE were proved to be a kind of stronger apoptosis inducers than nonphosphated CR. There was a negative correlation between proliferation and apoptosis. In conclusion, the CR, CP, and CPE could effectively inhibit growth by down-regulated expression of PCNA, and induce apoptosis in Hela cells. The efficiency of the modified CP and CPE preceded nonmodified CR compounds. The CP and CPE may be a new potential anti-cancer drug for therapy of human cervical carcinoma.

  19. Sensitization of interferon-γ induced apoptosis in human osteosarcoma cells by extracellular S100A4

    PubMed Central

    Pedersen, Kjetil Boye; Andersen, Kristin; Fodstad, Øystein; Mælandsmo, Gunhild Mari

    2004-01-01

    Background S100A4 is a small Ca2+-binding protein of the S100 family with metastasis-promoting properties. Recently, secreted S100A4 protein has been shown to possess a number of functions, including induction of angiogenesis, stimulation of cell motility and neurite extension. Methods Cell cultures from two human osteosarcoma cell lines, OHS and its anti-S100A4 ribozyme transfected counterpart II-11b, was treated with IFN-γ and recombinant S100A4 in order to study the sensitizing effects of extracellular S100A4 on IFN-γ mediated apoptosis. Induction of apoptosis was demonstrated by DNA fragmentation, cleavage of poly (ADP-ribose) polymerase and Lamin B. Results In the present work, we found that the S100A4-expressing human osteosarcoma cell line OHS was more sensitive to IFN-γ-mediated apoptosis than the II-11b cells. S100A4 protein was detected in conditioned medium from OHS cells, but not from II-11b cells, and addition of recombinant S100A4 to the cell medium sensitized II-11b cells to apoptosis induced by IFN-γ. The S100A4/IFN-γ-mediated induction of apoptosis was shown to be independent of caspase activation, but dependent on the formation of reactive oxygen species. Furthermore, addition of extracellular S100A4 was demonstrated to activate nuclear factor-κB (NF-κB). Conclusion In conclusion, we have shown that S100A4 sensitizes osteosarcoma cells to IFN-γ-mediated induction of apoptosis. Additionally, extracellular S100A4 activates NF-κB, but whether these events are causally related remains unknown. PMID:15318945

  20. Inhibition of apoptosis using exosomes in Chinese hamster ovary cell culture.

    PubMed

    Han, Seora; Rhee, Won Jong

    2018-05-01

    Animal cell culture technology for therapeutic protein production has shown significant improvement over the last few decades. Chinese hamster ovary (CHO) cells have been widely adapted for the production of biopharmaceutical drugs. In the biopharmaceutical industry, it is crucial to develop cell culture media and culturing conditions to achieve the highest productivity and quality. However, CHO cells are significantly affected by apoptosis in the bioreactors, resulting in a substantial decrease in product quantity and quality. Thus, to overcome the obstacle of apoptosis in CHO cell culture, it is critical to develop a novel method that does not have minimal concern of safety or cost. Herein, we showed for the first time that exosomes, which are nano-sized extracellular vesicles, derived from CHO cells inhibited apoptosis in CHO cell culture when supplemented to the culture medium. Flow cytometric and microscopic analyses revealed that substantial amounts of exosomes were delivered to CHO cells. Higher cell viability after staurosporine treatment was observed by exosome supplementation (67.3%) as compared to control (41.1%). Furthermore, exosomes prevented the mitochondrial membrane potential loss and caspase-3 activation, meaning that the exosomes enhanced cellular activities under pro-apoptotic condition. As the exosomes supplements are derived from CHO cells themselves, it is not only beneficial for the biopharmaceutical productivity of CHO cell culture to inhibit apoptosis, but also from a regulatory standpoint to diminish any safety concerns. Thus, we conclude that the method developed in this research may contribute to the biopharmaceutical industry where minimizing apoptosis in CHO cell culture is beneficial. © 2018 Wiley Periodicals, Inc.