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Sample records for human a549 non-small

  1. Enhancement of Radiosensitivity by Roscovitine Pretreatment in Human Non-small Cell Lung Cancer A549 Cells

    OpenAIRE

    Feng, Zhang; Tao, Zhang; Zhong-Ping, GU; Yong-An, Zhou; yong, Han; Xiao-Fei, LI; Xiao-Ping, WANG; Qing-Shu, CHENG; Qi-Bing, MEI; Department of Thoracic Surgery, Tangdu Hospital, The Fourth Military Medical University; Department of Pharmacology, The Fourth Military Medical University

    2008-01-01

    Roscovitine has been reported to have anti-proliferative properties and is in process of undergoing clinical trials. In addition to its intrinsic anticancer properties, it has recently been suggested that roscovitine may also enhance the activity of traditional chemo- and radio-therapies in certain cancer cell lines. The purpose of this study was to define the activity of roscovitine in increasing radiosensitivity of human non-small cell lung cancer (NSCLC) cell line A549 cells in vitro. A549...

  2. Enhancement of radiosensitivity by CpG-oligodeoxyribonucleotide-7909 in human non-small cell lung cancer A549 cells.

    Science.gov (United States)

    Zha, Lin; Qiao, Tiankui; Yuan, Sujuan; Lei, Linjie

    2010-04-01

    CpG-oligodeoxyribonucleotides (CpG-ODNs), which induce signaling through the toll-like receptor 9, are currently under investigation as immunity stimulators against cancer. It has recently been suggested that CpG-ODNs may also enhance sensitivity to traditional therapies including chemotherapy in certain cancer-cell lines. The purpose of this study was to define the activity of CpG-ODN7909 in increasing radiosensitivity of the human non-small cell lung cancer cell line A549 in vitro. First, a dose- and time-dependent inhibitory effect on cell viability was observed after A549 cells were treated with different concentrations of CpG-ODN7909 (5, 10, 30, and 60 microg/mL). Second, decreased cell clonogenic survival, enhanced cell apoptotic index, accumulated percentage of cells in the G2/M phase, and increased tumor necrosis factor (TNF)-alpha secretion were found after combined treatments with 10 microg/mL of CpG-ODN7909 and radiation compared to either treatment alone (p CpG-ODN7909 can increase the radiosensitivity of human non-small cell lung cancer A549 cells, which may be associated with reduced cell clonogenic survival, enhanced apoptosis, prolonged cell-cycle arrest in G2/M, and stimulation of TNF-alpha secretion.

  3. Oleifolioside B-mediated autophagy promotes apoptosis in A549 human non-small cell lung cancer cells.

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    Jin, Cheng-Yun; Yu, Hai Yang; Park, Cheol; Han, Min Ho; Hong, Su Hyun; Kim, Kyoung-Sook; Lee, Young-Choon; Chang, Young-Chae; Cheong, Jaehun; Moon, Sung-Kwon; Kim, Gi-Young; Moon, Hyung-In; Kim, Wun-Jae; Lee, Jai-Heon; Choi, Yung Hyun

    2013-12-01

    The biochemical mechanisms of cell death by oleifolioside B (OB), a cycloartane-type triterpene glycoside isolated from Dendropanax morbifera Leveille, were investigated in A549 human lung carcinoma cells. Our data indicated that exposure to OB led to caspase activation and typical features of apoptosis; however, apoptotic cell death was not prevented by z-VAD-fmk, a pan-caspase inhibitor, demonstrating that OB-induced apoptosis was independent of caspase activation. Subsequently, we found that OB increased autophagy, as indicated by an increase in monodansylcadaverine fluorescent dye-labeled autophagosome formation and in the levels of the autophagic form of microtubule-associated protein 1 light chain 3 and Atg3, an autophagy-specific gene, which is associated with inhibiting phospho-nuclear factor erythroid 2-related factor 2 (Nrf2) expression. However, pretreatment with bafilomycin A1, an autophagy inhibitor, attenuated OB-induced apoptosis and dephosphorylation of Nrf2. The data suggest that OB-induced autophagy functions as a death mechanism in A549 cells and OB has potential as a novel anticancer agent capable of targeting apoptotic and autophagic cell death and the Nrf2 signaling pathway.

  4. 卡瓦胡椒素B对人非小细胞肺癌A549细胞的抑制作用%Inhibitory Effect of Flavokawain B on Human Non-small Cell Lung Cancer A549 Cells

    Institute of Scientific and Technical Information of China (English)

    王晶; 安君霞; 朱启彧; 马玉玲; 裴哲; 魏枭; 孙健; 唐亚雄

    2012-01-01

    The anti-lung tumor potential of flavokawain B, one of active chalcones isolated from Kawa was investigated. Flavokawain B's action was assessed on proliferation, apoptosis, cell cycle and molecular mechanisms in human non-small cell lung cancer (NSCLC) A549 cells in vitro. The results demonstrated that flavokawain B significantly inhibited the growth of A549 cells in a dose- and time-dependent manner. Meanwhile, flavokawain B induced cell apoptosis and cell cycle G2-M phase arrest in A549 cells. Mechanistically, flavokawain B could activate JNK signaling pathway, down-regulate the expression of survivin protein, and activate the cleavage of PARP, leading to marked inhibitory effect on A549 cells. These findings suggest that flavokawain B may be a potential usefulness for preventing and treatment of NSCLC. Fig 5, Ref 16%卡瓦胡椒素B是药用植物卡瓦胡椒根中的一种天然查耳酮类化合物,研究了其对人非小细胞肺癌A549细胞的增殖抑制作用及其诱导细胞凋亡的分子机制.实验结果显示,卡瓦胡椒素B能显著抑制非小细胞肺癌A549细胞的增殖,且随着药物浓度的增加、处理时间的延长其抑制作用呈明显的剂量时间效应;同时,卡瓦胡椒素B能显著诱导A549细胞凋亡、细胞周期阻滞于G2-M期;分子机制研究表明,卡瓦胡椒素B能通过活化JNK激酶活性、下调凋亡抑制蛋白survivin的表达以及激活PARP活性从而导致其对A549细胞的增殖抑制作用.结果表明卡瓦胡椒素B对人非小细胞肺癌的预防与治疗可能具有潜在价值.

  5. Green tea catechin, epigallocatechin-3-gallate, attenuates the cell viability of human non-small-cell lung cancer A549 cells via reducing Bcl-xL expression.

    Science.gov (United States)

    Sonoda, Jun-Ichiro; Ikeda, Ryuji; Baba, Yasutaka; Narumi, Keiko; Kawachi, Akio; Tomishige, Erisa; Nishihara, Kazuya; Takeda, Yasuo; Yamada, Katsushi; Sato, Keizo; Motoya, Toshiro

    2014-07-01

    Clinical and epidemiological studies have indicated that the consumption of green tea has a number of beneficial effects on health. Epigallocatechin-3-gallate (EGCg), the major polyphenolic compound present in green tea, has received much attention as an active ingredient. Among the numerous promising profiles of EGCg, the present study focused on the anticancer effects. Apoptosis induced by EGCg and subsequent cell growth suppression have been demonstrated in a number of cell culture studies. However, the underlying mechanism of apoptotic cell death remains unclear. Thus, the aim of the present study was to identify the major molecule that mediates proapoptotic cell death by EGCg. The effect of EGCg on cell proliferation and the induction of mRNA that modulates apoptotic cell death was evaluated in the A549 human non-small-cell lung cancer cell line. In addition, morphological changes were assessed by microscopy in A549 cells that had been treated with 100 μM EGCg for 24 h. The MTT assay revealed that cell proliferation was significantly reduced by EGCg in a dose-dependent manner (3-100 μM). The mRNA expression level of B-cell lymphoma-extra large (Bcl-xL) was decreased in A549 cells following 24 h incubation with 100 μM EGCg. Therefore, the results indicated that the inhibition of cell proliferation by EGCg may be achieved via suppressing the expression of the cell death-inhibiting gene, Bcl-xL.

  6. Molecular switch role of Akt in Polygonatum odoratum lectin-induced apoptosis and autophagy in human non-small cell lung cancer A549 cells.

    Science.gov (United States)

    Li, Chunyang; Chen, Jie; Lu, Bangmin; Shi, Zheng; Wang, Hailian; Zhang, Bin; Zhao, Kailiang; Qi, Wei; Bao, Jinku; Wang, Yi

    2014-01-01

    Polygonatum odoratum lectin (POL), isolated from traditional Chinese medicine herb (Mill.) Druce, has drawn rising attention due to its wide biological activities. In the present study, anti-tumor effects, including apoptosis- and autophagy-inducing properties of POL, were determined by a series of cell biology methods such as MTT, cellular morphology observation, flow cytometry, immunoblotting. Herein, we found that POL could simultaneously induce apoptosis and autophagy in human non-small cell lung cancer A549 cells. POL initiated apoptosis through inhibiting Akt-NF-κB pathway, while POL triggered autophagy via suppressing Akt-mTOR pathway, suggesting the molecular switch role of Akt in regulating between POL-induced apoptosis and autophagy. Moreover, ROS was involved in POL-induced inhibition of Akt expression, and might therefore mediate both apoptosis and autophagy in A549 cells. In addition, POL displayed no significant cytotoxicity toward normal human embryonic lung fibroblast HELF cells. Due to the anti-tumor activities, POL might become a potent anti-cancer drug in future therapy, which might pave the way for exploring GNA-related lectins into effective drugs in cancer treatment.

  7. Human Noxin is an anti-apoptotic protein in response to DNA damage of A549 non-small cell lung carcinoma.

    Science.gov (United States)

    Won, Kyoung-Jae; Im, Joo-Young; Yun, Chae-Ok; Chung, Kyung-Sook; Kim, Young Joo; Lee, Jung-Sun; Jung, Young-Jin; Kim, Bo-Kyung; Song, Kyung Bin; Kim, Young-Ho; Chun, Ho-Kyung; Jung, Kyeong Eun; Kim, Moon-Hee; Won, Misun

    2014-06-01

    Human Noxin (hNoxin, C11Orf82), a homolog of mouse noxin, is highly expressed in colorectal and lung cancer tissues. hNoxin contains a DNA-binding C-domain in RPA1, which mediates DNA metabolic processes, such as DNA replication and DNA repair. Expression of hNoxin is associated with S phase in cancer cells and in normal cells. Expression of hNoxin was induced by ultraviolet (UV) irradiation. Knockdown of hNoxin caused growth inhibition of colorectal and lung cancer cells. The comet assay and western blot analysis revealed that hNoxin knockdown induced apoptosis through activation of p38 mitogen-activated protein kinase (MAPK)/p53 in non-small cell lung carcinoma A549 cells. Furthermore, simultaneous hNoxin knockdown and treatment with DNA-damaging agents, such as camptothecin (CPT) and UV irradiation, enhanced apoptosis, whereas Trichostatin A (TSA) did not. However, transient overexpression of hNoxin rescued cells from DNA damage-induced apoptosis but did not block apoptosis in the absence of DNA damage. These results suggest that hNoxin may be associated with inhibition of apoptosis in response to DNA damage. An adenovirus expressing a short hairpin RNA against hNoxin transcripts significantly suppressed the growth of A549 tumor xenografts, indicating that hNoxin knockdown has in vivo anti-tumor efficacy. Thus, hNoxin is a DNA damage-induced anti-apoptotic protein and potential therapeutic target in cancer.

  8. Andrographolide down-regulates hypoxia-inducible factor-1α in human non-small cell lung cancer A549 cells.

    Science.gov (United States)

    Lin, Hui-Hsuan; Tsai, Chia-Wen; Chou, Fen-Pi; Wang, Chau-Jong; Hsuan, Shu-Wen; Wang, Cheng-Kun; Chen, Jing-Hsien

    2011-02-01

    Andrographolide (Andro), a diterpenoid lactone isolated from a traditional herbal medicine Andrographis paniculata, is known to possess multiple pharmacological activities. In our previous study, Andro had been shown to inhibit non-small cell lung cancer (NSCLC) A549 cell migration and invasion via down-regulation of phosphatidylinositol 3-kinase (PI3K)/Akt signaling pathway. Here we demonstrated that Andro inhibited the expression of hypoxia-inducible factor-1α (HIF-1α) in A549 cells. HIF-1α plays an important role in tumor growth, angiogenesis and lymph node metastasis of NSCLC. The Andro-induced decrease of cellular protein level of HIF-1α was correlated with a rapid ubiquitin-dependent degradation of HIF-1α, and was accompanied by increased expressions of hydroxyl-HIF-1α and prolyl hydroxylase (PHD2), and a later decrease of vascular endothelial growth factor (VEGF) upon the treatment of Andro. The Andro-inhibited VEGF expression appeared to be a consequence of HIF-1α inactivation, because its DNA binding activity was suppressed by Andro. Molecular data showed that all these effects of Andro might be mediated via TGFβ1/PHD2/HIF-1α pathway, as demonstrated by the transfection of TGFβ1 overexpression vector and PHD2 siRNA, and the usage of a pharmacological MG132 inhibitor. Furthermore, we elucidated the involvement of Andro in HIF-1α transduced VEGF expression in A549 cells and other NSCLC cell lines. In conclusion, these results highlighted the potential effects of Andro, which may be developed as a chemotherapeutic or an anti-angiogenesis agent for NSCLC in the future.

  9. Inhibitory Effects of Salinomycin on Cell Survival, Colony Growth, Migration, and Invasion of Human Non-Small Cell Lung Cancer A549 and LNM35: Involvement of NAG-1.

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    Arafat, Kholoud; Iratni, Rabah; Takahashi, Takashi; Parekh, Khatija; Al Dhaheri, Yusra; Adrian, Thomas E; Attoub, Samir

    2013-01-01

    A major challenge for oncologists and pharmacologists is to develop more potent and less toxic drugs that will decrease the tumor growth and improve the survival of lung cancer patients. Salinomycin is a polyether antibiotic used to kill gram-positive bacteria including mycobacteria, protozoans such as plasmodium falciparum, and the parasites responsible for the poultry disease coccidiosis. This old agent is now a serious anti-cancer drug candidate that selectively inhibits the growth of cancer stem cells. We investigated the impact of salinomycin on survival, colony growth, migration and invasion of the differentiated human non-small cell lung cancer lines LNM35 and A549. Salinomycin caused concentration- and time-dependent reduction in viability of LNM35 and A549 cells through a caspase 3/7-associated cell death pathway. Similarly, salinomycin (2.5-5 µM for 7 days) significantly decreased the growth of LNM35 and A549 colonies in soft agar. Metastasis is the main cause of death related to lung cancer. In this context, salinomycin induced a time- and concentration-dependent inhibition of cell migration and invasion. We also demonstrated for the first time that salinomycin induced a marked increase in the expression of the pro-apoptotic protein NAG-1 leading to the inhibition of lung cancer cell invasion but not cell survival. These findings identify salinomycin as a promising novel therapeutic agent for lung cancer.

  10. Inhibitory Effects of Salinomycin on Cell Survival, Colony Growth, Migration, and Invasion of Human Non-Small Cell Lung Cancer A549 and LNM35: Involvement of NAG-1.

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    Kholoud Arafat

    Full Text Available A major challenge for oncologists and pharmacologists is to develop more potent and less toxic drugs that will decrease the tumor growth and improve the survival of lung cancer patients. Salinomycin is a polyether antibiotic used to kill gram-positive bacteria including mycobacteria, protozoans such as plasmodium falciparum, and the parasites responsible for the poultry disease coccidiosis. This old agent is now a serious anti-cancer drug candidate that selectively inhibits the growth of cancer stem cells. We investigated the impact of salinomycin on survival, colony growth, migration and invasion of the differentiated human non-small cell lung cancer lines LNM35 and A549. Salinomycin caused concentration- and time-dependent reduction in viability of LNM35 and A549 cells through a caspase 3/7-associated cell death pathway. Similarly, salinomycin (2.5-5 µM for 7 days significantly decreased the growth of LNM35 and A549 colonies in soft agar. Metastasis is the main cause of death related to lung cancer. In this context, salinomycin induced a time- and concentration-dependent inhibition of cell migration and invasion. We also demonstrated for the first time that salinomycin induced a marked increase in the expression of the pro-apoptotic protein NAG-1 leading to the inhibition of lung cancer cell invasion but not cell survival. These findings identify salinomycin as a promising novel therapeutic agent for lung cancer.

  11. 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

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    Jun Xie

    2015-11-01

    Full Text Available 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.

  12. Inhibitory effect of gallic acid on proliferation of human non-small cell lung cancer A549 cells%没食子酸对人非小细胞肺癌A549细胞增殖的抑制作用

    Institute of Scientific and Technical Information of China (English)

    郗艳丽; 许娜; 李澍; 王迪; 王舒然; 牛凤兰

    2016-01-01

    组间比较差异无统计学意义(P>0.05);中和高剂量没食子酸组 A549细胞中 GSH-Px活性低于对照组,但组间比较差异亦无统计学意义(P>0.05);随着没食子酸剂量的不断增加,GSH-Px活性反而降低。低、中和高剂量没食子酸组间A549细胞中CAT、SOD和 GSH-Px活性比较差异无统计学意义(P>0.05)。结论:没食子酸可能是通过诱导氧化损伤的方式抑制肺癌细胞增殖,Fas/FasL信号通路可能是其诱导细胞凋亡的重要作用机制之一。%Objective:To detect the inhibitory effect of gallic acid on the proliferation of human non-small cell lung cancer A549 cells,and to investigate the molecular mechanisms of its drug toxicity.Methods:The human non-small cell lung cancer A549 cells were cultured invitro.The cells were divided into control group,low,middle, and high dosages of gallic acid groups (0,300,500 and 750μmol·L-1 ).The survival rates of cells were tested by MTT method;the morphology of A549 cells were tested by Switzerland and Giemsa staining;the apoptotic rates of A549 cells and the levels of reactive oxygen species (ROS)in A549 cells were analyzed by FCM;the expression levels of Fas and FasL in the A549 cells in various groups were detected by Western blotting method;the activities of catalase (CAT),superoxide dismutase (SOD)and glutathione peroxidase (GSH-Px)in the A549 cells in various groups were analyzed by spectrophotometry. Results:Compared with gallic acid treatment for 6 h, the survival rates of A549 cells treated for 12 and 24 h in different dosages of gallic acid groups were significantly decreased (P0.05);the activities of CAT in A549 cells in different dosages of gallic acid groups were reduced with the increasing of gallic acid dosage. The activity of SOD in low dosage of gallic acid group was higher than that in control group (P0.05).The activities of SOD were reduced with the increasing of gallic acid dosages.The activity of GSH

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

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    Lu, Xiangyi; Liu, Wei; Wu, Junhua; Li, Mengxian [Key Laboratory of Food Nutrition and Safety, Ministry of Education, School of Food Engineering and Biotechnology, Tianjin University of Science and Technology, Tianjin 300457 (China); Wang, Juncheng; Wu, Jihui [School of Life Science, University of Science and Technology of China, Hefei 230022 (China); Luo, Cheng, E-mail: Luo58@yahoo.com [Key Laboratory of Food Nutrition and Safety, Ministry of Education, School of Food Engineering and Biotechnology, Tianjin University of Science and Technology, Tianjin 300457 (China)

    2013-01-11

    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 as 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.

  14. 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.

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    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

  15. 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

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    Pan ST

    2015-02-01

    Full Text Available Shu-Ting Pan,1,* Zhi-Wei Zhou,2,3,* Zhi-Xu He,3 Xueji Zhang,4 Tianxin Yang,5 Yin-Xue Yang,6 Dong Wang,7 Jia-Xuan Qiu,1 Shu-Feng Zhou2 1Department of Oral and Maxillofacial Surgery, The First Affiliated Hospital of Nanchang University, Nanchang, Jiangxi, People’s Republic of China; 2Department of Pharmaceutical Sciences, College of Pharmacy, University of South Florida, Tampa, FL, USA; 3Guizhou Provincial Key Laboratory for Regenerative Medicine, Stem Cell and Tissue Engineering Research Center and Sino-US Joint Laboratory for Medical Sciences, Guiyang Medical University, Guiyang, 4Research Center for Bioengineering and Sensing Technology, University of Science and Technology Beijing, Beijing, People’s Republic of China; 5Department of Internal Medicine, University of Utah and Salt Lake Veterans Affairs Medical Center, Salt Lake City, UT, USA; 6Department of Colorectal Surgery, General Hospital of Ningxia Medical University, Yinchuan, 7Cancer Center, Daping Hospital and Research Institute of Surgery, Third Military Medical University, Chongqing, People’s Republic of China *These two authors contributed equally to this work Abstract: 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

  16. Epithelial mesenchymal transition of non-small-cell lung cancer cells A549 induced by SPHK1

    Institute of Scientific and Technical Information of China (English)

    Min Ni; Xiao-Lei Shi; Zhi-Gang Qu; Hong Jiang; Zi-Qian Chen; Jun Hu

    2015-01-01

    Objective:To explore the effect and molecular mechanism ofSPHK1 in the invasion and metastasis process of non-small-cell lung cancer cells(A549).Methods:Recombinant retrovirus was used to mediate the production ofA549/vector,A549/SPHK1,A549/scramble, andA549/SPHK1/RNAi that stably expressed or silencedSPHK1.The invasion and migration capacities of A549 cells overexpressing or silencingSPHK1 were determined usingTranswell invasion assay and scratch wound repair experiment.The protein and mRNA expression levels ofE-cadherin, fibronectin, vimentin inA549/vector,A549/SPHK1,A549/scramble,A549/SPHK1/RNAi were detected withWestern blot(WB) and quantitativePCR(QPCR) methods, respectively.Results:Transwell invasion assay and scratch wound repair experiments showed that over-expression of SPHK1 obviously enhanced the invasion and migration capacities ofA549 cells.WB andQPCR detection results showed that, the expression ofE-cadherin(a molecular marker of epithelial cells) and fibronectin, vimentin(molecular markers of mesenchymal cells) inA549 cells was upregulated after overexpression ofSPHK1; whileSPHK1 silencing significantly reduced the invasion and metastasis capacities ofA549cells, upregulated the expression of molecular marker of epithelial cells, and downregulated the expression of molecular marker of mesenchymal cells. Conclusions:SPHK1 promotes epithelial mesenchymal transition of non-small-cell lung cancer cells and affects the invasion and metastasis capacities of these cells.

  17. Anti-Proliferative and Apoptosis-Inducing Effect of Theabrownin against Non-small Cell Lung Adenocarcinoma A549 Cells

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    Wu, Feifei; Zhou, Li; Jin, Wangdong; Yang, Weiji; Wang, Ying; Yan, Bo; Du, Wenlin; Zhang, Qiang; Zhang, Lei; Guo, Yonghua; Zhang, Jin; Shan, Letian; Efferth, Thomas

    2016-01-01

    With the highest cancer incidence rate, lung cancer, especially non-small cell lung cancer (NSCLC), is the leading cause of cancer death in the world. Tea (leaves of Camellia sinensis) has been widely used as a traditional beverage beneficial to human health, including anti-NSCLC activity. Theabrownin (TB) is one major kind of tea pigment responsible for the beneficial effects of tea liquor. However, its effect on NSCLC is unknown. The aim of the present study was to evaluate anti-proliferative and apoptosis-inducing effect of TB on NSCLC (A549) cells, using MTT assay, morphological observation (DAPI staining), in situ terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) assay, and annexin-V/PI flow cytometry. Subsequently, the expression of several genes associated with cell proliferation and apoptosis were detected by real time PCR assay to explore its potential underlying mechanism. TB was revealed to inhibit cell proliferation of A549 cells in a concentration-dependent and time-dependent manner. Morphological observation, TUNEL assay and flow cytometric analysis evidenced an apoptosis-inducing effect of TB on A549 cells in a concentration-dependent manner. The real time PCR assay demonstrated that TB down-regulated the expression of TOPO I, TOPO II, and BCL-2, and up-regulated the expression of E2F1, P53, GADD45, BAX, BIM, and CASP 3,7,8,9, which suggests an activation of P53-mediated apoptotic (caspase-dependent) pathway in response to TB treatment. The western blot analysis showed a similar trend for the corresponding protein expression (P53, Bax, Bcl-2, caspase 3,9, and PARP) and further revealed DNA damage as a trigger of the apoptosis (phosphorylation of histone H2A.X). Accordingly, TB can be speculated as a DNA damage inducer and topoisomerase (Topo I and Topo II) inhibitor that can up-regulate P53 expression and subsequently modulate the expression of the downstream genes to induce cell proliferation inhibition and apoptosis of A549 cells

  18. Anti-proliferative and apoptosis-inducing effect of theabrownin against non-small cell lung adenocarcinoma A549 cells

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    Feifei Wu

    2016-12-01

    Full Text Available With the highest cancer incidence rate, lung cancer, especially non-small cell lung cancer (NSCLC, is the leading cause of cancer death in the world. Tea (leaves of Camellia sinensis has been widely used as a traditional beverage beneficial to human health, including anti-NSCLC activity. Theabrownin (TB is one major kind of tea pigment responsible for the beneficial effects of tea liquor. However, its effect on NSCLC is unknown. The aim of the present study was to evaluate anti-proliferative and apoptosis-inducing effect of TB on NSCLC (A549 cells, using MTT assay, morphological observation (DAPI staining, in situ terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL assay, and annexin V/PI flow cytometry. Subsequently, the expression of several genes associated with cell proliferation and apoptosis were detected by real time PCR assay to explore its potential underlying mechanism. TB was revealed to inhibit cell proliferation of A549 cells in a concentration-dependent and time-dependent manner. Morphological observation, TUNEL assay and flow cytometric analysis evidenced an apoptosis-inducing effect of TB on A549 cells in a concentration-dependent manner. The real time PCR assay demonstrated that TB down-regulated the expression of TOPO I, TOPO II, and BCL-2, and up-regulated the expression of E2F1, P53, GADD45, BAX, BIM, and CASP 3,7,8,9, which suggests an activation of P53-mediated apoptotic (caspase-dependent pathway in response to TB treatment. The western blot analysis showed a similar trend for the corresponding protein expression (P53, Bax, Bcl-2, caspase 3,9, and PARP and further revealed DNA damage as a trigger of the apoptosis (phosphorylation of histone H2A.X. Accordingly, TB can be speculated as a DNA damage inducer and topoisomerase (Topo I and Topo II inhibitor that can up-regulate P53 expression and subsequently modulate the expression of the downstream genes to induce cell proliferation inhibition and apoptosis of

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

    Science.gov (United States)

    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.

  20. 线粒体靶向MPG基因重组体对人非小细胞肺癌多药耐药细胞A549/DDP增殖的抑制作用%Inhibitory Effect of Human Mitochondria-targeted MPG Recombinant on Proliferation of Human Non-small Cell Lung Cancer Multidrug-resistant Cell Line A549/DDP

    Institute of Scientific and Technical Information of China (English)

    余时沧; 钱桂生; 李玉英; 陆卫忠; 李瑾; 黄桂君

    2006-01-01

    背景与目的:多药耐药是影响肺癌化疗效果的重要因素.本研究拟构建线粒体靶向人N-甲基化嘌呤DNA糖基化酶(MPG)基因真核表达载体,观察其在稳定转染的人非小细胞肺癌多药耐药细胞线粒体内的表达情况,并研究其对多药耐药细胞增殖的抑制作用.方法:应用重叠延伸剪接技术重组锰超氧化物歧化酶(MnSOD)线粒体靶向序列-MPG融合基因(mito-MPG);构建pCMV-Script/mitoMPG重组真核表达载体;脂质体将其转染至人非小细胞肺癌多药耐药细胞A549/DDP;G418筛选稳定表达的转染细胞;RT-PCR检测mito-MPG基因mRNA的表达水平;分离线粒体蛋白后应用Western blot检测MPG在线粒体内的表达水平;台盼蓝拒染法检测细胞增殖能力;流式细胞术检测细胞周期分布.结果:构建的融合基因经过DNA测序分析;构建的重组载体经限制性酶切分析及DNA测序分析证实为pCMV-Script/mito-MPG重组真核表达载体;转染pCMV-Script/mito-MPG载体组(MPG组)检测到mito-MPG mRNA的表达,转染pCMV-Script载体组(P组)及未转染组(C组)细胞内则未检测到;MPG组细胞线粒体内检测到MPG,P组及C组则未检测到;MPG组细胞增殖能力明显下降,P组及C组细胞增殖能力无明显差异,倍增时间分别为72.6h(C组)、73.5 h(P组)、98.9 h(MPG组);分裂增殖指数分别为51.3%(C组)、54.3%(P组)、26.1%(MPG组),MPG组出现亚二倍体峰.结论:成功构建了线粒体靶向人MPG基因表达载体,MPG在MnSOD线粒体靶向序列的引导下,顺利地进入了A549/DDP细胞线粒体内,并导致其增殖能力下降,部分细胞死亡.

  1. SIRT1 Influences the Sensitivity of A549 Non-small Cell Lung Cancer Cell Line to 
Cisplatin via Modulating the Noxa Expression

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    Bin CAO

    2016-02-01

    Full Text Available Background and objective The resistance of non-small cell lung cancer cells to cisplant is a common clinical phenomenon which could induce a poor therapeutic effect and should be difficult problem to be solved. SIRT1 and Noxa expression are associated with the chemotherapy for tumors. The present study focused on how SIRT1 expression influence the senstivity of non-small cell lung cancer cells and dissected the potential mechanism involved with Noxa. Methods The difference of SIRT1 and Noxa expression between A549 cells and A549/DDP cells was detected by real-time quantitative PCR (qRT-PCR and Western blot. SIRT1 targeted siRNA was uesed to inhibit the SIRT1 expression in A549/DDP, after transfection, Cell Titer Blue assay, flow cytometry were performed to analyze the cell viability, cell cycle and cell apoptosis in order to reveal the effect of inhibition of SIRT1 on sensitivity of A549/DDP cells to cisplant. Moreover, the expression changes of Noxa in A549/DDP cells after siRNA treatment were detected by qRT-PCR and Western blot. Results There was a significant difference in senstivity to cisplant between A549 and A549/DDP cells. Compared with A549 cells, the A549/DDP cells showed a higher SIRT1 expression and lower Noxa expression. After transfected with SIRT1 targeted siRNA, the cell viability decreased accompanied with a increasing apoptosis rate, meanwhile, higher percent of G2/M phase was detected after the 4 μg/mL cisplant treatment. Further more, inhibition of SIRT1 could induce the Noxa expression in A549/DDP cells. Conclusion Higher SIRT1 expression may induce resistance to cisplant in A549 cells. SIRT1 inhibition may improve the sensitivity of A549/DDP cells to cisplantin though modulating the Noxa expression.

  2. Effects of miR-424 on Proliferation and Migration Abilities in Non-small Cell Lung Cancer A549 Cells and Its Molecular Mechanism

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    Hongmin LI

    2016-09-01

    Full Text Available Background and objective The inhibitory ability of miR-424 on the proliferation of renal carcinoma cell and the migration and invasion of cancer cells has been widely explored and demonstrated. However, the effects of miR-424 on non-small cell lung cancer (NSCLC have not been systematically examined. In this study, detected the growth and invasion effect of miR-424 in NSCLC A549 cell. The migration and molecular mechanism of this cell are also detected. Methods NSCLC A549 cell was transfected with miR-424 and its inhibitor. After transfection, the proliferation ability of A549 cell was detectedby CCK8 assay. Then, the migration ability in A549 cell was detected by migration assays. Furthermore, the expression level of MMP2 and MMP9 in A549 was detected by Western blot and immune fluorescence. The 3'UTR of E2F6 was cloned into luciferase reporter vector and its enzymatic activitywas detected to verify whether miR-424 can target E2F6. The expression level of E2F6 in a549 cell after transfecing with miR-424 was detected by Western blot. Results After transfection of miR-424, the proliferation and migration abilities were remarkably decreased and the expression level of MMP-2 and MMP-9 were down-regulated in A549. Moreover, MiR-424 inhibited the enzymatic activity of luviferase reporter vector of E2F6. Specifically, the expression level of E2F6 was down-regulated in A549. Conclusion miR-424 can inhibit the proliferation and migration abilities of A549 by negatively regulating the expression of E2F6.

  3. Xanthatin Induces Cell Cycle Arrest at G2/M Checkpoint and Apoptosis via Disrupting NF-κB Pathway in A549 Non-Small-Cell Lung Cancer Cells

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    Yin Lu

    2012-03-01

    Full Text Available Xanthatin, a natural sesquiterpene lactone, has significant antitumor activity against a variety of cancer cells, yet little is known about its anticancer mechanism. In this study, we demonstrated that xanthatin had obvious dose-/time-dependent cytotoxicity against the human non-small-cell lung cancer (NSCLC cell line A549. Flow cytometry analysis showed xanthatin induced cell cycle arrest at G2/M phase. Xanthatin also had pro-apoptotic effects on A549 cells as evidenced by Hoechst 33258 staining and annexin V-FITC staining. Mechanistic data revealed that xanthatin downregulated Chk1, Chk2, and phosphorylation of CDC2, which contributed to the cell cycle arrest. Xathatin also increased total p53 protein levels, decreased Bcl-2/Bax ratio and expression of the downstream factors procaspase-9 and procaspase-3, which triggered the intrinsic apoptosis pathway. Furthermore, xanthatin blocked phosphorylation of NF-κB (p65 and IκBa, which might also contribute to its pro-apoptotic effects on A549 cells. Xanthatin also inhibited TNFa induced NF-κB (p65 translocation. We conclude that xanthatin displays significant antitumor effects through cell cycle arrest and apoptosis induction in A549 cells. These effects were associated with intrinsic apoptosis pathway and disrupted NF-κB signaling. These results suggested that xanthatin may have therapeutic potential against NSCLC.

  4. Differential alterations of positive and negative regulators of beta catenin enhance endogenous expression and activity of beta catenin in A549 non small cell lung cancer (NSCLC cells

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    Supratim Ghatak

    2016-12-01

    Full Text Available Beta catenin has been well documented in previous studies to be involved in non small cell lung cancer (NSCLC. Beta catenin abundance and transcriptional activity are significantly regulated by several factors. Though it is well known that Akt and Gsk3 beta are respective positive and negative regulators of beta catenin, however, no single study has so far documented how the expression and activity of both positive as well as negative regulators play favorable role on beta catenin expression and activity in NSCLC. In this study, we compared expression and activity of beta catenin and its regulators in normal lung cell WI38 and NSCLC cell A549 by western blot, qRT-PCR and luciferase assay. We observed that beta catenin positive regulators (Akt and Hsp90 and negative regulators (Gsk3 beta and microRNA-214 have differential expression and/or activity in NSCLC cell A549. However the differentially altered statuses of both the positive and negative regulators rendered cumulative positive effect on beta catenin expression and activity in A549. Our study thus suggests that chemotherapeutic modulations of regulating factors are crucial when abrogation and/or inhibition of key oncogenic proteins are necessary for cancer chemotherapy.

  5. Non-small-cell lung cancer cell lines A549 and NCI-H460 express hypoxanthine guanine phosphoribosyltransferase on the plasma membrane

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    Townsend MH

    2017-03-01

    Full Text Available Michelle H Townsend, Michael D Anderson, Evita G Weagel, Edwin J Velazquez, K Scott Weber, Richard A Robison, Kim L O’Neill Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT, USA Abstract: In both males and females, lung cancer is one of the most lethal cancers worldwide and accounts for >30% of cancer-related deaths. Despite advances in biomarker analysis and tumor characterization, there remains a need to find suitable biomarker antigen targets for treatment in late-stage lung cancer. Previous research on the salvage pathway enzyme TK1 shows a unique relationship with cancer patients as serum levels are raised according to cancer grade. To expand this analysis, the other salvage pathway enzymes were evaluated for possible upregulation within lung cancer. Adenine phosphoribosyltransferase, deoxycytidine kinase, and hypoxanthine guanine phosphoribosyltransferase (HPRT were assessed for their presentation on two non-small-cell lung cancer cell lines NCI-H460 and A549. In the present study, we show that deoxycytidine kinase and adenine phosphoribosyltransferase have no significant relationship with the membrane of NCI-H460 cells. However, we found significant localization of HPRT to the membrane of NCI-H460 and A549 cells. When treated with anti-HPRT antibodies, the average fluorescence of the cell population increased by 24.3% and 12.9% in NCI-H460 and A549 cells, respectively, in comparison with controls. To ensure that expression was not attributed to cytoplasmic HPRT, confocal microscopy was performed to visualize HPRT binding on the plasma membrane. After staining NCI-H460 cells treated with both fluorescent antibodies and a membrane-specific dye, we observed direct overlap between HPRT and the membrane of the cancer cells. Additionally, gold-conjugated antibodies were used to label and quantify the amount of HPRT on the cell surface using scanning electron microscopy and energy-dispersive analysis X

  6. Antiproliferative and antimetastatic action of quercetin on A549 non-small cell lung cancer cells through its effect on the cytoskeleton.

    Science.gov (United States)

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

    2017-03-01

    To our knowledge, this study is the first to investigate the effect of the dietary flavonoid quercetin on the main cytoskeletal elements, namely microfilaments, microtubules and vimentin intermediate filaments, as well as cytoskeleton-driven processes in A549 non-small cell lung cancer cells. The methyl-thiazol-diphenyl-tetrazolium assay, annexin V/propidium iodide test, electron microscopic examination, cell cycle analysis based on DNA content, real-time PCR assays, in vitro scratch wound-healing assay, fluorescence staining of F-actin, β-tubulin and vimentin were performed to assess the effects of quercetin on A549 cells. Our results showed that quercetin triggered BCL2/BAX-mediated apoptosis, as well as necrosis and mitotic catastrophe, and inhibited the migratory potential of A549 cells. The disassembling effect of quercetin on microfilaments, microtubules and vimentin filaments along with its inhibitory impact on vimentin and N-cadherin expression might account for the decreased migration of A549 cells in response to quercetin treatment. We also suggest that the possible mechanism underlying quercetin-induced mitotic catastrophe involves the perturbation of mitotic microtubules leading to monopolar spindle formation, and, consequently, to the failure of cytokinesis. We further propose that cytokinesis failure could also be a result of the depletion of actin filaments by quercetin. These findings are important to our further understanding of the detailed mechanism of the antitumor activity of quercetin and render this flavonoid a potentially useful candidate for combination therapy with conventional antimicrotubule drugs, nucleic acid-directed agents or novel cytoskeletal-directed agents.

  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.

    Science.gov (United States)

    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. Non-small-cell lung cancer cell lines A549 and NCI-H460 express hypoxanthine guanine phosphoribosyltransferase on the plasma membrane.

    Science.gov (United States)

    Townsend, Michelle H; Anderson, Michael D; Weagel, Evita G; Velazquez, Edwin J; Weber, K Scott; Robison, Richard A; O'Neill, Kim L

    2017-01-01

    In both males and females, lung cancer is one of the most lethal cancers worldwide and accounts for >30% of cancer-related deaths. Despite advances in biomarker analysis and tumor characterization, there remains a need to find suitable biomarker antigen targets for treatment in late-stage lung cancer. Previous research on the salvage pathway enzyme TK1 shows a unique relationship with cancer patients as serum levels are raised according to cancer grade. To expand this analysis, the other salvage pathway enzymes were evaluated for possible upregulation within lung cancer. Adenine phosphoribosyltransferase, deoxycytidine kinase, and hypoxanthine guanine phosphoribosyltransferase (HPRT) were assessed for their presentation on two non-small-cell lung cancer cell lines NCI-H460 and A549. In the present study, we show that deoxycytidine kinase and adenine phosphoribosyltransferase have no significant relationship with the membrane of NCI-H460 cells. However, we found significant localization of HPRT to the membrane of NCI-H460 and A549 cells. When treated with anti-HPRT antibodies, the average fluorescence of the cell population increased by 24.3% and 12.9% in NCI-H460 and A549 cells, respectively, in comparison with controls. To ensure that expression was not attributed to cytoplasmic HPRT, confocal microscopy was performed to visualize HPRT binding on the plasma membrane. After staining NCI-H460 cells treated with both fluorescent antibodies and a membrane-specific dye, we observed direct overlap between HPRT and the membrane of the cancer cells. Additionally, gold-conjugated antibodies were used to label and quantify the amount of HPRT on the cell surface using scanning electron microscopy and energy-dispersive analysis X-ray. Further confirming HPRT presence, the gold weight percentage of the sample increased significantly when NCI-H460 cells were exposed to HPRT antibody (P=0.012) in comparison with isotype controls. Our results show that HPRT is localized on the

  9. Non-small-cell lung cancer cell lines A549 and NCI-H460 express hypoxanthine guanine phosphoribosyltransferase on the plasma membrane

    Science.gov (United States)

    Townsend, Michelle H; Anderson, Michael D; Weagel, Evita G; Velazquez, Edwin J; Weber, K Scott; Robison, Richard A; O’Neill, Kim L

    2017-01-01

    In both males and females, lung cancer is one of the most lethal cancers worldwide and accounts for >30% of cancer-related deaths. Despite advances in biomarker analysis and tumor characterization, there remains a need to find suitable biomarker antigen targets for treatment in late-stage lung cancer. Previous research on the salvage pathway enzyme TK1 shows a unique relationship with cancer patients as serum levels are raised according to cancer grade. To expand this analysis, the other salvage pathway enzymes were evaluated for possible upregulation within lung cancer. Adenine phosphoribosyltransferase, deoxycytidine kinase, and hypoxanthine guanine phosphoribosyltransferase (HPRT) were assessed for their presentation on two non-small-cell lung cancer cell lines NCI-H460 and A549. In the present study, we show that deoxycytidine kinase and adenine phosphoribosyltransferase have no significant relationship with the membrane of NCI-H460 cells. However, we found significant localization of HPRT to the membrane of NCI-H460 and A549 cells. When treated with anti-HPRT antibodies, the average fluorescence of the cell population increased by 24.3% and 12.9% in NCI-H460 and A549 cells, respectively, in comparison with controls. To ensure that expression was not attributed to cytoplasmic HPRT, confocal microscopy was performed to visualize HPRT binding on the plasma membrane. After staining NCI-H460 cells treated with both fluorescent antibodies and a membrane-specific dye, we observed direct overlap between HPRT and the membrane of the cancer cells. Additionally, gold-conjugated antibodies were used to label and quantify the amount of HPRT on the cell surface using scanning electron microscopy and energy-dispersive analysis X-ray. Further confirming HPRT presence, the gold weight percentage of the sample increased significantly when NCI-H460 cells were exposed to HPRT antibody (P=0.012) in comparison with isotype controls. Our results show that HPRT is localized on the

  10. Molecular mode of action and role of TP53 in the sensitivity to the novel epothilone sagopilone (ZK-EPO in A549 non-small cell lung cancer cells.

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    Sebastian Winsel

    Full Text Available Sagopilone, an optimized fully synthetic epothilone, is a microtubule-stabilizing compound that has shown high in vitro and in vivo activity against a broad range of human tumor models. We analyzed the differential mechanism of action of sagopilone in non-small cell lung cancer cell lines in vitro. Sagopilone inhibited proliferation of non-small cell lung cancer cell lines at lower nanomolar concentration. The treatment with sagopilone caused strong disturbances of cellular cytoskeletal organization. Two concentration-dependent phenotypes were observed. At 2.5 nM sagopilone or 4 nM paclitaxel an aneuploid phenotype occur whereas a mitotic arrest phenotype was induced by 40 nM sagopilone or paclitaxel. Interestingly, treatment with 2.5 nM of sagopilone effectively inhibited cell proliferation, but--compared to high concentrations (40 nM--only marginally induced apoptosis. Treatment with a high versus a low concentration of sagopilone or paclitaxel regulates a non-overlapping set of genes, indicating that both phenotypes substantially differ from each other. Genes involved in G2/M phase transition and the spindle assembly checkpoint, like Cyclin B1 and BUBR1 were upregulated by treatment with 40 nM sagopilone. Unexpectedly, also genes involved in DNA damage response were upregulated under that treatment. In contrast, treatment of A549 cells with a low concentration of sagopilone revealed an upregulation of direct transcriptional target genes of TP53, like CDKN1A, MDM2, GADD45A, FAS. Knockdown of TP53, which inhibited the transcriptional induction of TP53 target genes, led to a significant increase in apoptosis induction in A549 cells when treated with a low concentration of sagopilone. The results indicate that activation of TP53 and its downstream effectors like CDKN1A by low concentrations of sagopilone is responsible for the relative apoptosis resistance of A549 cells and might represent a mechanism of resistance to sagopilone.

  11. Studies on cytotoxic constituents from the leaves of Elaeagnus oldhamii Maxim. in non-small cell lung cancer A549 cells.

    Science.gov (United States)

    Liao, Chi-Ren; Kuo, Yueh-Hsiung; Ho, Yu-Ling; Wang, Ching-Ying; Yang, Chang-Syun; Lin, Cheng-Wen; Chang, Yuan-Shiun

    2014-07-04

    Elaeagnus oldhamii Maxim. is a commonly used traditional herbal medicine. In Taiwan the leaves of E. oldhamii Maxim. are mainly used for treating lung disorders. Twenty five compounds were isolated from the leaves of E. oldhamii Maxim. in the present study. These included oleanolic acid (1), 3-O-(Z)-coumaroyl oleanolic acid (2), 3-O-(E)-coumaroyl oleanolic acid (3), 3-O-caffeoyl oleanolic acid (4), ursolic acid (5), 3-O-(Z)-coumaroyl ursolic acid (6), 3-O-(E)-coumaroyl ursolic acid (7), 3-O-caffeoyl ursolic acid (8), 3β, 13β-dihydroxyolean-11-en-28-oic acid (9), 3β, 13β-dihydroxyurs-11-en-28-oic acid (10), uvaol (11), betulin (12), lupeol (13), kaempferol (14), aromadendrin (15), epigallocatechin (16), cis-tiliroside (17), trans-tiliroside (18), isoamericanol B (19), trans-p-coumaric acid (20), protocatechuic acid (21), salicylic acid (22), trans-ferulic acid (23), syringic acid (24) and 3-O-methylgallic acid (25). Of the 25 isolated compounds, 21 compounds were identified for the first time in E. oldhamii Maxim. These included compounds 1, 4, 5 and 8-25. These 25 compounds were evaluated for their inhibitory activity against the growth of non-small cell lung cancer A549 cells by the MTT assay, and the corresponding structure-activity relationships were discussed. Among these 25 compounds, compound 6 displayed the best activity against the A549 cell line in vitro (CC50=8.56±0.57 μg/mL, at 48 h of MTT asssay). Furthermore, compound 2, 4, 8 and 18 exhibited in vitro cytotoxicity against the A549 cell line with the CC50 values of less than 20 μg/mL at 48 h of MTT asssay. These five compounds 2, 4, 6, 8 and 18 exhibited better cytotoxic activity compared with cisplatin (positive control, CC50 value of 14.87±1.94 μg/mL, at 48 h of MTT asssay). The result suggested that the five compounds might be responsible for its clinical anti-lung cancer effect.

  12. Studies on Cytotoxic Constituents from the Leaves of Elaeagnus oldhamii Maxim. in Non-Small Cell Lung Cancer A549 Cells

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    Chi-Ren Liao

    2014-07-01

    Full Text Available Elaeagnus oldhamii Maxim. is a commonly used traditional herbal medicine. In Taiwan the leaves of E. oldhamii Maxim. are mainly used for treating lung disorders. Twenty five compounds were isolated from the leaves of E. oldhamii Maxim. in the present study. These included oleanolic acid (1, 3-O-(Z-coumaroyl oleanolic acid (2, 3-O-(E-coumaroyl oleanolic acid (3, 3-O-caffeoyl oleanolic acid (4, ursolic acid (5, 3-O-(Z-coumaroyl ursolic acid (6, 3-O-(E-coumaroyl ursolic acid (7, 3-O-caffeoyl ursolic acid (8, 3β, 13β-dihydroxyolean-11-en-28-oic acid (9, 3β, 13β-dihydroxyurs-11-en-28-oic acid (10, uvaol (11, betulin (12, lupeol (13, kaempferol (14, aromadendrin (15, epigallocatechin (16, cis-tiliroside (17, trans-tiliroside (18, isoamericanol B (19, trans-p-coumaric acid (20, protocatechuic acid (21, salicylic acid (22, trans-ferulic acid (23, syringic acid (24 and 3-O-methylgallic acid (25. Of the 25 isolated compounds, 21 compounds were identified for the first time in E. oldhamii Maxim. These included compounds 1, 4, 5 and 8–25. These 25 compounds were evaluated for their inhibitory activity against the growth of non-small cell lung cancer A549 cells by the MTT assay, and the corresponding structure-activity relationships were discussed. Among these 25 compounds, compound 6 displayed the best activity against the A549 cell line in vitro (CC50 = 8.56 ± 0.57 μg/mL, at 48 h of MTT asssay. Furthermore, compound 2, 4, 8 and 18 exhibited in vitro cytotoxicity against the A549 cell line with the CC50 values of less than 20 μg/mL at 48 h of MTT asssay. These five compounds 2, 4, 6, 8 and 18 exhibited better cytotoxic activity compared with cisplatin (positive control, CC50 value of 14.87 ± 1.94 μg/mL, at 48 h of MTT asssay. The result suggested that the five compounds might be responsible for its clinical anti-lung cancer effect.

  13. Cedrol induces autophagy and apoptotic cell death in A549 non-small cell lung carcinoma cells through the P13K/Akt signaling pathway, the loss of mitochondrial transmembrane potential and the generation of ROS.

    Science.gov (United States)

    Zhang, Shi-Yi; Li, Xue-Bo; Hou, Sheng-Guang; Sun, Yao; Shi, Yi-Ran; Lin, Song-Sen

    2016-07-01

    The objective of the present study was to determine the anticancer effects of cedrol in A549 human non-small cell lung cancer cells by examining the effects of cedrol on apoptosis induction, the phosphatidylinositol 3'-kinase (PI3K)/Akt signaling pathway, autophagy, reactive oxygen species (ROS) generation and mitochondrial transmembrane potential (MTP). The anticancer effects of cedrol were examined using A549 human lung carcinoma cells as an in vitro model. Cell viability was determined using MTT and lactate dehydrogenase (LDH) assays, and an inverted phase contrast microscope was used to examine the morphological changes in these cells. Cedrol‑triggered autophagy was confirmed by transmission electron microscopy (TEM) analysis of the cells, as well as by western blot analysis of microtubule-associated protein light-chain 3 (LC3)B expression. Intracellular ROS generation was measured by flow cytometry using 5-(6)-carboxy-2',7'-dichlorodihydrofluorescein diacetate (CM-DCFH2-DA) staining and MTP was measured using flow cytometry. The results demonstrated that cedrol reduced cell viability and induced cell apoptosis in a dose-dependent manner. Mechanistic evaluations indicated that cedrol induced apoptosis by reducing the MTP and by decreasing the levels of phosphorylated (p-)PI3K and p-Akt. Cedrol induced autophagy, which was confirmed by TEM analysis, by increasing intracellular ROS formation in a concentration-dependent manner, which was almost completely reversed by N-acetyl-L-cysteine (NAC) and tocopherol. Taken together, these findings reveal that cedrol inhibits cell proliferation and induces apoptosis in A549 cells through mitochondrial and PI3K/Akt signaling pathways. Our findings also reveal that cedrol induced pro-death autophagy by increasing intracellular ROS production.

  14. Empirical studies about quercetin increasing chemosensitivity on human lung adenocarcinoma cell line A549

    Institute of Scientific and Technical Information of China (English)

    Xuejun Zhan; Runxiang Zhang; Yanping Xu; Shuhua Yang; Daze Xie; Liwei Tan

    2012-01-01

    Objective: The present study was designed to investigate whether quercetin exerts increasing chemosensitivity on human lung adenocarcinoma cells when quercetin combined with cisplatin (DDP) and vincristine (VCR) in vitro respectively and its possible antitumor mechanism. To provide experimental proof for clinical combination application. Methods: Using intermittent administration of high dose VCR, human lung adenocarcinoma sensitive cell line (A549/S) was induced to VCR-resistant human lung adenocarcinoma cell line (A549/VCR). MTT assay was adapted for examing the 50% inhibition (IC50) value of DDP and VCR on A549/S and A549/VCR when quercetin combined with DDP and VCR respectively. Results: IC50 of DDP on A549/S and A549/VCR was 10.18 and 12.35 mg/L, and the IC50 of VCR on the two cell lines was 1.21 and 12.77 mg/L, respectively. The resistance fold of A549/VCR on VCR and DDP was 10.55 and 1.21, respectively. When quercetin at concentration of 50, 100 and 200 μmol/L in combination with DDP and VCR respectively, the IC50 of DDP and VCR on A549/S and A549/VCR were obvious decreased (P < 0.05 – P < 0.01). Conclusion: The experiment results suggested that quercetin could increase the chemosensitivity and partly revise the resistance of A549/VCR.

  15. Human decorin regulates proliferation and migration of human lung cancer A549 cells

    Institute of Scientific and Technical Information of China (English)

    LIANG Shuo; XU Jin-fu; CAO Wei-jun; LI Hui-ping; HU Cheng-ping

    2013-01-01

    Background Decorin is a small leucine-rich proteoglycan and it plays an important role in regulation of cell growth and migration in various tumor cell lines.Decorin was found down-regulated in non-small cell lung cancer tissue and may be involved in regulation of lung cancer development.Methods In this study,lentivirus-mediated RNA interference and over expression were employed to change the expression levels of decorin in lung cancer A549 cells.We tested the cell cycle of A549 cells and the expression of transforming growth factor (TGF)-β1,cyclin D1,epidermal growth factor receptor (EGFR),P53,and P21.Results We found that up-regulation of decorin could inhibit proliferation,block cell cycle at G1 and decrease invasive activity of A549 cells.Moreover,we also show that up-regulation of decorin induced significant decreases of TGF-β1,cyclin D1 expression,phosphorylation of EGFR,and increases of P53 and P21 expression.Opposite results were observed in A549 cells with down-regulation of decorin.Conclusion Our results suggest that decorin is a key regulator involved in proliferation and migration ofA549 cells.

  16. Inhibiting Effect and Its Mechanism of Ibandronate on the Proliferation of Humanized NSCLC A549 Cells in Vitro

    Institute of Scientific and Technical Information of China (English)

    YAO Qiang; HUA Dong

    2014-01-01

    Objective:To explore the effect of ibandronate on the proliferation and the expression of human telomerase reverse transcriptase (hTERT) of non-small cell lung cancer (NSCLC) A549 cell line in vitro. Methods: Methyl thiazolyl tetrazolium (MTT) assay, microscope, flow cytometry (FCM) and semi-quantitative RT-PCR were employed to detect the cell proliferation, cell cycle as well as the morphological change and the expression of hTERT mRNA of A549 cell line. Results:The data showed that ibandronate could effectively inhibit the proliferation of A549 cell line in time-and concentration-dependent. Under the microscope, the lfoating cells increased gradually as the drug concentration increasing. FCM detection showed that ibandronate could induce the cell cycle stopped in G0/G1 phase and downregulation expression of hTERT. Conclusion:Ibandronate can inhibit the proliferation of A549 cell line in vitro, whose mechanism may be associated with cell cycle arrestted in phase G0/G1 and downregulation expression of hTERT.

  17. Coptisine-induced cell cycle arrest at G2/M phase and reactive oxygen species-dependent mitochondria-mediated apoptosis in non-small-cell lung cancer A549 cells.

    Science.gov (United States)

    Rao, Poorna Chandra; Begum, Sajeli; Sahai, Mahendra; Sriram, D Saketh

    2017-03-01

    This study aimed to explore the effect of coptisine on non-small-cell lung cancer and its mechanism through various in vitro cellular models (A549). Results claimed significant inhibition of proliferation by coptisine against A549, H460, and H2170 cells with IC50 values of 18.09, 29.50, and 21.60 µM, respectively. Also, coptisine exhibited upregulation of pH2AX, cell cycle arrest at G2/M phase, and downregulation of the expression of cyclin B1, cdc2, and cdc25C and upregulation of p21 dose dependently. Furthermore, induction of apoptosis in A549 cells by coptisine was characterized by the activation of caspase 9, caspase 8, and caspase 3, and cleavage of poly adenosine diphosphate ribose polymerase. In addition, coptisine was found to increase reactive oxygen species generation, upregulate Bax/Bcl-2 ratio, disrupt mitochondrial membrane potential, and cause cytochrome c release into the cytosol. Besides, treatment with a reactive oxygen species inhibitor (N-acetyl cysteine) abrogated coptisine-induced growth inhibition, apoptosis, reactive oxygen species generation, and mitochondrial dysfunction. Thus, the mediation of reactive oxygen species in the apoptosis-induced effect of coptisine in A549 cells was corroborated. These findings have offered new insights into the effect and mechanisms of action of coptisine against non-small-cell lung cancer.

  18. [SIRT1 Influences the Sensitivity of A549 Non-small Cell Lung Cancer Cell Line to 
Cisplatin via Modulating the Noxa Expression].

    Science.gov (United States)

    Cao, Bin; He, Xiaofeng; Wang, Wengong; Shi, Minke

    2016-02-01

    背景与目的 非小细胞肺癌的顺铂耐药是常见的临床现象,严重制约了患者的化疗效果,是亟待解决的问题。SIRT1和Noxa的表达变化影响肿瘤细胞对化疗药物的敏感性。本研究旨在研究SIRT1表达对非小细胞肺癌对顺铂的敏感性的影响,并探讨其涉及Noxa表达的机制,以求为提高非小细胞肺癌细胞对顺铂敏感性提供希望。方法 利用实时荧光定量PCR和Western blot分析A549细胞及顺铂耐药的A549/DDP细胞SIRT1及Noxa mRNA和蛋白水平的表达差异。利用siRNA干扰技术抑制A549/DDP细胞的SIRT1表达,进而使用Cell Titer Blue试验、流式细胞术从细胞增殖、细胞周期和细胞凋亡方面分析SIRT1沉默对A549/DPP细胞顺铂敏感性的影响。同时利用实时荧光定量PCR和Western blot分析SIRT1抑制对A549/DPP细胞Noxa表达的影响。结果 A549细胞和A549/DDP细胞对顺铂的敏感性有显著差异,与A549细胞相比,A549/DDP细胞的SIRT1表达较高,但Noxa表达较低。使用siRNA抑制A549/DPP细胞的SIRT1表达后,与未抑制SIRT1细胞相比,4 μg/mL顺铂处理后的细胞存活率降低,G2期/M期阻滞比例增加,凋亡率提高。同时,SIRT1沉默导致A549/DPP细胞的Noxa表达增加。结论 较高的SIRT1可能引起A549细胞对顺铂的耐药性,抑制SIRT1可以提高A549/DDP细胞对顺铂的敏感性,其机制可能涉及SIRT1对Noxa的调节。.

  19. Cytotoxicity of withasteroids: withametelin induces cell cycle arrest at G2/M phase and mitochondria-mediated apoptosis in non-small cell lung cancer A549 cells.

    Science.gov (United States)

    Rao, Poorna Chandra; Begum, Sajeli; Jahromi, Mohammad Ali Farboodniay; Jahromi, Zahra Hosseini; Sriram, Saketh; Sahai, Mahendra

    2016-09-01

    Considerable interest has been gained by withasteroids because of their structural uniqueness and wide spectrum of biological activities. However, limited systematic studies for proving their cytotoxic potential have so far been reported. Hence, an attempt was made to test the cytotoxicity of six withasteroids viz., withametelin (WM), withaphysalin D, withaphysalin E, 12-deoxywithastramonolide, Withaperuvin B, and physalolactone against A549, HT-29, and MDA-MB-231 cancer cell lines. Significant cytotoxic effect of WM against A549 cells (IC50 value of 6.0 μM), MDA-MB-231 cells (IC50 value of 7.6 μM), and HT-29 cells (IC50 value of 8.2 μM) was observed. Withaperuvin B and physalolactone were found to be effective against MDA-MB-231 cells. The significantly active WM arrested the A549 cells at G2/M phase and downregulated the expression of G2/M regulatory proteins such as cdc2, cyclin B1, and cdc25C. Apoptosis induced by WM in A549 cells was associated with the generation of ROS and depletion of MMP. Furthermore, WM treatment resulted in Bax upregulation, Bcl-2 downregulation, translocation of cytochrome c to mitochondria, activation of caspase-9 and -3, and PARP cleavage corroborating the apoptosis induction through intrinsic apoptotic pathway. Thus, WM possessing broader cytotoxic effect is a promising lead molecule which has the potential to be developed as a new therapeutic agent for NSCLC.

  20. 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

    Energy Technology Data Exchange (ETDEWEB)

    Deng, Xuefeng, E-mail: dengxfdoctor@hotmail.com [Department of Thoracic Surgery, Tangdu Hospital, Forth Military Medical University (China); Department of Cardio-thoracic Surgery, Affiliated Hospital of Academy of Military Medical Sciences (China); Ma, Qunfeng [Department of Cardio-thoracic Surgery, Affiliated Hospital of Academy of Military Medical Sciences (China); Zhang, Bo [Department of Pathology, Affiliated Hospital of Academy of Military Medical Sciences (China); Jiang, Hong [College of Life Sciences and Bioengineering, Beijing Jiaotong University (China); Zhang, Zhipei; Wang, Yunjie [Department of Thoracic Surgery, Tangdu Hospital, Forth Military Medical University (China)

    2013-10-15

    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 of 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.

  1. Trichomonas vaginalis induces cytopathic effect on human lung alveolar basal carcinoma epithelial cell line A549.

    Science.gov (United States)

    Salvador-Membreve, Daile Meek C; Jacinto, Sonia D; Rivera, Windell L

    2014-12-01

    Trichomonas vaginalis, the causative agent of trichomoniasis is generally known to inhabit the genitourinary tract. However, several case reports with supporting molecular and immunological identifications have documented its occurrence in the respiratory tract of neonates and adults. In addition, the reports have documented that its occurrence is associated with respiratory failures. The medical significance or consequence of this association is unclear. Thus, to establish the possible outcome from the interaction of T. vaginalis with lung cells, the cytopathic effects of the parasites were evaluated using monolayer cultures of the human lung alveolar basal carcinoma epithelial cell line A549. The possible effect of association of T. vaginalis with A549 epithelial cells was analyzed using phase-contrast, scanning electron microscopy and fluorescence microscopy. MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide), crystal-violet and TUNEL (terminal deoxynucleotidyltransferase-mediated dUTP nick-end labelling) assays were conducted for cytotoxicity testing. The results demonstrate that T. vaginalis: (1) adheres to A549 epithelial cells, suggesting a density-dependent parasite-cell association; (2) adherence on A549 is through flagella, membrane and axostyle; (3) causes cell detachment and cytotoxicity (50-72.4%) to A549 and this effect is a function of parasite density; and (4) induces apoptosis in A549 about 20% after 6 h of incubation. These observations indicate that T. vaginalis causes cytopathic effects on A549 cell. To date, this is the first report showing a possible interaction of T. vaginalis with the lung cells using A549 monolayer cultures. Further studies are recommended to completely elucidate this association.

  2. Transcription Activity of Ectogenic Human Carcinoembryonic Antigen Promoter in Lung Adenocarcinoma Cells A549

    Institute of Scientific and Technical Information of China (English)

    XIONG Weining; FANG Huijuan; XU Yongjian; XIONG Shendao; CAO Yong; SONG Qingfeng; ZENG Daxiong; ZHANG Huilan

    2006-01-01

    The transcription activity of ectogenic human carcinoembryonic antigen (CEA) promoter in lung adenocarcinoma cells A549 was investigated for the further gene-targeting therapy. The reporter gene green fluorescent protein (GFP) driven by CEA promoter and human cytomegalovirus (CMV) promoter were relatively constructed and named plasmid pCEA-EGFP and pCMV-GFP respectively. The intensity of fluorescence was detected by fluorescence microscope and flow cytometry analysis after the pCEA-GFP and pSNAV-GFP plasmids were transfected into A549 cells through liposome respectively. The results showed (4.08±0.63) % of the A549 cells transfected with pCEA-AFP plasmid expressed, significantly lower than that of the A549 cells transfected with pCMV-GFP [(43.27±3.54) %]. It was suggested that ectogenic human CEA promoter in lung adenocarcinoma cells A549 was weakly expressed. The distinct specificity of CEA promoter in CEA high expression cells was regarded as a tool in selective gene therapy, but the transcription activity of ectogenic human CEA promoter was needed to increase in the future.

  3. Integrin α11 regulates IGF2 expression in fibroblasts to enhance tumorigenicity of human non-small-cell lung cancer cells

    Science.gov (United States)

    Zhu, Chang-Qi; Popova, Svetlana N.; Brown, Ewan R. S.; Barsyte-Lovejoy, Dalia; Navab, Roya; Shih, Warren; Li, Ming; Lu, Ming; Jurisica, Igor; Penn, Linda Z.; Gullberg, Donald; Tsao, Ming-Sound

    2007-01-01

    Integrin α11 (ITGA11/α11) is localized to stromal fibroblasts and commonly overexpressed in non-small-cell lung carcinoma (NSCLC). We hypothesized that stromal α11 could be important for the tumorigenicity of NSCLC cells. SV40 immortalized mouse embryonic fibroblasts established from wild-type (WT) and Itga11-deficient [knockout (KO)] mice were tested for their tumorigenicity in immune-deficient mice when implanted alone or coimplanted with the A549 human lung adenocarcinoma cells. A549 coimplanted with the fibroblasts showed a markedly enhanced tumor growth rate compared with A549, WT, or KO, which alone formed only small tumors. Importantly, the growth was significantly greater for A549+WT compared with A549+KO tumors. Reexpression of human α11 cDNA in KO cells rescued a tumor growth rate to that comparable with the A549+WT tumors. These findings were validated in two other NSCLC cell lines, NCI-H460 and NCI-H520. Gene expression profiling indicated that IGF2 mRNA expression level was >200 times lower in A549+KO compared with A549+WT tumors. Stable short-hairpin RNA (shRNA) down-regulation of IGF2 in WT (WTshIGF2) fibroblasts resulted in a decreased growth rate of A549+WTshIGF2, compared with A549+WT tumors. The results indicate that α11 is an important stromal factor in NSCLC and propose a paradigm for carcinoma–stromal interaction indirectly through interaction between the matrix collagen and stromal fibroblasts to stimulate cancer cell growth. PMID:17600088

  4. Human lung epithelial cell A549 proteome data after treatment with titanium dioxide and carbon black.

    Science.gov (United States)

    Vuong, Ngoc Q; Goegan, Patrick; Mohottalage, Susantha; Breznan, Dalibor; Ariganello, Marianne; Williams, Andrew; Elisma, Fred; Karthikeyan, Subramanian; Vincent, Renaud; Kumarathasan, Premkumari

    2016-09-01

    Here, we have described the dataset relevant to the A549 cellular proteome changes after exposure to either titanium dioxide or carbon black particles as compared to the non-exposed controls, "Proteomic changes in human lung epithelial cells (A549) in response to carbon black and titanium dioxide exposures" (Vuong et al., 2016) [1]. Detailed methodologies on the separation of cellular proteins by 2D-GE and the subsequent mass spectrometry analyses using MALDI-TOF-TOF-MS are documented. Particle exposure-specific protein expression changes were measured via 2D-GE spot volume analysis. Protein identification was done by querying mass spectrometry data against SwissProt and RefSeq protein databases using Mascot search engine. Two-way ANOVA analysis data provided information on statistically significant A549 protein expression changes associated with particle exposures.

  5. Human lung epithelial cell A549 proteome data after treatment with titanium dioxide and carbon black

    Directory of Open Access Journals (Sweden)

    Ngoc Q. Vuong

    2016-09-01

    Full Text Available Here, we have described the dataset relevant to the A549 cellular proteome changes after exposure to either titanium dioxide or carbon black particles as compared to the non-exposed controls, “Proteomic changes in human lung epithelial cells (A549 in response to carbon black and titanium dioxide exposures” (Vuong et al., 2016 [1]. Detailed methodologies on the separation of cellular proteins by 2D-GE and the subsequent mass spectrometry analyses using MALDI-TOF-TOF-MS are documented. Particle exposure-specific protein expression changes were measured via 2D-GE spot volume analysis. Protein identification was done by querying mass spectrometry data against SwissProt and RefSeq protein databases using Mascot search engine. Two-way ANOVA analysis data provided information on statistically significant A549 protein expression changes associated with particle exposures.

  6. β-Elemonic acid inhibits the cell proliferation of human lung adenocarcinoma A549 cells: The role of MAPK, ROS activation and glutathione depletion.

    Science.gov (United States)

    Wu, Tsu-Tuan; Lu, Chien-Lin; Lin, Hen-I; Chen, Bing-Fang; Jow, Guey-Mei

    2016-01-01

    β-elemonic acid, a known triterpene, exhibits anti-inflammatory effects, yet research on the pharmacological effects of β-elemonic acid is rare. We investigated the anticancer effects and the related molecular mechanisms of β-elemonic acid on human non-small cell lung cancer (NSCLC) A549 cells. The effects of β-elemonic acid on the growth of A549 cells were studied using a 3-(4,5)-2,5-diphenyltetrazolium bromide (MTT) assay. Apoptosis was detected using Annexin V staining. The effect of β-elemonic acid on the cell cycle of A549 cells was assessed using the propidium iodide method. The change in reactive oxygen species (ROS) was detected using a dichlorodihydrofluorescein diacetate (DCFH-DA) assay with microscopic examination. The expression levels of Bcl-2 family proteins, mitogen-activated protein kinase (MAPK) family proteins and cyclooxygenase 2 (COX-2) were detected using western blot analysis. Our data revealed that β-elemonic acid strongly induced human A549 lung cancer cell death in a dose- and time-dependent manner as determined by the MTT assay. β-elemonic acid-induced cell death was considered to be apoptotic when the phosphatidylserine exposure was observed using Annexin V staining. The death of human A549 lung cancer cells was caused by apoptosis induced by activation of ROS activity, increase in the sub-G1 proportion, downregulation of Bcl-2 expression, upregulation of Bax expression and inhibition of the MAPK signaling pathways. These results clearly demonstrated that β-elemonic acid inhibits proliferation by inducing hypoploid cells and cell apoptosis. Moreover, the anticancer effects of β-elemonic acid were related to the MAPK signaling pathway, ROS activation and glutathione depletion in human A549 lung cancer cells.

  7. Oleanolic acid-induced apoptosis and its relation with intracellular calcium in human lung adenocarcinoma A549 cells

    Institute of Scientific and Technical Information of China (English)

    Asmitanand; Thakur

    2010-01-01

    Objective To investigate the effect of oleanolic acid (OA) on apoptosis,correlation between apoptosis and intracellular calcium,and its mechanism in human lung adenocarcinoma cell line A549. Methods Human lung adenocarcinoma A549 cells were incubated in vitro and assigned with OA concentrations of 0,10,20 and 40μg/mL. The apoptosis status of A549 cell line was detected with Annexin V-FITC/PI by flow cytometry (FCM); fluorescence intensity (FI) of A549 cells was assessed and the level of intracellular calciu...

  8. Aression of TLR9 in human pulmonary adenocarcinoma cell line A549%TLR9在人肺腺癌细胞A549中的表达

    Institute of Scientific and Technical Information of China (English)

    Jun Yu; Tiecheng Pan; Xiang Wei; Ligang Liu; Min Hu; Fang Yuan; Jiaduo Li

    2009-01-01

    Objective: Being considered as a bridge between the innate immunity and acquired immunity, Toll-like receptors (TRLs) are very important innate immunity moleculars. Recent researchs on the innate immunity have focused on the relation- ship between TLRs and human tumor. This paper investgated the expression and significance of TLR9 in human pulmonary adenocarcinoma cell (A549 cell) and human bronchial epithelial cell (HBE cell). Methods: After culturing A549 cell and HBE cell in vitro, the expression of TLR9 mRNA and protein in both cells were detected by immunocytochemistry, Real-time Quantitative Reverse Transcriptase-Polymerase Chain Reaction (Real-time Quantitative PCR) and Western blot, respectively. Results: By immunocytochemistry staining, TLR9 was mainly expressed in both cells' cell membrane and endochylema as brown-yellow material. It showed that the expressions of TLR9 mRNA and protein in A549 cell were stronger than those in HBE cell (P < 0.01). Conclusion: The results suggest TLR9 might cause the progression of human pulmonary adenocaroinoma, and the mechanism needs to be further investgatied.

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

    Energy Technology Data Exchange (ETDEWEB)

    Huang, Chunrong; Zheng, Haichong; He, Wanmei; Lu, Guifang; Li, Xia [Department of Medical Intensive Care Unit, The First Affiliated Hospital, Sun Yat-Sen University, Guangzhou 510080 (China); Deng, Yubin, E-mail: dengyub@mail.sysu.edu.cn [Research Center of Translational Medicine, The First Affiliated Hospital, Sun Yat-Sen University, Guangzhou 510080 (China); Zeng, Mian, E-mail: zengmian2004@163.com [Department of Medical Intensive Care Unit, The First Affiliated Hospital, Sun Yat-Sen University, Guangzhou 510080 (China)

    2016-05-20

    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 activated 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.

  10. EM011 activates a survivin-dependent apoptotic program in human non-small cell lung cancer cells

    Directory of Open Access Journals (Sweden)

    Yates Clayton

    2009-10-01

    Full Text Available Abstract Background Lung cancer remains a leading cause of cancer death among both men and women in the United States. Treatment modalities available for this malignancy are inadequate and thus new drugs with improved pharmacological profiles and superior therapeutic indices are being continually explored. Noscapinoids constitute an emerging class of anticancer agents that bind tubulin but do not significantly alter the monomer/polymer ratio of tubulin. EM011, a rationally-designed member of this class of non-toxic agents, is more potent than the lead molecule, noscapine. Results Here we report that EM011 inhibited proliferation of a comprehensive panel of lung cancer cells with IC50's ranging from 4-50 μM. In A549 human non-small cell lung cancer cells, the antiproliferative activity was mediated through blockage of cell-cycle progression by induction of a transient but robust mitotic arrest accompanied by activation of the spindle assembly checkpoint. The mitotically-arrested A549 cells then override the activated mitotic checkpoint and aberrantly exit mitosis without cytokinesis resulting in pseudo G1-like multinucleated cells that either succumb directly to apoptosis or continue another round of the cell-cycle. The accumulated enormous DNA perhaps acts as genotoxic stress to trigger cell death. EM011-induced apoptotic cell death in A549 cells was associated with a decrease of the Bcl2/BAX ratio, activation of caspase-3 and cleavage of PARP. Furthermore, EM011 induced downregulation of survivin expression over time of treatment. Abrogation of survivin led to an increase of cell death whereas, overexpression caused decreased apoptosis. Conclusion These in vitro data suggest that EM011 mediates antiproliferative and proapoptotic activity in non-small cell A549 lung cancer cells by impeding cell-cycle progression and attenuating antiapoptotic signaling circuitries (viz. Bcl2, survivin. The study provides evidence for the potential usefulness of

  11. Anti-tumor activity of CrTX in human lung adenocarcinoma cell line A549

    Institute of Scientific and Technical Information of China (English)

    Bin YE; Yan XIE; Zheng-hong QIN; Jun-chao WU; Rong HAN; Jing-kang HE

    2011-01-01

    Aim:To assess the cytotoxic effect of crotoxin (CrTX),a potent neurotoxin extracted from the venom of the pit viper Crotalus durissus terrificus,in human lung adenocarcinoma A549 cells and investigated the underlying mechanisms.Methods:A549 cells were treated with gradient concentrations of CrTX,and the cell cycle and apoptosis were analyzed using a flow cytometric assay.The changes of cellular effectors p53,caspase-3 and cleaved caspase-3,total P38MAPK and pP38MAPK were investigated using Western blot assays.A549 xenograft model was used to examine the inhibition of CrTX on tumor growth in vivo.Results:Treatment of A549 cells with CrTX (25-200 μg/mL) for 48 h significantly inhibited the cell growth in a dose-dependent manner (IC50=78 μg/mL).Treatment with CrTX (25 iJg/mL) for 24 h caused G1 arrest and induced cell apoptosis.CrTX (25 μg/mL) significantly increased the expression of wt p53,cleaved caspase-3 and phospho-P38MAPK.Pretreatment with the specific P38MAPK inhibitor SB203580 (5 μmol/L) significantly reduced CrTX-induced apoptosis and cleaved caspase-3 level,but G1 arrest remained unchanged and highly expressed p53 sustained.Intraperitoneal injection of CrTX (10 μg/kg,twice a week for 4 weeks) significantly inhibited A549 tumor xenograft growth,and decreased MVD and VEGF levels.Conclusion:CrTX produced significant anti-tumor effects by inducing cell apoptosis probably due to activation of P38MAPK and caspase-3,and by cell cycle arrest mediated by increased wt p53 expression.In addition,CrTX displayed anti-angiogenic effects in vivo.

  12. Teroxirone inhibited growth of human non-small cell lung cancer cells by activating p53

    Energy Technology Data Exchange (ETDEWEB)

    Wang, Jing-Ping; Lin, Kai-Han; Liu, Chun-Yen; Yu, Ya-Chu; Wu, Pei-Tsun [Department of Life Science, National Taiwan Normal University, Taipei, Taiwan (China); Chiu, Chien-Chih [Department of Biotechnology, Kaohsiung Medical University, Kaohsiung, Taiwan (China); Su, Chun-Li [Department of Human Development and Family Studies, National Taiwan Normal University, Taipei, Taiwan (China); Chen, Kwun-Min [Department of Chemistry, National Taiwan Normal University, Taipei, Taiwan (China); Fang, Kang, E-mail: kangfang@ntnu.edu.tw [Department of Life Science, National Taiwan Normal University, Taipei, Taiwan (China)

    2013-11-15

    In this work, we demonstrated that the growth of human non-small-cell-lung-cancer cells H460 and A549 cells can be inhibited by low concentrations of an epoxide derivative, teroxirone, in both in vitro and in vivo models. The cytotoxicity was mediated by apoptotic cell death through DNA damage. The onset of ultimate apoptosis is dependent on the status of p53. Teroxirone caused transient elevation of p53 that activates downstream p21 and procaspase-3 cleavage. The presence of caspase-3 inhibitor reverted apoptotic phenotype. Furthermore, we showed the cytotoxicity of teroxirone in H1299 cells with stable ectopic expression of p53, but not those of mutant p53. A siRNA-mediated knockdown of p53 expression attenuated drug sensitivity. The in vivo experiments demonstrated that teroxirone suppressed growth of xenograft tumors in nude mice. Being a potential therapeutic agent by restraining cell growth through apoptotic death at low concentrations, teroxirone provides a feasible perspective in reversing tumorigenic phenotype of human lung cancer cells. - Highlights: • Teroxirone repressed tumor cell growth in nude mice of human lung cancer cells. • The apoptotic cell death reverted by caspase-3 inhibitor is related to p53 status. • Teroxirone provides a good candidate for lung cancer treatment.

  13. 人肺腺癌细胞株A549中HIF-1α对Survivin的表达调控%Regulation of survivin expression by hypoxia-inducible factor-1α in non-small cell lung cancer

    Institute of Scientific and Technical Information of China (English)

    李伟; 陈余清; 孙艳; 赵成岭; 王效静

    2011-01-01

    Background and purpose: Survivin gene is a unique member of the inhibitor of apoptosis protein (LAP) family. It plays an important role, not only in regulating mitosis but also in inhibiting apoptosis. It is highly expressed in almost all types of human tumors and fetal tissues but rarely detectable in normal adult tissues. High levels of survivin expression have been associated with tumor progression, resistance to radiation and drug treatments and poor survival rates in cancer patients. The current literature contains few reports on the transcriptional regulation of survivin expression in lung cancer. Previous studies have found that there are also 2 putative binding sites for hypoxia-inducible factor- la(HIF- la) in the core promoter region of survivin gene. Survivin promoter-luciferase reporter vectors Pgl3-SVP230-luc have been constructed early. The purpose of this study was to investigate the mechanism of (HIF-la)on transcriptional regulation of survivin in A549 cells by hypoxia. Methods: (l)Double labeling immunofluorescence method was used to detect co-expression of survivin/HIF-lα protein; (2)RT-polymerase chain reaction (RT-PCR) and Western blot was used to examine the level of survivin Mrna and protein in A549 cells transfected by HIF-lα expression plasmid and HIF-lα siRNA; (3)Luciferase activity was detected in A549 cells following cotransfection with Pgl3-SVP230-luc as well as HIF-la expression plasmid or HIF-lα siRNA to value the transcriptional activity of survivin. (4)Electrophoretic mobility shift assay (EMS A) was performed to test the nuclear extract of the A549 cells binding to the r-32P labeled probes containing survivin promoter squences. Results: (l)Survivin/HIF-lα proteins co-expressed in A549 cell; (2)Compared with control groups, the level of survivin Mrna and protein is markedly increased in A549 cells transfected with HIF-lα expression plasmid, but decreased in the HIF-lα siRNA group(P<0.01); (3)The relative activity of Pgl3-SVP

  14. Melatonin inhibits the migration of human lung adenocarcinoma A549 cell lines involving JNK/MAPK pathway.

    Directory of Open Access Journals (Sweden)

    Qiaoyun Zhou

    Full Text Available OBJECTIVE: Melatonin, an indolamine produced and secreted predominately by the pineal gland, exhibits a variety of physiological functions, possesses antioxidant and antitumor properties. But, the mechanisms for the anti-cancer effects are unknown. The present study explored the effects of melatonin on the migration of human lung adenocarcinoma A549 cells and its mechanism. METHODS: MTT assay was employed to measure the viability of A549 cells treated with different concentrations of melatonin. The effect of melatonin on the migration of A549 cells was analyzed by wound healing assay. Occludin location was observed by immunofluorescence. The expression of occludin, osteopontin (OPN, myosin light chain kinase (MLCK and phosphorylation of myosin light chain (MLC, JNK were detected by western blots. RESULTS: After A549 cells were treated with melatonin, the viability and migration of the cells were inhibited significantly. The relative migration rate of A549 cells treated with melatonin was only about 20% at 24 h. The expression level of OPN, MLCK and phosphorylation of MLC of A549 cells were reduced, while the expression of occludin was conversely elevated, and occludin located on the cell surface was obviously increased. The phosphorylation status of JNK in A549 cells was also reduced when cells were treated by melatonin. CONCLUSIONS: Melatonin significantly inhibits the migration of A549 cells, and this may be associated with the down-regulation of the expression of OPN, MLCK, phosphorylation of MLC, and up-regulation of the expression of occludin involving JNK/MAPK pathway.

  15. Antitumor activity of paederosidic acid in human non-small cell lung cancer cells via inducing mitochondria-mediated apoptosis.

    Science.gov (United States)

    Yu, Ping; Shi, Lifeng; Song, Meiyan; Meng, Yu

    2017-05-01

    This study was aimed to investigate antitumor activity of paederosidic acid (PA) in human non-small cell lung cancer cells and explore the related mechanisms. The anti-proliferative effects of PA on A549 cells were evaluated by MTT method and the IC50 values were calculated. Furthermore, the PA-induced apoptosis in A549 cells was determined by fluorescence microscope via staining with DAPI and by flow cytometer via staining with FITC conjugated Annexin V/PI. The expression of apoptosis-related or signaling proteins was investigated by Western blotting. Our results demonstrated that PA showed significant anti-tumor activity on lung cancer in vitro; the mechanisms were involved in inducing mitochondria-mediated apoptosis via up-regulation of caspase-3, caspase-8, caspase-9, Bid, Bax, down-regulation of Bcl-2 and stimulating the release of Cyto-C from mitochondria. In addition, JNK phosphorylation levels significantly increased concomitantly with decrease in Akt phosphorylation after treatment with PA in A549 cells. However, JNK siRNA-transfected cells diminished PA-induced caspase-3, 8 and 9, Bid and Bax activaton while enhanced the Bcl-2 activation. Collectively, these results indicated that PA-induced JNK activation played an important functional role in apoptosis. Copyright © 2017 Elsevier B.V. All rights reserved.

  16. Combination therapy in A549 cells

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    Yuan Menghui [Department of Nuclear Medicine, Xijing Hospital, Fourth Military Medical University, Xi' an 710038 (China); Wang Jing [Department of Nuclear Medicine, Xijing Hospital, Fourth Military Medical University, Xi' an 710038 (China)], E-mail: wangjing_fmmu@yahoo.com.cn; Deng Jinglan; Wang Zhe; Yang Weidong; Li Guoquan; Ren Bingxiu [Department of Nuclear Medicine, Xijing Hospital, Fourth Military Medical University, Xi' an 710038 (China)

    2010-04-15

    Background and aim: We investigated the anti-tumor effect induced by the combination of the radiotherapeutic agent {sup 131}I-RC-160 and the prodrug 5-FC in human non-small cell lung cancer (NSCLC) A549 cells that were co-expressing the human somatostatin receptor 2 gene (hSSTR2) and E. coli cytosine deaminase gene (CD). Methods: We cloned both hSSTR2 and CD into a bicistronic mammalian expression plasmid and stably transfected it into A549 cells (pCIS-A549 cells). After antibiotic selection, SSTR expression in stable clones was determined by reverse transcription and polymerase chain reaction (RT-PCR), Western blot, flow cytometry and immunofluorescence analyses. To assess the in vivo targeting efficiency of the 'engineered' A549 cells, the cells were subcutaneously injected into nude mice and the biodistribution of {sup 99m}Tc-RC-160 was assessed at different time points. The tumor inhibitory effects of {sup 131}I-RC-160 and/or 5-FC were evaluated by measurement of tumor growth and immunohistochemical analysis. Results: Multiple analyses demonstrated the successful expression of hSSTR2 in A549 cells. In vivo radioimaging revealed specific targeting of RC-160 to the tumors derived from pCIS-A549 cells when compared to those from control A549 cells. The tumor inhibitory rate of pCIS-A549 tumors in the {sup 131}I-RC-160 plus 5-FC-treated group was significantly higher than that in the single agent-treated group, control group and control tumors. Conclusion: Co-expression of the hSSTR2 and CD genes in tumor cells can selectively sensitize these cells to the infra-additive effects of radioisotope-labeled RC-160 and 5-FC in vivo. This approach offers a potential therapeutic strategy for the treatment of lung cancer.

  17. Effect of P2X7R agonist BzATP on cell growth and apoptosis in non-small cell lung cancer A549 cells%P2X7R激动剂BzATP对非小细胞肺癌A549细胞生长和凋亡的影响

    Institute of Scientific and Technical Information of China (English)

    曾康华; 茹琴; 熊琪; 艾永循

    2016-01-01

    目的 研究配体门控离子通道P2X7受体(P2X7R)在非小细胞肺癌A549细胞中的表达,观察P2X7R激动剂2'-3'-O-(4-苯甲酰-苯甲酰)腺苷三磷酸三乙烷胺盐(BzATP)对A549细胞生长及凋亡的影响,并探究相关作用机制.方法 采用免疫荧光法检测P2X7R在A549细胞中的表达.用不同浓度的BzATP(150、300、600 μmol/L)处理,未用BzATP干预的细胞作为对照组.采用四甲基偶氮唑蓝(MTT)和Hoest33342染色法分别检测细胞存活率与凋亡情况,酶联免疫吸附试验检测上清液中肿瘤坏死因子-α(TNF-α)的浓度,Western blotting检测核转录因子-κB(NF-κB) p65、NF-κB抑制因子α(IκBα)及磷酸化NF-κB抑制因子α(phospho-IκBα)蛋白的表达.结果 P2X7R在A549细胞膜上表达.在300、600 μmol/L BzATP作用下A549细胞存活率分别为(67.87±8.98)%、(44.73±6.92)%,较对照组(98.60±1.44)%明显下降,差异均具有统计学意义(=4.481,P=0.027;t =3.920,P=0.038).BzATP可促进细胞凋亡,并且300、600 μmol/L BzATP可上调细胞培养上清中TNF-α浓度,分别为(57.35±6.41)pg/ml、(78.63±11.33) pg/ml,与对照组(42.56±0.37) pg/ml比较差异具有统计学意义(t=6.410,P=0.035;t=11.330,P=0.005).此外,BzATP可下调NF-κB p65的表达,上调IκBα的表达,对phospho-IκBα的表达无明显作用.结论 P2X7R表达于A549细胞膜,BzATP能够抑制细胞增殖,促进细胞凋亡,其作用机制可能与促进细胞中TNF-α的释放,抑制NF-κB通路有关.%Objective To investigate the expression of P2X7 receptor (P2X7R) and the effect of P2X7R agonist 2'-3'-O-(4-benzoyl-benzoyl) ethane adenosine triphosphate three amine salt (BzATP) on cell growth and apoptosis in non-small cell lung cancer A549 cells,and to explore the related mechanism.Methods The expression of P2X7R in A549 cells was detected by immunofluorescence.Cells were treated with different concentrations (150,300,600 μmol/L) of BzATP.Cells untreated with BzATP were used as control

  18. 仙人掌多糖对人肺癌A549细胞形态结构的影响%The influence of shape and structure in human lung cancer A549 cell by purification of Cactus polysaccharides

    Institute of Scientific and Technical Information of China (English)

    车加祥; 邵淑丽; 徐君懿; 陈闯; 张伟伟; 徐兴军

    2014-01-01

    以人肺癌A549细胞为研究对象,探讨仙人掌多糖组分OP1对人肺癌A549细胞生长抑制,以及对其形态、结构的影响.采用台盼蓝拒染法测定人肺癌A549细胞生长抑制曲线,通过倒置显微镜观察不同质量浓度的仙人掌多糖组分OP1作用于人肺癌A549细胞引发的形态变化.结果表明,仙人掌多糖组分OP1能抑制人肺癌A549细胞增殖,诱导人肺癌A549细胞凋亡,在一定范围内,呈时间、剂量依赖性,作用48 h的IC50为(597.55±28.97)μg/mL.经仙人掌多糖组分OP1诱导后,人肺癌A549细胞形态结构出现典型的凋亡特征.%Study to human lung cancer A549 cells as the research object,explore purification OP1 of Cactus polysaccharides on morphological structure of human lung cancer A549 cells.The inhibitory ratio of cells was measured by trypan blue stain assay.By inverted microscope to observe different concentrations of purification OP1 of cactus polysaccharides to act on human lung cancer A549 cell morphological changes.The result shown that purification OP1 of Cactus polysaccharides can inhibit proliferation of human lung cancer A549,and induce apoptosis of human lung cancer A549,and in a range of time,dose-dependent,the IC50 is (597.55±28.97)μg/mL.Purification OP1 of Cactus polysaccharides can make human lung cancer A549 DNA fragmentation.

  19. Telomere shortening and cell senescence induced by perylene derivatives in A549 human lung cancer cells.

    Science.gov (United States)

    Taka, Thanachai; Huang, Liming; Wongnoppavich, Ariyaphong; Tam-Chang, Suk-Wah; Lee, T Randall; Tuntiwechapikul, Wirote

    2013-02-15

    Cancer cells evade replicative senescence by re-expressing telomerase, which maintains telomere length and hence chromosomal integrity. Telomerase inhibition would lead cancer cells to senesce and therefore prevent cancer cells from growing indefinitely. G-quadruplex ligands can attenuate telomerase activity by inducing G-quadruplex formation at the 3'-overhang of telomere and at the human telomerase reverse transcriptase (hTERT) promoter; the former prevents telomerase from accessing the telomere, and the latter acts as a transcriptional silencer. The present investigation found that perylene derivatives PM2 and PIPER induced G-quadruplex formation from both telomeric DNA and the hTERT promoter region in vitro. Further, TRAP assay showed that these compounds inhibited telomerase in a dose-dependent manner. When A549 human lung cancer cells were treated with these compounds, hTERT expression was down-regulated. Moreover, the crude protein extract from these treated cells exhibited less telomerase activity. In the long-term treatment of A549 lung cancer cells with sub-cytotoxic dose of these perylenes, telomere shortening, reduction of cell proliferation and tumorigenicity, and cell senescence were observed. The results of this study indicate that perylene derivatives warrant further consideration as effective agents for cancer therapy.

  20. Integrin αv promotes proliferation by activating ERK 1/2 in the human lung cancer cell line A549.

    Science.gov (United States)

    Fu, Shijie; Fan, Limin; Pan, Xufeng; Sun, Yifeng; Zhao, Heng

    2015-02-01

    Lung cancer is a leading cause of cancer-related death worldwide, and non-small cell lung cancer (NSCLC) constitutes ~85% of lung cancers. However, the mechanisms underlying the progression of NSCLC remain unclear. In this study, we found the mRNA and protein expression levels of integrin αv are both increased in NSCLC tissues compared to healthy ones, which indicates that integrin αv may play an important role in NSCLC progression. To further investigate the roles of integrin αv in NSCLC, we overexpressed the integrin αv gene in the NSCLC cell line A549, and found that the cell proliferative ability increased. The apoptosis of A549 cells was inhibited with overexpression of integrin αv. To elucidate the molecular mechanism underlying the role of integrin αv in promoting NSCLC progression, we studied the expression of proteins from a number of important pathways associated with tumorigenesis, and found that the extracellular signal regulated protein kinase (ERK)1/2 signaling pathway may be involved in the mediation of the observed integrin αv effects. component of an important pathway for tumorigenesis, the ERK 1/2. Following inhibition of ERK 1/2 signaling, the proliferation of A549 cells induced by integrin αv was reduced, while the inhibition of apoptosis was attenuated. Our findings demonstrate that integrin αv promotes the proliferation of the human lung cancer cell line A549 by activating the ERK 1/2 signaling pathway, which suggests that this pathway may be a promising target for the treatment of human lung cancer.

  1. Previous heat shock treatment inhibits Mayaro virus replication in human lung adenocarcinoma (A549) cells.

    Science.gov (United States)

    Virgilio, P L; Godinho-Netto, M C; Carvalho Mda, G

    1997-01-01

    Human lung adenocarcinoma cells (A549) were submitted to mild or severe heat shock (42 degrees C or 44 degrees C) for 1 h, while another group of cells was double-heat-shocked (submitted to 42 degrees C for 1 h, returned to 37 degrees C for 3 h, then exposed to 44 degrees C for 1 h). After each heat treatment, the cells were infected with Mayaro virus for 24 h and incubated at 37 degrees C. The results showed that the double-heat-shocked thermotolerant cells exhibited a 10(4)-fold virus titre inhibition, despite the recovery of protein synthesis and original morphology 24 h post-infection. In contrast, cells submitted to mild or severe heat shock exhibited weaker inhibition of Mayaro virus titre (10(2)-fold). The mildly heat-shocked cells also presented a full recovery in protein synthesis, which was not observed in severely heat-shocked cells. These results indicate that exposure of A549 cells to a mild or to a double heat shock treatment before Mayaro virus infection induces an antiviral state.

  2. 低氧对人肺腺癌A549细胞迁移和黏附的影响%Effect of hypoxia on migration, invasion and adhesion to endothelium of human pulmonary adenocarcinoma A549 cells

    Institute of Scientific and Technical Information of China (English)

    Weigan Shen; Jun Zhu; Zhiyong Yu; Qingyu Xue

    2008-01-01

    Objective:To evaluate the effect of hypoxia on migration,invasion and adhesion to endothelium of human pulmonary adenocarcinoma A549 cells.Methods:Wound-healing and Transwell invasion assays were performed to study the effect of hypoxia on migration and invasion of A549 cells,and A549 cells were added to a monolayer of human umbilical vein endothelial cells (HUVECs) to test the ability to adhere to endothelium.Immunofluorescence assay and luciferase reporter gene assay were also used to evaluate the effect of hypoxia on distribution of E-cadherin,β-catenin,and actin,and hypoxia-inducible factor-1 (HIF-1)-dependent transcription,respectively.Results:Hypoxia facilitated A549 cell migration,invasion,and A549 cell-endothelial cells adhesion,and modulated the distribution of E-cadherin and β-catenin,and actin cytoskeleton rearrangement,and up-regulated HIF-1-dependent reporter gene expression in A549 cells.Conclusion:Promotion of A549 cell migration,invasion,and adhesion on endothelium by hypoxia might be modulated through its up-regulating HIF-l-dependent gene expression,which then induced the redistribution of E-cadherin and β-catenin,and the actin cytoskeletal reorganization.

  3. PRDM14 Promotes the Migration of Human Non-small Cell Lung Cancer Through Extracellular Matrix Degradation in vitro

    Institute of Scientific and Technical Information of China (English)

    Hong-Xia Bi; Han-Bing Shi; Ting Zhang; Ge Cui

    2015-01-01

    Background:As a novel molecular markerof non-small cell lung cancer (NSCLC),PRDI-BF1 and RIZ homology domain containing protein 14 (PRDM 14) is over-expressed in NSCLC tumor tissues.Extracellular matrix degradation mediated by the balance between matrix metalloproteinases (MMPs) and tissue inhibitor of metalloproteinases (TIMPs) is one of the most important mechanism in lung cancer metastasis.This study aimed to determine if PRDM 14 promoted the migration of NSCLC cells through extracellular matrix degradation mediated by change of MMP/TIMP expression.Methods:The expression of PRDM14 was down-regulated in human cell line A 549 after transfection with lentiviral vector-mediated short-hairpin ribonucleic acids (shRNAs) which targeted the PRDM 14 promoter.Cellular migration ofshRNA-infected cells was detected by a scratch wound healing assay and transwell cell migration assay.Expression levels of MMP1,MMP2,TIMP1,and TIMP2 were measured by quantitative real-time polymerase chain reaction (RT-PCR).Results:Migration ofPRDM 14-shRNA-infected cells was significantly inhibited relative to control cells as measured by the scratch wound healing (P < 0.05) and transwell cell migration assays (P < 0.01).The expression of MMP1 in A549 cells infected by PRDM14-shRNA was down-regulated significantly (P < 0.01),whereas the expression of TIMPl and TIMP2 was up-regulated significantly (P < 0.01).Conclusions:PRDM 14 accelerates A549 cells migration in vitro through extracellular matrix degradation.PRDM 14 is considered as a potential therapeutic target in metastatic NSCLC.

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

    Science.gov (United States)

    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.

  5. The human lung cell line A549 does not develop adaptive protection against the DNA-damaging action of formaldehyde.

    Science.gov (United States)

    Speit, Günter; Neuss, Simone; Schmid, Oliver

    2010-03-01

    The alkaline comet assay was used to further characterize the induction of DNA-protein crosslinks (DPX) by formaldehyde (FA) and their removal in the human lung cell line A549. DPX were indirectly measured as the reduction of gamma ray-induced DNA migration. Repeated treatments of A549 cells with low FA concentrations (up to 100 microM) did not lead to significant differences in the induction of DPX in comparison with a single treatment. Pretreatment with higher FA-concentrations (200 microM and above) enhanced the crosslinking effect. There was no indication for an adaptive protection against the induction of DPX by FA. These findings are in agreement with RT-PCR measurements of the expression of genes that encode the main enzymes involved in FA detoxification. A549 cells exposed to FA (50-300 microM) for 1, 4, or 24 hr did not reveal altered expression of the GSH-dependent formaldehyde dehydrogenase (FDH, which is identical to alcohol dehydrogenase 3; ADH3), the cytosolic aldehyde dehydrogenase 1 (ALDH1A1) and the mitochondrial ALDH2. Pretreatment of A549 cells with a low FA concentration (50 microM) also did not enhance the removal of DPX induced by higher FA concentrations. Taken together, these results suggest that A549 cells do not develop adaptive protection against the genotoxic action of FA. Neither metabolic inactivation of FA nor the repair of FA-induced DPX seems to be enhanced in cells pretreated with FA.

  6. Schisandrin B inhibits the proliferation of human lung adenocarcinoma A549 cells by inducing cycle arrest and apoptosis.

    Science.gov (United States)

    Lv, Xue-Jiao; Zhao, Li-Jing; Hao, Yu-Qiu; Su, Zhen-Zhong; Li, Jun-Yao; Du, Yan-Wei; Zhang, Jie

    2015-01-01

    Lung cancer is the leading cause of cancer death in the world. Schizandrin B (Sch B) is one of the main dibenzocyclooctadiene lignans present in the fruit of Schisandra chinensis (Schisandraceae). Sch B has multiple functions against cancer. The aim of this study was to determine the effect of Sch B on the proliferation, cell cycling, apoptosis and invasion of lung adenocarcinoma A549 cells by MTT, flow cytometry, wound healing and transwell invasion assays. Treatment with Sch B inhibited the proliferation of A549 cells in a dose-dependent manner. Sch B induced cell cycle arrest at G0/G1 phase by down-regulating the expression of cyclin D1, cyclin-dependent kinase (CDK)4, and CDK6, but up-regulating p53 and p21 expression in A549 cells. Furthermore, Sch B triggered A549 cell apoptosis by increasing Bax, cleaved caspase-3, 9, Cyto C, but decreasing Bcl-2 and PCNA expression. In addition, Sch B inhibited the invasion and migration of A549 cells by down-regulating the expressions of HIF-1, VEGF, MMP-9 and MMP-2. Therefore, Sch B has potent anti-tumor activity and may be a promising traditional Chinese medicine for human lung carcinoma.

  7. Effects of herpes simplex virus thymidine kinase/acyclovir system on growth of human pulmonary adenocarcinoma A549 cell line in vitro and in vivo

    Institute of Scientific and Technical Information of China (English)

    HE Xiang-liang; HE Dong-hua; GUO Xian-jian; QIAN Gui-sheng; HUANG Gui-jun; CHEN Wei-zhong; LI Shu-ping

    2002-01-01

    Objective: To observe the effect of anciclovir (ACV) treatment on tumors induced by inoculation of TK gene-transfected human pulmonary adenocarcinoma A549 cells in nude mice. Methods: A recombinant plasmid containing TK gene was constructed and transfected into A549 cells by electroporation. The sensitivity of the transgenic cells (A549-TK) to ACV was examined by MTT assay in vitro and for in vivo observation, inoculation of A549-TK and A-549 cells into nude mice was separately performed to induce tumor growth, the response of which to ACV treatment was observed, and the tumor tissues were pathologically examined. Results: A recombinant plasmid containing TK gene was successfully constructed and transfected into A549 cells. The sensitivity of A549-TK cells to ACV was 43 times higher than that of A549 cells. The tumors induced by A549-TK cells showed no significant increase in size after ACV treatment (P>0. 05), and light microscopy revealed local tissue necrosis, karyoklasis, and nuclei disappearance. Conclusion: A549-TK cells acquires sensitivity to ACV both in vitro and in vivo, and ACV can inhibit the growth of tumors induced by A549-TK cell inoculation in nude mice.

  8. Establishment of a Multidrug Resistance Cell Line A549/cDDP of Human Lung Adenocarcinoma and Expression Analysis of Multidrug Resistance-Associated Genes

    Directory of Open Access Journals (Sweden)

    Yongcheng PAN

    2009-03-01

    Full Text Available Background and objective It has been proven that chemotherapy failure caused by multidrug resistance in lung tumor cells is the main cause for the patient's survival rate. The aim of this study is to establish a multidrug resistance cell line of human lung adenocarcinoma and study the mechanism of multidrug resistance. Methods Human lung adenocarcinoma cell line A549 was induced to multidrug resistance cell line A549/cDDP by intermittentadministration of high dose of cisplatin (cDDP. The multidrug resistance was detected by using MTT assay. The levels of expression of MDR-1 gene-coded P-glycoportein (P-gp, multidrug resistance-associated protein (MRP, and GSH/GST were examined by flow cytometric assay. The levels of expression of MDR and MRP gene were also detected by RTPCR in both A549/cDDP and A549 cell lines. Results A549/cDDP was resistant to many anti-tumor agents. The IC50 of A549/cDDP was 16.87 times higher than that of A549. The expressions of P-gp and MRP in A549/cDDP were increased significantly to (70.5±4.9% and (29.4±2.9%, respectively, vs (42.4±5.6% and (21.4±3.5% in A549. There was no difference of the GSH/GST expression between A549/cDDPand A549 cells. Conclusion A549/cDDP is a model with multidrug resistance and the levels of MDR and MRP mRNA expressions are remarkably higher in A549/cDDP than those in A549.

  9. 4-Nitroquinoline-1-oxide effects human lung adenocarcinoma A549 cells by regulating the expression of POLD4

    OpenAIRE

    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 expres...

  10. Direct and in vitro observation of growth hormone receptor molecules in A549 human lung epithelial cells by nanodiamond labeling

    Science.gov (United States)

    Cheng, C.-Y.; Perevedentseva, E.; Tu, J.-S.; Chung, P.-H.; Cheng, C.-L.; Liu, K.-K.; Chao, J.-I.; Chen, P.-H.; Chang, C.-C.

    2007-04-01

    This letter presents direct observation of growth hormone receptor in one single cancer cell using nanodiamond-growth hormone complex as a specific probe. The interaction of surface growth hormone receptor of A549 human lung epithelial cells with growth hormone was observed using nanodiamond's unique spectroscopic signal via confocal Raman mapping. The growth hormone molecules were covalent conjugated to 100nm diameter carboxylated nanodiamonds, which can be recognized specifically by the growth hormone receptors of A549 cell. The Raman spectroscopic signal of diamond provides direct and in vitro observation of growth hormone receptors in physiology condition in a single cell level.

  11. Effects of the Spider Venom on proliferation of Human Lung Adenocarcinoma Cell A549

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    Zengxiang HU

    2010-10-01

    Full Text Available Background and objective The spider venom may inspire new drugs to treat cancer. The aim of this study is to investigate the effects and mechanisms of spider venom on lung adenocarcinoma cell A549. Methods The proliferation of lung adenocarcinoma A549 cells was detected by MTT. The apoptosis rate was observed with MTT assay and flow cytometer. The activity of catalase was detected by colorimetry. The malondialdehyde (MDA content was determined by improved thiobarbituric acid fluorometric method. The expression of P38MAPK protein was analyzed with Western blot. Results Spider venom can remarkably inhibite the proliferation of lung adenocarcinoma A549 cells, increased activity of catalase and MDA content, down-regulated expression of P38MAPK compared with the control group. Conclusion The reduced proliferation of lung adenocarcinoma A549 cells by spider venom is may be associated with the increased of activity of catalase and MDA content and decreased expression of P38MAPK.

  12. Role of Autophagy in the Radiosensitivity of Human Lung Adenocarcinoma A549 Cells

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    Liyao XU

    2016-12-01

    Full Text Available Background and objective Radiotherapy is an important treatment for lung cancer. The poor prognosis of lung cancer is largely caused by the high recurrence rate and metastasis of the tumor. Autophagy, which can be induced by radiotherapy, might be associated with DNA repair. The aim of this study is to investigate whether activating autophagy using rapamycin can enhance the radiosensitivity of lung cancer cells and clarify the association of autophagy with DNA repair. Methods The human adenocarcinoma A549 cell line was selected as the experimental subject. The specimens were divided into three groups: control (N, radiation (R, and Rapamycin and radiation (R+RAPA. The protein levels of γ-H2AX, Rad51, Ku70/Ku80, p62, and LC3 were determined by Western blot. Autophagosome was observed under a transmission electron microscope, and SF was determined by colony formation assay. Results Compared with group R, the activity of autophagy and the protein expression levels of Rad51 and Ku70/80 were remarkably increased in group R+RAPA. Conclusion The radiosensitivity of lung cancer can be promoted by activating autophagy via treatment with Rapamycin, and the process may be associated with DNA repair.

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

    Science.gov (United States)

    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.

  14. SIRT1通过调节Noxa表达影响非小细胞肺癌细胞株A549对顺铂的敏感性%SIRT1 Influences the Sensitivity of A549 Non-small Cell Lung Cancer Cell Line to Cisplatin via Modulating the Noxa Expression

    Institute of Scientific and Technical Information of China (English)

    曹彬; 何晓峰; 王文公; 史敏科

    2016-01-01

    背景与目的 非小细胞肺癌的顺铂耐药是常见的临床现象,严重制约了患者的化疗效果,是亟待解决的问题.SIRT1和Noxa的表达变化影响肿瘤细胞对化疗药物的敏感性.本研究旨在研究SIRT1表达对非小细胞肺癌对顺铂的敏感性的影响,并探讨其涉及Noxa表达的机制,以求为提高非小细胞肺癌细胞对顺铂敏感性提供希望.方法 利用实时荧光定量PCR和Westem blot分析A549细胞及顺铂耐药的A549/DDP细胞SIRT1及Noxa mRNA和蛋白水平的表达差异.利用siRNA干扰技术抑制A549/DDP细胞的SIRT1表达,进而使用Cell Titer Blue试验、流式细胞术从细胞增殖、细胞周期和细胞凋亡方面分析SIRT1沉默对A549/DPP细胞顺铂敏感性的影响.同时利用实时荧光定量PCR和Western blot分析SIRT1抑制对A549/DPP细胞Noxa表达的影响.结果 A549细胞和A549/DDP细胞对顺铂的敏感性有显著差异,与A549细胞相比,A549/DDP细胞的SIRT1表达较高,但Noxa表达较低.使用siRN抑制A549/DPP细胞的SIRT1表达后,与未抑制SIRT1细胞相比,4μg/mL顺铂处理后的细胞存活率降低,G2期/M期阻滞比例增加,凋亡率提高.同时,SIRT1沉默导致A549/DPP细胞的Noxa表达增加.结论 较高的SIRT1可能引起A549细胞对顺铂的耐药性,抑制SIRT1可以提高A549/DDP细胞对顺铂的敏感性,其机制可能涉及SIRT1对Noxa的调节.

  15. Doxycycline decreases production of interleukin-8 in a549 human lung epithelial cells

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    Hoyt JC

    2013-03-01

    Full Text Available Doxycycline is an antibiotic that possess anti-inflammatory properties. These anti-inflammatory properties make doxycycline an attractive candidate for possible treatments for a variety of common chronic obstructive airway diseases. Interleukin-8 (IL-8 is a major inflammatory chemokine and a powerful chemo-attractant for both neutrophils and monocytes. We hypothesized that doxycycline might exert its anti-inflammatory effects, at least in part, by modulating IL-8 production. To test this hypothesis, A549 human lung epithelial cells were stimulated with cytomix (IL-1beta, TNF-alpha and gamma-IFN in the presence or absence of varying concentrations of doxycycline. Doxycycline decreased IL-8 protein production in a concentration- and time-dependent manner. In the presence of 30 microg/ml doxycycline IL-8 protein production was decreased by 63% through out a 30 hr time course. In chemotaxis assays monocyte and neutrophil migration was decreased by 55% and 57% respectively. Reverse transcriptase-polymerase chain reaction (RT-PCR experiments suggest that doxycycline does not decrease expression of IL-8 mRNA and that use of the RNA polymerase II inhibitor DRB indicates that doxycycline does not effect stability of this mRNA. In the presence of doxycycline p38-alpha mitogen-activated protein kinase (MAPK expression is decreased by 36% in cytomix-stimulated cells. These data demonstrate that doxycycline can modulate IL-8 release and suggest that it has potential as an anti-inflammatory in those disorders where IL-8 is an important inflammatory mediator.

  16. Inonotus obliquus-derived polysaccharide inhibits the migration and invasion of human non-small cell lung carcinoma cells via suppression of MMP-2 and MMP-9.

    Science.gov (United States)

    Lee, Ki Rim; Lee, Jong Seok; Song, Jeong Eun; Ha, Suk Jin; Hong, Eock Kee

    2014-12-01

    Polysaccharides isolated from the fruiting body of Inonotus obliquus (PFIO) are known to possess various pharmacological properties including antitumor activity. However, the anti-metastatic effect and its underlying mechanistic signaling pathway involved these polysaccharides in human non-small cell lung carcinoma remain unknown. The present study therefore aimed to determine the anti-metastatic potential and signaling pathways of PFIO in the highly metastatic A549 cells. We found that PFIO suppressed the migration and invasive ability of A549 cells while decreasing the expression levels and activity of matrix metalloproteinase (MMP)-2 and MMP-9. Furthermore, PFIO decreased the phosphorylation levels of mitogen-activated protein kinases (MAPKs) and phosphoinositide 3-kinase (PI3K)/protein kinase B (AKT) as well as the expression level of COX-2, and inhibited the nuclear translocation of nuclear factor κB (NF-κB) in A549 cells. These results suggested that PFIO could suppress the invasion and migration of human lung carcinoma by reducing the expression levels and activity of MMP-2 and MMP-9 via suppression of MAPKs, PI3K/AKT, and NF-κB signaling pathways.

  17. Capsaicin-induced genotoxic stress does not promote apoptosis in A549 human lung and DU145 prostate cancer cells.

    Science.gov (United States)

    Lewinska, Anna; Jarosz, Paulina; Czech, Joanna; Rzeszutek, Iwona; Bielak-Zmijewska, Anna; Grabowska, Wioleta; Wnuk, Maciej

    2015-02-01

    Capsaicin is the major pungent component of the hot chili peppers of the genus Capsicum, which are consumed worldwide as a food additive. More recently, the selective action of capsaicin against cancer cells has been reported. Capsaicin was found to induce apoptosis and inhibit proliferation of a wide range of cancer cells in vitro, whereas being inactive against normal cells. As data on capsaicin-induced genotoxicity are limited and the effects of capsaicin against human lung A549 and DU145 prostate cancer cells were not explored in detail, we were interested in determining whether capsaicin-associated genotoxicity may also provoke A549 and DU145 cell death. Capsaicin-induced decrease in metabolic activity and cell proliferation, and changes in the cell cycle were limited to high concentrations used (≥ 100 μM), whereas, at lower concentrations, capsaicin stimulated both DNA double strand breaks and micronuclei production. Capsaicin was unable to provoke apoptotic cell death when used up to 250 μM concentrations. Capsaicin induced oxidative stress, but was ineffective in provoking the dissipation of the mitochondrial inner transmembrane potential. A different magnitude of p53 binding protein 1 (53BP1) recruitment contributed to diverse capsaicin-induced genotoxic effects in DU145 and A549 cells. Capsaicin was also found to be a DNA hypermethylating agent in A549 cells. In summary, we have shown that genotoxic effects of capsaicin may contribute to limited susceptibility of DU145 and A549 cancer cells to apoptosis in vitro, which may question the usefulness of capsaicin-based anticancer therapy, at least in a case of lung and prostate cancer.

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

    OpenAIRE

    Kathiravan, Govindarajan; Sureban, Sripathi M.

    2010-01-01

    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 ch...

  19. Cytotoxicity, oxidative stress and genotoxicity induced by glass fibers on human alveolar epithelial cell line A549.

    Science.gov (United States)

    Rapisarda, Venerando; Loreto, Carla; Ledda, Caterina; Musumeci, Giuseppe; Bracci, Massimo; Santarelli, Lory; Renis, Marcella; Ferrante, Margherita; Cardile, Venera

    2015-04-01

    Man-made vitreous fibers have been widely used as insulation material as asbestos substitutes; however their morphology and composition raises concerns. In 1988 the International Agency for Research on Cancer classified fiberglass, rock wool, slag wool, and ceramic fibers as Group 2B, i.e. possibly carcinogenic to humans. In 2002 it reassigned fiberglass, rock and slag wool, and continuous glass filaments to Group 3, not classifiable as carcinogenic to humans. The aim of this study was to verify the cytotoxic and genotoxic effects and oxidative stress production induced by in vitro exposure of human alveolar epithelial cells A549 to glass fibers with a predominant diameter 5 μm (93%). A549 cells were incubated with 5, 50, or 100 μg/ml (2.1, 21, and 42 μg/cm(2), respectively) of glass fibers for 72 h. Cytotoxicity and DNA damage were tested by the MTT and the Comet assay, respectively. Oxidative stress was determined by measuring inducible nitric oxide synthase (iNOS) expression by Western blotting, production of nitric oxide (NO) with Griess reagent, and concentration of reactive oxygen species by fluorescent quantitative analysis with 2',7'-dichlorofluorescein-diacetate (DCFH-DA). The results showed that glass fiber exposure significantly reduced cell viability and increased DNA damage and oxidative stress production in a concentration-dependent manner, demonstrating that glass fibers exert cytotoxic and genotoxic effects related to increased oxidative stress on the human alveolar cell line A549.

  20. Safrole oxide induces apoptosis by activating caspase-3, -8, and -9 in A549 human lung cancer cells.

    Science.gov (United States)

    Du, Aiying; Zhao, Baoxiang; Yin, Deling; Zhang, Shangli; Miao, Junying

    2006-01-01

    Previously we found that 3,4-(methylenedioxy)-1-(2',3'-epoxypropyl)-benzene (safrole oxide) induced a typical apoptosis in A549 human lung cancer cells. In this study, we further investigated which caspases were activated by safrole oxide during the apoptosis. The data showed that the activity of caspase-3, -8, and -9 was significantly enhanced by the compound, which suggested that safrole oxide might be used as a caspase promoter to initiate lung cancer cell apoptosis.

  1. Solitary Lung Tumors and Their Spontaneous Metastasis in Athymic Nude Mice Orthotopically Implanted with Human Non-Small Cell Lung Cancer

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    Takeshi Yamaura

    2000-07-01

    Full Text Available We examined the tumorigenic and metastatic potentials of three human non-small cell lung cancer (NSCLC cell lines. PC-14, A549 or Lu-99 cell lines suspended in Matrigel-containing phosphate-buffered saline were orthotopically implanted into the lungs of nude mice. The formation of a solitary tumor nodule in the lung was observed after the implantation of all cell lines. Intrapulmonary implantation of PC-14 or Lu-99 cells resulted in spontaneous distant metastases. In contrast, A549 cells caused multiple intrapulmonary metastases to the right and left lobes of the lung without producing visible lymphatic metastasis. We also investigated the expression of matrix metal loproteinases (MMPs, urokinase-type plasminogen activator (u-PA, u-PA receptor (u-PAR and c-MET in these cell lines in vitro and in vivo. Reverse transcription polymerase chain reaction (RT-PCR analysis showed that the expression of MMP-2 and membrane-type 1 MMP (MT1-MMP was elevated in PC-14 as compared with the other two cell lines. In contrast, stronger expression of c-METwas observed in A549 than in PC-14 or Lu-99. These results indicate that differential patterns of metastasis of lung cancer might be associated with differential expression of metastasis-associated molecules. Our orthotopic implantation models display clinical features resembling those of NSCLC, may provide a useful basis for lung cancer research.

  2. Cardenolide-Induced Lysosomal Membrane Permeabilization Demonstrates Therapeutic Benefits in Experimental Human Non-Small Cell Lung Cancers

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    Tatjana Mijatovic

    2006-05-01

    Full Text Available Non-small cell lung cancers (NSCLCs are the leading cause of cancer deaths in most developed countries. Targeting heat shock protein 70 (Hsp70 expression and function, together with the induction of lysosomal membrane permeabilization (LMP, could overcome the multiple anti-cell death mechanisms evidenced in NSCLCs that are responsible for the failure of currently used chemotherapeutic drugs. Because cardenolides bind to the sodium pump, they affect multiple signaling pathways and thus have a number of marked effects on tumor cell behavior. The aim of the present study was to characterize in vitro and in vivo the antitumor effects of a new cardenolide (UNBS1450 on experimental human NSCLCs. UNBS1450 is a potent source of in vivo antitumor activity in the case of paclitaxeland oxaliplatin-resistant subcutaneous human NCIH727 and orthotopic A549 xenografts in nude mice. In vitro UNBS1450-mediated antitumor activity results from the induction of nonapoptotic cell death. UNBS1450 mediates the decrease of Hsp70 at both mRNA and protein levels, and this is at least partly due to UNBS1450-induced downregulation of NFAT5/ TonEBP (a factor responsible for the transcriptional control of Hsp70. These effects were paralleled by the induction of LMP, as evidenced by acridine orange staining and immunofluorescence analysis for cathepsin B accumulation.

  3. The targeted inhibitory effects of human amniotic fluid stem cells carrying CXCR4 promoter and DAL-1 on non-small cell lung carcinoma growth.

    Science.gov (United States)

    Li, L; Li, S; Cai, T; Wang, H; Xie, X; Liu, Z; Zhang, Y

    2016-02-01

    The differentially expressed in adenocarcinoma of the lung-1 (DAL-1) protein has been demonstrated to be suppressive to various types of tumors including lung cancer. This study aimed to determine the targeted effects of human amniotic fluid stem cells (hAFS cells) carrying CXCR4 promoter driven conditionally replicable adenovirus vector overexpressing DAL-1 (Ad-CXCR4-DAL-1) on non-small cell lung carcinoma (NSCLC) growth. The apoptotic effects of virus vectors were assessed using flow cytometry, and the cytotoxicity analyzed by CCK-8 assay. In vivo imaging system was used to determine the homing capability of hAFS cells. A549 cell xenograft mouse model was created to assess the in vivo effect of DAL-1 overexpression on NSCLC growth. We found that infection of Ad-CXCR4-DAL-1 increased the apoptosis of A549 NSCLC cells but not 16HBE normal human bronchial epithelial cells. Ad-CXCR4-DAL-1 administered via intratumoral injection led to significant reduced growth and greater necrosis of A549 xenograft tumors comparing to null vector treated animals. When infused via tail vein, hAFS cells carrying Ad-CXCR4-DAL-1 homed to lung cancer xenografts, caused virus replication and DAL-1 overexpression, and led to significant lower growth and greater necrosis of A549 cell xenografts comparing to non-treatment control. In conclusion, hAFS cells are capable of carrying Ad-CXCR4-DAL-1 vectors, specifically targeting to lung cancer, and causing oncolytic effects when administered in vivo.

  4. Function of Lycium Barbarum Polysaccharide on Proliferation and Apoptosis of Human Lung Cancer A549 Cells%枸杞多糖对人肺腺癌A549细胞增殖和凋亡的影响

    Institute of Scientific and Technical Information of China (English)

    孟凡珍; 江涛

    2012-01-01

    Objective To investigate the growth inhibition and proliferation of human lung adenocarcinoma AS49 cells induced by Lycium bar-barum polysaccharide( LBP) in vitro,and the possible underlying mechanisms. Methods A549 cells cultured in vitro were divided into control group and experimental group ( I/ZICjo for 48 hours) by the different concentrations of LBP treatment. The inhibitory effects of LBP on proliferation of A549 were determined by MTT assay at 24 h ,48 h ,72 h after the addition of LBP to A549 culture. Growth curve were generated by MTT assay,doubling time were calculated by cell counting,Cell cycle and apoptosis were examined by Flow cytometry(FCM) ,Sur-vivin mRNA changes were detected by RT-PCR,CyclinBl protein changes were detected by Western blot ,Tramwell assay in vitro was utilized to evaluate the invasive activity. Results MTT assay demonstrated that different concentrations of LBP significantly inhibited the proliferation of A549 cells in a dose-dependent manner,the doubling time and the rate of apoptosis of cells in experimental group is dramatically different from the control group. ( P < 0. 05 ) , It was found that LBP arrested A549 cells at G2 phase; Survivin mRNA expression and CyclinBl protein expression were lower,compared with that in the control group( P <0. 05 ) . Conclusion The present study suggests that LBP can significantly inhibit the proliferation of A549 cells. The mechanism may be related to an arrest effect on cell cycle,reducing Survivin mRNA and CyclinBl protein expression and inhibiting tumour invasion.%目的 探讨枸杞多糖(LBP)对体外培养的人肺腺癌细胞A549的增殖抑制作用及其可能的作用机制.方法 用不同浓度的LBP处理A549细胞,MTT法检测24、48、72h时间点LBP对A549细胞的生长抑制率,实验设为对照组和实验组(1/2IC50作用48小时),MTT法绘制生长曲线、细胞计数计算倍增时间、流式细胞仪检测凋亡率及其细胞周期、RT-PcR检测Survivin m

  5. Differential Regulation of Gene Expression of Alveolar Epithelial Cell Markers in Human Lung Adenocarcinoma-Derived A549 Clones

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    Hiroshi Kondo

    2015-01-01

    Full Text Available Stem cell therapy appears to be promising for restoring damaged or irreparable lung tissue. However, establishing a simple and reproducible protocol for preparing lung progenitor populations is difficult because the molecular basis for alveolar epithelial cell differentiation is not fully understood. We investigated an in vitro system to analyze the regulatory mechanisms of alveolus-specific gene expression using a human alveolar epithelial type II (ATII cell line, A549. After cloning A549 subpopulations, each clone was classified into five groups according to cell morphology and marker gene expression. Two clones (B7 and H12 were further analyzed. Under serum-free culture conditions, surfactant protein C (SPC, an ATII marker, was upregulated in both H12 and B7. Aquaporin 5 (AQP5, an ATI marker, was upregulated in H12 and significantly induced in B7. When the RAS/MAPK pathway was inhibited, SPC and thyroid transcription factor-1 (TTF-1 expression levels were enhanced. After treatment with dexamethasone (DEX, 8-bromoadenosine 3′5′-cyclic monophosphate (8-Br-cAMP, 3-isobutyl-1-methylxanthine (IBMX, and keratinocyte growth factor (KGF, surfactant protein B and TTF-1 expression levels were enhanced. We found that A549-derived clones have plasticity in gene expression of alveolar epithelial differentiation markers and could be useful in studying ATII maintenance and differentiation.

  6. A new dammarane-type saponin from Gynostemma pentaphyllum induces apoptosis in A549 human lung carcinoma cells.

    Science.gov (United States)

    Xing, Shao-Fang; Jang, Moonhee; Wang, Yu-Rong; Piao, Xiang-Lan

    2016-04-01

    Gynostemma pentaphyllum has been widely used as a traditional herb for its antioxidant and immunostimulatory activities. We have previously reported several useful dammarane-type saponins with cytotoxicity against A549 human lung cancer cells from heat-processed G. pentaphyllum. In this study, a new dammarane-type saponin, 20(S)-2α,3β,12β-tetrahydroxydammar-3-O-β-d-glucopyranoside (namely gypenoside Jh1), was isolated from the ethanol extract of heat-processed G. pentaphyllum using column chromatography and semi-preparative HPLC. Gypenoside Jh1 exhibited strong cytotoxicity against A549 cells in a concentration-dependent manner, which was associated with apoptotic cell death characterized by morphological changes, Hoechst 33258 nuclear staining, Annexin V and propidium iodide binding and mitochondrial potentials assay. Quantitative analysis using flow cytometry also showed that the proportion of apoptotic cells was increased after gypenoside Jh1 treatment. These findings indicated that gypenoside Jh1 showed antiproliferative effects on A549 cells and mitochondrial-dependent pathway is involved in gypenoside Jh1-induced apoptosis. Copyright © 2016 Elsevier Ltd. All rights reserved.

  7. Vitamin D Analogs Potentiate the Antitumor Effect of Imatinib Mesylate in a Human A549 Lung Tumor Model.

    Science.gov (United States)

    Maj, Ewa; Filip-Psurska, Beata; Świtalska, Marta; Kutner, Andrzej; Wietrzyk, Joanna

    2015-11-13

    In previous papers, we presented data on studies on the anticancer activity of the vitamin D₃ analogs, named PRI-2191 and PRI-2205, in different cancer models. In this study, we showed the improved antiproliferative activity of a combination of imatinib mesylate (Gleevec, GV) and cytostatic agents in in vitro studies, when used with a third compound, namely PRI-2191, in an A549 human lung cancer model. Furthermore, we analyzed the influence of both PRI-2191, as well as PRI-2205 on the anticancer activity of GV in mice bearing A549 tumors. The route of PRI-2191 analog administration showed a significant impact on the outcome of GV treatment: subcutaneous injection was more efficient and less toxic than oral gavage. Moreover, both vitamin D compounds increased the anticancer activity of GV; however, they might also potentiate some adverse effects. We also evaluated in tumor tissue the expression of VEGF, PDGF-BB, vitamin D receptor, CYP27B1, CYP24, p53 and Bcl-2, as well as PDGF receptors: α and β. We observed the upregulation of p53 expression and the downregulation of Bcl-2, as well as VEGF in A549 tumors as a result of the tested treatment. However, vitamin D analogs did not significantly influence the expression of these proteins.

  8. Vitamin D Analogs Potentiate the Antitumor Effect of Imatinib Mesylate in a Human A549 Lung Tumor Model

    Science.gov (United States)

    Maj, Ewa; Filip-Psurska, Beata; Świtalska, Marta; Kutner, Andrzej; Wietrzyk, Joanna

    2015-01-01

    In previous papers, we presented data on studies on the anticancer activity of the vitamin D3 analogs, named PRI-2191 and PRI-2205, in different cancer models. In this study, we showed the improved antiproliferative activity of a combination of imatinib mesylate (Gleevec, GV) and cytostatic agents in in vitro studies, when used with a third compound, namely PRI-2191, in an A549 human lung cancer model. Furthermore, we analyzed the influence of both PRI-2191, as well as PRI-2205 on the anticancer activity of GV in mice bearing A549 tumors. The route of PRI-2191 analog administration showed a significant impact on the outcome of GV treatment: subcutaneous injection was more efficient and less toxic than oral gavage. Moreover, both vitamin D compounds increased the anticancer activity of GV; however, they might also potentiate some adverse effects. We also evaluated in tumor tissue the expression of VEGF, PDGF-BB, vitamin D receptor, CYP27B1, CYP24, p53 and Bcl-2, as well as PDGF receptors: α and β. We observed the upregulation of p53 expression and the downregulation of Bcl-2, as well as VEGF in A549 tumors as a result of the tested treatment. However, vitamin D analogs did not significantly influence the expression of these proteins. PMID:26580599

  9. Vitamin D Analogs Potentiate the Antitumor Effect of Imatinib Mesylate in a Human A549 Lung Tumor Model

    Directory of Open Access Journals (Sweden)

    Ewa Maj

    2015-11-01

    Full Text Available In previous papers, we presented data on studies on the anticancer activity of the vitamin D3 analogs, named PRI-2191 and PRI-2205, in different cancer models. In this study, we showed the improved antiproliferative activity of a combination of imatinib mesylate (Gleevec, GV and cytostatic agents in in vitro studies, when used with a third compound, namely PRI-2191, in an A549 human lung cancer model. Furthermore, we analyzed the influence of both PRI-2191, as well as PRI-2205 on the anticancer activity of GV in mice bearing A549 tumors. The route of PRI-2191 analog administration showed a significant impact on the outcome of GV treatment: subcutaneous injection was more efficient and less toxic than oral gavage. Moreover, both vitamin D compounds increased the anticancer activity of GV; however, they might also potentiate some adverse effects. We also evaluated in tumor tissue the expression of VEGF, PDGF-BB, vitamin D receptor, CYP27B1, CYP24, p53 and Bcl-2, as well as PDGF receptors: α and β. We observed the upregulation of p53 expression and the downregulation of Bcl-2, as well as VEGF in A549 tumors as a result of the tested treatment. However, vitamin D analogs did not significantly influence the expression of these proteins.

  10. Genomic signature and toxicogenomics comparison of polycationic gene delivery nanosystems in human alveolar epithelial A549 cells

    Directory of Open Access Journals (Sweden)

    J Barar

    2009-10-01

    Full Text Available "nBackground and the purpose of the study: Of the gene delivery systems, non-viral polycationic gene delivery nanosystems have been alternatively exploited as a relatively safe delivery reagents compared to viral vectors. However, little is known about the genomic impacts of these delivery systems in target cells/tissues. In this study, the toxicogenomics and genotoxicity potential of some selected polycationic lipid/polymer based nanostructures (i.e., Oligofectamine® (OF, starburst polyamidoamine Polyfect® (PF and diaminobutane (DAB dendrimers were investigated in human alveolar epithelial A549 cells. "nMethods: To study the nature and the ontology of the gene expression changes in A549 cells upon treatment with polycationic nanostructures, MTT assay and microarray gene expression profiling methodology were employed. For microarray analysis, cyanine (Cy3/Cy5 labeled cDNA samples from treated and untreated cells were hybridized on target arrays housing 200 genes. "nResults and major conclusions: The polycationic nanosystems induced significant gene expression changes belonging to different genomic ontologies such as cell defence and apoptosis pathways. These data suggest that polycationic nanosystems can elicit multiple gene expression changes in A549 cells upon their chemical structures and interactions with cellular/subcellular components. Such impacts may interfere with the main goals of the desired genemedicine.

  11. In vitro and in vivo therapeutic efficacy of CXCR4 antagonist BKT140 against human non-small cell lung cancer.

    Science.gov (United States)

    Fahham, Duha; Weiss, Ido D; Abraham, Michal; Beider, Katia; Hanna, Wald; Shlomai, Zippora; Eizenberg, Orly; Zamir, Gideon; Izhar, Uzi; Shapira, Oz M; Peled, Amnon; Wald, Ori

    2012-11-01

    CXCR4/CXCL12 interactions promote non-small cell lung cancer (NSCLC) growth and dissemination. Furthermore, this axis might promote NSCLC resistance to chemotherapy and/or radiotherapy. Therefore, the CXCR4/CXCL12 axis constitutes an attractive therapeutic target for the treatment of NSCLC. We aimed to characterize the therapeutic efficacy of the novel CXCR4 antagonist BKT140 against human NSCLC. We determined the CXCR4 expression in 5 NSCLC cell lines (H358, A549, H460, H1299, and L4). We then tested the colony-forming capacity and proliferation of these cells in the presence of CXCL12 and BKT140. Next, we measured the in vivo growth of A549 and H460 xenografts with or without BKT140 treatment. Finally, we examined, in vitro, the potential antiproliferative effect of BKT140 combined with cisplatin or paclitaxel and after irradiation of NSCLC cells. All tested cell lines expressed CXCR4 and showed increased colony formation in response to CXCL12 stimulation. BKT140 reduced the colony-forming capacity of NSCLC cells. Proliferation assays demonstrated both cytotoxic and cytostatic properties for this peptide. H460 cells were the most sensitive to BKT140 and A549 cells the least. Subcutaneous administration of BKT140 significantly delayed the development of H460 xenografts and showed a similar trend for A549 xenografts. Finally, the antiproliferative effects of BKT140 appears to be additive to those of chemotherapeutic drugs and radiotherapy. Targeting the CXCL12/CXCR4 axis with BKT140 attenuated NSCLC cells tumor growth and augmented the effects of chemotherapy and radiotherapy. Future research will benefit from delineating the downstream mechanism of BKT140 action and defining BKT140 susceptibility markers. Crown Copyright © 2012. Published by Mosby, Inc. All rights reserved.

  12. A novel fluorinated thiosemicarbazone derivative- 2-(3,4-difluorobenzylidene) hydrazinecarbothioamide induces apoptosis in human A549 lung cancer cells via ROS-mediated mitochondria-dependent pathway.

    Science.gov (United States)

    Zhao, Yue; Guo, Chuanlong; Wang, Lijun; Wang, Shuaiyu; Li, Xiangqian; Jiang, Bo; Wu, Ning; Guo, Shuju; Zhang, Renshuai; Liu, Kun; Shi, Dayong

    2017-09-09

    Thiosemicarbazone, a class of compounds with excellent biological activity, especially antitumor activity, have attracted wide attention. In this study, a novel fluorinated thiosemicarbazone derivative, 2-(3,4-difluorobenzylidene) hydrazinecarbothioamide (compound 1) was synthesized and its antitumor activities were further investigated on a non-small cell lung cancer cell line (A549) along with its underlying mechanisms. Compound 1 showed significant anti-proliferative activity on A549 cells, which was further proved by colony formation experiment. Compound 1 also inhibits the invasion of A549 cells in a trans-well culture system. Moreover, compound 1 markedly induced apoptosis on A549 cells, and the ratio of Bcl-2/Bax was decreased while the amount of p53, Cleaved-Caspase 3 and Cleaved-PARP expression were increased significantly. Compound 1 decreased the mitochondrial membrane potential, while the content of reactive oxygen was increased obviously. It is revealed that compound 1 mediated cell cycle arrest in G0/G1 phase by reducing G1 phase dependent proteins, CDK4 and Cyclin D1. As a result, it is indicated that compound 1 induced apoptosis on A549 cells was realized by regulating ROS-mediated mitochondria-dependent signaling pathway. Copyright © 2017 Elsevier Inc. All rights reserved.

  13. 北冬虫夏草水提物对人肺腺癌细胞A549的作用%Effects of Cordyceps militaris extract on human pulmonary adenocarcinoma cell line A549

    Institute of Scientific and Technical Information of China (English)

    颜晶晶; 唐永范; 陆魏杰; 张鑫; 孙加源; 韩宝惠

    2011-01-01

    目的 探讨北冬虫夏草水提物(CME)对人肺腺癌细胞A549的作用.方法 以不同质量浓度的CME处理A549细胞24、48、72 h,CCK8法检测细胞生长抑制率.以0.5 mg/mL CME处理A549细胞(干预组)12、24和48 h,流式细胞术检测细胞周期和细胞凋亡率,Western blotting检测A549细胞增殖相关蛋白(p-ERK和p-Akt)和凋亡相关蛋白(caspase-3)的表达;以未予CME处理的A549细胞作为对照组.结果 A549细胞生长抑制率随CME质量浓度的升高和处理时间的延长而上升,呈明显的浓度-时间依赖性(P<0.01).与对照组比较,干预组G2/M期细胞比例明显增加(P<0.01);干预组细胞凋亡率显著高于对照组(P<0.01),且与CME处理时间呈显著正相关(r=0.995,P<0.01);干预组p-ERK和p-Akt表达明显低于对照组,且与CME处理时间呈显著负相关(r=-0.881,P<0.01;r=-0.932,P<0.01);干预组caspase-3表达显著高于对照组(P<0.05),且与CME处理时间呈显著正相关(r=0.681,P<0.01).结论 CME对A549细胞生长的抑制作用可能与其使细胞阻滞于G2/M期并诱导细胞凋亡有关;CME有望成为肺腺癌辅助治疗药物.%Objective To investigate the effects of Cordyceps militaris extract ( CME) on human pulmonary adenocarcinoma cell line A549. Methods A549 cells were treated with CME of different mass concentrations for 24 h, 48 h and 72 h, and the inhibition rates of A549 cell proliferation were measured by CCK8 assay. A549 cells were treated with 0.5 nig/mL of CME (treatment group) for 12 h, 24 h and 48 h, cell cycle and cell apoptosis rates were determined by flow cytometry, and the expression of proliferation-related proteins (p-ERK and p-Akt) and apoptosis related protein (caspase-3) of A549 cells was determined by Western blotting. A549 cells without treatment with CME were served as control group. Results The inhibition rates of A549 cell proliferation increased with the mass concentrations of CME and time of treatment with CME

  14. Claudin-18 inhibits cell proliferation and motility mediated by inhibition of phosphorylation of PDK1 and Akt in human lung adenocarcinoma A549 cells.

    Science.gov (United States)

    Shimobaba, Shun; Taga, Saeko; Akizuki, Risa; Hichino, Asami; Endo, Satoshi; Matsunaga, Toshiyuki; Watanabe, Ryo; Yamaguchi, Masahiko; Yamazaki, Yasuhiro; Sugatani, Junko; Ikari, Akira

    2016-06-01

    Abnormal expression of claudin subtypes has been reported in various cancers. However, the pathological role of each claudin has not been clarified in detail. Claudin-18 was absent in human non-small cell and small cell lung cancers, although it is expressed in normal lung tissues. Here, we examined the effect of claudin-18 expression on the expression of junctional proteins, cell proliferation, and cell motility using human lung adenocarcinoma A549 cells. Real-time PCR and western blotting showed that exogenous expression of claudin-18 had no effect on the expression of junctional proteins including claudin-1, zonula occludens-1 (ZO-1), occludin, and E-cadherin. Claudin-18 was mainly distributed in cell-cell contact areas concomitant with ZO-1. Cell proliferation was significantly decreased at 48 and 72h after seeding of claudin 18-expressing cells. Claudin-18 suppressed cell motility, whereas it increased cell death in anoikis. Claudin-18 decreased phosphorylated (p)-3-phosphoinositide-dependent protein kinase-1 (PDK1) and p-Akt levels without affecting p-epidermal growth factor receptor and p-phosphatidylinositol-3 kinase (PI3K) levels. Furthermore, claudin-18 was bound with PDK1 and suppressed the nuclear localization of PDK1. We suggest that claudin-18 suppresses the abnormal proliferation and motility of lung epithelial cells mediated by inhibition of the PI3K/PDK1/Akt signaling pathway.

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

    Energy Technology Data Exchange (ETDEWEB)

    Zhang, Jian, E-mail: zhangjian197011@yahoo.com [Department of Respiratory Medicine, Xijing Hospital, The Fourth Military Medical University, Xi' an 710032 (China); Zhang, Tao [Department of Thoracic Surgery, Tangdu Hospital, The Fourth Military Medical University, Xi' an 710038 (China); Ti, Xinyu; Shi, Jieran; Wu, Changgui; Ren, Xinling [Department of Respiratory Medicine, Xijing Hospital, The Fourth Military Medical University, Xi' an 710032 (China); Yin, Hong, E-mail: yinnhong@yahoo.com [The Medical Image Center, Xijing Hospital, The Fourth Military Medical University, Xi' an 710032 (China)

    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 of 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.

  16. Extract of menispermum dauricum induces apoptosis of human lung cancer cell line A549%蝙蝠葛提取物对人肺癌细胞系A549诱导凋亡作用的研究

    Institute of Scientific and Technical Information of China (English)

    王永刚; 孙抒; 杨万山; 孙凤丹; 刘全

    2011-01-01

    目的 探讨蝙蝠葛提取物对人肺癌细胞系A549诱导凋亡和抗增殖作用及其机制,为开发抗肿瘤新中药提供实验依据.方法 应用MTT法测定蝙蝠葛提取物对人肺癌细胞系A549的生长抑制作用;通过倒置显微镜、光学显微镜观察肿瘤细胞凋亡的形态学变化;采用流式细胞术检测A549细胞的凋亡率;应用免疫组织化学技术检测药物处理前后凋亡相关蛋白酶caspase-3、caspase-8、caspase-9的表达.结果 (1) MTT法检测结果:蝙蝠葛提取物对人肺癌细胞系A549有明显的抑制生长的作用,且呈现出浓度的依赖性;(2)倒置显微镜观察结果:实验组肿瘤细胞体积变小、变圆,核染色质凝集,细胞间连接疏松,贴壁能力减弱;(3) HE染色观察结果:实验组肿瘤细胞体积变小、变圆,核染色质浓缩或染色质块形成,有的细胞膜起泡形成凋亡小体;(4)流式细胞术检测结果显示:蝙蝠葛提取物可以诱导A549细胞发生凋亡,加药组出现亚二倍体峰.结论 (1)蝙蝠葛提取物在体外对人肺癌细胞系A549有显著的诱导凋亡作用;(2)蝙蝠葛提取物诱导凋亡作用机制可能通过上调caspase-3、caspase-8和caspase-9蛋白表达,经由细胞凋亡的死亡受体和线粒体通路完成凋亡的启动和执行;(3)蝙蝠葛提取物具有显著的体外抗肿瘤作用,有望开发成一种新的抗肿瘤药物.%Objective To investigate the effects of menispermum dauricum extract on apoptosis of human lung cancer cell line A349. Methods MTI assay was used to determine the effects of menispermum dauricum extract on the growth inhibition of human lung cancer A549 cells. Morphological changes of the apoptosis of A549 cells were observed by using inverted microscope. Flow cytometry was used to detected apoptotic rate of A549 cells. caspase-3, caspase-8 and caspase-9 proteins were determined by immunocytochemical S-P staining technique. Results MTT test showed that menispermum dauricum extract

  17. CDK-associated Cullin 1 promotes cell proliferation with activation of ERK1/2 in human lung cancer A549 cells

    Energy Technology Data Exchange (ETDEWEB)

    Chen, Tian Jun [Respiratory Department, The First Affiliated Hospital, Xi’an Jiaotong University College of Medicine, Xi’an 710061 (China); Gao, Fei [Hua-shan Central Hospital of Xi’an, Xi’an 710043 (China); Yang, Tian; Thakur, Asmitanand; Ren, Hui; Li, Yang; Zhang, Shuo; Wang, Ting [Respiratory Department, The First Affiliated Hospital, Xi’an Jiaotong University College of Medicine, Xi’an 710061 (China); Chen, Ming Wei, E-mail: xjtucmw@163.com [Respiratory Department, The First Affiliated Hospital, Xi’an Jiaotong University College of Medicine, Xi’an 710061 (China)

    2013-07-19

    Highlights: •CDK-associated Cullin 1 (CAC1) expression increases in human lung carcinoma. •CAC1 promotes the proliferation of lung cancer A549 cells. •CAC1 promotes human lung cancer A549 cell proliferation with activation of ERK1/2. -- Abstract: Lung cancer is one of the most common causes of cancer-related death in the world, but the mechanisms remain unknown. In this study, we investigated the expression of CDK-associated Cullin 1 (CAC1) in lung cancer, the effect of CAC1 on the proliferation of human lung cancer A549 cells, and the activation of signaling pathways of mitogen-activated protein kinases (MAPKs). Results showed that CAC1 expression was higher levels in human lung carcinoma than normal lung tissue, and CAC1 siRNA reduced the proliferation of lung cancer A549 cells by decreasing cell activity and cell division in vitro. The proportion of cells treated with CAC1 siRNA increased in the G1 phase and decreased in the S and G2/M phase, indicative of G1 cell cycle arrest. Furthermore, the proportions of early/late apoptosis in lung cancer A549 cells were enhanced with CAC1 siRNA treatment. It was also found that activation of extracellular signal-regulated protein kinase (ERK) and p38 signaling pathways were involved in the proliferation of A549 cells. After CAC1 siRNA treatment, p-ERK1/2 levels decreased, and meanwhile p-p38 level increased, A549 cell proliferation increased when ERK1/2 signaling is activated by PMA. Our findings demonstrated that CAC1 promoted the proliferation of human lung cancer A549 cells with activation of ERK1/2 signaling pathways, suggesting a potential cure target for treatment of human lung cancer.

  18. The Study of CpG Island Methylation of BRCA1 Gene Promoter in a Taxol Induced Drug-resistant Human Lung Aadenocarcinoma Cell Line A549%耐紫杉醇人肺腺癌A549细胞株中BRCA1基因启动子CpG岛甲基化的研究

    Institute of Scientific and Technical Information of China (English)

    尹红英; 王红兵

    2012-01-01

    目的 检测耐紫杉醇人肺腺癌A549细胞株(A549/Taxol)中BRCA1基因启动子CpG岛甲基化状态,探讨A549/Taxol细胞对紫杉醇的耐药机制.方法 应用甲基化特异性聚合酶链反应(MSP)技术,检测耐紫杉醇人肺腺癌A549细胞株BRCA1基因启动子CpG岛甲基化状态.结果 A549/Taxol细胞存在BRCA1基因异常甲基化,呈部分甲基化.结论 A549/Taxol细胞存在BRCA1基因异常甲基化,可能是A549/Taxol细胞对紫杉醇耐药的机制之一.%Objective To detect the CpG island methylation status of BRCA1 gene promoter in the Taxol induced drug-resistant human lung adenocarcinoma cell line A549 ( A549/Taxol ), and to explore the resistance mechanisms of A549/Taxol. Methods A549/Taxol were examined CpG island methylation of BRCA1 gene promoter by methylation specific PCR ( MSP ). Results IBRCA1 gene aberrant methylation of A549/Taxol cells is part of methylation. Conclusion BRCA1 gene aberrant methylation of A549/Taxol may be one of the resistance mechanisms of taxol in A549/Taxol.

  19. Proteasome Inhibitor YSY01A Abrogates Constitutive STAT3 Signaling via Down-regulation of Gp130 and JAK2 in Human A549 Lung Cancer Cells

    Directory of Open Access Journals (Sweden)

    Wei Huang

    2017-08-01

    Full Text Available Proteasome inhibition interfering with many cell signaling pathways has been extensively explored as a therapeutic strategy for cancers. Proteasome inhibitor YSY01A is a novel agent that has shown remarkable anti-tumor effects; however, its mechanisms of action are not fully understood. Here we report that YSY01A is capable of suppressing cancer cell survival by induction of apoptosis. Paradoxically, we find that YSY01A abrogates constitutive activation of STAT3 via proteasome-independent degradation of gp130 and JAK2, but not transcriptional regulation, in human A549 non-small cell lung cancer cells. The reduction in gp130 and JAK2 can be restored by co-treatment with 3-methyladenine, an early-stage autophagy lysosome and type I/III PI3K inhibitor. YSY01A also effectively inhibits cancer cell migration and lung xenograft tumor growth with little adverse effect on animals. Thus, our findings suggest that YSY01A represents a promising candidate for further development of novel anticancer therapeutics targeting the proteasome.

  20. Grape proanthocyanidins induce apoptosis by loss of mitochondrial membrane potential of human non-small cell lung cancer cells in vitro and in vivo.

    Directory of Open Access Journals (Sweden)

    Tripti Singh

    Full Text Available Lung cancer remains the leading cause of cancer-related deaths worldwide, and non-small cell lung cancer (NSCLC represents approximately 80% of total lung cancer cases. The use of non-toxic dietary phytochemicals can be considered as a chemotherapeutic strategy for the management of the NSCLC. Here, we report that grape seed proanthocyanidins (GSPs induce apoptosis of NSCLC cells, A549 and H1299, in vitro which is mediated through increased expression of pro-apoptotic protein Bax, decreased expression of anti-apoptotic proteins Bcl2 and Bcl-xl, disruption of mitochondrial membrane potential, and activation of caspases 9, 3 and poly (ADP-ribose polymerase (PARP. Pre-treatment of A549 and H1299 cells with the caspase-3 inhibitor (z-DEVD-fmk significantly blocked the GSPs-induced apoptosis of these cells confirmed that GSPs-induced apoptosis is mediated through activation of caspases-3. Treatments of A549 and H1299 cells with GSPs resulted in an increase in G1 arrest. G0/G1 phase of the cell cycle is known to be controlled by cyclin dependent kinases (Cdk, cyclin-dependent kinase inhibitors (Cdki and cyclins. Our western blot analyses showed that GSPs-induced G1 cell cycle arrest was mediated through the increased expression of Cdki proteins (Cip1/p21 and Kip1/p27, and a simultaneous decrease in the levels of Cdk2, Cdk4, Cdk6 and cyclins. Further, administration of 50, 100 or 200 mg GSPs/kg body weight of mice by oral gavage (5 d/week markedly inhibited the growth of s.c. A549 and H1299 lung tumor xenografts in athymic nude mice, which was associated with the induction of apoptotic cell death, increased expression of Bax, reduced expression of anti-apoptotic proteins and activation of caspase-3 in tumor xenograft cells. Based on the data obtained in animal study, human equivalent dose of GSPs was calculated, which seems affordable and attainable. Together, these results suggest that GSPs may represent a potential therapeutic agent for the non-small

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

    Science.gov (United States)

    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.

  2. Brazilian green propolis induced apoptosis in human lung cancer A549 cells through mitochondrial-mediated pathway.

    Science.gov (United States)

    Frión-Herrera, Yahima; Díaz-García, Alexis; Ruiz-Fuentes, Jenny; Rodríguez-Sánchez, Hermis; Sforcin, José Maurício

    2015-10-01

    Propolis effect on the growth and apoptosis of human lung adenocarcinoma (A549 cells) was investigated as well as its mechanisms. Cells were incubated with propolis for 72 h, and 3-(4,5- dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide and lactate dehydrogenase assays were employed to assess cell viability and the inhibitory concentration (IC). Apoptosis was detected by Acridine Orange/Ethidium Bromide and 4',6-diamidino-2-phenylindole staining after 24 and 48 h of incubation with ¼ IC50 of propolis by testing the mitochondrial membrane potential (ΔΨm) and the expression of apoptosis-related genes (p53, Caspase-3, Bax, Bcl-2, Bcl-XL , Noxa, Puma and p21) by reverse transcription polymerase chain reaction. Propolis displayed antiproliferative and cytotoxic effects on A549 cells in a dose- and time-dependent manner, but it did not suppress the growth of normal Vero cells. An enhanced apoptosis was seen in A549 propolis-treated cells after 48 h compared with the control cells. Propolis decreased mitochondrial membrane potential by overexpression of pro-apoptotic genes (Bax and Noxa) and reduction of the antiapoptotic gene Bcl-XL . The expression level of other genes remained unchanged (p53, Caspse-3 and Bax), whereas p21 expression was increased. Propolis induced caspase-independent apoptosis through a p53-independent mitochondrial pathway, and cell cycle arrest by upregulation of p21. Although propolis induces apoptosis mainly by p53-independent manner, it may be induced by another pathway, and new insights may arise for preventing or treating lung cancer. © 2015 Royal Pharmaceutical Society.

  3. Nanoparticles of Selaginella doederleinii leaf extract inhibit human lung cancer cells A549

    Science.gov (United States)

    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.

  4. THE EFFECT OF IRISQUINONE ON THE GLUTATHIONE SYSTEM AND MRP EXPRESSION OF CISPLATIN-RESISTANT HUMAN LUNG ADENOCARCINOMA CELL LINE (A549DDP)

    Institute of Scientific and Technical Information of China (English)

    LIANG; li

    2001-01-01

    [1] Li DH. A novel radiosensitizer "ANKA" for tumor (Irisquinone) [J]. Chin J Clin Oncol 1999; 26:153.[2]Bordow SB, Haber M, Madafiglio J, et al. Expression of the multidrug resistance-associated protein (MRP) gene correlates with amplification and overexpression of the N-myc oncogene in childhood neuroblastoma [J]. Cancer Res 1994; 54:5036.[3]Cai P, Liu XY, Han FS, et al. Establishment human lung adenocarcinoma cisplatin-resistant cell line A549DDP and the mechanism of its drug resistance [J]. Chin J Clin Oncol 1995; 22:582.[4]Cai P, Liu XY, Wang P. The value of glutathione reductase recycling assay measurement of content of glutathione in human plasma during tumor chemotherapy [J]. Chin J Clin Oncol l994; 21:717.[5]Zhan MC, Liu XY, Cai P, et al. Mechanism of resistance of human cell line A549DDP to cisplatin [J]. Chin J Clin Oncol 1998; 25:726.[6]Wang J, Liu XY, Wu MN, et al. Expression and reversion of drug resistance- and apoptosis- related genes of a DDP-resistant lung adeno-carcinoma cell line A549DDP [J]. Chin J Oncol 1999; 21:422.[7]Ishikawa T. The ATP-dependent glutathione S-conjugate export pump [J]. Treads Biol Sci 1992; 17:463.[8]Goto S, Yoshida K, Morikawa T, et al. Augmen-tation of transport for cisplatin-glutathione adduct in cisplatin-resistant cancer cells [J]. Cancer Res 1995; 55:4297.[9]Fujil R, Mutoh M, Sumizama T, et al. Adenosine triphosphate-dependent transport of leukotriene C4 by membrane vesicles prepared from cis-platinum-resistant human epidermoid carcinoma tumor cells [J]. JNCI 1994; 86:1781.[10]Ishikawa T, Ali-Osman F. Glutathion-associated cis-diamminedichloroplatinum (II) metabolism and ATP-dependent efflux from leukemia cells [J]. J Biol Chem 1993; 268:20116.[11]Ishikawa T, Wrighe CE, Ishizuka H. GS-X pumq is function ally overexpressed in cis-diammine-dichloroplatinum (II)-resistant human leukemia HL-60 cells and downregulated by cell differentiation [J]. J Biol Chem 1994; 269: 29085.

  5. Toxicity of engineered nanomaterials and their transformation products following wastewater treatment on A549 human lung epithelial cells

    Directory of Open Access Journals (Sweden)

    Yanjun Ma

    2014-01-01

    Full Text Available Here we characterize the toxicity of environmentally-relevant forms of engineered nanomaterials (ENMs, which can transform during wastewater treatment and persist in aqueous effluents and biosolids. In an aerosol exposure scenario, cytotoxicity and genotoxicity of effluents and biosolids from lab-scale sequencing batch reactors (SBRs to A549 human lung epithelial cells were examined. The SBRs were dosed with nanoAg, nano zero-valent iron (NZVI, nanoTiO2 and nanoCeO2 at sequentially increasing concentrations from 0.1 to 20 mg/L. Toxicities were compared to outputs from SBRs dosed with ionic/bulk analogs, undosed SBRs, and pristine ENMs. Pristine nanoAg and NZVI showed significant cytotoxicity to A549 cells in a dose-dependent manner from 1 to 67 μg/mL, while nanoTiO2 and nanoCeO2 only exerted cytotoxicity at 67 μg/mL. Only nanoAg induced a genotoxic response, at 9, 33 and 53 μg/mL. However, no significant cytotoxic or genotoxic effects of the SBR effluents or biosolids containing nanomaterials were observed.

  6. LMO4基因沉默对Snail诱导的非小细胞肺癌A549细胞上皮-间质转化的影响研究%Impact of LMO4 Gene Silencing on Snail-induced EMT of Non-small Cell Lung Cancer A549 Cells

    Institute of Scientific and Technical Information of China (English)

    王雯珺; 伍思培; 列璞怡; 郭敏章; 何建行

    2016-01-01

    目的 探讨LMO4基因沉默对Snail诱导的非小细胞肺癌A549细胞上皮-间质转化(EMT)的影响.方法 体外培养非小细胞肺癌A549细胞,取对数生长期细胞进行实验,根据转染慢病毒分为sh-NC组(转染sh-NC慢病毒)、sh-LMO4组(转染sh-LMO4慢病毒)、Snail组(转染Snail慢病毒)和Snail+ sh-LMO4组(转染Snail慢病毒和sh-LMO4慢病毒).采用实时定量RT-PCR检测sh-NC组和sh-LMO4组LMO4mRNA相对表达量,采用Western Bloting法检测sh-NC组和sh-LMO4组LMO4蛋白相对表达量及4组N-eadherin、Vimentin、E-cadherin蛋白相对表达量,采用Transwell实验检测4组细胞侵袭个数,采用细胞划痕实验检测4组细胞迁移距离.结果 sh-LMO4组LMO4 mRNA相对表达量和LMO4蛋白相对表达量均低于sh-NC组(P<0.05).Snail组侵袭细胞个数多于sh-NC组,sh-LMO4组侵袭细胞个数少于sh-NC组,Snail+ sh-LMO4组侵袭细胞个数少于Snail组、多于sh-LMO4组(P<0.05).培养24 h后Snail组细胞迁移距离长于sh-NC组,sh-LMO4组细胞迁移距离短于sh-NC组,而Snail+ sh-LMO4组细胞迁移距离短于Snail组、长于sh-LMO4组(P<0.05).Snail组N-cadherin和Vimentin蛋白相对表达量高于sh-NC组,E-eadherin蛋白相对表达量低于sh-NC组(P<0.05);sh-LMO4组N-cadherin和Vimentin蛋白相对表达量低于sh-NC组,E-eadherin蛋白相对表达量高于sh-NC组(P<0.05);Snail+ sh-LMO4组N-cadherin和Vimentin蛋白相对表达量低于Snail组,E-cadherin蛋白相对表达量高于Snail组(P<0.05);Snail+ sh-LMO4组N-cadherin和Vimentin蛋白相对表达量高于sh-LMO4组,E-cadherin蛋白相对表达量低于sh-LMO4组(P<0.05).结论 LMO4基因沉默可逆转Snail诱导的非小细胞肺癌A549细胞EMT.

  7. The difference between multi-drug resistant cell line A549/Gem and its parental cell A549%多药耐药细胞株A549/Gem及其亲代细胞A549之间的区别研究

    Institute of Scientific and Technical Information of China (English)

    Weixia Wang; Xiaoqing Liu; Chuanhao Tang

    2009-01-01

    Objective: To discuss the difference between multi-drug resistant cell line A549/Gem and its parental cell A549 on the basis of establishment of human gemcitabine-resistant cell line A549/Gem so as to elaborate the possible mechanisms of gemcitabine resistance. Methods: Human gemcitabine-resistant non-small cell lung cancer cell line A549/Gem was estab-lished by the method of repeated clinical serous peak concentration plus gradually increasing concentration of gemcitabine from its parental cell human lung adenocaroinoma cell line A549 which was sensitive to gemcitabine. During the course of inducement, we had monitored their morphology, checked their resistance indexes and resistant pedigree by MTT method, gathered their growth curves and calculated their doubling time, examined their DNA contents and cell cycles by FCM; at the same time, we had measured their expressions of P53, EGFR, Cerb-B-2, PTEN, PCNA, c-myc, VEGF, MDR-1, Bcl-2, nm23, MMP-9, TIMP-1, and CD44v6 proteins via immunocytochemistry staining, RRM1 and ERCC1 mRNA by real-time fluorescent quantitative-PCR. Results: The resistance index of A549/Gem' cells (the deputy of cells in the process of inducement) to gemcitabine was 163.228, and the cell line also exhibited cross-resistance to vinorelbine, taxotere, fluorouraci, etoposide and cisplatin, but kept sensitivity to paclitaxol and oxaliplatin. The doubling time of A549/Gem' was shorter and figures in G0-G1 phases were increased than A549 cells. Compared with A549 cells, A549/Gem' cells achieved EGFR and c-myc proteins expressions, nm23 protein expression enhanced, P53, Cerb-B-2 and Bcl-2 proteins expressions reduced, PTEN ,PCNA and MDR-1 proteins expressions vanished, but those of MMP-9, VEGF, CD44v6 and TIMP-1 proteins changed trivially. Meanwhile, expressions of RRM1 and ERCC1 mRNA were augmented markedly. The resistance index of A549/Gem cells to gemcitabine was 129.783, and the cell line also held cross-resistance to vinorelbine, taxotere

  8. Alteration of membrane lipid biophysical properties and resistance of human lung adenocarcinoma A549 cells to cisplatin

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Alterations of membrane lipid biophysical properties of sensitiveA549 and resistant A549/DDP cells to the Cis-dichlorodiammine platinum (Cisplatin) were performed by measurements of fluorescence and flow cytometry approaches using fluorescence dyes of DPH, N-AS and Merocyanine 540 (MC 540) respectively. Fatty acids of membrane lipid of the two cell lines were analyzed by gas chromatography. The results indicated clearly that fluorescence polarization (P) of the DPH probe is 0.169 for the sensitive A549 cell and 0.194 for the resistant A549/DDP cells. Statistical analysis showed significant difference between the two cell lines. The polarizations of 2-AS and 7-AS which reflect the fluidity of surface and middle of lipid bilayer are 0.134 and 0.144 for the sensitive A549 cells as well as 0.171 and 0.178 for the resistant A549/DDP cells respectively, but there is no significant difference of the polarization of 12-AS between the two cell lines. This shows that altera-tions of the membrane fluidity of both cells were mainly located on the surface and middle of the lipid bilayer. In addition, the packing density of phospholipid molecules in the membrane of the two cell lines detected by MC540 probe indicated that lipid packing of A549 cell membranes was looser than that of the A549/DDP cells. And unsaturation degree of plasma membrane fatty acids of the A549/DDP cells was also lower than that of A549 cells. Taken together, it was proposed that the al-teration of membrane lipid biophysical state may be involved in the resistance of A549/DDP cells to cisplatin.

  9. Low Dose Hyper-radiosensitivity in Human Lung Cancer Cell Line A549 and Its Possible Mechanisms

    Institute of Scientific and Technical Information of China (English)

    Xiaofang DAI; Dan TAO; Hongge WU; Jing CHENG

    2009-01-01

    The low dose hyper-radiosensitivity (HRS) in human lung cancer cell line A549 was in-vestigated,the changes of ATM kinase,cell cycle and apoptosis of cells at different doses of radiation were observed,and the possible mechanisms were discussed.A549 cells in logarithmic growth phase were irradiated with 60Co γ-rays at doses of 0-2 Gy.Together with flow cytometry for precise cell sorting,cell survival fraction was measured by means of conventional colony-formation assay.The expression of ATM1981Ser-P protein was examined by Western blot 1 h after radiation.Apoptosis was detected by Hoechst 33258 fluorescent staining,and Annexin V-FITC/PI staining flow cytometry 24 h after radiation.Cell cycle distribution was observed by flow cytometly 6,12 and 24 h after ra-diation.The results showed that the expression of ATM1981Ser-P protein was observed at 0.2 Gy,followed by an increase at >0.2 Gy,and reached the peak at 0.5 Gy,with little further increase as the dose exceeded 0.5 Gy.Twenty-four h after radiation,partial cells presented the characteristic mor-phological changes of apoptosis,and the cell apoptosis curve was coincident with the survival curve.As compared with control group,the cell cycle almost had no changes after exposure to 0.1 and 0.2 Gy radiation (P>0.05).After exposure to 0.3,0.4 and 0.5 Cry radiation,G2/M phase arrest occurred 6 and 12 h after radiation (P<0.05),and the ratio of G2/M phase cells was decreased 24 h after radiation (P<0.05).It was concluded that A549 cells displayed the phenomenon of HRS/IRR.The mode of cell death was mainly apoptosis.The activity of ATM and cell cycle change may take an important role in HRS/IRR.

  10. Short-Course Treatment With Gefitinib Enhances Curative Potential of Radiation Therapy in a Mouse Model of Human Non-Small Cell Lung Cancer

    Energy Technology Data Exchange (ETDEWEB)

    Bokobza, Sivan M.; Jiang, Yanyan; Weber, Anika M.; Devery, Aoife M.; Ryan, Anderson J., E-mail: anderson.ryan@oncology.ox.ac.uk

    2014-03-15

    Purpose: To evaluate the combination of radiation and an epidermal growth factor receptor (EGFR) tyrosine kinase inhibitor (TKI) in preclinical models of human non-small cell lung cancer. Methods and Materials: Sensitivity to an EGFR TKI (gefitinib) or radiation was assessed using proliferation assays and clonogenic survival assays. Effects on receptor signal transduction pathways (pEGFR, pAKT, pMAPK) and apoptosis (percentage of cleaved PARP Poly (ADP-ribose) polymerase (PARP)) were assessed by Western blotting. Radiation-induced DNA damage was assessed by γH2AX immunofluorescence. Established (≥100 mm{sup 3}) EGFR-mutated (HCC287) or EGFR wild-type (A549) subcutaneous xenografts were treated with radiation (10 Gy, day 1) or gefitinib (50 mg/kg, orally, on days 1-3) or both. Results: In non-small cell lung cancer (NSCLC) cell lines with activating EGFR mutations (PC9 or HCC827), gefitinib treatment markedly reduced pEGFR, pAKT, and pMAPK levels and was associated with an increase in cleaved PARP but not in γH2AX foci. Radiation treatment increased the mean number of γH2AX foci per cell but did not significantly affect EGFR signaling. In contrast, NSCLC cell lines with EGFR T790M (H1975) or wild-type EGFR (A549) were insensitive to gefitinib treatment. The combination of gefitinib and radiation treatment in cell culture produced additive cell killing with no evidence of synergy. In xenograft models, a short course of gefitinib (3 days) did not significantly increase the activity of radiation treatment in wild-type EGFR (A549) tumors (P=.27), whereas this combination markedly increased the activity of radiation (P<.001) or gefitinib alone (P=.002) in EGFR-mutated HCC827 tumors, producing sustained tumor regressions. Conclusions: Gefitinib treatment increases clonogenic cell killing by radiation but only in cell lines sensitive to gefitinib alone. Our data suggest additive rather than synergistic interactions between gefitinib and radiation and that a

  11. TNF-α pro-inflammatory cytokinemodulates CD44 expression in human lung epithelial cell line (A549 treated with Temporin-Ra

    Directory of Open Access Journals (Sweden)

    Maryam Hooshmand

    2016-03-01

    Full Text Available Antimicrobial peptides (AMPs are molecules present in innate immune systems of vertebrates and invertebrates. These small peptides inhibit the growth of pathogens invading host's body. In this study, the toxicity effect of Temporin-Ra (T-Ra antimicrobial peptide on A549 cell line was investigated by MTT assay. Furthermore, the toxicity of T-Ra peptide on human's red and white blood cells was investigated. Gene expression levels of TNF-α (tumor necrosis factor-alphaaspro-inflammatory cytokine, and CD44 cancer marker were investigated by real time- PCR, 48 h after treatment of A549 cells by different concentrations of peptide. Moreover, the production of reactive oxygen species was studied by flow cytometer. According to our results, T-Ra viability of A549 cells decreased up to 15%, while had no hemolytic and cytotoxic effects on human blood cells. In addition, T-Ra increased the expression of pro- inflammatory cytokine (TNF-α at the highest concentration (30 µg/ml, while decreased gene expression of CD44 cancer marker. Production of reactive oxygen species (ROS in A549 cells treated by T-Ra significantly increased. In conclusion, our results revealed that T-Ra could induce the expression of TNF-α, which consequently decrease CD44 expression, increase the production of reactive oxygen species, and as a result induced death in A549 cell lines.

  12. Differential replication of avian influenza H9N2 viruses in human alveolar epithelial A549 cells

    Directory of Open Access Journals (Sweden)

    Peiris Malik

    2010-03-01

    Full Text Available Abstract Avian influenza virus H9N2 isolates cause a mild influenza-like illness in humans. However, the pathogenesis of the H9N2 subtypes in human remains to be investigated. Using a human alveolar epithelial cell line A549 as host, we found that A/Quail/Hong Kong/G1/97 (H9N2/G1, which shares 6 viral "internal genes" with the lethal A/Hong Kong/156/97 (H5N1/97 virus, replicates efficiently whereas other H9N2 viruses, A/Duck/Hong Kong/Y280/97 (H9N2/Y280 and A/Chicken/Hong Kong/G9/97 (H9N2/G9, replicate poorly. Interestingly, we found that there is a difference in the translation of viral protein but not in the infectivity or transcription of viral genes of these H9N2 viruses in the infected cells. This difference may possibly be explained by H9N2/G1 being more efficient on viral protein production in specific cell types. These findings suggest that the H9N2/G1 virus like its counterpart H5N1/97 may be better adapted to the human host and replicates efficiently in human alveolar epithelial cells.

  13. Digoxin Downregulates NDRG1 and VEGF through the Inhibition of HIF-1α under Hypoxic Conditions in Human Lung Adenocarcinoma A549 Cells

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    Dong Wei

    2013-04-01

    Full Text Available Digoxin, an inhibitor of Na+/K+ ATPase, has been used in the treatment of heart-related diseases (such as congestive heart failure and atrial arrhythmia for decades. Recently, it was reported that digoxin is also an effective HIF-1α inhibitor. We investigated whether digoxin could suppress tumor cell growth through HIF-1α in non-small cell lung cancer cells (A549 cells under hypoxic conditions. An MTT assay was used to measure cell viability. RT-PCR and western blotting were performed to analyze the mRNA and protein expression of VEGF, NDRG1, and HIF-1α. HIF-1α nuclear translocation was then determined by EMSA. Digoxin was found to inhibit the proliferation of A549 cells under hypoxic conditions. Our results showed that hypoxia led to the upregulation of VEGF, NDRG1, and HIF-1α both at the mRNA and protein levels. We also found that the hypoxia-induced overexpression of VEGF, NDRG1, and HIF-1α was suppressed by digoxin in a concentration-dependent manner. As expected, our EMSA results demonstrated that under hypoxic conditions HIF-1α nuclear translocation was also markedly reduced by digoxin in a concentration-dependent manner. Our results suggest that digoxin downregulated hypoxia-induced overexpression of VEGF and NDRG1 at the transcriptional level probably through the inhibition of HIF-1α synthesis in A549 cells.

  14. Growth-inhibiting and apoptosis-inducing activities of Myricanol from the bark of Myrica rubra in human lung adenocarcinoma A549 cells.

    Science.gov (United States)

    Dai, G H; Meng, G M; Tong, Y L; Chen, X; Ren, Z M; Wang, K; Yang, F

    2014-09-25

    Myrica rubra (Lour.) Sieb. Et Zucc. is a myricaceae Myrica plant. It is a subtropical fruit tree in China and other Asian countries. The bark of M. rubra is used in Chinese folk medicine because of its antibacterial, antioxidant, anti-inflammatory, and anticancer activities. However, the mechanisms underlying such activities remain unclear. This study investigated whether or not Myricanol extracted from M. rubra bark elicits anti-cancer effects on human lung adenocarcinoma A549 cells by inducing apoptosis in vivo. Myricanol was extracted from M. rubra bark through system solvent extraction and silica gel layer column separation. The results of tritiated thymidine assay, colony formation assay, and flow cytometry indicated that Myricanol inhibited the growth of A549 cells. The effects of Myricanol on the expression of key apoptosis-related genes in A549 cells were evaluated by quantitative PCR and Western blot analyses. Myricanol significantly inhibited the growth of A549 cells in a dose-dependent manner, with a half maximal inhibitory concentration of 4.85 μg/ml. Myricanol significantly decreased colony formation and induced A549 cell apoptosis. Myricanol upregulated the expression of Caspase-3, Caspase-9, Bax, and p21 and downregulated the expression of Bcl-2 at the mRNA and protein levels. These changes were associated with apoptosis. Based on these results, we propose that Myricanol elicits growth inhibitory and cytotoxic effects on lung cancer cells. Therefore, Myricanol may be a clinical candidate for the prevention and treatment of lung cancer.

  15. Effects of matrine on the growth inhibition, c-myc and hTERT protein expression in human adenocarcinoma lung cancer cell line A549

    Directory of Open Access Journals (Sweden)

    Qiong CHEN

    2008-08-01

    Full Text Available Background and objective It was reported that telomerase was associated with the oncogenesis and progression of cancer, and to be the common targets of cancer therapy. The mechanism of matrine on lung cancer in vitro is not clear. We studied the effect of matrine on growth of human lung adenocarcinoma A549 cells and the mechanism related with telomerase. Methods MTT was used for measuring A549 cells viability, Hoechst 33342-propidium iodide fluorescent staining for observing apoptotic cells, flow cytometry (FCM for analyzing cell cycle and apoptosis, and immunocytochemistry for measuring the protein expressions of c-myc and hTERT in A549 cells. Results Matrine inhibited the proliferation of A549 cells with a time-dose-dependent manner (P<0.05. Hoechst 33342-propidium iodide staining showed apoptotic cells with chromatin condensation and fragmentation of nuclei. FCM analysis indicated elevating rate of cells in G0/G1 phase, lowering rate of that in S phase and the highering apoptotic rate. The levels of c-myc and hTERT protein expression in the matrine group was lower than that in the control group (P<0.05, and AOD of c-myc showed positive correlation with AOD of hTERT (r=0.633, P<0.01 Conclusion The inhibitory effect of matrine on A549 cells may be related to the lower expression of c-myc and hTERT.

  16. Inhibition of proliferation, migration and invasion of human non ...

    African Journals Online (AJOL)

    of human non-small cell lung cancer cell line A549 and explore the possible mechanisms. Methods: The .... Figure 1: Chemical structure of phlomisoside F. Cell culture. Human non-small ... optical microscope (Olympus, Japan). For transwell ...

  17. Interaction between fragile histamine triad and protein kinase C alpha in human non-small cell lung cancer tissues

    Institute of Scientific and Technical Information of China (English)

    Peng-hui Zhuang; Zhao-hui Liu; Xiao-gang Jiang; Cheng-en Pan

    2009-01-01

    Objective To investigate the interaction between fragile histamine triad (FHIT) and protein kinase C alpha (PKCα) in human non-small cell lung cancer tissues. Methods FHIT and PKC伪 double positive samples were screened by immunohistochemical staining from 13 human non-small cell lung cancer tissues. Co-immunoprecipitation was performed by using anti-FHIT and anti-PKCα. The immune precipitate was analyzed by SDS-PAGE and Western blot. Results Immune precipitate staining detection showed that 3 samples out of the 13 cases were double positive for FHIT and PKCα. FHIT protein was present in the immune precipitate of anti-PKCα while there was PKCα in the immune precipitate of anti-FHITmAb. Conclusion FHIT and PKCα exist as a complex in human non-small cell lung cancer tissues, which will provide a new route for studying the pathogenesis and immunotherapy of human non-small cell lung cancer.

  18. Raman spectroscopy identifies radiation response in human non-small cell lung cancer xenografts

    Science.gov (United States)

    Harder, Samantha J.; Isabelle, Martin; Devorkin, Lindsay; Smazynski, Julian; Beckham, Wayne; Brolo, Alexandre G.; Lum, Julian J.; Jirasek, Andrew

    2016-02-01

    External beam radiation therapy is a standard form of treatment for numerous cancers. Despite this, there are no approved methods to account for patient specific radiation sensitivity. In this report, Raman spectroscopy (RS) was used to identify radiation-induced biochemical changes in human non-small cell lung cancer xenografts. Chemometric analysis revealed unique radiation-related Raman signatures that were specific to nucleic acid, lipid, protein and carbohydrate spectral features. Among these changes was a dramatic shift in the accumulation of glycogen spectral bands for doses of 5 or 15 Gy when compared to unirradiated tumours. When spatial mapping was applied in this analysis there was considerable variability as we found substantial intra- and inter-tumour heterogeneity in the distribution of glycogen and other RS spectral features. Collectively, these data provide unique insight into the biochemical response of tumours, irradiated in vivo, and demonstrate the utility of RS for detecting distinct radiobiological responses in human tumour xenografts.

  19. CCL21/CCR7 prevents apoptosis via the ERK pathway in human non-small cell lung cancer cells.

    Directory of Open Access Journals (Sweden)

    Ying Xu

    Full Text Available Previously, we confirmed that C-C chemokine receptor 7 (CCR7 promotes cell proliferation via the extracellular signal-regulated kinase (ERK pathway, but its role in apoptosis of non-small cell lung cancer (NSCLC cell lines remains unknown. A549 and H460 cells of NSCLC were used to examine the effect of CCL21/CCR7 on apoptosis using flow cytometry. The results showed that activation of CCR7 by its specific ligand, exogenous chemokine ligand 21 (CCL21, was associated with a significant decline in the percent of apoptosis. Western blot and real-time PCR assays indicated that activation of CCR7 significantly caused upregulation of anti-apoptotic bcl-2 and downregulation of pro-apoptotic bax and caspase-3, but not p53, at both protein and mRNA levels. CCR7 small interfering RNA significantly attenuated these effects of exogenous CCL21. Besides, PD98059, a selective inhibitor of MEK that disrupts the activation of downstream ERK, significantly abolished these effects of CCL21/CCR7. Coimmunoprecipitation further confirmed that there was an interaction between p-ERK and bcl-2, bax, or caspase-3, particularly in the presence of CCL21. These results strongly suggest that CCL21/CCR7 prevents apoptosis by upregulating the expression of bcl-2 and by downregulating the expression of bax and caspase-3 potentially via the ERK pathway in A549 and H460 cells of NSCLC.

  20. Nanostructured delivery system for zinc phthalocyanine: preparation, characterization, and phototoxicity study against human lung adenocarcinoma A549 cells

    Directory of Open Access Journals (Sweden)

    Mariana da Volta Soares

    2011-01-01

    Full Text Available Mariana da Volta Soares1, Mainara Rangel Oliveira1, Elisabete Pereira dos Santos1, Lycia de Brito Gitirana2, Gleyce Moreno Barbosa3, Carla Holandino Quaresma3, Eduardo Ricci-Júnior11Department of Medicines, Laboratório de Desenvolvimento Galênico (LADEG, Faculty of Pharmacy, 2Laboratory of Animal and Comparative Histology, Glycobiology Research Program, Institute of Biomedical Science, 3Department of Medicines, Laboratório Multidisciplinar de Ciências Farmacêuticas, Faculty of Pharmacy, Federal University of Rio de Janeiro (UFRJ, Rio de Janeiro, BrazilAbstract: In this study, zinc phthalocyanine (ZnPc was loaded onto poly-ε-caprolactone (PCL nanoparticles (NPs using a solvent emulsification–evaporation method. The process yield and encapsulation efficiency were 74.2% ± 1.2% and 67.1% ± 0.9%, respectively. The NPs had a mean diameter of 187.4 ± 2.1 nm, narrow distribution size with a polydispersity index of 0.096 ± 0.004, zeta potential of -4.85 ± 0.21 mV, and spherical shape. ZnPc has sustained release, following Higuchi’s kinetics. The photobiological activity of the ZnPc-loaded NPs was evaluated on human lung adenocarcinoma A549 cells. Cells were incubated with free ZnPc or ZnPc-loaded NPs for 4 h and then washed with phosphate-buffered saline. Culture medium was added to the wells containing the cells. Finally, the cells were exposed to red light (660 nm with a light dose of 100 J/cm2. The cellular viability was determined after 24 h of incubation. ZnPc-loaded NPs and free photosensitizer eliminated about 95.9% ± 1.8% and 28.7% ± 2.2% of A549 cells, respectively. The phototoxicity was time dependent up to 4 h and concentration dependent at 0–5 µg ZnPc. The cells viability decreased with the increase of the light dose in the range of 10–100 J/cm2. Intense lysis was observed in the cells incubated with the ZnPc-loaded NPs and irradiated with red light. ZnPc-loaded PCL NPs are the release systems that promise photodynamic

  1. Effects of fatty acids on benzo[a]pyrene uptake and metabolism in human lung adenocarcinoma A549 cells.

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    Rola Barhoumi

    Full Text Available Dietary supplementation with natural chemoprotective agents is receiving considerable attention because of health benefits and lack of toxicity. In recent in vivo and in vitro experimental studies, diets rich in n-3 polyunsaturated fatty acids have been shown to provide significant anti-tumor action. In this investigation, the effects of control fatty acids (oleic acid (OA, linoleic acid (LA and n-3 PUFA, e.g., docosahexaenoic acid (DHA on the uptake and metabolism of the carcinogenic polycyclic aromatic hydrocarbon, benzo[a]pyrene (BaP was investigated in A549 cells, a human adenocarcinoma alveolar basal epithelial cell line. A549 cells activate BaP through the cytochrome P450 enzyme system to form reactive metabolites, a few of which covalently bind to DNA and proteins. Therefore, multiphoton microscopy spectral analysis combined with linear unmixing was used to identify the parent compound and BaP metabolites formed in cells, in the presence and absence of fatty acids. The relative abundance of select metabolites was associated with altered P450 activity as determined using ethoxyresorufin-O-deethylase activity in cells cultured in the presence of BSA-conjugated fatty acids. In addition, the parent compound within cellular membranes increases significantly in the presence of each of the fatty acids, with the greatest accumulation observed following DHA treatment. DHA treated cells exhibit significantly lower pyrene-like metabolites indicative of lower adducts including DNA adducts compared to control BSA, OA or LA treated cells. Further, DHA reduced the abundance of the proximate carcinogen BaP 7,8-dihydrodiol and the 3-hydroxybenzo[a]pyrene metabolites compared to other treatments. The significant changes in BaP metabolites in DHA treated cells may be mediated by the effects on the physicochemical properties of the membrane known to affect enzyme activity related to phase I and phase II metabolism. In summary, DHA is a highly bioactive chemo

  2. Effects of sodium lactate Ringer's injection on transfection of human protein kinase C-α antisense oligonucleotide in A549 lung cancer cells.

    Science.gov (United States)

    Wang, Z H; Sun, W W; Han, Y L; Ma, Z

    2016-08-26

    In the present study, we evaluated the effects of four solutions [Dulbecco's modified Eagle's medium (DMEM), sodium lactate Ringer's injection (SLRI), phosphate-buffered saline (PBS), and NaCl] on the transfection of the human protein kinase C-a antisense oligonucleotide (PKC-a ASO) aprinocarsen in human lung carcinoma A549 cells. Specifically, SLRI, DMEM, PBS, or NaCl were used as the growth solutions for A549 cells, and OPTI-MEM was used as the PKC-a ASO diluent for transfection. Additionally, SLRI, DMEM, PBS, or NaCl were used as both the growth solutions and diluents for transfection. The cell viability and transfection efficiency were determined. The results demonstrated that when SLRI was used as either the growth solution or both the growth solution and diluent for aprinocarsen transfection in A549 cells, the effects were close to the best effects observed with DMEM as the growth solution and OPTI-MEM as the diluent, which supported the transfection of aprinocarsen into the cells. Moreover, SLRI resulted in higher transfection efficiency than those of PBS and NaCl. In in vitro experiments, aprinocarsen effectively induced apoptosis in A549 cells. In conclusion, SLRI may replace PBS or NaCl in clinical trials as a transfection solution readily accepted by the human body. To our knowledge, this is the first report demonstrating the use of SLRI as a transfection solution in lung-cancer cell lines.

  3. A study of the effect of CCL21/CCR7 axis on VEGF-C expression in human lung adenocarcinoma A549 cells%CCL21/CCR7轴对人肺癌A549细胞VEGF-C表达影响的研究

    Institute of Scientific and Technical Information of China (English)

    郭学光; 陈正堂; 刘长庭

    2011-01-01

    目的 研究二级淋巴组织趋化因子/CC趋化因子受体7(CCL21/CCR7)轴对肺癌A549细胞血管内皮细胞生长因子-C(VEGF-C)表达的影响.方法 实时定量PCR及Western Blot法检测CCL21作用前后A549细胞及A549-CCR7细胞VEGF-C mRNA及蛋白的表达.结果 CCL21作用下无论是在mRNA水平还是蛋白水平A549-CCR7细胞VEGF-C的表达均较A549细胞高.结论 CCL21与其受体CCR7结合能够促进A549细胞VEGF-C的表达,CCL21/CCR7轴可能参与了肺癌淋巴结转移的过程.进一步研究CCL21/CCR7轴和VEGF-C 的关系可能有助于阐明肺癌淋巴结转移的机制.%Objective To investigate the effects of CCL21/CCR7 axis on VEGF - C expression of human lung adenocarcinoma A549 cells and A549 - CCR7 cells. Methods VEGF - C expression was detected using Real - Time RT - PCR and Western Blot in A549 cells and A549 -CCR7 cells before and after incuhation with CCL21 . Results Under the influence of CCL21 , the VEGF - C expression of the A549 - CCR7 cells ,in mRNA and protein levels, was significantly increased compared to that of A549 cells ( P <0. 01 ). Conclusion These data suggest that the bind of CCL21 to its receptor CCR7 leads to the increase of the VE(-JF - C expression of A549 cells, and CCL21/CCR7 axis may play a role in lymph node metastasis of lung cancer. To further study the relationship between CCL21/CCR7 axis and VEGF - C may help to elucidate the mechanism of lymph node metastasis in human lung cancer.

  4. Gliotoxin promotes Aspergillus fumigatus internalization into type II human pneumocyte A549 cells by inducing host phospholipase D activation.

    Science.gov (United States)

    Jia, Xiaodong; Chen, Fangyan; Pan, Weihua; Yu, Rentao; Tian, Shuguang; Han, Gaige; Fang, Haiqin; Wang, Shuo; Zhao, Jingya; Li, Xianping; Zheng, Dongyu; Tao, Sha; Liao, Wanqing; Han, Xuelin; Han, Li

    2014-06-01

    The internalization of Aspergillus fumigatus into lung epithelial cells is critical for the infection process in the host. Gliotoxin is the most potent toxin produced by A. fumigatus. However, its role in A. fumigatus internalization into the lung epithelial cells is still largely unknown. In the present study, the deletion of the gliP gene regulating the production of gliotoxin in A. fumigatus suppressed the internalization of conidia into the A549 lung epithelial cells, and this suppression could be rescued by the exogenous addition of gliotoxin. At lower concentrations, gliotoxin enhanced the internalization of the conidia of A. fumigatus into A549 cells; in contrast, it inhibited the phagocytosis of J774 macrophages in a dose-dependent manner. Under a concentration of 100 ng/ml, gliotoxin had no effect on A549 cell viability but attenuated ROS production in a dose-dependent manner. Gliotoxin significantly stimulated the phospholipase D activity in the A549 cells at a concentration of 50 ng/ml. This stimulation was blocked by the pretreatment of host cells with PLD1- but not PLD2-specific inhibitor. Morphological cell changes induced by gliotoxin were observed in the A549 cells accompanying with obvious actin cytoskeleton rearrangement and a moderate alteration of phospholipase D distribution. Our data indicated that gliotoxin might be responsible for modulating the A. fumigatus internalization into epithelial cells through phospholipase D1 activation and actin cytoskeleton rearrangement.

  5. Photodynamic Activity and Security of Five New Hypocrellins Derivatives in Human Lung Cancer A549 Cells%五种竹红菌素衍生物对A549细胞的光动力效应研究

    Institute of Scientific and Technical Information of China (English)

    张露勇; 顾瑛; 赵井泉; 邱海霞; 曾晶

    2013-01-01

    目的 比较五种新型竹红菌素衍生物分别为竹红菌素乙素(hypocrellin,HB)的二位ω-氨基磺酸衍生物THB、3HB和4HB,及十七位ω-氨基磺酸衍生物3SB和4SB对体外培养的人肺腺癌上皮细胞(A549)的光动力(photodynamic therapy,PDT)效应,筛选光动力活性和安全性较好的竹红菌素衍生物.方法 (1)杀伤效应.将0.94 nmol/ml的5种新型竹红菌素衍生物和HB分别与A549细胞孵育4h后,分别以波长630和532 nm激光照射,功率密度20 mW/cm2,照射时间1000s,能量密度20 J/cm2,照光后继续避光孵育24 h后采用MTT法测定细胞存活率.(2)安全系数.分别以波长532和630nm激光照射,以血卟啉(hematoporph-yrin derivative,HpD)为对照光敏剂,研究17-4-amino-1-butane-sulfonic acid-hypocrellin B(4SB)对A549细胞的光动力效应及和暗毒性,并比较安全系数(暗毒性IC50/光毒性IC50).结果 (1)杀伤效应.五种竹红菌素衍生物中,4SB在630和532 nm激光照射下对A549的光动力杀伤作用强于其它衍生物,接近HB.(2)安全系数.波长532 nm激光照射,4SB的光毒性分别为103.86和84.16 ng/ml是HpD 960.14 ng/ml的10.53和11.4倍,但前两者之间差异无显著意义(P>0.05);波长630 nm激光照射下,4SB光毒性的IC50为50.7 ng/ml,HpDIC50为1 069.88 ng/ml,暗毒性HpD、4SB分别为7.84、21.93μg/ml,安全系数4SB(432.5)>HpD(7.3).HpD在532和630 nm两波长下的光毒性IC50差异无显著意义(P>0.05),而4SB在532和630hm两波长下的光毒性差异有显著意义(P<0.05).结论 5种衍生物可能成为有价值的光敏剂,值得进一步深入研究.%Objective To study the photodynamic activity of five new hypocrellins derivatives in human lung cancer A549 cells line,filtrate these hypocrellin derivatives,discuss their metabolism and transport in vivo,and study the combination of hypocrellin derivative molecule with HSA for the sake of driving photosensitive drug development.Methods (1) Spectral properties of hypocrellin

  6. Inhibitory Effect of Cantharidin on Proliferation of A549 Cells

    Institute of Scientific and Technical Information of China (English)

    WANG Xiao-hua; YIN Yuan-qin; SUI Cheng-guang; MENG Fan-dong; MA Ping; JIANG You-hong

    2007-01-01

    Objective: To study the inhibition of Cantharidin against the proliferation of human lung cancer A549 cells and its mechanism. Methods: MTT assay was employed to determine the inhibition of Cantharidin against proliferation of A549 cells and flow Cytometry was applied to analyze A549 cell cycle and the effect of Cantharidin on cell cycle. Results: Cantharidin showed inhibition against the proliferation of A549 cells, and the inhibition was mediated by blocking A549 cell cycle at G2/M phase significantly. Conclusion: Cantharidin exhibits inhibition against the proliferation of human lung cancer A549 cells.

  7. Safrole oxide induces apoptosis by up-regulating Fas and FasL instead of integrin beta4 in A549 human lung cancer cells.

    Science.gov (United States)

    Du, AiYing; Zhao, BaoXiang; Miao, JunYing; Yin, DeLing; Zhang, ShangLi

    2006-04-01

    Previously, we found that 3,4-(methylenedioxy)-1-(2',3'-epoxypropyl)-benzene (safrole oxide) induced a typical apoptosis in A549 human lung cancer cells by activating caspase-3, -8, and -9. In this study, we further investigated which upstream pathways were activated by safrole oxide during the apoptosis. Immunofluorescence assay combined with laser scanning confocal microscopy revealed that both Fas and Fas ligand (FasL) were up-regulated by the small molecule. In addition, Fas protein distribution was altered, showing a clustering distribution instead of a homogeneous one. Subsequently, Western blot analysis confirmed the up-regulations of Fas and its membrane-binding form of FasL (m-FasL), as well as P53 protein. Conversely, safrole oxide hardly affected integrin beta4 subunit expression or distribution, which was reflected from the data obtained by immunofluorescence assay combined with laser scanning confocal microscopy. The results suggested that Fas/FasL pathway might be involved in safrole oxide-induced apoptosis of A549 cells, while integrin beta4 might be irrelevant to the apoptosis. Nevertheless, we first found the strong expression of integrin beta4 in A549 cells. The study first suggested that safrole oxide might be used as a small molecular promoter of Fas/FasL pathway to elicit apoptosis in A549 cells, which would lay the foundation for us to insight into the new strategies for lung cancer therapy.

  8. 莪术油对人肺腺癌细胞A549增殖的影响%Effect of Zedoary Turmeric Oil on Proliferation in Human Lung Adenocarcinoma Cell Line A549

    Institute of Scientific and Technical Information of China (English)

    王晓波; 牛建昭; 崔巍; 刘飒; 杨长福; 赵丕文; 唐炳华

    2011-01-01

    目的 探讨莪术油对人肺腺癌细胞A549增殖的抑制作用.方法 体外培养肺腺癌细胞A549,MTT比色法测定莪术油对A549细胞作用24、48、72 h后抑制率;流式细胞术分析莪术油对A549细胞作用24 h后细胞周期的变化;Annexin V-FITC/PI双染检测莪术油对A549细胞作用24 h后细胞凋亡与坏死情况.结果 莪术油对A549细胞增殖的抑制率随时间延长明显升高,随药物浓度增加抑制作用增强;莪术油对A549细胞作用24 h后,细胞周期停滞在G0/G1期,阻止其进入S期;细胞的早期凋亡、晚期凋亡和坏死比例随着莪术油浓度的增加而增加,且坏死细胞的比例高于凋亡细胞.结论 莪术油对A549细胞的增殖具有抑制作用,并呈时间、浓度依赖,其作用是通过阻滞细胞周期及诱导凋亡和坏死采实现的.%Objective To explore the inhibiting effect of Zedoary turmeric oil on the proliferation of A549 cell line. Methods Lung adenocarcinoma cell line A549 was cultured in vitro. The inhibition rate of Zedoary turmeric oil on the proliferation of lung adenocarcinoma cell line A549 for 24, 48, 72 h were determined by MTT colorimetric assay. The cell cycle of lung adenocarcinoma cell line A549 stimulated by Zedoary turmeric oil for 24 h was analyzed by flow cytometry. The apoptosis and necrosis of lung adenocarcinoma cell line A549 stimulated by Zedoary turmeric oil for 24 h was tested by Annexin V-FITC/PI assay. Results MTT assay indicated that the inhibition rate of Zedoary turmeric oil on the proliferation of lung adenocarcinoma cell line A549 increased significantly with the growing of time and concentration. Further analysis by flow cytometry indicated that Zedoary turmeric oil stimulating the A549 cells for 24 h led to Go/Gi phase arrest and blocked S phase entry. Meanwhile cells in early apoptosis, late apoptosis and necrosis were increased, and the percentage of necrotic cells was more than apoptotic cells with the increase of

  9. In vitro antiproliferative effect of trastuzumab (Herceptin(®)) combined with cetuximab (Erbitux(®)) in a model of human non-small cell lung cancer expressing EGFR and HER2.

    Science.gov (United States)

    Privitera, G; Luca, T; Musso, N; Vancheri, C; Crimi, N; Barresi, V; Condorelli, D; Castorina, S

    2016-05-01

    Lung cancer is the leading cause of cancer death. For this reason, new therapies are needed for the treatment of this devastating disease. In this study, we investigated the effects of combining cetuximab and the trastuzumab on the growth of a model of human non-small cell lung carcinoma cell line (A549). The results were compared with those obtained from a human lung squamous carcinoma cell line (NCI-H226). Both cell lines were treated with cetuximab and trastuzumab, alone or in combination, at various concentrations, for 24, 48 and 72 h. Cell proliferation was measured by the 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide assay. EGFR and HER-2 mRNA expression was detected by reverse transcription polymerase chain reaction, and the gene amplification status of receptors was evaluated by fluorescence in situ hybridisation. The colorimetric proliferation assay showed that trastuzumab combined with cetuximab significantly inhibited A549 cells at a dose of 40 μg/ml after 72 h of treatment (p < 0.05), while no time-dose dependent inhibition was observed in NCI-H226 cells. The combined treatment influenced both levels of EGFR and HER-2 mRNA in A549 cells and only EGFR mRNA levels in NCI-H226 cells. Fluorescence in situ hybridisation showed that both cell lines were aneuploid for the two genes with equally increased EGFR and CEN7 signals, as well as HER-2 and CEN17 signals, indicating a condition of polysomy without amplification. The preliminary results of this study encourage further investigations to elucidate the downstream events involved and to understand how these mechanisms influence non-small cell lung cancers growth.

  10. Winter fine particulate matter from Milan induces morphological and functional alterations in human pulmonary epithelial cells (A549).

    Science.gov (United States)

    Gualtieri, Maurizio; Mantecca, Paride; Corvaja, Viviana; Longhin, Eleonora; Perrone, Maria Grazia; Bolzacchini, Ezio; Camatini, Marina

    2009-07-10

    Samples of PM(2.5) were gravimetrically collected during the winter 2005/2006 in the urban area of Milan (North Italy). Samples were chemically characterized and the particles were detached from filters to determine their cytotoxic effects on the A549 cell line. Based on the potential toxicological relevance of its components, Milan winter PM(2.5) contained high concentrations of pro-oxidant transition metals and PAHs, while re-suspended particles showed a relatively high frequency of dimensional classes ranging from 40 nm to 300 nm. A549 cells exposed to particle suspensions showed a concentration-dependent decrease in viability, starting from 10 microg/cm(2). Phagocytosis of particles by A549 cells and particle aggregates were morphologically characterized and seemed to depend on both particle concentration and exposure time, with the majority of particles being engulfed in membrane-bound vacuoles after 24h of exposure. The ability of ultrafine particles to penetrate and spread throughout the cells was also verified. Cell membrane lysis and mitochondrial ultrastructural disruption appeared to be the main modifications induced by PM(2.5) on A549 cells. Concomitantly to the adverse effects observed in terms of cell mortality and ultrastructural lesions, a significant intracellular production of reactive oxygen species (ROS) was observed, suggesting that the cytotoxicity, exerted by the winter PM(2.5) in Milan, derived also from its oxidative potential, probably associated with particle-adsorbed metals and PAHs.

  11. CCL21/CCR7轴促进人肺癌A549细胞的趋化与侵袭%CCL21/CCR7 axis promotes chemotaxis and invasion of human lung adenocarcinoma A549 cells

    Institute of Scientific and Technical Information of China (English)

    郭学光; 陈正堂

    2007-01-01

    目的:研究CCL21/CCR7轴对肺癌A549细胞定向趋化与侵袭活性的影响.方法:RT-PCR法从临床肺腺癌标本中扩增出CCR7编码区序列,定向克隆至载体pEGFP-N1中,稳定转染A549细胞,Boyden小室法检测转染前后A549细胞对CCL21的趋化和侵袭活性的改变.结果:CCL21作用下多聚碳酸酯膜背面的转染后A549细胞数明显多于转染前的A549细胞数.结论:CCL21/CCR7轴能够促进A549细胞的趋化与侵袭,其可能参与了肺癌淋巴结转移的过程.进一步研究CCL21/CCR7在肺癌中的作用将有助于阐明肺癌淋巴结转移的机制.

  12. Neuropilin 1 expression correlates with the Radio-resistance of human non-small-cell lung cancer cells.

    Science.gov (United States)

    Dong, Juan Cong; Gao, Hui; Zuo, Si Yao; Zhang, Hai Qin; Zhao, Gang; Sun, Shi Long; Han, Hai Ling; Jin, Lin Lin; Shao, Li Hong; Wei, Wei; Jin, Shun Zi

    2015-09-01

    The purpose of this study was to determine the correlation between over-expression of the neuropilin 1 (NRP1) gene and growth, survival, and radio-sensitivity of non-small cell lung carcinoma (NSCLC) cells. 3-[4,5-dimethylthylthiazol-2-yl]-2,5 diphenyltetrazolium broide (MTT) and colony assays were then performed to determine the effect of NRP1 inhibition on the in vitro growth of NSCLC cells. The Annexin V-Fluorescein Isothiocyanate (FITC) apoptosis detection assay was performed to analyse the effect of NRP1 enhancement on apoptosis of NSCLC cells. Transwell invasion and migration assays were employed to examine the metastatic ability of A549 cells post X-ray irradiation. In addition, Western blot assays were carried out to detect the protein level of VEGFR2, PI3K and NF-κB. Finally, to examine the effect of shNRP1 on proliferation and radio-sensitivity in vivo, a subcutaneous tumour formation assay in nude mice was performed. Microvessel density in tumour tissues was assessed by immunohistochemistry. The stable transfected cell line (shNRP1-A549) showed a significant reduction in colony-forming ability and proliferation not only in vitro, but also in vivo. Moreover, shRNA-mediated NRP1 inhibition also significantly enhanced the radio-sensitivity of NSCLC cells both in vitro and in vivo. The over-expression of NRP1 was correlated with growth, survival and radio-resistance of NSCLC cells via the VEGF-PI3K- NF-κB pathway, and NRP1 may be a molecular therapeutic target for gene therapy or radio-sensitization of NSCLC.

  13. Establishment and biological characteristics of a multi-drug resistant cell line A549/Gem

    Directory of Open Access Journals (Sweden)

    Yunfeng ZHU

    2008-02-01

    Full Text Available Background and objective Multi-drug resistance is one of the most important reason why the survival time of non-small cell lung cancer patients is so short. The aim of this study is to establish multi-drug resistant cell line A549/Gem and discuss its biological characters so as to elaborate the possible mechanisms of gemcitabine resistance. Methods Human gemcitabine-resistant non-small cell lung cancer cell line A549/Gem was established by repeated clinical serous peak concentration then low but gradually increasing concentration of gemcitabine from its parental cell human lung adenocarcinoma cell line A549 which is sensitive to gemcitabine. During the course of inducement, monitored its morphology, checked its resistance index and resistant pedigree by MTT method, gathered its growth curve and calculated its doubling time, examined its DNA contents and cell cycles by flow cytometry; at the same time, measured its expression of P53, EGFR, c-erb-B-2, PTEN, PCNA, c-myc, VEGF, MDR-1, Bcl-2, nm23, MMP-9, TIMP-1, CD44v6 Proteins, and RRM1 mRNA. Results The resistance index of A549/Gem?to gemcitabine was 163.228, and the cell line also exhibited cross-resistance to vinorelbine, taxotere, fluorouraci, etoposide and cisplatin, but kept sensitivity to paclitaxol and oxaliplatin. The doubling time of it was shorter and figures in G0-G1 phase were increased than A549. Compared with A549, A549/Gem?achieved EGFR and c-myc protein expression, nm23 protein expression enhanced, p53, Cerb-B-2 and bcl-2 protein expression reduced, PTEN, PCNA and MDR-1 protein expression vanished, but that of MMP-9, VEGF, CD44v6 and TIMP-1 protein changed trivially. Meanwhile, the expression of RRM1 mRNA was augmented markedly. The resistance index of A549/Gem to gemcitabine was 129.783, and the cell line also held cross-resistance to vinorelbine, taxotere, etoposide, cisplatin and sensitivity to paclitaxol. But the resistance to fluorouracil and sensitivity to oxaliplatin

  14. Anti-Inflammatory Effects of Ginsenoside Rg3 via NF-κB Pathway in A549 Cells and Human Asthmatic Lung Tissue

    Directory of Open Access Journals (Sweden)

    In-Seung Lee

    2016-01-01

    Full Text Available Objective. There is limited information of the anti-inflammatory effects of Rg3 on inflamed lung cells and tissues. Therefore, we confirmed the anti-inflammatory mechanism of ginsenoside Rg3 in inflamed human airway epithelial cells (A549 and tissues whether Rg3 regulates nuclear factor kappa B (NF-κB activity. Methods. To induce the inflammation, IL-1β (10 ng/ml was treated to A549 cells for 4 h. The effects of Rg3 on NF-κB activity and COX-2 expression were evaluated by western blotting analysis in both IL-1β-induced inflamed A549 cell and human asthmatic airway epithelial tissues. Using multiplex cytokines assay, the secretion levels of NF-κB-mediated cytokines/chemokines were measured. Result. Rg3 showed the significant inhibition of NF-κB activity thereby reduced COX-2 expression was determined in both IL-1β-induced inflamed A549 cell and human asthmatic airway epithelial tissues. In addition, among NF-κB-mediated cytokines, the secretion levels of IL-4, TNF-α, and eotaxin were significantly decreased by Rg3 in asthma tissues. Even though there was no significant difference, IL-6, IL-9, and IL-13 secretion showed a lower tendency compared to saline-treated human asthmatic airway epithelial tissues. Conclusion. The results from this study demonstrate the potential of Rg3 as an anti-inflammatory agent through regulating NF-κB activity and reducing the secretion of NF-κB-mediated cytokines/chemokines.

  15. Interaction between fragile histamine triad and protein kinase C alpha in human non-small cell lung cancer tissues

    Institute of Scientific and Technical Information of China (English)

    2009-01-01

    Objective To investigate the interaction between fragile histamine triad (FHIT) and protein kinase C alpha (PKCα) in human non-small cell lung cancer tissues. Methods FHIT and PKCα double positive samples were screened by immunohistochemical staining from 13 human non-small cell lung cancer tissues. Co-immunoprecipitation was performed by using anti-FHIT and anti-PKCα. The immune precipitate was analyzed by SDS-PAGE and Western blot. Results Immune precipitate staining detection showed that 3 samples out of...

  16. Human microRNA-24 modulates highly pathogenic avian-origin H5N1 influenza A virus infection in A549 cells by targeting secretory pathway furin.

    Science.gov (United States)

    Loveday, Emma-Kate; Diederich, Sandra; Pasick, John; Jean, François

    2015-01-01

    A common critical cellular event that many human enveloped viruses share is the requirement for proteolytic cleavage of the viral glycoprotein by furin in the host secretory pathway. For example, the furin-dependent proteolytic activation of highly pathogenic (HP) influenza A (infA) H5 and H7 haemagglutinin precursor (HA0) subtypes is critical for yielding fusion-competent infectious virions. In this study, we hypothesized that viral hijacking of the furin pathway by HP infA viruses to permit cleavage of HA0 could represent a novel molecular mechanism controlling the dynamic production of fusion-competent infectious virus particles during the viral life cycle. We explored the biological role of a newly identified furin-directed human microRNA, miR-24, in this process as a potential post-transcriptional regulator of the furin-mediated activation of HA0 and production of fusion-competent virions in the host secretory pathway. We report that miR-24 and furin are differentially expressed in human A549 cells infected with HP avian-origin infA H5N1. Using miR-24 mimics, we demonstrated a robust decrease in both furin mRNA levels and intracellular furin activity in A549 cells. Importantly, pretreatment of A549 cells with miR-24 mimicked these results: a robust decrease of H5N1 infectious virions and a complete block of H5N1 virus spread that was not observed in A549 cells infected with low-pathogenicity swine-origin infA H1N1 virus. Our results suggest that viral-specific downregulation of furin-directed microRNAs such as miR-24 during the life cycle of HP infA viruses may represent a novel regulatory mechanism that governs furin-mediated proteolytic activation of HA0 glycoproteins and production of infectious virions.

  17. Increased Midkine and Estrogen Receptor-β Expression in Human Non-Small Cell Lung Cancer

    Institute of Scientific and Technical Information of China (English)

    Shi-hua Zhang; Guang-feng Zhao; Ya-hong Huang; Kai-hua Lu; Ya-yi Hou

    2009-01-01

    Objective: Midkine (MK), a new member of the heparin-binding growth factor family, has been found recently to have a high expression level in many tumor specimens including lung carcinoma. Estrogens may be involved in lung carcinogenesis, and estrogen receptors, mainly estrogen receptor-β (ER-β), are present and functional in normal lung and tumor cell lines and tissues. In addition, estrogens and growth factors may promote the progression of human non-small cell lung cancer (NSCLC). Previously, we have immunohistochemically demonstrated that MK and ER-β proteins were overexpressed in NSCLC and their expression levels were both significantly negatively correlated with the pathological classification. The purpose of this study was to further verify their expression and its correlation with NSCLC.Methods: Taking NSCLC tissues and their corresponding paraneoplastic and normal lung as research objects, we further examined the expression of MK and ER-β by meas of RT-PCR, in situ hybridization and Western blot analyses at the levels of messenger RNA (mRNA) and protein, respectively.Results: The increased MK and ER-β mRNA expression was found in NSCLC by RT-PCR and in situ hybridization analyses. Furthermore, Western blot analysis also displayed increased expression of MK and ER-β proteins in NSCLC. Finally, their correlation analysis at the levels of mRNA and protein expression in NSCLC demonstrated that MK protein level was significantly correlated to estrogen receptor-β (P0.05, r_s=0.178).Conclusion: All these results in the present study confirmed that MK and ER-β were overexpressed in human NSCLC.

  18. Effects of resveratrol on cell cycle regulatory processes of human lung adenocarcinoma A549 cells and its mechanism%白藜芦醇对人肺腺癌A549细胞周期的影响及其机制研究

    Institute of Scientific and Technical Information of China (English)

    陈加顺; 吕俊明; 束永前

    2011-01-01

    目的 研究白藜芦醇对人肺腺癌A549细胞周期的影响,并探讨其分子机制.方法 MTT法检测白藜芦醇对人肺腺癌A549细胞增殖的影响;PI单标流式细胞术检测白藜芦醇对A549细胞周期分布的影响;Western blotting法检测白藜芦醇对A549细胞cyclin D1和p21cip1蛋白表达的影响.结果 MTT法检测显示,白藜芦醇能明显抑制人肺腺癌A549细胞的增殖,其抑制作用呈时效和量效关系,100μmol/L白藜芦醇在作用48h时抑制率最高,为(76.54±1.33)%.流式细胞术检测提示不同浓度白藜芦醇作用24h细胞周期发生改变,G0/G1期细胞比例明显增加(P<0.01),且呈剂量依赖关系.Western blotting法检测显示,白藜芦醇以时间依赖方式下调周期蛋白cyclin D1的表达,上调p21cip1蛋白表达.结论 白藜芦醇能明显抑制人肺腺癌A549细胞的增殖.白藜芦醇可通过调节细胞周期蛋白cyclin D1、p21cip1的表达调控细胞周期进程,抑制细胞增殖.%Objective To investigate the anti-cancer activities of resveratrol on human lung adenocarcinoma A549 cells and its molecular mechanism involved. Methods The effects of resveratrol on the growth of human lung carcinoma A549 cell lines were studied by MTT assay. The effect of resveratrol on the cell cycle phase distribution of A549 cells was analyzed using flow cytometry by a propidium iodide method. The effect of resveratrol on the expression of cyclin D1 and p21cipl protein was studied by Western blotting analysis. Results MTT assay showed that resveratrol could significantly inhibit the growth of human lung adenocarcinoma A549 cells.It inhibited the proliferation of A549 cells in a time and dose dependent manner, and the highest inhibitory rate was ( 76. 54 ± 1.33 ) %at the concentration of 100μmol/L when cells were cultured for 48h. Meanwhile, different concentrations of resveratrol treated A549 cells for 24h resulted in an increase of G0/G1 phase cells (P < 0. 01 ). The expression of

  19. Oxidative Stress, DNA Damage, and Inflammation Induced by Ambient Air and Wood Smoke Particulate Matter in Human A549 and THP-1 Cell Lines

    DEFF Research Database (Denmark)

    Danielsen, Pernille Høgh; Møller, Peter; Jensen, Keld Alstrup

    2011-01-01

    PM (WSPM) is poorly assessed. We assessed a wide spectrum of toxicity end points in human A549 lung epithelial and THP-1 monocytic cell lines comparingWSPM from high or low oxygen combustion and ambient PM collected in a village with many operating wood stoves and from a rural background area....... In both cell types, all extensively characterized PM samples (1.25-100 μg/mL) induced dose-dependent formation of reactive oxygen species and DNA damage in terms of strand breaks and formamidopyrimidine DNA glycosylase sites assessed by the comet assay with WSPM being most potent. The WSPM contained more...... polycyclic aromatic hydrocarbons (PAH), less soluble metals, and expectedly also had a smaller particle size than PM collected from ambient air. All four types of PM combined increased the levels of 8-oxo-7,8-dihydro-20-deoxyguanosine dose-dependently in A549 cells, whereas there was no change in the levels...

  20. Monitoring of TGF-β 1-Induced Human Lung Adenocarcinoma A549 Cells Epithelial-Mesenchymal Transformation Process by Measuring Cell Adhesion Force with a Microfluidic Device.

    Science.gov (United States)

    Li, Yuan; Gao, AnXiu; Yu, Ling

    2016-01-01

    The epithelial-mesenchymal transition (EMT) is a process in which epithelial cells lose their cell polarity and cell-cell adhesion, and gain migratory and invasive properties. It is believed that EMT is associated with initiation and completion of the invasion-metastasis cascade. In this study, an economic approach was developed to fabricate a microfluidic device with less instrumentation requirement for the investigation of EMT by quantifying cell adhesion force. Fluid shear force was precisely controlled by a homemade microfluidic perfusion apparatus and interface. The adhesion capability of the human lung adenocarcinoma cell line A549 on different types of extracellular matrix protein was studied. In addition, effects of transforming growth factor-β (TGF-β) on EMT in A549 cells were investigated by characterizing the adhesion force changes and on-chip fluorescent staining. The results demonstrate that the microfluidic device is a potential tool to characterize the epithelial-mesenchymal transition process by measuring cell adhesion force.

  1. 肉桂酸对人肺腺癌A549细胞相关基因表达的影响%Effects of cinnamic acid on correlative gene expression of A549 cells in human lung cancer

    Institute of Scientific and Technical Information of China (English)

    范天黎; 许培荣; 杨静; 王霞; 杨观瑞

    2006-01-01

    目的 探讨肉桂酸(CINN)诱导肺癌A549细胞分化的能力及其分子机制.方法 采用原位杂交和完整细胞原位斑点印迹等技术,研究CINN对人肺腺癌A549细胞分化相关蛋白质分子CD15,相关基因c-myc、EGFR、wtp53、wtp16等表达的影响.结果 CINN可上调wtp53、wtp16基因,抑制CD15、c-myc、EGFR基因表达.结论 CINN的上述作用可能是其诱导肺癌A549细胞分化的机制之一.

  2. Erucin, a new promising cancer chemopreventive agent from rocket salads, shows anti-proliferative activity on human lung carcinoma A549 cells.

    Science.gov (United States)

    Melchini, A; Costa, C; Traka, M; Miceli, N; Mithen, R; De Pasquale, R; Trovato, A

    2009-07-01

    Erucin (ER) is a dietary isothiocyanate present in cruciferous vegetables, such as rocket salads (Erucasativa Mill., Diplotaxis sp.), that has been recently considered a promising cancer chemopreventive phytochemical. Biological activity of ER was investigated on human lung adenocarcinoma A549 cells, analyzing its effects on molecular pathways involved in apoptosis and cell cycle arrest, such as PARP-1 cleavage, p53 and p21 protein expression. Our results show that ER affects the A549 cell proliferation, enhancing significantly p53 and p21 protein expression in a dose-dependent manner (pinduction of p53, p21 and PARP-1 cleavage may participate in the anti-proliferative activity of ER in human lung adenocarcinoma A549 cells. Comparison of data with those obtained with the isothiocyanate sulforaphane (SF), structurally related to ER, underlines the strong relationship between structural analogy of ITCs and their biological activity. The ability of dietary compounds to modulate molecular mechanisms that affect cancer cell proliferation is certainly a key point of the cancer prevention potential by functional foods.

  3. Highly expressed N1-acetylpolyamine oxidase detoxifies polyamine analogue N1-cyclopropylmethyl-N11-ethylnorspermine in human lung cancer cell line A549

    Institute of Scientific and Technical Information of China (English)

    HAN Yu; REN Yu-san; CAO Chun-yu; REN Dong-ming; ZHOU Yong-qin; WANG Yan-lin

    2009-01-01

    Background The critical roles of polyamines in cell growth and differentiation have made polyamine metabolic pathway a promising target for antitumor therapy. Recent studies have demonstrated in vitro that some antitumor polyamine analogues could be used as substrates and oxidized by purified recombinant human N1-acetylpolyamine oxidase (APAO, an enzyme that catabolizes natural polyamines), indicating a potential role of APAO in determining the sensitivity of cancer cells to specific antitumor analogues. This study evaluated, in vivo, the effect of APAO on cytotoxicity of antitumor polyamine analogue, N1-cyclopropylmethyI-N11-ethylnorspermine (CPENS) and its mechanism when highly expressed in human lung cancer line A549.Methods A clone with high expression of APAO was obtained by transfecting A549 lung cancer cell line with pcDNA3.1/APAO plasmid and selecting with quantitative realtime PCR and APAO activity assay. Cell proliferation was determined by MTT method and apoptosis related events were evaluated by DNA fragmentation, sub-G1/flow cytometric assay, western blotting (for cytochrome C and Bax) and colorimetric assay (for casapse-3 activity). Results A clone highly expressing APAO was obtained. High expression of APAO in A549 cells inhibited accumulation of CPENS, decreased their sensitivity to the toxicity of CPENS and prevented CPENS induced apoptosis. Conclusion These results indicate a new drug resisting, mechanism in the tumor cells. High expression of APAO can greatly decrease the sensitivity of tumor cells to the specific polyamine analogues by detoxitying those analogues and prevent analogue induced apoptosis.

  4. Progesterone and estrogen receptor expression and activity in human non-small cell lung cancer.

    Science.gov (United States)

    Marquez-Garban, Diana C; Mah, Vei; Alavi, Mohammad; Maresh, Erin L; Chen, Hsiao-Wang; Bagryanova, Lora; Horvath, Steve; Chia, David; Garon, Edward; Goodglick, Lee; Pietras, Richard J

    2011-08-01

    Lung cancer is the most common cause of cancer mortality in male and female patients in the US. Although it is clear that tobacco smoking is a major cause of lung cancer, about half of all women with lung cancer worldwide are never-smokers. Despite a declining smoking population, the incidence of non-small cell lung cancer (NSCLC), the predominant form of lung cancer, has reached epidemic proportions particularly in women. Emerging data suggest that factors other than tobacco, namely endogenous and exogenous female sex hormones, have a role in stimulating NSCLC progression. Aromatase, a key enzyme for estrogen biosynthesis, is expressed in NSCLC. Clinical data show that women with high levels of tumor aromatase (and high intratumoral estrogen) have worse survival than those with low aromatase. The present and previous studies also reveal significant expression and activity of estrogen receptors (ERα, ERβ) in both extranuclear and nuclear sites in most NSCLC. We now report further on the expression of progesterone receptor (PR) transcripts and protein in NSCLC. PR transcripts were significantly lower in cancerous as compared to non-malignant tissue. Using immunohistochemistry, expression of PR was observed in the nucleus and/or extranuclear compartments in the majority of human tumor specimens examined. Combinations of estrogen and progestins administered in vitro cooperate in promoting tumor secretion of vascular endothelial growth factor and, consequently, support tumor-associated angiogenesis. Further, dual treatment with estradiol and progestin increased the numbers of putative tumor stem/progenitor cells. Thus, ER- and/or PR-targeted therapies may offer new approaches to manage NSCLC.

  5. Detection of human papillomavirus in non-small cell carcinoma of the lung.

    Science.gov (United States)

    Chang, Sing Yun; Keeney, Michael; Law, Mark; Donovan, Janis; Aubry, Marie-Christine; Garcia, Joaquin

    2015-11-01

    High-risk human papillomavirus (hrHPV) is an etiologic agent in squamous cell carcinoma (SqCC) arising in the oropharynx and cervix, and a proven prognostic factor in oropharyngeal SqCC. Many studies have found HPV in non-small cell lung carcinoma (NSCLC). Recent studies advocate the detection of messenger RNA transcripts of E6/E7 as more reliable evidence of transcriptively active HPV in tumor cells. The clinical significance of finding HPV remains unclear in NSCLC. This study sought to determine the prevalence of biologically active HPV infection in NSCLC comparing different methodologies. Surgical pathology material from resected primary lung adenocarcinoma (ADC; n=100) and SqCC (n=96) were retrieved to construct tissue microarrays. In situ hybridization (ISH) for hrHPV DNA (DNA-ISH), hrHPV E6/E7 RNA (RNA-ISH), and p16 immunohistochemistry were performed. Cases of oropharyngeal SqCC with known HPV infection were used as positive controls. Expression of p16 was scored as positive if at least 70% of tumor cells showed diffuse and strong nuclear and cytoplasmic staining. Punctate nuclear hybridization signals by DNA-ISH in the malignant cells defined an HPV-positive carcinoma. Of the 196 patients (range, 33-87 years; 108 men), p16 was positive in 19 ADCs and 9 SqCCs, but HPV DNA-ISH and RNA-ISH were negative in all cases. Our study did not detect HPV infection by DNA-ISH or RNA-ISH in any cases of primary NSCLC despite positive p16 expression in a portion of ADC and SqCC. p16 should therefore not be used as a surrogate marker for HPV infection in NSCLC.

  6. Effects of ultrafine petrol exhaust particles on cytotoxicity, oxidative stress generation, DNA damage and inflammation in human A549 lung cells and murine RAW 264.7 macrophages.

    Science.gov (United States)

    Durga, Mohan; Nathiya, Soundararajan; Rajasekar, Abbu; Devasena, Thiyagarajan

    2014-09-01

    Air pollution has persistently been the major cause of respiratory-related illness and death. Environmental pollutants such as diesel and petrol exhaust particles (PEPs) are the major contributors to urban air pollution. The aim of the present study was to characterize and investigate the in vitro cytotoxicity, oxidative stress, DNA damage and inflammation induced by PEPs. Cultured type II epithelium cells (human A549 lung cells) and alveolar macrophages (murine RAW 264.7 cells) were exposed to control, vehicle control and to different concentrations of PEPs for up to 24h. Each treatment was evaluated by cell viability, cytotoxicity, oxidative stress, DNA damage and inflammatory parameters. Overall in vitro studies demonstrated that both cell lines showed similar patterns in response to the above studies induced by petrol exhaust nanoparticles (PENPs). Vehicle control showed no changes compared with the control. In both cell lines, significant changes at the dose of 20 and 50μg/mL (A549 cell lines) and 10and 20μg/mL (macrophages) for PENPs were found. The reactive oxygen species production in both cell lines shot up in minutes, reached the maximum within an hour and came down after 4h. Hence, exposure to PENPs resulted in dose-dependent toxicity in cultured A549 cells and RAW 264.7 cells and was closely correlated to increased oxidative stress, DNA damage and inflammation. Copyright © 2014 Elsevier B.V. All rights reserved.

  7. The role of reactive oxygen species (ROS) production on diallyl disulfide (DADS) induced apoptosis and cell cycle arrest in human A549 lung carcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Wu Xinjiang [Institute of Indoor and Environmental Toxicology, Faculty of Medicine, Justus-Liebig-University of Giessen, Aulweg 123, D-35385 Giessen (Germany); Kassie, Fekadu [Institute of Indoor and Environmental Toxicology, Faculty of Medicine, Justus-Liebig-University of Giessen, Aulweg 123, D-35385 Giessen (Germany); Mersch-Sundermann, Volker [Institute of Indoor and Environmental Toxicology, Faculty of Medicine, Justus-Liebig-University of Giessen, Aulweg 123, D-35385 Giessen (Germany)]. E-mail: Volker.mersch-sundermann@uniklinikum-giessen.de

    2005-11-11

    Diallyl disulfide (DADS), an oil soluble constituent of garlic (Allium sativum), has been reported to cause antimutagentic and anticarcinogenic effects in vitro and in vivo by modulating phases I and II enzyme activities. In recent years, several studies suggested that the chemopreventive effects of DADS can also be attributed to induction of cell cycle arrest and apoptosis in cancer cells. In the present study, we reported that DADS-induced cell cycle arrest at G2/M and apoptosis in human A549 lung cancer cells in a time- and dose-dependent manner. Additionally, a significant increase of intracellular reactive oxygen species (ROS) was induced in A549 cells less than 0.5 h after DADS treatment, indicating that ROS may be an early event in DADS-modulated apoptosis. Treatment of A549 cells with N-acetyl cysteine (NAC) completely abrogated DADS-induced cell cycle arrest and apoptosis. The result indicated that oxidative stress modulates cell proliferation and cell death induced by DADS.

  8. Identification of epigallocatechin-3-gallate in green tea polyphenols as a potent inducer of p53-dependent apoptosis in the human lung cancer cell line A549.

    Science.gov (United States)

    Yamauchi, Rieko; Sasaki, Kaori; Yoshida, Kenichi

    2009-08-01

    The effects of green tea polyphenols on cultured cancer cells have been well characterized, especially the effects of epigallocatechin-3-gallate (EGCg), since EGCg suppresses oncogenic signaling pathways and induces cell cycle arrest or apoptosis by regulating cell cycle-associated proteins. In the present study, we attempted to identify signaling pathways or target molecules regulated by each of or a mixture of green tea polyphenols, including epicatechin (EC), epicatechin-3-gallate (ECg), epigallocatechin (EGC), and EGCg, in the human lung cancer cell line A549. ECg, EGC, and a catechin mixture, in addition to EGCg, significantly decreased cell viability. In contrast, caspase 3/7 activity, an apoptosis indicator, was specifically induced by EGCg. By conducting a series of luciferase-based reporter assays, we revealed that the catechin mixture only up-regulates the p53 reporter. EGCg was a more potent inducer of p53-dependent transcription, and this induction was further supported by the induced level of p53 protein. RNA interference (RNAi)-mediated p53 knockdown completely abolished EGCg-induced apoptosis. Finally, a proteome and western blot analysis using approximately 70 different antibodies failed to detect up-regulated proteins in catechin mixture-treated A549 cells. Taken together, these results indicate that EGCg, among several green tea polyphenols, is a potent apoptosis inducer that functions exclusively through a p53-dependent pathway in A549 cells.

  9. 雷氏大疣蛛蜘蛛毒素对人肺癌细胞A549增殖的影响%Effects of the Spider Venom on proliferation of Human Lung Adenocarcinoma Cell A549

    Institute of Scientific and Technical Information of China (English)

    胡增祥; 杜昱蕾; 刘全喜; 王媛; 李亮

    2010-01-01

    背景与目的 雷氏大疣蛛蜘蛛毒素作为新药有可能用于癌症的治疗,本研究旨在探讨雷氏大疣蛛蜘蛛毒索对人肺癌细胞4549的作用及机理.方法 应用MTT法检测雷氏大疣蛛蜘蛛毒素对人肺癌细胞A549增殖的影响,比色法检测过氧化氢酶活性,改良的硫代巴比妥酸荧光法检测丙二醛含量;流式细胞仪检测细胞凋亡率.采用免疫印迹分析A549细胞中P38MAPK蛋白的表达.结果 雷氏大疣蛛蜘蛛毒素可抑制A549细胞增殖,使CAT活性和MDA的形成增加,且使P38MAPK的表达较对照组明显增多.结论 雷氏大疣蛛蜘蛛毒素抑制A549细胞增殖可能与CAT活性和MDA的形成增加以及P38MAPK的表达降低相关.

  10. Effect of FGF-2 on the Proliferation of Human Lung Adenocarcinoma Cell line A549%FGF-2对人肺腺癌细胞株A549增殖的影响

    Institute of Scientific and Technical Information of China (English)

    李婷; 周建华

    2014-01-01

    目的:观察不同浓度成纤维细胞生长因子-2(fibroblast growth factor-2,FGF-2)对体外培养的人肺腺癌细胞株A549增殖的影响.方法:以体外培养的人肺腺癌细胞株A549为研究对象,采用MTT比色法测定不同浓度(0,12.5,25,50,75,100 ng/mL)的FGF-2对A549细胞增殖活性的影响.结果:与对照组相比,不同浓度的FGF-2处理组细胞的OD值、增殖比均明显增加,差异具有统计学意义(P<0.01).FGF-2浓度为75 ng/mL时细胞增殖比最高,为对照组的183%.结论:FGF-2促进A549细胞增殖,FGF-2对肺腺癌的发生发展具有重要作用.

  11. 托瑞米芬协同顺铂对人肺癌细胞株A549的影响%Synergistic effect of toremifene and cisplatin on human lung cancer cell line A549

    Institute of Scientific and Technical Information of China (English)

    张雪艳; 李强; 韩一平; 刘忠令

    2002-01-01

    目的研究托瑞米芬(TOR)对人肺腺癌细胞系A549的毒性作用及其与顺铂(DDP)联用的协同效应,探讨肺癌综合治疗的方向.方法用MTT显色法检测TOR及与DDP联用后对A549细胞的毒性作用,测定其吸光度(A)值.用流式细胞仪检测细胞DNA含量,Western blot 法检测p21蛋白表达.结果 TOR能直接抑制A549细胞的生长,≥5 μmol/L 的TOR可明显增强DDP的细胞毒性作用.TOR可加强DDP对S期、G2期及M期细胞的作用,且DDP+TOR后p21蛋白表达增加.结论≥5 μmol/L的TOR与DDP联用对A549细胞具有显著的协同抗肿瘤效应.

  12. Inhibition of Oridonin on Human Lung Adenocarcinoma A549 Cells and Its Mechanisms%冬凌草甲素诱导人肺腺癌细胞株A549凋亡及其机制研究

    Institute of Scientific and Technical Information of China (English)

    彭蕾; 顾振纶; 薛仁宇; 周颖; 蒋小岗; 郭次仪

    2010-01-01

    目的:探讨冬凌草甲素(oridnin)对人肺腺癌细胞株A549细胞凋亡的影响.方法:利用MTT法检测oridnin对A549细胞增殖作用的影响;Hoechst 33258染色观察给药后细胞形态改变;透射电镜观察给药后细胞超微结构改变;FITC-AnnexinV/PI双标记检测细胞凋亡率.结果:Hoechst 33258染色和透射电镜观察,oridonin给药后A549细胞出现空泡变性,染色质高度凝集;FTTC-AnnexinV/PI双标记检测oridnin(25,50,100μmol/L)作用细胞48 h后凋亡率分别为1.5%,6.2%,59.7%.结论:Oridonin对A549细胞具有抑制增殖和诱导凋亡作用.

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

    Science.gov (United States)

    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.

  14. Progesterone and estrogen receptor expression and activity in human non-small cell lung cancer

    OpenAIRE

    2011-01-01

    Lung cancer is the most common cause of cancer mortality in male and female patients in the US. Although it is clear that tobacco smoking is a major cause of lung cancer, about half of all women with lung cancer worldwide are never-smokers. Despite a declining smoking population, the incidence of non-small cell lung cancer (NSCLC), the predominant form of lung cancer, has reached epidemic proportions particularly in women. Emerging data suggest that factors other than tobacco, namely endogeno...

  15. 盐霉素抑制人肺腺癌耐顺铂细胞株A549/DDP增殖及诱导凋亡的机制%Salinomycin inhibited proliferation and induced apoptosis of cisplatin-resistant human lung adenocarcinoma cell line A549/DDP

    Institute of Scientific and Technical Information of China (English)

    曾葭; 刘成成; 祝爱珍; 陈小宇; 谭广销; 刘革修

    2012-01-01

    目的:探讨盐霉素对人肺腺癌耐药细胞株A549/DDP增殖的抑制作用及其可能机制.方法:采用MTT法检测盐霉素对A549/DDP细胞生长的抑制作用;流式细胞术检测盐霉素对A549/DDP细胞凋亡及线粒体膜电位(ΔΨm)的影响;比色法检测caspase-3、8和9活性;Western blotting分析细胞色素C、Bcl-2、Bax、β-catenin和磷酸化低密度脂蛋白受体相关蛋白6(p-LRP6)蛋白水平.结果:盐霉素对A549/DDP细胞生长具有剂量依赖性抑制作用.0.2 μmol/L盐霉素作用于A549/DDP细胞,ΔΨm显著下降,而细胞内活性氧和Ca2+浓度在短期显著升高,胞浆细胞色素C蛋白水平、caspase-3、8和9酶活性均显著增加,与对照组比较差异均有统计学意义(P<0.01);Bcl- 2 的表达下调,Bax 的表达明显增加,Bcl-2/Bax 比值显著降低.48 h时增殖抑制率为(34.61±1.97)%,细胞凋亡率为(18.74±2.08)%.盐霉素也减少A549/DDP细胞内β-catenin和p-LRP6蛋白水平.结论:盐霉素通过抑制Wnt信号通路抑制A549/DDP细胞增殖,通过Bcl-2/Bax途径和线粒体凋亡途径诱导人肺腺癌耐药细胞株A549/DDP凋亡.%AIM: To invesligale the effect of salinomycin on the proliferation and apoptosis of cisplalin - resist-ant human lung adenocarcinoma cell line A549/DDP. METHODS: The inhibitory effect of salinomycin on the growlh of A549/DDP cells was tesled by MTT method in vitro. The apoptosis and milochondrial membrane potential ( △ψm ) of A549/DDP cells were assayed by flow cylomelry. The aclivily of caspase -3,8 and 9 was determined by the melhod of col-orimeLry. The levels of cylochrome C, Bcl - 2 , Bax, β - catenin, and phosphorylaled low - densily lipoprolein receptor -relaled protein 6(p - LRP6) were measured by Weslern blotting. RESULTS: Salinomycin inhibited the growth of A549/ DDP cells in a dose - dependent manner. Salinomycin at concentration of 0. 2 μmol/L decreased △ψm level, and increased reactive oxygen species (ROS) , cytochrome C and

  16. DUAL INHIBITION OF PI3K/AKT AND mTOR SIGNALING IN HUMAN NON-SMALL CELL LUNG CANCER CELLS BY A DIETARY FLAVONOID FISETIN

    Science.gov (United States)

    Khan, Naghma; Afaq, Farrukh; Khusro, Fatima H.; Adhami, Vaqar Mustafa; Suh, Yewseok; Mukhtar, Hasan

    2011-01-01

    Lung cancer is one of the most commonly occurring malignancies. It has been reported that mTOR is phosphorylated in lung cancer and its activation was more frequent in tumors with over-expression of PI3K/Akt. Therefore, dual inhibitors of PI3K/Akt and mTOR signaling could be valuable agents for treating lung cancer. In the present study, we show that fisetin, a dietary tetrahydroxyflavone inhibits cell-growth with the concomitant suppression of PI3K/Akt and mTOR signaling in human non-small cell lung cancer (NSCLC) cells. Using autodock 4, we found that fisetin physically interacts with the mTOR complex at two sites. Fisetin treatment was also found to reduce the formation of A549 cell colonies in a dose-dependent manner. Treatment of cells with fisetin caused decrease in the protein expression of PI3K (p85 and p110), inhibition of phosphorylation of Akt, mTOR, p70S6K1, eIF-4E and 4E-BP1. Fisetin-treated cells also exhibited dose-dependent inhibition of the constituents of mTOR signaling complex like Rictor, Raptor, GβL and PRAS40. There was increase in the phosphorylation of AMPKα and decrease in the phosphorylation of TSC2 on treatment of cells with fisetin. We also found that treatment of cells with mTOR inhibitor rapamycin and mTOR-siRNA caused decrease in phosphorylation of mTOR and its target proteins which were further downregulated on treatment with fisetin, suggesting that these effects are mediated in part, through mTOR signaling. Our results show that fisetin suppressed PI3K/Akt and mTOR signaling in NSCLC cells and thus, could be developed as a chemotherapeutic agent against human lung cancer. PMID:21618507

  17. Ameliorative effects of dimetylthiourea and N-acetylcysteine on nanoparticles induced cyto-genotoxicity in human lung cancer cells-A549.

    Directory of Open Access Journals (Sweden)

    Ritesh Kumar Srivastava

    Full Text Available 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.

  18. 端粒酶催化亚单位基因启动子调控Hsv-tk/GCV系统对肺癌细胞A549选择性杀伤作用的研究%A study on selective killing effect of Hsv-tk/GCV driven by human telomerase catalytic subunit promoter on human lung cancer cell A549

    Institute of Scientific and Technical Information of China (English)

    唐小军; 王艳萍; 周清华; 车国卫; 陈小禾; 朱大兴

    2007-01-01

    目的 研究人端粒酶催化亚单位(human telomerase catalytic subunit,hTERT)基因启动子调控单纯疱疹胸苷激酶/更昔洛韦(herpes simplex virus-thymidine kinase/ganciclovir, Hsv-tk/GCV)治疗系统对人肺癌细胞株A549的选择体外杀伤作用.方法 (1)用脂质体法将hTERT启动子和sv40启动子调控的tk基因表达质粒(pGL3-hTp-tk和pGL3-sv40-tk)转染端粒酶阳性的人肺腺癌细胞株A549及端粒酶阴性的人胚肺成纤维细胞株MRC-5,用逆转录-PCR方法检测转染细胞中tk基因的表达情况; (2)用MTT法检测GCV对上述转染细胞体外增殖的抑制作用; (3) 用流式细胞仪检测GCV对上述转染细胞凋亡和细胞周期的影响. 结果 (1)转染pGL3-sv40-tk的细胞A549、MRC-5和转染pGL3-hTp-tk的A549均有tk mRNA表达,转染pGL3-hTp-tk的MRC-5无tk mRNA表达; (2)GCV对转染pGL3-sv40-tk的细胞A549、MRC-5和转染pGL3-hTp-tk的A549体外增殖均有明显抑制作用,对转染pGL3-hTp-tk的MRC-5无明显抑制作用; (3) 转染pGL3-sv40-tk的A549、MRC-5和转染pGL3-hTp-tk的A549细胞,GCV处理后细胞凋亡指数(21.58%、9.35%和23.19%)均显著高于转染pGL3-hTp的A549和MRC-5细胞(0.78%和0.55%)及空白对照A549和MRC-5细胞(2.17%和0.60%),转染pGL3-hTp-tk的MRC-5细胞凋亡指数(0.88%)无明显升高.结论 hTERT启动子调控Hsv-tk基因可以在肺癌细胞中选择性表达,hTERT调控的Hsv-tk/GCV治疗系统对肺癌细胞体外增殖具有靶向性抑制作用.

  19. Inhibitor-sensitive FGFR1 amplification in human non-small cell lung cancer.

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    Amit Dutt

    Full Text Available BACKGROUND: Squamous cell lung carcinomas account for approximately 25% of new lung carcinoma cases and 40,000 deaths per year in the United States. Although there are multiple genomically targeted therapies for lung adenocarcinoma, none has yet been reported in squamous cell lung carcinoma. METHODOLOGY/PRINCIPAL FINDINGS: Using SNP array analysis, we found that a region of chromosome segment 8p11-12 containing three genes-WHSC1L1, LETM2, and FGFR1-is amplified in 3% of lung adenocarcinomas and 21% of squamous cell lung carcinomas. Furthermore, we demonstrated that a non-small cell lung carcinoma cell line harboring focal amplification of FGFR1 is dependent on FGFR1 activity for cell growth, as treatment of this cell line either with FGFR1-specific shRNAs or with FGFR small molecule enzymatic inhibitors leads to cell growth inhibition. CONCLUSIONS/SIGNIFICANCE: These studies show that FGFR1 amplification is common in squamous cell lung cancer, and that FGFR1 may represent a promising therapeutic target in non-small cell lung cancer.

  20. Celecoxib enhances radiation response of secondary bone tumors of a human non-small cell lung cancer via antiangiogenesis in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Klenke, Frank Michael [Bern Univ. (Switzerland). Dept. of Orthopedic Surgery; Abdollahi, Amir [Deutsches Krebsforschungszentrum, Heidelberg (Germany). Dept. of Radiation Oncology; Tufts Univ. School of Medicine, Boston, MA (United States). Center of Cancer Systems Biology; Bischof, Marc; Huber, Peter E. [Deutsches Krebsforschungszentrum, Heidelberg (Germany). Dept. of Radiation Oncology; Gebhard, Martha-Maria [Heidelberg Univ. (Germany). Dept. of Experimental Surgery; Ewerbeck, Volker [Heidelberg Univ. (Germany). Dept. of Orthopedic Surgery; Sckell, Axel [Charite Univ. Medical Center, Berlin (Germany). Dept. of Orthopedic, Trauma and Reconstructive Surgery

    2011-01-15

    Purpose: Cyclooxygenase-2 (COX-2) inhibitors mediate a systemic antitumor activity via antiangiogenesis and seem to enhance the response of primary tumors to radiation. Radiosensitizing effects of COX-2 inhibition have not been reported for bone metastases. Therefore, the aim of this study was the investigation of the radiosensitizing effects of the selective COX-2 inhibitor celecoxib in secondary bone tumors of a non-small cell lung carcinoma in vivo. Materials and Methods: Human A549 lung carcinomas were implanted into a cranial window preparation in male SCID mice (n = 24). Animals were treated with either celecoxib or radiation (7 Gy single photon dose) alone or a combination of celecoxib and radiation, respectively. Untreated animals served as controls. The impact of radiation and COX-2 inhibition on angiogenesis, microcirculation, and tumor growth was analyzed over 28 days by means of intravital microscopy and histological methods. Results: Monotherapies with radiation as well as celecoxib had significant antitumor effects compared to untreated controls. Both therapies reduced tumor growth and vascularization to a similar extent. The simultaneous administration of celecoxib and radiation further enhanced the antitumor and antiangiogenic effects of single-beam radiation. With the combined treatment approach, tumor vascularization and tumor size were decreased by 57% and 51%, respectively, as compared to monotherapy with radiation. Conclusion: The combined application of radiation therapy and COX-2 inhibition showed synergistic effects concerning the inhibition of tumor growth and tumor angiogenesis. Therefore, the combination of radiation with COX-2 inhibitor therapy represents a promising approach to improve the therapeutic efficacy of radiotherapy of bone metastases. (orig.)

  1. Hyperoside induces both autophagy and apoptosis in non-small cell lung cancer cells in vitro.

    Science.gov (United States)

    Fu, Ting; Wang, Ling; Jin, Xiang-nan; Sui, Hai-juan; Liu, Zhou; Jin, Ying

    2016-04-01

    Hyperoside (quercetin-3-O-β-D-galactopyranoside) is a flavonol glycoside found in plants of the genera Hypericum and Crataegus, which exhibits anticancer, anti-oxidant, and anti-inflammatory activities. In this study we investigated whether autophagy was involved in the anticancer mechanisms of hyperoside in human non-small cell lung cancer cells in vitro. Human non-small cell lung cancer cell line A549 was tested, and human bronchial epithelial cell line BEAS-2B was used for comparison. The expression of LC3-II, apoptotic and signaling proteins was measured using Western blotting. Autophagosomes were observed with MDC staining, LC3 immunocytochemistry, and GFP-LC3 fusion protein techniques. Cell viability was assessed using MTT assay. Hyperoside (0.5, 1, 2 mmol/L) dose-dependently increased the expression of LC3-II and autophagosome numbers in A549 cells, but had no such effects in BEAS-2B cells. Moreover, hyperoside dose-dependently inhibited the phosphorylation of Akt, mTOR, p70S6K and 4E-BP1, but increased the phosphorylation of ERK1/2 in A549 cells. Insulin (200 nmol/L) markedly enhanced the phosphorylation of Akt and decreased LC3-II expression in A549 cells, which were reversed by pretreatment with hyperoside, whereas the MEK1/2 inhibitor U0126 (20 μmol/L) did not blocked hyperoside-induced LC3-II expression. Finally, hyperoside dose-dependently suppressed the cell viability and induced apoptosis in A549 cells, which were significantly attenuated by pretreatment with the autophagy inhibitor 3-methyladenine (2.5 mmol/L). Hyperoside induces both autophagy and apoptosis in human non-small cell lung cancer cells in vitro. The autophagy is induced through inhibiting the Akt/mTOR/p70S6K signal pathways, which contributes to anticancer actions of hyperoside.

  2. Inhibition of nuclear factor-κB activity enhanced chemosensitivity to cisplatin in human lung adeno-carcinoma A549 cells under chemical hypoxia conditions

    Institute of Scientific and Technical Information of China (English)

    LI Fang; HUANG Li; SU Xiao-li; GU Qi-hua; HU Cheng-ping

    2013-01-01

    Background Tumor hypoxia,one of the features of solid tumors,is associated with chemo-resistance.Recently,nuclear factor-KB (NF-kB) was found to be activated during hypoxia.However,the impact of NF-kB activation on chemo-resistance during hypoxia remains unknown.Methods Human lung adenocarcinoma A549 cells were transfected with NF-kB p65siRNA and treated with cobalt chloride (CoCl2) to mimic hypoxia in the presence or absence of cisplatin.NF-kB expression was measured by Western blotting,immune-fluorescence and real-time PCR.Hypoxia-inducible factor-1α (HIF-1α) and Bcl-2 expression were determined by Western blotting.Cell apoptosis and survival with half-maximum inhibitory concentration (IC50) of cisplatin were determined by Annexin V-FITC/PI and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT),respectively.Results Exposure ofA549 cells to CoCl2 increased nuclear HIF-1α protein expression,and enhanced NF-kB p65 protein nuclear accumulation (the mark of NF-kB activation) in a time and dose dependant manner.CoCl2 did not promote apoptosis in A549 cells; on the contrary,it reduced cisplatin-induced apoptosis and increased the IC50 of cisplatin.However,when we inhibited CoCl2-induced activation of NF-kB through NF-kB p65siRNA,cisplatin-induced apoptosis was increased and IC50 of cisplatin was reduced to levels similar to those in control cells.Meanwhile,CoCl2-induced Bcl-2 overexpression was down-regulated in the presence of cisplatin when NF-KB activity was inhibited.Conclusion Up-regulating Bcl-2 might be involved in NF-kB activation induced resistance to cisplatin in A549 cells under CoCl2-induced chemical hypoxia.

  3. HIF-1对乏氧人肺腺癌 A549细胞侵袭、迁徙能力的影响及其机制%Impact of hypoxia inducible factor-1 on tumor invasion and metastasis in human lung adenocarcinoma A549 cells under hypoxia

    Institute of Scientific and Technical Information of China (English)

    洪昆; 李芳; 黄瓅; 胡成平

    2015-01-01

    Objective To investigate the role of hypoxia inducible factor-1 (HIF-1 )in tumor metastasis and invasion in human lung adenocarcinoma A549 cells under hypoxia.Methods CoCl2 was used to establish chemical hypoxia model of human lung adenocarcinoma A549 cells.HIF-1-specific inhibitor YC-1 was added to block the expression of HIF-1.Western blot,immune-chemistry and real time-PCR were used to examine the protein and mRNA expressions of HIF-1αand S100A4.Capability of cellular invasion and migration were detected by transwell booth model.Results Compared with normoxic group,HIF-1αand S100A4 were over-expressed,capability of cellular invasion and migration increased in A549 cells of CoCl2 group.However,when HIF-1α expression was specifically inhibited by YC-1,the expression of S100A4 was depressed both at protein and mRNA level,the ability of invasion and migration attenuated in A549 cells of YC-1 group.Conclusions HIF-1 may be able to promote invasion and metastasis of lung adenocarcinoma cells through up-regulating S100A4.%目的:探索低氧诱导因子-1(HIF-1)对乏氧人肺腺癌 A549细胞侵袭及迁徙能力的影响及可能机制。方法采用氯化钴建立人肺腺癌 A549细胞体外化学乏氧模型。给予 HIF-1α特异性抑制剂YC-1抑制 A549细胞 HIF-1α的表达;通过 Western blot、免疫细胞化学、real time-PCR 检测 HIF-1α、S100A4蛋白及 mRNA 的表达;Transwell 小室模型检测 A549细胞 侵 袭 及 迁 徙 能 力。结果乏氧组A549细胞 HIF-1α、S100A4表达较常氧组增加,侵袭及迁徙能力较常氧组增强。YC-1干扰 HIF-1α表达后,YC-1组 A549细胞侵袭及迁徙能力减弱,S100A4蛋白及 mRNA 表达均降低。结论 HIF-1可能通过上调 S100A4的表达促进乏氧人肺腺癌 A549细胞侵袭及迁徙的能力。

  4. Tea extracts inhibit the human lung cancer cells (A549) growth in vitro%茶叶提取物对抗肺癌细胞(A549)体外试验研究

    Institute of Scientific and Technical Information of China (English)

    李伟; 屠幼英

    2006-01-01

    观察表没食子儿茶素没食子酸酯(EGCG)、茶黄素-3、3'-双没食子酸酯(TFDG)、茶黄素(TF)、茶黄素复合物(TFs)、葡萄籽提取物(Grape Seed Extract,GSE)、松树皮提取物(Pine Bark Extract,PBE)、咖啡因(Caf)、槲寄生(VCE)和茶氨酸(The)的体外抗癌活性,通过人肺癌细胞(A549)进行体外试验,结果表明除咖啡因、槲寄生、茶氨酸的作用较小以外其他几种均有很强的促进人肺癌细胞(A549)凋亡的作用,并且EGCG的作用最强,顺序为EGCG、TFs>GSE>PBE、TFDG>TF.用作图法求得EGCG、GSE、PBE、TFDG和TF的IC50值分别为1.01μM、21.91μM、31.01μM、32.87 μM和279.67 μM.

  5. Biological impacts of TiO2 on human lung cell lines A549 and H1299: particle size distribution effects

    Science.gov (United States)

    Tedja, Roslyn; Marquis, Christopher; Lim, May; Amal, Rose

    2011-09-01

    Increasing use of titanium dioxide (TiO2) nanoparticles in many commercial applications has led to emerging concerns regarding the safety and environmental impact of these materials. In this study, we have investigated the biological impact of nano-TiO2 (with particle primary size of 20 nm Aeroxide P25) on human lung cell lines in vitro and also the effect of particle size distribution on the particle uptake and apparent toxicity. The biological impact of nano-TiO2 is shown to be influenced by the concentration and particle size distribution of the TiO2 and the impact was shown to differ between the two cell lines (A549 and H1299) investigated herein. A549 cell line was shown to be relatively resistant to the total amount of TiO2 particles uptaken, as measured by cell viability and metabolic assays, while H1299 had a much higher capacity to ingest TiO2 particles and aggregates, with consequent evidence of impact at concentrations as low as 30-150 μg/mL TiO2. Evidence gathered from this study suggests that both viability and metabolic assays (measuring metabolic and mitochondrial activities and also cellular ATP level) should be carried out collectively to gain a true assessment of the impact of exposure to TiO2 particles.

  6. Apigenin induces caspase-dependent apoptosis in human lung cancer A549 cells through Bax- and Bcl-2-triggered mitochondrial pathway.

    Science.gov (United States)

    Lu, Hsu-Feng; Chie, Yu-Jie; Yang, Ming-Sung; Lee, Ching-Sung; Fu, Jene-John; Yang, Jai-Sing; Tan, Tzu-Wei; Wu, Shin-Hwar; Ma, Yi-Shih; Ip, Siu-Wan; Chung, Jing-Gung

    2010-06-01

    The molecular mechanism and possible signaling pathway of apigenin-induced cytotoxicity and apoptosis in human lung cancer cells has not been reported. We investigated the role of ROS, Ca2+, caspases and Bax proteins and mitochondria membrane potential in apigenin-induced apoptosis in A549 cells. Cells were incubated with different concentrations of apigenin then cell morphological changes, DNA damage, cell viability and apoptosis were determined by Comet assay, and flow cytometric analysis. Sub-G1 phase was also examined. Western blot analysis was used to determined the levels of Bax and Bcl-2 and apoptosis associated proteins, and confocal laser microscope for examining the translocation of associated protein after exposed to apigenin. The results indicated that apigenin induced morphological changes, decreased percentage of viable cells and induced apoptosis dose- and time-dependently. DAPI staining and Comet assay also confirmed that apigenin-induced DNA condensation and damage. The levels of caspase-3, -8 and -9 involved in apigenin-induced apoptosis indicating caspase-dependent pathway was induced by apigenin. Western blotting showed that apigenin promoted cytochrome c levels and also induced dysfunction of mitochondria leading to the release of cytochrome c, AIF and Endo G, causing the activation of caspase-9 and -3, then apoptosis in A549 cells.

  7. Deletion and Mutation of WWOX Exons 6-8 in Human Non-small Cell Lung Cancer

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    To examine the deletion and point mutation of WWOX (WW domain containing oxidoreductase) exons 6-8 in human non-small cell lung cancer and their possible relationship with pathological stages, tumor tissues and the corresponding normal tissues were obtained from 44 Chinese patients who had undergone surgery for non-small cell lung cancer. RNA was extracted from each sample and deletion and mutation of WWOX exons 6-8 were analyzed by RT-PCR and DNA sequencing. Our results showed that 28 of 44 (63.6 %) lung cancer samples showed loss of WWOX exons 6-8 transcript and the deletion was detected in only 3 of 44 (6.8 %) corresponding adjacent normal tissues (P<0.05). The transcript sequencing analyses of the 16 lung cancer samples without transcript loss of WWOX exons 6-8 revealed no difference from the sequence of GenBank. Moreover, the deletion of WWOX exons 6-8 was significantly higher in the smokers when compared with the non-smokers. It is also higher in the men and squamous carcinomas than in women and adenocarcinomas (P<0.05). The deletion, however, was not found to be associated with pathological stages of the tumors. Our study documented a high incidence of deletion of WWOX exons 6-8 in non-small cell lung cancer in Chinese patients and suggested that the frequent loss of WWOX exons 6-8 might play an important role in the tumorigenesis of non-small cell lung cancer in Chinese. WWOX exons 6-8 may serves as a candidate molecular target of smoking carcinogenesis, and point mutation is not a predominant way of alteration of WWOX exons 6-8.

  8. Secretion of beta-human chorionic gonadotropin by non-small cell lung cancer: a case report

    Directory of Open Access Journals (Sweden)

    Varma Seema

    2011-01-01

    Full Text Available Abstract Introduction We describe a case of non-small cell lung cancer that was found to stain positive for beta-human chorionic gonadotropin on immunohistochemistry. Only a few case reports have described lung cancers that secrete beta-human chorionic gonadotropin. Case presentation A 68-year-old Caucasian man presented with symptoms of weakness, fatigue and weight loss for the past two months. On examination, he was found to have generalized lymphadenopathy, and radiologic workup revealed numerous metastases in the lungs, liver and kidneys. Biopsy of the supraclavicular lymph node revealed metastatic large cell lung cancer with beta-human chorionic gonadotropin hormone positivity. The serum beta-human chorionic gonadotropin level was 11,286 mIU/ml (upper limit of normal, 0.5 mIU/ml in non-pregnant females. He was diagnosed with stage 4 lung non-small cell lung cancer. The patient refused chemotherapy. He was discharged home with hospice care. Conclusion The markedly elevated serum values of beta-human chorionic gonadotropin initially prompted the medical team to investigate germinal tumors. In the presence of a negative testicular ultrasound, workup was performed to find an extratesticular source of the tumor. Finally, the diagnosis was made with a tissue biopsy. This case illustrates that atypical markers can be seen in many cancers, emphasizing the role of immunohistochemistry and tissue biopsy in establishing the diagnosis.

  9. Toxic Effects of the Major Components of Diesel Exhaust in Human Alveolar Basal Epithelial Cells (A549

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    Pavel Rossner

    2016-08-01

    Full Text Available We investigated the toxicity of benzo[a]pyrene (B[a]P, 1-nitropyrene (1-NP and 3-nitrobenzanthrone (3-NBA in A549 cells. Cells were treated for 4 h and 24 h with: B[a]P (0.1 and 1 μM, 1-NP (1 and 10 μM and 3-NBA (0.5 and 5 μM. Bulky DNA adducts, lipid peroxidation, DNA and protein oxidation and mRNA expression of CYP1A1, CYP1B1, NQO1, POR, AKR1C2 and COX2 were analyzed. Bulky DNA adducts were induced after both treatment periods; the effect of 1-NP was weak. 3-NBA induced high levels of bulky DNA adducts even after 4-h treatment, suggesting rapid metabolic activation. Oxidative DNA damage was not affected. 1-NP caused protein oxidation and weak induction of lipid peroxidation after 4-h incubation. 3-NBA induced lipid peroxidation after 24-h treatment. Unlike B[a]P, induction of the aryl hydrocarbon receptor, measured as mRNA expression levels of CYP1A1 and CYP1B1, was low after treatment with polycyclic aromatic hydrocarbon (PAH nitro-derivatives. All test compounds induced mRNA expression of NQO1, POR, and AKR1C2 after 24-h treatment. AKR1C2 expression indicates involvement of processes associated with reactive oxygen species generation. This was supported further by COX2 expression induced by 24-h treatment with 1-NP. In summary, 3-NBA was the most potent genotoxicant, whereas 1-NP exhibited the strongest oxidative properties.

  10. Toxicity of wood smoke particles in human A549 lung epithelial cells: the role of PAHs, soot and zinc.

    Science.gov (United States)

    Dilger, Marco; Orasche, Jürgen; Zimmermann, Ralf; Paur, Hanns-Rudolf; Diabaté, Silvia; Weiss, Carsten

    2016-12-01

    Indoor air pollution is associated with increased morbidity and mortality. Specifically, the health impact of emissions from domestic burning of biomass and coal is most relevant and is estimated to contribute to over 4 million premature deaths per year worldwide. Wood is the main fuel source for biomass combustion and the shift towards renewable energy sources will further increase emissions from wood combustion even in developed countries. However, little is known about the constituents of wood smoke and biological mechanisms that are responsible for adverse health effects. We exposed A549 lung epithelial cells to collected wood smoke particles and found an increase in cellular reactive oxygen species as well as a response to bioavailable polycyclic aromatic hydrocarbons. In contrast, cell vitality and regulation of the pro-inflammatory cytokine interleukin-8 were not affected. Using a candidate approach, we could recapitulate WSP toxicity by the combined actions of its constituents soot, metals and PAHs. The soot fraction and metals were found to be the most important factors for ROS formation, whereas the PAH response can be mimicked by the model PAH benzo[a]pyrene. Strikingly, PAHs adsorbed to WSPs were even more potent in activating target gene expression than B[a]P individually applied in suspension. As PAHs initiate multiple adverse outcome pathways and are prominent carcinogens, their role as key pollutants in wood smoke and its health effects warrants further investigation. The presented results suggest that each of the investigated constituents soot, metals and PAHs are major contributors to WSP toxicity. Mitigation strategies to prevent adverse health effects of wood combustion should therefore not only aim at reducing the emitted soot and PAHs but also the metal content, through the use of more efficient combustion appliances, and particle precipitation techniques, respectively.

  11. Responses of genes involved in cell cycle control to diverse DNA damaging chemicals in human lung adenocarcinoma A549 cells

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    Gooderham Nigel J

    2005-08-01

    Full Text Available Abstract Background Many anticancer agents and carcinogens are DNA damaging chemicals and exposure to such chemicals results in the deregulation of cell cycle progression. The molecular mechanisms of DNA damage-induced cell cycle alteration are not well understood. We have studied the effects of etoposide (an anticancer agent, cryptolepine (CLP, a cytotoxic alkaloid, benzo [a]pyrene (BaP, a carcinogenic polycyclic aromatic hydrocarbon and 2-amino-1-methyl-6-phenylimidazo [4,5-b]pyridine (PhIP, a cooked-meat derived carcinogen on the expression of cell cycle regulatory genes to understand the molecular mechanisms of the cell cycle disturbance. Results A549 cells were treated with DMSO or chemicals for up to 72 h and periodically sampled for cell cycle analysis, mRNA and protein expression. DMSO treated cells showed a dominant G1 peak in cell cycle at all times examined. Etoposide and CLP both induced G2/M phase arrest yet the former altered the expression of genes functioning at multiple phases, whilst the latter was more effective in inhibiting the expression of genes in G2-M transition. Both etoposide and CLP induced an accumulation of p53 protein and upregulation of p53 transcriptional target genes. Neither BaP nor PhIP had substantial phase-specific cell cycle effect, however, they induced distinctive changes in gene expression. BaP upregulated the expression of CYP1B1 at 6–24 h and downregulated many cell cycle regulatory genes at 48–72 h. By contrast, PhIP increased the expression of many cell cycle regulatory genes. Changes in the expression of key mRNAs were confirmed at protein level. Conclusion Our experiments show that DNA damaging agents with different mechanisms of action induced distinctive changes in the expression pattern of a panel of cell cycle regulatory genes. We suggest that examining the genomic response to chemical exposure provides an exceptional opportunity to understand the molecular mechanism involved in cellular

  12. NBM-T-BMX-OS01, an Osthole Derivative, Sensitizes Human Lung Cancer A549 Cells to Cisplatin through AMPK-Dependent Inhibition of ERK and Akt Pathway

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    Tian-Jun Chen

    2015-06-01

    Full Text Available Background: Drug combination therapies using cisplatin and natural products are common practice in the treatment of human lung cancer. Osthole is a natural compound extracted from a number of medicinal plants and has been shown to exert strong anticancer activities with low toxicity. Methods: In the present study, NBM-T-BMX-OS01 (BMX, derived from the semi-synthesis of osthole, was evaluated in cisplatin treated A549 cells to investigate its effect on cisplatin resistance in human lung cancer. The anticancer effect of BMX were measured by cell viablity‚ colony formation‚ TUNEL staining‚ flow cytometry and cell cycle assay. The fluorescence staining was performed to detect intracellular and mitochondrial reactive oxygen species (ROS generation. Western blot analysis, antagonists pretreatment and small interfering RNA (siRNA transfection were used to determine the potential mechanism. Results: It was found that, in comparison with single cisplatin treatment, the combination of BMX and cisplatin resulted in greater efficacy in inhibition of proliferation and colony formation, apoptosis induction and cell cycle arrest. The results of fluorescence staining showed that the combination effect of BMX and cisplatin was due to oxidative stress induced by mitochondrial ROS generation. In addition, BMX significantly attenuated the phosphorylation of ERK and Akt, two important pro-survival kinases. In contrast, BMX inhibited the activation of AMPK, and knockdown of AMPK using specific siRNA partially reversed BMX-induced inhibition of ERK and Akt, as well as its synthetic effects on cisplatin induced anticancer activity in A549 cells. Conclusion: Taken together, this study provides that BMX might modulate cisplatin resistance through AMPK-ERK and AMPK-Akt pathways. These results also support the role of BMX as a potential drug candidate for use in combination with cisplatin in the treatment of human lung cancer.

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

    Science.gov (United States)

    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.

  14. Effects of IL-13 on IL-13Rα2 Expression,Proliferation and Migration in Human Lung Adenocarcinoma A549 Cells%IL-13对 A549细胞 IL-13Rα2表达及增殖和迁移的影响

    Institute of Scientific and Technical Information of China (English)

    陈厚文; 余群芳; 何晓燕; 金旗; 蔡震宇; 熊绍恒; 熊丽霞

    2016-01-01

    目的:探讨 IL-13对人肺腺癌细胞(A549)IL-13受体α2(IL-13Rα2)表达及增殖、迁移的影响。方法采用定量逆转录聚合酶链式反应(qRT-PCR)、Western blot 法检测 IL-13刺激 A549细胞后,检测 IL-13Rα2的转录和蛋白质表达水平;酶联免疫吸附实验(ELISA)检测细胞培养上清液中的可溶型 IL-13受体α2(sIL-13Rα2)含量;流式细胞术(FCM)检测胞膜型 IL-13受体α2(memIL-13Rα2)、胞内型 IL-13受体 Rα2(iIL-13Rα2)的细胞数。MTT 法、Transwell 实验和细胞划痕实验观察 IL-13对 A549细 胞 增 殖 及 迁 移 作 用 的 影 响。结果 A549细 胞 可 表 达IL-13Rα2,并且 IL-13在较低质量浓度(10、20 ng·mL-1)时能上调 A549细胞 IL-13Rα2的表达水平,而对增殖和迁移无显著影响;在较高质量浓度(50、100 ng·mL-1)时对 IL-13Rα2的上调作用不明显,但能促进细胞增殖和迁移。另外低质量浓度 IL-13(10、20 ng·mL-1)对 IL-13Rα2整体水平、sIL-13Rα2、memIL-13Rα2以及 iIL-13Rα2的表达均有不同程度的上调(P <0.05);高质量浓度 IL-13(50、100 ng·mL-1)刺激条件下 IL-13Rα2整体水平的上调不明显且不再有剂量依赖性,sIL-13Rα2水平无明显改变、memIL-13Rα2表达略有增加、iIL-13Rα2水平明显下调(P <0.05)。结论IL-13对人肺腺癌 A549细胞 IL-13Rα2表达水平及增殖、迁移的影响具有双相性和差异性;IL-13Rα2在肺腺癌 A549中发挥抑制 IL-13促细胞增殖和迁移的作用。%ABSTRACT:Objective To investigate the effects of interleukin 13(IL-13)on IL-13 receptor α2 (IL-13Rα2),proliferation and migration in human lung adenocarcinoma A549 cells.Methods The expression of IL-13Rα2 mRNA and protein in A549 cells was determined after IL-13 stimulation by qRT-PCR and Western blot,respectively.The content of soluble IL-13Rα2(sIL-13Rα2)in cell culture supernatant was measured by ELISA

  15. 应用A549细胞单层模型研究蛋白多肽类药物肺部吸收的特性%Transport of proteins and peptides across human cultured alveolar A549 cell monolayers

    Institute of Scientific and Technical Information of China (English)

    王智瑛; 张悦; 张强

    2004-01-01

    Aim An in vitro cultured monolayer system of alveolar epithelial cells was used as a model to investigate the transport pathway peptides or proteins, salmon calcitonin (sCT), insulin (INS), recombinant hirudin (rHAV2), and recombinant human growth hormone (rhGH), in pulmonary epithelium in vivo. Methods Human lung adenocareinoma A549 cells formed continuous monolayers with growing polycarbonate filters of Transwell plate. Transport studies of macromolecules in the monolayer system were carried out after 6 days in culture. The transport of peptides or proteins with MW 3 400 - 22 000 was studied in cultured human lung adenocareinoma A549 cell monolayers at different conditions. Results The results showed that the apparent permeability coefficients (Papp) of these macromolecules across A549 cell monolayers ranged from 2×10-6 to 5×10-6 cm·s-1 and exhibited good inverse correlation with molecule weight. No concentration, direction and temperature dependence were observed in the permeation of sCT, INS and rHAV2. While the Papp of rhGH in the BA direction (2.25×10-6 cm·s-1) was significantly less than that in the reverse direction. ThePapp values of rhGH were concentration and temperature independent in the AB direction. Conclusion These findings suggest that the hydrophilic peptides and proteins, salmon calcitonin, insulin, recombinant hirudin, and recombinant human growth hormone used in this study, appeared to penetrate the A549 cell monolayers via a paracellular pathway by passive diffusion mechanism.

  16. CCL21/CCR7 promotes G2/M phase progression via the ERK pathway in human non-small cell lung cancer cells.

    Directory of Open Access Journals (Sweden)

    Ying Xu

    Full Text Available C-C chemokine receptor 7 (CCR7 contributes to the survival of certain cancer cell lines, but its role in the proliferation of human non-small cell lung cancer (NSCLC cells remains vague. Proliferation assays performed on A549 and H460 NSCLC cells using Cell Counting Kit-8 indicated that activation of CCR7 by its specific ligand, exogenous chemokine ligand 21 (CCL21, was associated with a significant linear increase in cell proliferation with duration of exposure to CCL21. The CCL21/CCR7 interaction significantly increased the fraction of cells in the G(2/M phase of the cell cycle as measured by flow cytometry. In contrast, CCL21/CCR7 had no significant influence on the G(0/G(1 and S phases. Western blot and real-time PCR indicated that CCL21/CCR7 significantly upregulated expression of cyclin A, cyclin B1, and cyclin-dependent kinase 1 (CDK1, which are related to the G(2/M phase transition. The expression of cyclin D1 and cyclin E, which are related to the G(0/G(1 and G(1/S transitions, was not altered. The CCL21/CCR7 interaction significantly enhanced phosphorylation of extracellular signal-regulated kinase (P-ERK but not Akt, as measured by Western blot. LY294002, a selective inhibitor of PI3K that prevents activation of the downstream Akt, did not weaken the effect of CCL21/CCR7 on P-ERK. Coimmunoprecipitation further confirmed that there was an interaction between P-ERK and cyclin A, cyclin B1, or CDK1, particularly in the presence of CCL21. CCR7 small interfering RNA or PD98059, a selective inhibitor of MEK that disrupts the activation of downstream ERK, significantly abolished the effects of exogenous CCL21. These results suggest that CCL21/CCR7 contributes to the time-dependent proliferation of human NSCLC cells by upregulating cyclin A, cyclin B1, and CDK1 potentially via the ERK pathway.

  17. Role of Metallothionein1H in Cisplatin Resistance of Non-Small Cell Lung Cancer Cells

    Institute of Scientific and Technical Information of China (English)

    Xin-fang Hou; Qing-xia Fan; Liu-xing Wang; Shi-xin Lu

    2009-01-01

    Objective: Despite platinum-based adjuvant chemotherapy has improved greatly patients' outcomes, drug resistance poses a major impediment to the successful use of such an effective agent. Metallothioneins(MTs) are known to play putative roles in cancer cell proliferation, apoptosis, differentiation, drug resistance and prognosis. The present studiy was to investigte the role of metallethioein1H(MT1H) in cisplatin resistance of human non-small cell lung cancer(NSCLC) cell lines in vitro or its possible molecular mechanisms. Methods: MT1H mRNA expression in A549 and A549/DDP cells was detected by RT-PCR. A recombinant eukaryotic expression plasmid pcDNA3.1(-)-MT1H was constructed and transfected into A549 cells which express no MT1H. MT1H siRNA was transfected into A549/DDP cells which express MT1H highly. MT1H expression was detected by RT-PCR and Immunoblot. The chemosensitivity to cisplatin was assessed by MTT assay. Apoptosis rate was determined by Tunel and FCM. Bcl-2 and Bax were determined by immunohistochemistry. Results: MT1H mRNA was expressed in A549/DDP but not in A549. After transfection of MT1H, MT1H expression was enhanced and the chemosensitivity to cisplatin was decreased in A549 cells. Inversely, after transfection of MT1H siRNA, MT1H expression was decreased and the chemosensitivity to cisplatin was increased in A549/DDP. The apoptosis rate induced by cisplatin was increased and Bcl-2 was down-regulated but Bax showed little change in A549/DDP cells interferred with MT1H siRNA. Conclusion: MT1H overexpression can promote drug resistance in A549 cells . Down-regulation of MT1H interfered with siRNA can effectively reverses the drug resistance in A549/DDP cells by down-regulating the expression of Bcl-2 and increasing cisplatin induced apoptosis. SiRNA targeting MT1H combined with chemotherapy may be a very promising strategy for treatment of lung cancer.

  18. Vaccine research on biological characteristics of human dendritic cells and A-549 lung cancer cell fusion%人树突状细胞与肺癌细胞 A-549融合疫苗生物学特性研究

    Institute of Scientific and Technical Information of China (English)

    王佳烈; 马国强

    2014-01-01

    目的:探讨人树突状细胞(DC)与人肺癌细胞 A-549融合所得疫苗在制备过程中的生物学特性,总结高效制备融合疫苗的方法。方法应用 GM-CSF 和 IL-4优化的方法制备肺癌患者人外周血单核细胞以获得 DC,寻找 DC 制备率最高时间段;同时应用 PKH672GL(绿色荧光)和 PKH262GL(红荧光)分别标记 DC 和肺癌细胞 A-549细胞,筛查最佳的融合比例。结果应用 GM-CSF 和 IL-4优化法进行 DC 制备第7天所得百分率为(66.26±5.13)%,高于其他时间( P <0.05);通过对比不同融合比例 DC 与人肺癌细胞 A-549,显示1∶1时所取得的融合百分率为(35.15±2.16)%,高于其他比例( P <0.05)。结论在 DC 制备过程中制备第7天所得 DC 百分率最高,应选取此时作为提取 DC 的最佳时间;同时 DC 与人肺癌细胞 A-549以1∶1比例相融合所得疫苗百分率最高。%Objective To explore the human dendritic cells (DC) and A-549 in human lung cancer cell fusion vac -cine in the biological characteristics of the process of preparation , summarize the methods of efficient preparation of fusion vac -cine.Methods By using of GM-CSF and IL-4 optimization method for preparing patients with lung cancer in human peripher -al blood mononuclear cells for DC, DC looking for the highest rate of preparation time ; while applying PKH672GL (green flu-orescence) and PKH262GL (red fluorescence) and lung cancer cells were labeled DC and A -549 cells, screening the best blend ratio.Results Application of GM-CSF and IL-4 optimization method for the first 7 days resulting percentage was (66.26 ±5.13)%, higher than at other times ( P <0.05) DC preparation; By comparing different fusion the proportion of DC with human lung cancer cell A -549, showing the percentage of fusion was obtained (35.15 ±2.16)%, higher than the other ratios ( P <0.05).Conclusion The seventh days 'percentage of DC is the best in

  19. Panduratin A, a Possible Inhibitor in Metastasized A549 Cells through Inhibition of NF-Kappa B Translocation and Chemoinvasion

    Directory of Open Access Journals (Sweden)

    Mohd. Rais Mustafa

    2013-07-01

    Full Text Available In the present study, we investigated the effects of panduratin A (PA, isolated from Boesenbergia rotunda, on apoptosis and chemoinvasion in A549 human non-small cell lung cancer cells. Activation of the executioner procaspase-3 by PA was found to be dose-dependent. Caspase-3 activity was significantly elevated at the 5 µg/mL level of PA treatment and progressed to a maximal level. However, no significant elevated level was detected on procaspase-8. These findings suggest that PA activated caspase-3 but not caspase-8. Numerous nuclei of PA treated A549 cells stained brightly by anti-cleaved PARP antibody through High Content Screening. This result further confirmed that PA induced apoptotic cell death was mediated through activation of caspase-3 and eventually led to PARP cleavage. Treatment of A549 cells with PA resulted in a strong inhibition of NF-κB activation, which was consistent with a decrease in nuclear levels of NF-κB/p65 and NF-κB/p50 and the elevation of p53 and p21. Besides that, we also showed that PA significantly inhibited the invasion of A549 cells in a dose-dependent manner through reducing the secretion of MMP-2 of A549 cells gelatin zymography assay. Our findings not only provide the effects of PA, but may also be important in the design of therapeutic protocols that involve targeting of either p53 or NF-κB.

  20. Up-Regulation of Pro-Inflammatory Cytokines and Chemokine Production in Avian Influenza H9N2 Virus-Infected Human Lung Epithelial Cell Line (A549).

    Science.gov (United States)

    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 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.

  1. Differential effect of grape seed extract against human non-small-cell lung cancer cells: the role of reactive oxygen species and apoptosis induction.

    Science.gov (United States)

    Tyagi, Alpna; Raina, Komal; Gangar, Subhash; Kaur, Manjinder; Agarwal, Rajesh; Agarwal, Chapla

    2013-01-01

    The present study examines grape seed extract (GSE) efficacy against a series of non-small-cell lung cancer (NSCLC) cell lines that differ in their Kras and p53 status to establish GSE potential as a cytotoxic agent against a wide range of lung cancer cells. GSE suppressed growth and induced apoptotic death in NSCLC cells irrespective of their k-Ras status, with more sensitivity toward H460 and H322 (wt k-Ras) than A549 and H1299 cells (mutated k-Ras). Mechanistic studies in A549 and H460 cells, selected, based on comparative efficacy of GSE at higher and lower doses, respectively, showed that apoptotic death involves cytochrome c release associated caspases 9 and 3 activation, and poly (ADP-ribosyl) polymerase cleavage, strong phosphorylation of ERK1/2 and JNK1/2, downregulation of cell survival proteins, and upregulated proapoptotic Bak expression. Importantly, GSE treatment caused a strong superoxide radical-associated oxidative stress, significantly decreased intracellular reduced glutathione levels, suggesting, for the first time, the involvement of GSE-caused oxidative stress in its apoptotic inducing activity in these cells. Because GSE is a widely-consumed dietary agent with no known untoward effects, our results support future studies to establish GSE efficacy and usefulness against NSCLC control.

  2. The combination effect of gemcitabine with mitomycin on human lung cancer A549 cell line in vitro%吉西他滨和丝裂霉素联合应用对人肺癌细胞A549的作用

    Institute of Scientific and Technical Information of China (English)

    赵可新; 李军霞; 姜杉; 王伟刚; 胡建平; 单靖珊

    2011-01-01

    目的 观察吉西他滨和丝裂霉素联合应用对人肺癌细胞A549的抑制效应,并探讨其作用机制.方法 用MTT法检测化疗药物对人肺癌细胞A549生长的抑制作用;流式细胞术(FCM)检测细胞周期分布和凋亡率.结果 (1)丝裂霉素和吉西他滨单独和联合用药可浓度依赖性抑制A549细胞的生长.(2)丝裂霉素和吉西他滨在合用72 h后,当Fa >0.18时,丝裂霉素和吉西他滨合用指数CI均1外,其余均1外,其余均<1.(5)丝裂霉素和吉西他滨单用和合用时对细胞周期和凋亡均有影响.结论 丝裂霉素和吉西他滨单独和联合用药可浓度依赖性抑制A549细胞的生长.不同用药次序和药物浓度比例也影响药物联合作用.%Objective To investigate the combination effect of gemcitabine with mitomycin on human lung cancer A549 cell line in vitro,and to explore its action mechanism. Methods MTT assay was used to analyze the inhibition effect of chemotherapy drugs on the growth of human lung cancer A549 cell line, and the cell cycle and cell apoptosis were detected by flow cytometry. Results The gemcitabine and mitomycin alone or combination application could inhibit the growth of human lung cancer A549 cell line in a time and concentration dependent mode. The combination index (CI) was less than 1 at 72h when Fa >0.18,which indicated that gemcitabine and mitomycin had synergistic effect. However,the combination index was more than 1 at 72h when Fa < 0.18,which indicated that gemcitabine and mitomycin had antagonistic effect. The combination application of gemcitabine with mitomycin could get the highest inhibitory rate, next, the moderate inhibitory rate was got when mitomycin was given prior to gemcitabine,and the lowest inhibitory rate was got when gemcitabine was given prior to mitocymin. The CI was all less than 1 when mitomycin was combined with gemcitabine at different concentrations. The gemcitabine and.mitomycin alone or combination application had

  3. Osteopontin knockdown suppresses non-small cell lung cancer cell invasion and metastasis

    Institute of Scientific and Technical Information of China (English)

    SUN Bing-sheng; YOU Jian; LI Yue; ZHANG Zhen-fa; WANG Chang-li

    2013-01-01

    Background Osteopontin (OPN) was identified as one of the leading genes that promote the metastasis of malignant tumor.However,the mechanism by which OPN mediates metastasis in non-small cell lung cancer (NSCLC) remains unknown.The aim of the study is to investigate the biological significance and the related molecular mechanism of OPN expression in lung cancer cell line.Methods Lentiviral-mediated RNA interference was applied to inhibit OPN expression in metastatic human NSCLC cell line (A549).The invasion,proliferation,and metastasis were evaluated OPN-silenced in A549 cells in vitro and in vivo.The related mechanism was further investigated.Results Interestingly,OPN knockdown significantly suppressed the invasiveness of A549 cells,but had only a minor effect on the cellular migration and proliferation.Moreover,we demonstrated that OPN knockdown significantly reduced the levels of matrix metalloproteinase (MMP)-2 and urokinase plasminogen activator (uPA),and led to an obviousinhibition of both in vitro invasion and in vivo lung metastasis of A549 cells (P <0.001).Conclusions Our data demonstrate that OPN contributes to A549 cell metastasis by stimulating cell invasion,independent of cellular migration and proliferation.OPN could be a new treatment target of NSCLC.

  4. Effect of XPA expression on the chemotherapy sensitivity of A549/DDP cells%着色性干皮病A基因表达对A549/DDP化疗敏感性的影响

    Institute of Scientific and Technical Information of China (English)

    张强; 吴金香; 魏玉平; 郝俊青; 黄山英; 董亮

    2012-01-01

    目的:探讨沉默着色性干皮病A(XPA)基因表达在非小细胞肺癌耐药细胞株顺铂化疗敏感性的影响.方法:采用免疫组化法、实时定量PCR(qPCR)及Western blot方法检测非小细胞肺癌患者肿瘤组织中XPA的表达情况.应用qPCR及Western blot方法检测A549/DDP细胞经XPA-shRNA转染后XPA-mRNA及其蛋白表达.通过MTT法检测沉默XPA基因后A549/DDP细胞凋亡情况及其对顺铂的敏感性.结果:肺癌组织XPA表达水平明显高于癌旁组织;沉默XPA基因能够促进A549/DDP细胞凋亡,并能提高A549/DDP对顺铂的药物敏感性.结论:沉默XPA基因表达能够逆转肺癌A549/DDP细胞对顺铂的耐药性.%AIM; To investigate the influence on platinum-based chemotherapy sensitivity by silencing xeroderma pigmentosum group A (XPA) gene expression in non-small cell lung cancer (NSCLC) drug resistance cell lines (A549/ DDP). METHODS; We detected the expression of XPA in lung normal and tumor tissues by immunohistochemistry, quantitative real-time PCR (qPCR) and Western blotting. We silenced XPA expression in A549/DDP cells by XPA-shRNA transfection, and detected the expression of XPA by qPCR and Western blotting. The cell sensitivity to cisplatin and the apoptosis of A549/DDP cells transfected with XPA-shRNA were determined by MTT assay. RESULTS: The expression of XPA was higher in NSCLC tissues than that in normal lung tissues. Silencing XPA gene increased the apoptosis and sensitivity of A549/DDP cells to cisplatin. CONCLUSION: Silencing XPA gene can partly reverse the cisplatin resistance in human cisplatin-resistant NSCLC cell line A549/DDP.

  5. 曲古抑菌素A下调人肺腺癌细胞A549内IDO表达的分子机制%Molecular Mechanism of TSA-induced Down-regulation of IDO in Human Lung Adenocarcinoma Cell Line A549

    Institute of Scientific and Technical Information of China (English)

    李玲玲; 江冠民; 杜军

    2011-01-01

    [Objective] To investigate the molecular mechanism of TSA-induced indoleamine 2, 3-dioxygenase (IDO) down-regulation in human lung adenocarcinoma cell line A549. [Methods] The roles of TSA on the IFN-7 induced IDO expression in A549 cell, the phosphorylation of signal transducer and activator of transcription 1 (STAT1) and the activation of interferon regulatory factor 1 (IRF-l)were examined by western blotting. Effect of TSA on STAT1 nuclear translocation was observed by a confocal laser-scanning microscope. The luciferase activity of the activation of 7-interferon activated sites (GAS) , interferon stimulated response element (ISRE) and nuclear factor-kB (NF-kB) was measured by dual luciferase reporter assay system. [Results] TSA concentration-dependently reduced IFN-7 induced IDO expression, inhibited STAT1 phosphorylation at Tyr-701 and nuclear translocation in A549 cell. Dual luciferase reporter assay and Western blotting results showed that TSA blocked IFN-"y -induced activation of GAS, ISRE, and IRF-1, but not NF-kB. [Conclusions] TSA can down-regulate IFN-7 induced IDO expression in A549 cell, which may be associated with the repression of phosphorylation and nuclear translocation of STAT1 and its binding to GAS.%[目的]研究曲古抑菌素A (TSA)抑制γ干扰素(IFN-γ)诱导的人A549细胞内吲哚胺2,3-双加氧酶(IDO)表达的分子机制.[方法]采用蛋白质免疫印迹技术检测TSA在IFN-γ诱导的A549细胞中IDO的表达、信号转导及转录激活子1 (STAT1)的磷酸化和干扰素调节因子1(IRF-1)的激活等过程中的作用,在激光共聚焦显微镜下观察TSA对STAT1核转位的影响,利用双荧光素酶报告基因系统检测TSA对γ-干扰素激活位点(GAS)、干扰素刺激应答元件(ISRE)和核因子-κB(NF-κB)的激活的影响.[结果]TSA以浓度依赖方式下调A549细胞中IFN-γ诱导的IDO表达,并能明显抑制STAT1第701位酪氨酸的磷酸化和STAT1的核转位.双荧光

  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.

    Science.gov (United States)

    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. Novel functional view of the crocidolite asbestos-treated A549 human lung epithelial transcriptome reveals an intricate network of pathways with opposing functions

    Directory of Open Access Journals (Sweden)

    Stevens John R

    2008-08-01

    Full Text Available Abstract Background Although exposure to asbestos is now regulated, patients continue to be diagnosed with mesothelioma, asbestosis, fibrosis and lung carcinoma because of the long latent period between exposure and clinical disease. Asbestosis is observed in approximately 200,000 patients annually and asbestos-related deaths are estimated at 4,000 annually1. Although advances have been made using single gene/gene product or pathway studies, the complexity of the response to asbestos and the many unanswered questions suggested the need for a systems biology approach. The objective of this study was to generate a comprehensive view of the transcriptional changes induced by crocidolite asbestos in A549 human lung epithelial cells. Results A statistically robust, comprehensive data set documenting the crocidolite-induced changes in the A549 transcriptome was collected. A systems biology approach involving global observations from gene ontological analyses coupled with functional network analyses was used to explore the effects of crocidolite in the context of known molecular interactions. The analyses uniquely document a transcriptome with function-based networks in cell death, cancer, cell cycle, cellular growth, proliferation, and gene expression. These functional modules show signs of a complex interplay between signaling pathways consisting of both novel and previously described asbestos-related genes/gene products. These networks allowed for the identification of novel, putative crocidolite-related genes, leading to several new hypotheses regarding genes that are important for the asbestos response. The global analysis revealed a transcriptome that bears signatures of both apoptosis/cell death and cell survival/proliferation. Conclusion Our analyses demonstrate the power of combining a statistically robust, comprehensive dataset and a functional network genomics approach to 1 identify and explore relationships between genes of known importance

  8. Changes in gene expression induced by polycyclic aromatic hydrocarbons in the human cell lines HepG2 and A549.

    Science.gov (United States)

    Castorena-Torres, Fabiola; Bermúdez de León, Mario; Cisneros, Bulmaro; Zapata-Pérez, Omar; Salinas, Juan E; Albores, Arnulfo

    2008-03-01

    Polycyclic aromatic hydrocarbons (PAH) are the main components of emissions generated by coke oven factories and many of these chemicals are carcinogenic. The goal of this study was to examine changes in gene expression in two human cell lines, HepG2 and A549, induced by exposure to a soil extract containing PAH using microarry technology. Soil samples were obtained from the vicinity of a coke oven factory in northeastern Mexico. For comparison, the gene expression pattern induced by Benz[a]pyrene (BaP) was also analyzed. The number of altered genes by both treatments was 2-fold higher in hepatic than in pulmonary cells. Differentially-modulated genes in the two cell lines were identified and grouped by biological function using genomic databases. A group of nine genes up- and down-regulated by either the PAH extract or BaP were selected for validation by real-time PCR. The cellular functions of these PAH-responsive genes included: xenobiotic metabolism (CYP1A1 and CYP1B1), DNA repair (ERCC5), oxidative stress response and cell proliferation (FTH1 and PRDX1), protein degradation (PSMD7), ion transport (FXYD3), steroid biosynthesis (FDFT1), and signaling pathways (PTGER3). The real-time PCR analysis confirmed most of the microarray data with significant correlation. Additional studies are required to determine the mechanisms involved in the PAH-mediated modulation of these genes and to associate these changes with human health.

  9. MicroRNA-429 induces tumorigenesis of human non-small cell lung cancer cells and targets multiple tumor suppressor genes

    Energy Technology Data Exchange (ETDEWEB)

    Lang, Yaoguo; Xu, Shidong; Ma, Jianqun; Wu, Jun [Department of Thoracic Surgery, Harbin Medical University Cancer Hospital, 150 Haping Road, Harbin, Heilongjiang 150081 (China); Jin, Shi; Cao, Shoubo [Department of Medical Oncology, Harbin Medical University Cancer Hospital, 150 Haping Road, Harbin, Heilongjiang 150081 (China); Yu, Yan, E-mail: yuyan@hrbmu.edu.cn [Department of Medical Oncology, Harbin Medical University Cancer Hospital, 150 Haping Road, Harbin, Heilongjiang 150081 (China)

    2014-07-18

    Highlights: • MiR-429 expression is upregulated in non-small cell lung cancer (NSCLC). • MiR-429 inhibits PTEN, RASSF8 and TIMP2 expression. • MiR-429 promotes metastasis and proliferation. • We report important regulatory mechanisms involved in NSCLC progression. • MiR-429 is a potential therapeutic target and diagnostic marker. - Abstract: Lung cancer is the major cause of cancer death globally. MicroRNAs are evolutionally conserved small noncoding RNAs that are critical for the regulation of gene expression. Aberrant expression of microRNA (miRNA) has been implicated in cancer initiation and progression. In this study, we demonstrated that the expression of miR-429 are often upregulated in non-small cell lung cancer (NSCLC) compared with normal lung tissues, and its expression level is also increased in NSCLC cell lines compared with normal lung cells. Overexpression of miR-429 in A549 NSCLC cells significantly promoted cell proliferation, migration and invasion, whereas inhibition of miR-429 inhibits these effects. Furthermore, we demonstrated that miR-429 down-regulates PTEN, RASSF8 and TIMP2 expression by directly targeting the 3′-untranslated region of these target genes. Taken together, our results suggest that miR-429 plays an important role in promoting the proliferation and metastasis of NSCLC cells and is a potential target for NSCLC therapy.

  10. The histone demethylase PHF8 is an oncogenic protein in human non-small cell lung cancer

    Energy Technology Data Exchange (ETDEWEB)

    Shen, Yuzhou; Pan, Xufeng; Zhao, Heng, E-mail: hengzhao1966@sina.com

    2014-08-15

    Highlights: • PHF8 overexpresses in human NSCLC and predicts poor survival. • PHF8 regulates lung cancer cell growth and transformation. • PHF8 regulates apoptosis in human lung cancer cells. • PHF8 promotes miR-21 expression in human lung cancer. • MiR-21 is critically essential for PHF8 function in human lung cancer cells. - Abstract: PHF8 is a JmjC domain-containing protein and erases repressive histone marks including H4K20me1 and H3K9me1/2. It binds to H3K4me3, an active histone mark usually located at transcription start sites (TSSs), through its plant homeo-domain, and is thus recruited and enriched in gene promoters. PHF8 is involved in the development of several types of cancer, including leukemia, prostate cancer, and esophageal squamous cell carcinoma. Herein we report that PHF8 is an oncogenic protein in human non-small cell lung cancer (NSCLC). PHF8 is up-regulated in human NSCLC tissues, and high PHF8 expression predicts poor survival. Our in vitro and in vivo evidence demonstrate that PHF8 regulates lung cancer cell proliferation and cellular transformation. We found that PHF8 knockdown induces DNA damage and apoptosis in lung cancer cells. PHF8 promotes miR-21 expression in human lung cancer, and miR-21 knockdown blocks the effects of PHF8 on proliferation and apoptosis of lung cancer cells. In summary, PHF8 promotes lung cancer cell growth and survival by regulating miR-21.

  11. Alteration of membrane lipid biophysical properties and resistance of human lung adenocarcinoma A549 cells to cisplatin

    Institute of Scientific and Technical Information of China (English)

    LIANG; Xingjie; (

    2001-01-01

    [1]Simon, S. M., Schindler, M., Cell biological mechanisms of multidrug resistance in tumors, Proc. Natl. Acad. Sci. USA, 1994, 91: 3497.[2]Ambudkar, S. V., Dey, S., Hrycyna, C. A. et al., Biochemical, cellular, and pharmacological aspects of the multidrug trans-porter, Annu. Rev. Pharmacol. Toxicol., 1999, 39: 361.[3]Dudeja, P. K., Anderson, K. M., Harris, J. S. et al., Reversal of multidrug resistance phenotype by surfactants: Relationship to membrane lipid fluidity, Arch. Biochem. Biophys., 1995, 319 (1): 8309.[4]Collins, J. M., Scott, R. B., Grogan, W. M., Plasma membrane fluidity gradients of human peripheral blood leukocytes, J. Cell Physiol., 1990, 144: 42.[5]Collins, J. M., Dominey, R. N., Grogan, W. M., Shape of the fluidity gradient in the plasma membrane of living Hela cells, J. Lipid Res., 1990, 31: 261.[6]Ashman, R. F., Peckham, D., Alhasan, S. et al., Membrane unpacking and the rapid disposal of apoptotic cells, Immunol. Lett., 1995, 48(3): 159.[7]Sentjurc, M., Zorec, M., Cemazar, M. et al., Effect of vinblastine on cell membrane fluidity in vinblastine-sensitive and -resistant HeLa cells, Cancer Lett., 1998, 130(1-2):183.[8]Regev, R., Assaraf, Y. G., Eytan, G. D. et al., Membrane fluidization by ether, other anesthetics, and certain agents abolish-es P-glycoprotein ATPase activity and modulates efflux from multidrug-resistant cells, Eur. J. Biochem., 1999, 259(1-2): 18.[9]Robert, A. S., Mariamme, S., Katherine, L. S., Altered lipid packing identifies apoptotic thymocytes, Immunol. Lett., 1993, 36: 283.[10] Lagerberg, J. W., Kallen, K. J., Haest, C. W. et al., Factors affecting the amount and the mode of merocyanine 540 binding to the membrane of human erythrocytes, Biochim. Biophys. Acta, 1995, 1235(2): 428.[11] Stillwell, W., Wassall, S. R., Dumaual, A. C. et al., Use of merocyanine (MC540) in quantifying lipid domains and pack-ing in phospholipid vesicles and tumor cells, Biochem. Biophys. Acta, 1993

  12. Evaluation of cytotoxic, oxidative stress, proinflammatory and genotoxic responses of micro- and nano-particles of dolomite on human lung epithelial cells A(549).

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    Patil, Govil; Khan, Mohd Imran; Patel, Devendra Kumar; Sultana, Sarwat; Prasad, Rajendra; Ahmad, Iqbal

    2012-09-01

    Dolomite is a natural mineral of great industrial importance and used worldwide, thus millions of workers are at risk of occupational exposure. Its toxicity is however, meagerly documented. In the present investigation, a dolomite powder obtained from its milling unit was analyzed by some standard methods namely, optical microscopy, transmission electron microscopy and dynamic light scattering. Results showed that dolomite powder contained particles of different shapes and size both microparticles (MPs) and nanoparticles (NPs), suggesting potential occupational exposure of these particles. An attempt was therefore, made to investigate dolomite toxicity in a particle size-dependent manner in human lung epithelial cells A(549). The comparative toxicity evaluation of MPs and NPs was carried out by assessing their effects on cell viability, membrane damage, glutathione, reactive oxygen species (ROS), lipid peroxidation (LPO), micronucleus (MN) and proinflammatory cytokines, namely tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β) and interleukin-6 (IL-6). These markers of cytotoxicity, genotoxicity and inflammation were assayed in cells exposed to MPs and NPs in a dose-and time-dependent manner. Invariably, their toxic effects were dose-and time-dependent while NPs in general were significantly more toxic. Notably, NPs caused oxidative stress, genotoxicity and inflammatory responses, as seen by significant induction of ROS, LPO, MN, TNF-α, IL-1β and IL-6. Thus, the study tends to suggest that separate health safety standards would be required for micrometer and nanometer scale particles of dolomite.

  13. Identification of a Short Cell-Penetrating Peptide from Bovine Lactoferricin for Intracellular Delivery of DNA in Human A549 Cells.

    Science.gov (United States)

    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.

  14. Mimulone-induced autophagy through p53-mediated AMPK/mTOR pathway increases caspase-mediated apoptotic cell death in A549 human lung cancer cells.

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    Hyun-Kyu An

    Full Text Available Anticancer properties and mechanisms of mimulone (MML, C-geranylflavonoid isolated from the Paulownia tomentosa fruits, were firstly elucidated in this study. MML prevented cell proliferation in a dose- and time-dependent way and triggered apoptosis through the extrinsic pathway in A549 human lung adenocarcinoma cells. Furthermore, MML-treated cells displayed autophagic features, such as the formation of autophagic vacuoles, a primary morphological feature of autophagy, and the accumulation of microtubule-associated protein 1 light chain 3 (LC3 puncta, another typical maker of autophagy, as determined by FITC-conjugated immunostaining and monodansylcadaverine (MDC staining, respectively. The expression levels of LC3-I and LC3-II, specific markers of autophagy, were also augmented by MML treatment. Autophagy inhibition by 3-methyladenine (3-MA, pharmacological autophagy inhibitor, and shRNA knockdown of Beclin-1 reduced apoptotic cell death induced by MML. Autophagic flux was not significantly affected by MML treatment and lysosomal inhibitor, chloroquine (CQ suppressed MML-induced autophagy and apoptosis. MML-induced autophagy was promoted by decreases in p53 and p-mTOR levels and increase of p-AMPK. Moreover, inhibition of p53 transactivation by pifithrin-α (PFT-α and knockdown of p53 enhanced induction of autophagy and finally promoted apoptotic cell death. Overall, the results demonstrate that autophagy contributes to the cytotoxicity of MML in cancer cells harboring wild-type p53. This study strongly suggests that MML is a potential candidate for an anticancer agent targeting both autophagy and apoptotic cell death in human lung cancer. Moreover, co-treatment of MML and p53 inhibitor would be more effective in human lung cancer therapy.

  15. Mimulone-Induced Autophagy through p53-Mediated AMPK/mTOR Pathway Increases Caspase-Mediated Apoptotic Cell Death in A549 Human Lung Cancer Cells

    Science.gov (United States)

    Lee, Ji-Won; Park, Mi-Hyun; Moon, Hyung-In; Park, Shin-Ji; Baik, Ji-Sue; Kim, Cheorl-Ho; Lee, Young-Choon

    2014-01-01

    Anticancer properties and mechanisms of mimulone (MML), C-geranylflavonoid isolated from the Paulownia tomentosa fruits, were firstly elucidated in this study. MML prevented cell proliferation in a dose- and time-dependent way and triggered apoptosis through the extrinsic pathway in A549 human lung adenocarcinoma cells. Furthermore, MML-treated cells displayed autophagic features, such as the formation of autophagic vacuoles, a primary morphological feature of autophagy, and the accumulation of microtubule-associated protein 1 light chain 3 (LC3) puncta, another typical maker of autophagy, as determined by FITC-conjugated immunostaining and monodansylcadaverine (MDC) staining, respectively. The expression levels of LC3-I and LC3-II, specific markers of autophagy, were also augmented by MML treatment. Autophagy inhibition by 3-methyladenine (3-MA), pharmacological autophagy inhibitor, and shRNA knockdown of Beclin-1 reduced apoptotic cell death induced by MML. Autophagic flux was not significantly affected by MML treatment and lysosomal inhibitor, chloroquine (CQ) suppressed MML-induced autophagy and apoptosis. MML-induced autophagy was promoted by decreases in p53 and p-mTOR levels and increase of p-AMPK. Moreover, inhibition of p53 transactivation by pifithrin-α (PFT-α) and knockdown of p53 enhanced induction of autophagy and finally promoted apoptotic cell death. Overall, the results demonstrate that autophagy contributes to the cytotoxicity of MML in cancer cells harboring wild-type p53. This study strongly suggests that MML is a potential candidate for an anticancer agent targeting both autophagy and apoptotic cell death in human lung cancer. Moreover, co-treatment of MML and p53 inhibitor would be more effective in human lung cancer therapy. PMID:25490748

  16. Kaempferol modulates the metastasis of human non-small cell lung cancer cells by inhibiting epithelial-mesenchymal transition

    Directory of Open Access Journals (Sweden)

    Meng Hang

    2015-06-01

    Full Text Available The present study was done to determine whether kaempferol, a natural polyphenol of the flavonoid family, affects Epithelial-Mesenchymal Transition (EMT in non-small cell lung cancer cells. Kaempferol not only inhibited cancer cell proliferation and migration in a dose-dependent manner but also modulated the expression of EMT-related proteins E-cadherin and vimentin which are indispensible to cellular motility, invasiveness and metastasis. These results indicate that kaempferol suppresses non-small cell lung cancer migration by modulating the expression of EMT proteins. Therefore, kaempferol may be useful as a potential anticancer agent for non-small cell lung cancer.

  17. Astaxanthin down-regulates Rad51 expression via inactivation of AKT kinase to enhance mitomycin C-induced cytotoxicity in human non-small cell lung cancer cells.

    Science.gov (United States)

    Ko, Jen-Chung; Chen, Jyh-Cheng; Wang, Tai-Jing; Zheng, Hao-Yu; Chen, Wen-Ching; Chang, Po-Yuan; Lin, Yun-Wei

    2016-04-01

    Astaxanthin has been demonstrated to exhibit a wide range of beneficial effects, including anti-inflammatory and anti-cancer properties. However, the molecular mechanism of astaxanthin-induced cytotoxicity in non-small cell lung cancer (NSCLC) cells has not been identified. Rad51 plays a central role in homologous recombination, and studies show that chemo-resistant carcinomas exhibit high levels of Rad51 expression. In this study, astaxanthin treatment inhibited cell viability and proliferation of two NSCLC cells, A549 and H1703. Astaxanthin treatment (2.5-20 μM) decreased Rad51 expression and phospho-AKT(Ser473) protein level in a time and dose-dependent manner. Furthermore, expression of constitutively active AKT (AKT-CA) vector rescued the decreased Rad51 mRNA and protein levels in astaxanthin-treated NSCLC cells. Combined treatment with phosphatidylinositol 3-kinase (PI3K) inhibitors (LY294002 or wortmannin) further decreased the Rad51 expression in astaxanthin-exposed A549 and H1703 cells. Knockdown of Rad51 expression by transfection with si-Rad51 RNA or cotreatment with LY294002 further enhanced the cytotoxicity and cell growth inhibition of astaxanthin. Additionally, mitomycin C (MMC) as an anti-tumor antibiotic is widely used in clinical NSCLC chemotherapy. Combination of MMC and astaxanthin synergistically resulted in cytotoxicity and cell growth inhibition in NSCLC cells, accompanied with reduced phospho-AKT(Ser473) level and Rad51 expression. Overexpression of AKT-CA or Flag-tagged Rad51 reversed the astaxanthin and MMC-induced synergistic cytotoxicity. In contrast, pretreatment with LY294002 further decreased the cell viability in astaxanthin and MMC co-treated cells. In conclusion, astaxanthin enhances MMC-induced cytotoxicity by decreasing Rad51 expression and AKT activation. These findings may provide rationale to combine astaxanthin with MMC for the treatment of NSCLC.

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

    Science.gov (United States)

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

    2016-07-13

    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.

  19. Curcumin Promoted the Apoptosis of Cisplain-resistant Human Lung Carcinoma Cells A549/DDP through Down-regulating miR-186*

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    Jian ZHANG

    2010-04-01

    Full Text Available Background and objective Curcumin, a natural compound, is derived from the rthizom of Curcuma longa. In vitro and in vivo preclinical studies have shown its anti-inflammatory, antioxidant, anticancer activities and so on. miR-186*, which was found by microarray technology, was highly expressed in lung carcinoma cells A549/DDP. The aim of this study is to illustrate whether Curcumin could promote the apoptosis of A549/DDP cells through regulating the expression of miR-186*. Methods An oligonucleotide microarray chip was used to profile microRNA (miRNA expressions in A549/DDP cells treated with and without Curcumin. The significantly differentially expressed miRNA, which was selected from microarray chip, validated by quantitative real-time PCR. Ultimately, the remarkably expressed miRNA modulated the apoptosis assaying by flow cytometry expriments and the survival rate was measured by MTT method. Results The microarray chip results demonstrated: Curcumin altered the expression level of miRNAs compared with untreated control in A549/DDP cell line, miR-186* was significantly down-regulated after Curcumin treatment, which confirmed by quantitative real-time PCR. Downregulation of miR-186* expression by curcumin elevated the apoptosis, and the survival rate of A549/DDP cells decreased; but up-regulation of miR-186* expression by transfection its mimics restrained the apoptosis, the survival rate of A549/DDP cells increased, which were assayed by flow cytometry expriments and MTT method. Conclusion Modulation of miRNAs expression may be an important mechanism underlying the biological roles of Curcumin.

  20. Trichostatin A Induced Bcl-2 Protein Level Decrease Mediated A549/CDDP Cells Apoptosis by Mitochondria Pathway

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    Jun WU

    2009-11-01

    Full Text Available Background and objective The use of platinum-based combination chemotherpy remains the standard treatment for non-small cell lung cancer. However, the resistance to platinum limits further treatment clinically. Trichostatin A (TSA is one of histone deacetylase (HDAC inhibitors. It inhibits tumor cell proliferation and acts as a chemosensitizer. The aim of this study is to investigate the action mechanism of TSA on cisplatin-resistant human lung adenocarcinoma cell line A549/CDDP. Methods Cytotoxicity and cell viability was assayed by Neutral Red method. Morphologic assessment of apoptosis was determined by fluorescence microscope; cell cycle and mitochondrial membrane potential were detected by flow cytometry. In addition, A549/CDDP cells were transfected with Bcl-2 expression Vector and siRNA-bcl-2. Results A549/CDDP cells treated with TSA showed apparently cytotoxicity, IC50 of TSA was (446.59±27.32 nmol/L. The growth curve showed the ratio of growth decreased with the increase of concentration of TSA. The apoptosis appeared 24 hours after treated by (125-500 nmol/L TSA, morphologic changes including nuclear chromatin condensation. Fluorescence strength was observed with fluorescence microscope. Treated by TSA, mitochondrial membrane potential was decreased and cells were arrested at S phase. Western blotting analyses showed that the levels of Bcl-2 decreased, while expression of Bax increased. Simultaneously caspase-3 was activated. Over expression of Bcl-2 can inhibit TSA-induced A549/CDDP cell apoptosis, while the decrease of Bcl-2 enhanced the sensitivity of A549/CDDP cell to TSA. Conclusion TSA induce A549/CDDP cell apoptosis by mitochondria pathway.

  1. The significance of PIWI family expression in human lung embryogenesis and non-small cell lung cancer.

    Science.gov (United States)

    Navarro, Alfons; Tejero, Rut; Viñolas, Nuria; Cordeiro, Anna; Marrades, Ramon M; Fuster, Dolors; Caritg, Oriol; Moises, Jorge; Muñoz, Carmen; Molins, Laureano; Ramirez, Josep; Monzo, Mariano

    2015-10-13

    The expression of Piwi-interacting RNAs, small RNAs that bind to PIWI proteins, was until recently believed to be limited to germinal stem cells. We have studied the expression of PIWI genes during human lung embryogenesis and in paired tumor and normal tissue prospectively collected from 71 resected non-small-cell lung cancer patients. The mRNA expression analysis showed that PIWIL1 was highly expressed in 7-week embryos and downregulated during the subsequent weeks of development. PIWIL1 was expressed in 11 of the tumor samples but in none of the normal tissue samples. These results were validated by immunohistochemistry, showing faint cytoplasmic reactivity in the PIWIL1-positive samples. Interestingly, the patients expressing PIWIL1 had a shorter time to relapse (TTR) (p = 0.006) and overall survival (OS) (p = 0.0076) than those without PIWIL1 expression. PIWIL2 and 4 were downregulated in tumor tissue in comparison to the normal tissue (p < 0.001) and the patients with lower levels of PIWIL4 had shorter TTR (p = 0.048) and OS (p = 0.033). In the multivariate analysis, PIWIL1 expression emerged as an independent prognostic marker. Using 5-Aza-dC treatment and bisulfite sequencing, we observed that PIWIL1 expression could be regulated in part by methylation. Finally, an in silico study identified a stem-cell expression signature associated with PIWIL1 expression.

  2. Isogambogenic acid induces apoptosis-independent autophagic cell death in human non-small-cell lung carcinoma cells.

    Science.gov (United States)

    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.

  3. Clinical significance of aberrant Wnt7a promoter methylation in human non-small cell lung cancer in Koreans.

    Science.gov (United States)

    Kim, Tae-Hyung; Moon, Ji-Yong; Kim, Sang-Heon; Paik, Seung Sam; Yoon, Ho Joo; Shin, Dong Ho; Park, Sung Soo; Sohn, Jang Won

    2015-02-01

    The Wnt signaling pathway has regulatory roles in cell proliferation, differentiation, and polarity. Aberrant Wnt pathway regulation can lead to abnormal cell proliferation and cancer, and loss of Wnt7a expression has been demonstrated in lung cancer cell lines. E-cadherin keeps intercellular integrity and prevents metastasis. Therefore, E-cadherin has been known as a prognostic factor in cancer. In the present study, we investigated the E-cadherin expression status by immunohistochemical stain and the Wnt7a promoter methylation status in human non-small cell lung carcinoma (NSCLC) by methylation-specific PCR. We also analyzed their correlations with clinicopathological factors. Methylation of the Wnt7a gene promoter was detected in the lung tissues of 32 of 121 (26.4%) patients with NSCLC. Wnt7a promoter methylation was correlated with advanced tumor stage (P = 0.036) and distant metastasis (P = 0.037). In addition, Wnt7a promoter methylation showed correlation with loss of E-cadherin expression (P promoter methylation was not closely related with gender, age, histological type, or smoking habit. Even though Wnt7a methylation could not show significant correlation with the long term survival of the patients with limited follow up data, these findings suggest that loss of the Wnt7a gene induced by promoter methylation might be another prognostic factor for NSCLC and that restoration of Wnt7a may be a promising treatment for NSCLC.

  4. Association of advanced glycation end products with A549 cells, a human pulmonary epithelial cell line, is mediated by a receptor distinct from the scavenger receptor family and RAGE.

    Science.gov (United States)

    Nakano, Nahoko; Fukuhara-Takaki, Kaori; Jono, Tadashi; Nakajou, Keisuke; Eto, Nobuaki; Horiuchi, Seikoh; Takeya, Motohiro; Nagai, Ryoji

    2006-05-01

    Cellular interactions with advanced glycation end products (AGE)-modified proteins are known to induce several biological responses, not only endocytic uptake and degradation, but also the induction of cytokines and growth factors, combined responses that may be linked to the development of diabetic vascular complications. In this study we demonstrate that A549 cells, a human pulmonary epithelial cell line, possess a specific binding site for AGE-modified bovine serum albumin (AGE-BSA) (K(d) = 27.8 nM), and additionally for EN-RAGE (extracellular newly identified RAGE binding protein) (K(d) = 118 nM). Western blot and RT-PCR analysis showed that RAGE (receptor for AGE) is highly expressed on A549 cells, while the expression of other known AGE-receptors such as galectin-3 and SR-A (class A scavenger receptor), are below the level of detection. The binding of (125)I-AGE-BSA to these cells is inhibited by unlabeled AGE-BSA, but not by EN-RAGE. In contrast, the binding of (125)I-EN-RAGE is significantly inhibited by unlabeled EN-RAGE and soluble RAGE, but not by AGE-BSA. Our results indicate that A549 cells possess at least two binding sites, one specific for EN-RAGE and the other specific for AGE-BSA. The latter receptor on A549 cells is distinct from the scavenger receptor family and RAGE.

  5. Anacardic acid, a histone acetyltransferase inhibitor, modulates LPS-induced IL-8 expression in a human alveolar epithelial cell line A549 [v1; ref status: indexed, http://f1000r.es/o7

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    Tetsuo Yasutake

    2013-03-01

    Full Text Available 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.

  6. Smad2/3-Regulated Expression of DLX2 Is Associated with Radiation-Induced Epithelial-Mesenchymal Transition and Radioresistance of A549 and MDA-MB-231 Human Cancer Cell Lines

    Science.gov (United States)

    Choi, Yeo-Jin; Baek, Ga-Young; Park, Hae-Ran; Jo, Sung-Kee; Jung, Uhee

    2016-01-01

    The control of radioresistance and metastatic potential of surviving cancer cells is important for improving cancer eradication by radiotheraphy. The distal-less homeobox2 (DLX2) gene encodes for a homeobox transcription factor involved in morphogenesis and its deregulation was found in human solid tumors and hematologic malignancies. Here we investigated the role of DLX2 in association with radiation-induced epithelial to mesenchymal transition (EMT) and stem cell-like properties and its regulation by Smad2/3 signaling in irradiated A549 and MDA-MB-231 human cancer cell lines. In irradiated A549 and MDA-MB-231 cells, EMT was induced as demonstrated by EMT marker expression, phosphorylation of Smad2/3, and migratory and invasive ability. Also, irradiated A549 and MDA-MB-231 cells showed increased cancer stem cells (CSCs) marker. Interestingly, DLX2 was overexpressed upon irradiation. Therefore, we examined the role of DLX2 in radiation-induced EMT and radioresistance. The overexpression of DLX2 alone induced EMT, migration and invasion, and CSC marker expression. The reduced colony-forming ability in irradiated cells was partially restored by DLX2 overexpression. On the other hand, the depletion of DLX2 using si-RNA abolished radiation-induced EMT, CSC marker expression, and phosphorylation of Smad2/3 in irradiated A549 and MDA-MB-231 cells. Also, depletion of DLX2 increased the radiation sensitivity in both cell lines. Moreover, knockdown of Smad2/3, a key activator of TGF-β1 pathway, abrogated the radiation-induced DLX2 expression, indicating that radiation-induced DLX2 expression is dependent on Smad2/3 signaling. These results demonstrated that DLX2 plays a crucial role in radioresistance, radiation-induced EMT and CSC marker expression, and the expression of DLX2 is regulated by Smad2/3 signaling in A549 and MDA-MB-231 cell lines. PMID:26799321

  7. Clinical significance of SOX9 in human non-small cell lung cancer progression and overall patient survival

    Directory of Open Access Journals (Sweden)

    Zhou Chun-Hui

    2012-03-01

    Full Text Available Abstract Background Sex determining region Y (SRY-related high mobility groupbox 9 (SOX9 is an important transcription factor required for development, which regulates the expression of target genes in the associated pathway. The aim of this study was to describe the expression of SOX9 in human non-small cell lung cancer (NSCLC and to investigate the association between SOX9 expression and progression of NSCLC. Methods SOX9 protein and mRNA expression in normal human pneumonocytes, lung cancer cell lines, and eight pairs of matched lung cancer tissues and their adjacent normal lung tissues were detected by Western blotting and real-time reverse transcription-polymerase chain reaction (RT-PCR. Immunohistochemistry was used to determine SOX9 protein expression in 142 cases of histologically characterized NSCLC. Statistical analyses were applied to test for prognostic and diagnostic associations. Results SOX9 in lung cancer cell lines was upregulated at both mRNA and protein levels, and SOX9 mRNA and protein were also elevated in NSCLC tissues compared with levels in corresponding adjacent non-cancerous lung tissues. Immunohistochemical analysis demonstrated a high expression of SOX9 in 74/142 (52.1% paraffin-embedded archival lung cancer biopsies. Statistical analysis indicated that upregulation of SOX9 was significantly correlated with the histological stage of NSCLC (P = 0.017 and that patients with a high SOX9 level exhibited a shorter survival time (P Conclusions This work shows that SOX9 may serve as a novel and prognostic marker for NSCLC, and play a role during the development and progression of the disease.

  8. MiR-153 inhibits migration and invasion of human non-small-cell lung cancer by targeting ADAM19

    Energy Technology Data Exchange (ETDEWEB)

    Shan, Nianxi [Department of Oncology, Xiangya Hospital, Central South University, Changsha, Hunan 410008 (China); Institute of Medical Sciences, Xiangya Hospital, Central South University, Changsha, Hunan 410008 (China); Shen, Liangfang [Department of Oncology, Xiangya Hospital, Central South University, Changsha, Hunan 410008 (China); Wang, Jun; He, Dan [Institute of Medical Sciences, Xiangya Hospital, Central South University, Changsha, Hunan 410008 (China); Duan, Chaojun, E-mail: duancjxy@163.com [Department of Oncology, Xiangya Hospital, Central South University, Changsha, Hunan 410008 (China); Institute of Medical Sciences, Xiangya Hospital, Central South University, Changsha, Hunan 410008 (China)

    2015-01-02

    Highlights: • Decreased miR-153 and up-regulated ADAM19 are correlated with NSCLC pathology. • MiR-153 inhibits the proliferation and migration and invasion of NSCLC cells in vitro. • ADAM19 is a direct target of miR-153. • ADAM19 is involved in miR-153-suppressed migration and invasion of NSCLC cells. - Abstract: MiR-153 was reported to be dysregulated in some human cancers. However, the function and mechanism of miR-153 in lung cancer cells remains unknown. In this study, we investigated the role of miR-153 in human non-small-cell lung cancer (NSCLC). Using qRT-PCR, we demonstrated that miR-153 was significantly decreased in clinical NSCLC tissues and cell lines, and downregulation of miR-153 was significantly correlated with lymph node status. We further found that ectopic expression of miR-153 significantly inhibited the proliferation and migration and invasion of NSCLC cells in vitro, suggesting that miR-153 may be a novel tumor suppressor in NSCLC. Further integrated analysis revealed that ADAM19 is as a direct and functional target of miR-153. Luciferase reporter assay demonstrated that miR-153 directly targeted 3′UTR of ADAM19, and correlation analysis revealed an inverse correlation between miR-153 and ADAM19 mRNA levels in clinical NSCLC tissues. Knockdown of ADAM19 inhibited migration and invasion of NSCLC cells which was similar with effects of overexpression of miR-153, while overexpression of ADAM19 attenuated the function of miR-153 in NSCLC cells. Taken together, our results highlight the significance of miR-153 and ADAM19 in the development and progression of NSCLC.

  9. Study on proliferation inhibition and its mechanism of astragalus polysaccharide combined with cisplatin on human lung cancer A549 cells%黄芪多糖联合顺铂对人肺癌A549细胞增殖抑制作用及其机制研究

    Institute of Scientific and Technical Information of China (English)

    李蓉; 章运生; 谭小武

    2015-01-01

    Objective To observe the inhibitory effects of astragalus polysaccharides and cisplatin on the cell prolifera-tion,cycle,apoptosis induction of human lung cancer A549 cells,and initially discuss its action mechanism. Methods The ef-fects on cell proliferation of astragalus polysaccharides ,cisplatin and their combinations was detected by MTT assay. It analyzed the impact of cell cycle and apoptosis rate by flow cytometry. The A549 human lung cancer cells Bax ,Bcl-2 and Caspase-3 gene expression levels by reverse transcription polymerase chain reaction (RT-PCR) and Western blotting. Results Astragalus polysac-charide,cisplatin and two-drug combination on inhibition of proliferation of human lung cancer A549 cells had a time-concentra-tion-dependent proliferation. Compared with the negative control group ,the cell cycle blockage in G1 phase after astragalus polysaccharide action while the cell cycle blockage in S phase after cisplatin action and the application of two-drug combination appeared cell cycle blockage by flow cytometry both in G1 and S phases(P<0.01). The expression of Bax,Caspase-3 mRNA and pro-tein were upgraded in the expression of the two-drug combination group and lowered in Bcl-2 mRNA protein compared to the nega-tive control and monotherapy group. The difference had statistical significance (P<0.05). Conclusion The astragalus polysaccharide combined with cisplatin may strengthen human lung cancer A549 cells to enhance the pro-apoptosis,whose mechanism is associ-ated with upgrading the expression of Bax ,Caspase-3 and lowering the expression of Bcl-2mRNA.%目的:观察黄芪多糖与顺铂对人肺癌A549细胞增殖、细胞周期、诱导凋亡的影响,并对其作用机制进行初步探讨。方法采用四甲基偶氮唑盐法检测黄芪多糖、顺铂及两药联合对细胞增殖的影响;用流式细胞仪分析对细胞周期及凋亡率的影响;用反转录聚合酶链反应(RT-PCR)和Western blotting法检测对人肺癌A

  10. Role of Hypoxia-inducible factor-1 and its target genes in human lung adenocarcinoma cells after photon- versus carbon ion irradiation; Expression HIF-1-abhaengiger Gene in humanen Lungenadenokarzinom (A549)-Zellen und deren Regulation nach Photonen- und Schwerionenbestrahlung

    Energy Technology Data Exchange (ETDEWEB)

    Bill, Verena Maria

    2013-11-26

    Exposed to hypoxia tumor cells are notably resistant to photon irradiation. The hypoxiainducible transcription factor 1α (HIF-1α) seems to play a fundamental role in this resistance, while its role after heavy-ion beam remains unknown. The intention of this study was to determine how A549-cells (non-small-cell lung carcinoma) react in different oxygenation states after irradiation with photons or heavy ions, particularly in regards to their expression of HIF-1 target genes. Resistance of hypoxic A549 cells after photon irradiation was documented by cellular and clonogenic survival. In contrast, cellular survival after heavy-ion irradiation in hypoxic cells was not elevated to normoxic cells. Among the oxygen dependent regulation of HIF-1 target genes, gene expression analyses showed an increased expression of GLUT-1, LDH-A, PDK-1 and VEGF after photon irradiation but not after heavy-ion irradiation after 48 hours in normoxic cells. As expected, CDKN1A as inhibitor of cell cycle progression showed higher expression after both radiation forms; interestingly CDKN1A was also in an oxygen dependent manner lightly upregulated. In western blot analyses we demonstrated a significant increase of HIF-1 and GLUT-1 caused by hypoxia, but only a tendency of increased protein level in hypoxia after photon irradiation and no changes after heavy-ion irradiation. Significantly higher protein level of secreted VEGF-A could be measured 72 hours after photon irradiation in normoxic cells by ELISA analyses. Controversially discussed, I could not detect an association between HIF-1 and SCF or Trx-1 in A549-cells in this study. Whereas Trx-1-expression was neither influenced by changed oxygen partial pressure nor irradiation, I could show increased SCF mRNA by quantitative Real Time-PCR and secreted protein level by ELISA after photon irradiation independent of oxygen state. In summary, this study showed that HIF-1 and its target genes (GLUT-1, LDHA; PDK, VEGF) and also SCF was

  11. Combined therapy using suicide gef gene and paclitaxel enhances growth inhibition of multicellular tumour spheroids of A-549 human lung cancer cells.

    Science.gov (United States)

    Prados, Jose; Melguizo, Consolacion; Rama, Ana; Ortiz, Raul; Boulaiz, Houria; Rodriguez-Serrano, Fernando; Caba, Octavio; Rodriguez-Herva, Jose Juan; Ramos, Juan Luis; Aranega, Antonia

    2008-07-01

    The low efficiency of conventional therapies in achieving long-term survival of lung cancer patients calls for development of novel options. The potential use of combined gene therapy is under intensive study. One approach uses the expression of genes encoding cytotoxic proteins that affect cellular viability. The gef gene from E. coli, identified as a member of a gene family encoding homologous cell-killing functions, encodes for a membrane protein with a toxic domain which leads to a decrease in the rate of tumour cell growth. To improve the antitumoral effect of the paclitaxel in lung cancer cells, we investigated a combined suicide gene therapy using this drug and gef gene in vitro, using A-549 lung cancer cells in culture and forming multicellular tumour spheroids (MTS). Our results showed that gef expression in A-549 cells led to an ultrastructural changes, including dilated mitochondria with clear matrices and disrupted cristae and cell surface alterations such as reduction in length and number of microvilli and cytoplasmic membrane evaginations. The use of paclitaxel in A-549 lung cancer cells transfected with gef gene enhanced the chemotherapeutic effect of this drug. Volume analyses showed an 87.4% decrease in the A-549 MTS growth after 96 h in comparison with control MTS. This inhibition was greater than that obtained using the gene therapy or chemotherapy alone. In conclusion, gef gene has a cytotoxic effect in lung cancer cells and enhances cell growth inhibition when used with paclitaxel. These results indicate that this combined therapy may be of potential therapeutic value in lung cancer.

  12. Hedgehog Pathway Inhibition Radiosensitizes Non-Small Cell Lung Cancers

    Energy Technology Data Exchange (ETDEWEB)

    Zeng, Jing; Aziz, Khaled; Chettiar, Sivarajan T. [Department of Radiation Oncology and Molecular Radiation Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland (United States); Aftab, Blake T. [Department of Medical Oncology, The Johns Hopkins University School of Medicine, Baltimore, Maryland (United States); Armour, Michael; Gajula, Rajendra; Gandhi, Nishant; Salih, Tarek; Herman, Joseph M.; Wong, John [Department of Radiation Oncology and Molecular Radiation Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland (United States); Rudin, Charles M. [Department of Medical Oncology, The Johns Hopkins University School of Medicine, Baltimore, Maryland (United States); Tran, Phuoc T. [Department of Radiation Oncology and Molecular Radiation Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland (United States); Department of Medical Oncology, The Johns Hopkins University School of Medicine, Baltimore, Maryland (United States); Hales, Russell K., E-mail: rhales1@jhmi.edu [Department of Radiation Oncology and Molecular Radiation Sciences, The Johns Hopkins University School of Medicine, Baltimore, Maryland (United States)

    2013-05-01

    Purpose: Despite improvements in chemoradiation, local control remains a major clinical problem in locally advanced non-small cell lung cancer. The Hedgehog pathway has been implicated in tumor recurrence by promoting survival of tumorigenic precursors and through effects on tumor-associated stroma. Whether Hedgehog inhibition can affect radiation efficacy in vivo has not been reported. Methods and Materials: We evaluated the effects of a targeted Hedgehog inhibitor (HhAntag) and radiation on clonogenic survival of human non-small cell lung cancer lines in vitro. Using an A549 cell line xenograft model, we examined tumor growth, proliferation, apoptosis, and gene expression changes after concomitant HhAntag and radiation. In a transgenic mouse model of Kras{sup G12D}-induced and Twist1-induced lung adenocarcinoma, we assessed tumor response to radiation and HhAntag by serial micro-computed tomography (CT) scanning. Results: In 4 human lung cancer lines in vitro, HhAntag showed little or no effect on radiosensitivity. By contrast, in both the human tumor xenograft and murine inducible transgenic models, HhAntag enhanced radiation efficacy and delayed tumor growth. By use of the human xenograft model to differentiate tumor and stromal effects, mouse stromal cells, but not human tumor cells, showed significant and consistent downregulation of Hedgehog pathway gene expression. This was associated with increased tumor cell apoptosis. Conclusions: Targeted Hedgehog pathway inhibition can increase in vivo radiation efficacy in lung cancer preclinical models. This effect is associated with pathway suppression in tumor-associated stroma. These data support clinical testing of Hedgehog inhibitors as a component of multimodality therapy for locally advanced non-small cell lung cancer.

  13. Cytotoxic activity, DNA damage, cellular uptake, apoptosis and western blot analysis of ruthenium(II) polypyridyl complex against human lung decarcinoma A549 cell.

    Science.gov (United States)

    Lai, Shang-Hai; Jiang, Guang-Bin; Yao, Jun-Hua; Li, Wei; Han, Bing-Jie; Zhang, Cheng; Zeng, Chuan-Chuan; Liu, Yun-Jun

    2015-11-01

    A new ruthenium(II) polypyridyl complex [Ru(dmp)2(pddppn)](ClO4)2Ru1 was synthesized and characterized. The cytotoxic activity in vitro of the complex was evaluated by MTT method. Ru1 shows high effect on the inhibition of the cell growth against BEL-7402, HeLa, MG-63 and A549 cells with low IC50 values of 1.6±0.4, 9.0±0.8, 1.5±0.2 and 1.5±0.3 μM, respectively. The cellular uptake indicates that Ru1 can enter into the cytoplasm and accumulate in the cell nuclei. Ru1 can induce apoptosis in A549 cells and enhance the levels of reactive oxygen species (ROS) and induce the decrease of mitochondrial membrane potential. In addition, Ru1 can down-regulate the levels of Bcl-2, Bcl-x, Bak, and Bim expression and up-regulate the expression of Bag-1 and Bad. The complex induces apoptosis of A549 cells through an intrinsic ROS-mediated mitochondrial dysfunction pathway, which was accompanied by regulating the expression of caspases and Bcl-2 family proteins. Copyright © 2015 Elsevier Inc. All rights reserved.

  14. Construction of Eukaryotic Expression Vector of Human CC10 Gene and Expression of CC10 Protein in Lung Adenocarcinoma A549 Cell Line

    Institute of Scientific and Technical Information of China (English)

    2005-01-01

    A mammalian expression plasmid pcDNA3.1-hCC10 was constructed and identified, then CC10 protein expression in A549 lung cancer cell line was detected. A 273 bp cDNA fragment was amplified from the total RNA of normal lung tissue by using RT-PCR and cloned into expression plasmid cDNA3.1, and the recombinant plasmid was identified by employing double digestion restriction enzymes HindⅢ and BamH Ⅰ and the cDNA sequence was assayed by the Sanger dideoxymediated chain termination method. The segment was then transfected into the A549 lung cancer cell line. The protein expression of CC10 was detected by immunofluorescence and Western blot.Our results showed that the cDNA fragment included the entire coding region (273 bp). The recombinant eukaryotic cell expression vector of pcDNA3.1-hCC10 was successfully constructed, and the sequence of the insert was identical to the published sequence. A549 cells line transfected with the pcDNA3.1-hCC10 expressed high level of CC10 protein. The recombinant plasmid cDNA3. 1hCC10 may serve as an effective tool for the study of tumorogenesis and tumor treatment.

  15. Suppression of Type 1 Insulin-like Growth Factor Receptor Expression by Small Interfering RNA Inhibits A549 Human Lung Cancer Cell Invasion in vitro and Metastasis in Xenograft Nude Mice

    Institute of Scientific and Technical Information of China (English)

    Jianfang QIAN; Aiqiang DONG; Minjian KONG; Zhiyuan MA; Junqiang FAN; Guanyu JIANG

    2007-01-01

    Cancer invasion and metastasis, involving a variety of pathological processes and cytophysiological changes, contribute to the high mortality of lung cancer. The type 1 insulin-like growth factor receptor (IGF-1R), associated with cancer progression and invasion, is a potential anti-invasion and anti-metastasis target in lung cancer. To inhibit the invasive properties of lung cancer cells, we successfully down-regulated IGF-1R gene expression in A549 human lung cancer cells by small interfering RNA (siRNA)technology, and evaluated its effects on invasion-related gene expression, tumor cell in vitro invasion, and metastasis in xenograft nude mice. A549 cells transfected with a plasmid expressing hairpin siRNA for IGF-1R showed a significantly decreased IGF-1R expression at the mRNA level as well as the protein level. In biological assays, transfected A549 cells showed a significant reduction of cell-matrix adhesion,migration and invasion. Consistent with these results, we found that down-regulation of IGR-1R concomitantly accompanied by a large reduction in invasion-related gene expressions, including MMP-2,MMP-9, u-PA, and IGF-1R specific downstream p-Akt. Direct tail vein injections of plasmid expressing hairpin siRNA for IGF-1R significantly inhibited the formation of lung metastases in nude mice. Our results showed the therapeutic potential of siRNA as a method for gene therapy in inhibiting lung cancer invasion and metastasis.

  16. Ent-11α-Hydroxy-15-oxo-kaur-16-en-19-oic-acid Inhibits Growth of Human Lung Cancer A549 Cells by Arresting Cell Cycle and Triggering Apoptosis

    Institute of Scientific and Technical Information of China (English)

    Li Li; George G Chen; Ying-nian Lu; Yi Liu; Ke-feng Wu; Xian-ling Gong; Zhan-ping Gou; Ming-yue Li; Nian-ci Liang

    2012-01-01

    Objective:To examine the apoptotic effect of ent-11α-hydroxy-15-oxo-kaur-16-en-19-oic-acid (5F),a compound isolated from Pteris semipinnata L(PsL),in human lung cancer A549 cells.Methods:A549 cells were treated with 5F (0-80 μg/ml) for different time periods.Cytotoxicity was examined using a MTT method.Cell cycle was examined using propidium iodide staining.Apoptosis was examined using Hoechst 33258 staining,enzyme-linked immunosorbent assay (ELISA) and caspase-3 activity analysis.Expression of representative apoptosis-related proteins was evaluated by Western blot analysis.Reactive oxygen species (ROS) level was measured using standard protocols.Potential interaction of 5F with cisplatin was also examined.Results:5F inhibited the proliferation of A549 cells in a concentration- and time-dependent manner.5F increased the accumulation of cells in sub-G1 phase and arrested the cells in the G2 phase.Exposure to 5F induced morphological changes and DNA fragmentation that are characteristic of apoptosis.The expression of p21 was increased.5F exposure also increased Bax expression,release of cytochrome c and apoptosis inducing factor (AIF),and activation of caspase-3.5F significantly sensitized the cells to cisplatin toxicity Interestingly,treatment with 5F did not increase ROS,but reduced ROS production induced by cisplatin.Conclusion:SF could inhibit the proliferation of A549 cells by arresting the cells in G2 phase and by inducing mitochondrial-mediated apoptosis.

  17. Telmisartan Exerts Anti-Tumor Effects by Activating Peroxisome Proliferator-Activated Receptor-γ in Human Lung Adenocarcinoma A549 Cells

    Directory of Open Access Journals (Sweden)

    Juan Li

    2014-03-01

    Full Text Available Telmisartan, a member of the angiotensin II type 1 receptor blockers, is usually used for cardiovascular diseases. Recent studies have showed that telmisartan has the property of PPARγ activation. Meanwhile, PPARγ is essential for tumor proliferation, invasion and metastasis. In this work we explore whether telmisartan could exert anti-tumor effects through PPARγ activation in A549 cells. MTT and trypan blue exclusion assays were included to determine the survival rates and cell viabilities. RT-PCR and western blotting were used to analyze the expression of ICAM-1, MMP-9 and PPARγ. DNA binding activity of PPARγ was evaluated by EMSA. Our data showed that the survival rates and cell viabilities of A549 cells were all reduced by telmisartan in a time- and concentration-dependent manner. Meanwhile, our results also demonstrated that telmisartan dose-dependently inhibited the expression of ICAM-1 and MMP-9. Moreover, the cytotoxic and anti-proliferative effects, ICAM-1 and MMP-9 inhibitive properties of telmisartan were totally blunted by the PPARγ antagonist GW9662. Our findings also showed that the expression of PPARγ was up-regulated by telmisartan in a dose dependent manner. And, the EMSA results also figured out that DNA binding activity of PPARγ was dose-dependently increased by telmisartan. Additionally, our data also revealed that telmisartan-induced PPARγ activation was abrogated by GW9662. Taken together, our results indicated that telmisartan inhibited the expression of ICAM-1 and MMP-9 in A549 cells, very likely through the up-regulation of PPARγ synthesis.

  18. 用SILAC技术研究感染H5N1禽流感病毒后A549肺癌细胞蛋白质组的表达变化%Cellular Proteome Alterations in Highly Pathogenic H5N1 Avian In-fluenza Virus-Infected Human Lung Cell Line A549

    Institute of Scientific and Technical Information of China (English)

    王继峰; 李靖; 康晓平; 吴晓燕; 钱小红; 应万涛; 杨银辉

    2013-01-01

    Objective: To determine the cellular proteome responses of human lung A549 cell lines to the highly pathogenic H5N1 avian influenza virus infection, explore changes of specific molecular pathways and identify the key proteins involved in the infection. Methods: By using stable isotope labeling by amino acids in cell culture (SILAC) method to obtain“heavy”labeled cell lines which were infected with H5N1 virus and“light”labeled cell lines which were not infected, from which the cellular proteins were extracted and mixed in even amounts. Then the peptides derived from the mixed proteins digestion were identified by orthogonal reversed-phase chroma-tography coupled with mass spectroscopy and performed qualitative and quantitative analysis. Results: Of the total 3504 identified proteins and 2469 proteins with quantitative information, 72 were significantly up-regulated, 66 were significantly down-regulated. These proteins were involved in several molecular regulation pathways, including RNA splicesome, interferon inducible pathways, ubiquitin degradation pathway, insulin pathway and so on. Conclu-sion: We successfully established a strategy to explore the virus-host cell interactions with SILAC method. The identification of the key proteins involved in highly pathogenic H5N1 avian influenza virus infection, providedthe theoretical basis forunderstanding the molecular pathogenesis of H5N1 infection.%目的:鉴定高致病性H5N1禽流感病毒感染A549肺癌细胞后,细胞蛋白质组的表达变化,并鉴定特异分子通路的改变及其涉及的关键蛋白质分子。方法:利用稳定同位素标记氨基酸技术(SILAC)标记A549细胞,得到“重标”或“轻标”的A549细胞;“重标”细胞感染高致病性H5N1禽流感病毒24 h后提取细胞总蛋白,与从未感染病毒的“轻标”细胞中提取的总蛋白等量混合,酶解肽段,经正交反相色谱分离后用质谱鉴定,对数据进行定性和定量

  19. Minocycline enhances mitomycin C-induced cytotoxicity through down-regulating ERK1/2-mediated Rad51 expression in human non-small cell lung cancer cells.

    Science.gov (United States)

    Ko, Jen-Chung; Wang, Tai-Jing; Chang, Po-Yuan; Syu, Jhan-Jhang; Chen, Jyh-Cheng; Chen, Chien-Yu; Jian, Yun-Ting; Jian, Yi-Jun; Zheng, Hao-Yu; Chen, Wen-Ching; Lin, Yun-Wei

    2015-10-01

    Minocycline is a semisynthetic tetracycline derivative; it has anti-inflammatory and anti-cancer effects distinct from its antimicrobial function. However, the molecular mechanism of minocycline-induced cytotoxicity in non-small cell lung cancer (NSCLC) cells has not been identified. Rad51 plays a central role in homologous recombination and high levels of Rad51 expression are observed in chemo- or radioresistant carcinomas. Our previous studies have shown that the MKK1/2-ERK1/2 signal pathway maintains the expression of Rad51 in NSCLC cells. In this study, minocycline treatment inhibited cell viability and proliferation of two NSCLC cells, A549 and H1975. Treatment with minocycline decreased Rad51 mRNA and protein levels through MKK1/2-ERK1/2 inactivation. Furthermore, expression of constitutively active MKK1 (MKK1-CA) vectors significantly rescued the decreased Rad51 protein and mRNA levels in minocycline-treated NSCLC cells. However, combined treatment with MKK1/2 inhibitor U0126 and minocycline further decreased the Rad51 expression and cell viability of NSCLC cells. Knocking down Rad51 expression by transfection with small interfering RNA of Rad51 enhanced the cytotoxicity and cell growth inhibition of minocycline. Mitomycin C (MMC) is typically used as a first or second line regimen to treat NSCLC. Compared to a single agent alone, MMC combined with minocycline resulted in cytotoxicity and cell growth inhibition synergistically in NSCLC cells, accompanied with reduced activation of phospho-ERK1/2, and reduced Rad51 protein levels. Overexpression of MKK1-CA or Flag-tagged Rad51 could reverse the minocycline and MMC-induced synergistic cytotoxicity. These findings may have implications for the rational design of future drug regimens incorporating minocycline and MMC for the treatment of NSCLC.

  20. Ginsenoside Rg3 sensitizes human non-small cell lung cancer cells to γ-radiation by targeting the nuclear factor-κB pathway.

    Science.gov (United States)

    Wang, Lei; Li, Xiankui; Song, Yi-Min; Wang, Bin; Zhang, Fu-Rui; Yang, Rui; Wang, Hua-Qi; Zhang, Guo-Jun

    2015-07-01

    At present, it is elusive how non-small cell lung cancer (NSCLC) develops resistance to γ-radiation; however, the transcription factor nuclear factor-κB (NF-κB) and NF-κB-regulated gene products have been proposed as mediators. Ginsenoside Rg3 is a steroidal saponin, which was isolated from Panax ginseng. Ginsenoside Rg3 possesses high pharmacological activity and has previously been shown to suppress NF-κB activation in various types of tumor cell. Therefore, the present study aimed to determine whether Rg3 could suppress NF-κB activation in NSCLC cells and sensitize NSCLC to γ-radiation, using an NSCLC cell line and NSCLC xenograft. A clone formation assay and lung tumor xenograft experiment were used to assess the radiosensitizing effects of ginsenoside Rg3. NF-κB/inhibitor of NF-κB (IκB) modulation was ascertained using an electrophoretic mobility shift assay and western blot analysis. NF-κB-regulated gene products were monitored by western blot analysis. The present study demonstrated that ginsenoside Rg3 was able to sensitize A549 and H1299 lung carcinoma cells to γ-radiation and significantly enhance the efficacy of radiation therapy in C57BL/6 mice bearing a Lewis lung carcinoma cell xenograft tumor. Furthermore, ginsenoside Rg3 suppressed NF-κB activation, phosphorylation of IκB protein and expression of NF-κB-regulated gene products (cyclin D1, c-myc, B-cell lymphoma 2, cyclooxygenase-2, matrix metalloproteinase-9 and vascular endothelial growth factor), a number of which were induced by radiation therapy and mediate radioresistance. In conclusion, the results of the present study suggested that ginsenoside Rg3 may potentiate the antitumor effects of radiation therapy in NSCLC by suppressing NF-κB activity and NF-κB-regulated gene products, leading to the inhibition of tumor progression.

  1. Anticancer activity of Noscapine, an opioid alkaloid in combination with Cisplatin in human non-small cell lung cancer.

    Science.gov (United States)

    Chougule, Mahavir; Patel, Apurva R; Sachdeva, Pratik; Jackson, Tanise; Singh, Mandip

    2011-03-01

    The purpose of this study was to examine the efficacy of Noscapine (Nos) and Cisplatin (Cis) combination treatment in vitro in A549 and H460 lung cancer cells, in vivo in murine xenograft model and to investigate the underlying mechanism. The combination index values (< 0.6) suggested synergistic effects of Nos+Cis and resulted in the highest increase in percentage of apoptotic NSCLC cells and increased expression of p53, p21, caspase 3, cleaved caspase 3, cleaved PARP, Bax, and decreased expression of Bcl₂ and surviving proteins compared with treatment with either agent. Nos+Cis treatment reduced tumor volume by 78.1 ± 7.5% compared with 38.2 ± 6.8% by Cis or 35.4 ± 6.9% by Nos alone in murine xenograft lung cancer model. Nos+Cis treatment decreased expression of pAkt, Akt, cyclin D1, survivin, PARP, Bcl₂, and increased expression of p53, p21, Bax, cleaved PARP, caspase 3, cleaved caspase 3, cleaved caspase 8, caspase 8, cleaved caspase 9 and caspase 9 compared to single-agent treated and control groups. Our results suggest that Nos enhanced the anticancer activity of Cis in an additive to synergistic manner by activating multiple signaling pathways including apoptosis. These findings suggest potential benefit for use of Nos and Cis combination in treatment of lung cancer.

  2. Effect of an Albumin-Coated Mesoporous Silicon Nanoparticle Platform for Paclitaxel Delivery in Human Lung Cancer Cell Line A549

    Directory of Open Access Journals (Sweden)

    Yu Gao

    2016-01-01

    Full Text Available Albumin-coated paclitaxel-mesoporous silicon nanoparticles (APMSN were prepared to improve the anticancer effect in lung cancer by means of regulating the dissolution rate of paclitaxel (PTX. PTX was absorbed into the mesoporous structure of mesoporous silicon nanoparticles (MSN, which was defined as PMSN. PTX was proved to exist in an amorphous state in PMSN, which increased the dissolution rate of PTX. Albumin was coated on the surface of MSN to form AMSN; AMSN and PTX were mixed to form APMSN in order to achieve sustained release of PTX. Then, it was found that APMSN had more significant antiproliferate effects and induced more apoptotic proportion in comparison with PTX in A549 cells. Furthermore, the absorption mechanism of APMSN into A549 cells was investigated. Transmission electron microscopy (TEM and laser scanning confocal microscopy (LSCM showed that APMSN could cross the cell membrane and was taken into the cytoplasm quickly. Taken together, our results demonstrate that AMSN carriers have potential as nanodrug delivery systems in the treatment of lung cancer.

  3. Synergistic induction of apoptosis by sulindac and simvastatin in A549 human lung cancer cells via reactive oxygen species-dependent mitochondrial dysfunction.

    Science.gov (United States)

    Hwang, Ki-Eun; Park, Chul; Kwon, Su-Jin; Kim, Young-Suk; Park, Do-Sim; Lee, Mi-Kyung; Kim, Byoung-Ryun; Park, Seong-Hoon; Yoon, Kwon-Ha; Jeong, Eun-Taik; Kim, Hak-Ryul

    2013-07-01

    Prevention of lung cancer is more feasible and holds greater promise when different agents are used in combination to target multiple processes during carcinogenesis. The mechanisms by which non-steroidal anti-inflammatory drugs and statins inhibit cancer cell growth and induce apoptosis are not fully understood. This study was designed to investigate lung cancer chemoprevention through a mechanism-based approach using sulindac at low doses in combination with simvastatin. We found that sulindac-induced cytotoxicity was significantly enhanced in the presence of simvastatin. The combination of sulindac and simvastatin induced more extensive caspase-dependent apoptosis in A549 cells compared to that induced with either drug alone. The combination of sulindac and simvastatin also increased the loss of mitochondrial transmembrane potential (∆Ψm) and the cytosolic release of cytochrome c. In addition, ROS generation in cells treated with both sulindac and simvastatin was markedly increased compared to cells treated with either sulindac or simvastatin alone. The enhancement of ROS generation by sulindac and simvastatin was abrogated by pretreatment with NAC, which also prevented apoptosis and mitochondrial dysfunction induced by sulindac and simvastatin. These results suggest that sulindac and simvastatin-induced ROS generation in A549 lung cancer cells causes their accumulation in mitochondria, triggering the release of apoptogenic molecules from the mitochondria to the cytosol, and thus leading to caspase activation and cell death.

  4. Aspergillus fumigatus germ tube growth and not conidia ingestion induces expression of inflammatory mediator genes in the human lung epithelial cell line A549.

    Science.gov (United States)

    Bellanger, Anne-Pauline; Millon, Laurence; Khoufache, Khaled; Rivollet, Danièle; Bièche, Ivan; Laurendeau, Ingrid; Vidaud, Michel; Botterel, Françoise; Bretagne, Stéphane

    2009-02-01

    Inhalation of conidia is the main cause of invasive pulmonary aspergillosis (IPA) and the respiratory epithelium is the first line of defence. To explore the triggering factor for the inflammatory response to Aspergillus fumigatus, the species mainly responsible for IPA, this study analysed the differential expression of three inflammatory genes in A549 cells after challenge with live and killed conidia. The influence of steroids, one of the main risk factors for developing IPA, was also investigated. Quantification of mRNAs of the inflammatory mediator genes encoding interleukin (IL)-8, tumour necrosis factor (TNF)-alpha and granulocyte-monocyte colony-stimulating factor (GM-CSF) was carried out using real-time PCR. Ingestion rates were studied for the conidia of A. fumigatus and Penicillium chrysogenum using a fluorescence brightener. Similar results were obtained for both species, with ingestion rates ranging from 35 to 40 %. Exposure of A549 cells to live A. fumigatus conidia only induced a four- to fivefold increase in the mRNA levels of the three genes, starting 8 h after the initial contact. Both inactivation of live A. fumigatus conidia and treatment by dexamethasone (10(-7) M) prevented the overexpression of TNF-alpha, IL-8 and GM-CSF. Fungal growth, rather than conidia ingestion, appears to be the main stimulus for the production of inflammatory mediators by epithelial cells, and this production is inhibited by steroid therapy. These results underline the role that the epithelium plays in the innate response against IPA.

  5. PPAR-γ Activation Inhibits Angiogenesis by Blocking ELR+CXC Chemokine Production in Non-small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Venkateshwar G. Keshamouni

    2005-03-01

    Full Text Available Activation of peroxisome proliferator-activated receptor-γ (PPAR-γ results in inhibition of tumor growth in various types of cancers, but the mechanism(s by which PPAR-γ induces growth arrest has not been completely defined. In a recent study, we demonstrated that treatment of A549 (human non small cell lung cancer cell line tumor-bearing SCID mice with PPAR-γ ligands troglitazone (Tro and pioglitazone significantly inhibits primary tumor growth. In this study, immunohistochemical analysis of Tro-treated and Pio-treated tumors with factor VIII antibody revealed a significant reduction in blood vessel density compared to tumors in control animals, suggesting inhibition of angiogenesis. Further analysis showed that treatment of A549 cells in vitro with Tro or transient transfection of A549 cells with constitutively active PPAR-γ (VP16-PPAR-γ construct blocked the production of the angiogenic ELR +CXC chemokines IL-8 (CXCL8, ENA-78 (CXCL5, Gro-α (CXCL1. Similarly, an inhibitor of NF-ΚB activation (PDTC also blocked CXCL8, CXCL5, CXCL1 production, consistent with their NF-ΚB-dependent regulation. Conditioned media from A549 cells induce human microvascular endothelial cell (HMVEC chemotaxis. However, conditioned media from Tro-treated A549 cells induced significantly less HMVEC chemotaxis compared to untreated A549 cells. Furthermore, PPAR-γ activation inhibited NF-ΚB transcriptional activity, as assessed by TransAM reporter gene assay. Collectively, our data suggest that PPAR-γ ligands can inhibit tumor-associated angiogenesis by blocking the production of ELR+CXC chemokines, which is mediated through antagonizing NF-ΚB activation. These antiangiogenic effects likely contribute to the inhibition of primary tumor growth by PPAR-γ ligands.

  6. Empirical study on the anti-proliferation effect of siRNA against pokemon on human lung cancer cell line A549%siRNA干扰Pokemon基因影响A549细胞增殖的实验研究

    Institute of Scientific and Technical Information of China (English)

    谢勇; 江涛

    2012-01-01

    目的 研究siRNA干扰Pokemon基因对肺腺癌A549细胞增殖抑制效应的变化.方法 专业设计合成3条针对Pokemon的siRNA,分别转染A549细胞后,RT-PCR检测转录水平Pokemon mRNA表达的变化,筛选出其中最高效的1条siRNA;用MTT法检测该siRNA干扰Pokemon对A549细胞增殖的抑制作用;流式细胞技术检测该siRNA干扰Pokemon对A549细胞凋亡的影响.结果 3条siRNA均成功转染A549细胞,倒置荧光显微镜下观察细胞呈圆绿色.RT-PCR结果显示有2条siRNA使细胞中Pokemon的mRNA表达降低(P<0.05).MTT法结果显示siRNA干扰Pokemon后对A549细胞增殖有抑制作用(P<0.05),其中48 h抑制效率达(24.14±1.39)%.流式细胞技术检测结果显示该siRNA干扰Pokemon可增加A549细胞的凋亡,凋亡率为14.05%.结论 应用RNA干扰Pokemon基因能够抑制A549细胞的增殖,促进A549细胞的凋亡.Pokemon基因有可能成为肺癌治疗中的一个新靶点.

  7. PKM2 Thr454 phosphorylation increases its nuclear translocation and promotes xenograft tumor growth in A549 human lung cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Yu, Zhenhai, E-mail: tomsyu@163.com [Center for Reproductive Medicine, Affiliated Hospital of Weifang Medical University, Weifang, Shandong, 261031 (China); Huang, Liangqian [Institute of Health Sciences, Shanghai Institutes for Biological Sciences (SIBS), Chinese Academy of Sciences (CAS) & Shanghai Jiao Tong University School of Medicine -SJTUSM, Shanghai, 200025 (China); Qiao, Pengyun; Jiang, Aifang; Wang, Li; Yang, Tingting [Center for Reproductive Medicine, Affiliated Hospital of Weifang Medical University, Weifang, Shandong, 261031 (China); Tang, Shengjian; Zhang, Wei [Plastic Surgery Institute of Weifang Medical University, Weifang, Shandong, 261041 (China); Ren, Chune, E-mail: ren@wfmc.edu.cn [Center for Reproductive Medicine, Affiliated Hospital of Weifang Medical University, Weifang, Shandong, 261031 (China)

    2016-05-13

    Pyruvate kinase M2 (PKM2) is a key enzyme of glycolysis which is highly expressed in many tumor cells, and plays an important role in the Warburg effect. In previous study, we found PIM2 phosphorylates PKM2 at Thr454 residue (Yu, etl 2013). However, the functions of PKM2 Thr454 modification in cancer cells still remain unclear. Here we find PKM2 translocates into the nucleus after Thr454 phosphorylation. Replacement of wild type PKM2 with a mutant (T454A) enhances mitochondrial respiration, decreases pentose phosphate pathway, and enhances chemosensitivity in A549 cells. In addition, the mutant (T454A) PKM2 reduces xenograft tumor growth in nude mice. These findings demonstrate that PKM2 T454 phosphorylation is a potential therapeutic target in lung cancer.

  8. Overexpression of SAMD9 suppresses tumorigenesis and progression during non small cell lung cancer

    Energy Technology Data Exchange (ETDEWEB)

    Ma, Qing; Yu, Tao; Ren, Yao-Yao; Gong, Ting; Zhong, Dian-Sheng, E-mail: zhongdsyx@126.com

    2014-11-07

    Highlights: • SAMD9 is down-regulated in human non-small cell lung cancer (NSCLC). • Knockdown of SAMD9 expression is increased the invasion, migration and proliferation in H1299 cells in vitro. • Overexpression of SAMD9 suppressed proliferation and invasion in A549 cells in vitro. • Depletion of SAMD9 increases tumor formation in vivo. - Abstract: The Sterile Alpha Motif Domain-containing 9 (SAMD9) gene has been recently emphasized during the discovery that it is expressed at a lower level in aggressive fibromatosis and some cases of breast and colon cancer, however, the underlying mechanisms are poorly understood. Here, we found that SAMD9 is down-regulated in human non-small cell lung cancer (NSCLC). Furthermore, knockdown of SAMD9 expression is increased the invasion, migration and proliferation in H1299 cells in vitro and overexpression of SAMD9 suppressed proliferation and invasion in A549 cells. Finally, depletion of SAMD9 increases tumor formation in vivo. Our results may provide a strategy for blocking NSCLC tumorigenesis and progression.

  9. RNA干扰抑制Snail表达对A549细胞上皮-间充质转分化及体外侵袭的影响%Inhibition of Snail expression by RNA interference repress epithelial-mesenchymal transition and invasion ability of human lung carcinoma cell A549 in vitro

    Institute of Scientific and Technical Information of China (English)

    卓文磊; 张云嵩; 王彦; 敖绪军; 陈正堂

    2008-01-01

    目的:研究用RNA干扰(RNA interference,RNAi)技术抑制转录因子Snail表达后,对人肺腺癌A549细胞上皮-间充质转分化表型和体外侵袭能力的影响.方法:构建能表达针对Snail的小干扰RNA(Small interfering RNA,siRNA)的RNA干扰载体(Snail siRNA vector)和表达不针对任何已知mRNA的siRNA的阴性对照RNA干扰载体(control siRNA vector),分别转染A549细胞,新霉素抗性筛选得到Snail表达受抑制的A549-siSnail细胞和Snail表达未受影响的A549-siControl细胞.针对非转染(A549-nontransfection)、A549-siSnail、A549-siControl三组细胞,分别采用RT-PCR和Western blot技术检测Snail、α-平滑肌肌动蛋白(α-SMA)和E-钙粘素表达,用Boyden chamber模型检测细胞侵袭能力.结果: A549-nontransfection组:Snail和α-SMA表达强阳性,E-钙粘素表达弱阳性;A549-siSnail组:和A549-nontransfection组相比,Snail和α-SMA表达显著减弱(P0.05).结论: 通过RNA干扰阻滞Snail表达能有效地抑制A549细胞上皮-间充质转分化及体外侵袭能力.Snail可能在肺腺癌上皮-间充质转分化及侵袭过程中扮演重要角色,抑制Snail表达可能成肺腺癌治疗的可行策略.

  10. P型铜转运ATP酶(ATP7B)在肺腺癌细胞株A549中的表达与顺铂耐药的关系%Expression of Copper-Transporting P-Type Adenosine Triphosphatase(ATP7B) Correlates with Cisplatin-Resistance in Human Lung Adenocarcinoma Cell Line A549

    Institute of Scientific and Technical Information of China (English)

    高贵洲; 王建军; 石思恩

    2009-01-01

    背景与目的 顺铂作为肺癌的基础化疗药物,顺铂耐药是导致肺癌患者化疗失败的重要原因.本实验通过检测P型铜转运ATP酶在肺腺癌细胞A549不同水平耐药株中的表达,以评估ATP7B与A549细胞顺铂耐药的关系.方法采用逐步增加顺铂剂量的方法,诱导出3株不同水平耐顺铂A549细胞株A549/DDP0.5、A549/DDP1.0、A549/DDP2.0,MTT方法检测各组别细胞对顺铂敏感性,应用RT-PCR及Western Blot方法分别检测各组别细胞的ATP7B的mRNA及蛋白表达水平,分析A549细胞顺铂敏感性与ATP7B表达水平的关系.结果相对于亲本A549细胞,3组耐药细胞的顺铂耐药指数分别达到了1.7、3.2、5.2(P<0.001),与此相对应ATP7B的mRNA表达水平分别达到了亲本A549细胞的1.6、4.9、10.1倍(P<0.001),同样地ATP7B的蛋白表达水平也呈现出与顺铂耐药性相平行的递增性高表达.结论肺腺癌细胞A549的顺铂耐药与细胞ATP7B高表达有关,后者的高表达有可能促成了A549细胞的获得性顺铂耐药.

  11. 牛膝多糖硫酸酯和磷酸酯衍生物对人肺癌A549细胞的影响%Effect of the sulfated and phosphorylated derivative of Achyranthes bidentata polysaccharides on human lung cancer cell A549

    Institute of Scientific and Technical Information of China (English)

    薛胜霞; 金丽琴; 叶发青; 贾东明

    2007-01-01

    目的 探讨牛膝多糖硫酸酯(s-AbPS)和磷酸酯(P-AbPS)衍生物对人肺癌A549细胞的影响及其作用机制.方法 用四甲基偶氯唑盐(MTT)法检测牛膝多糖、S-AbPS和P-AbPS对A549细胞增殖的抑制作用;流式细胞仪观察S-AbPS和P-AbPS处理A549细胞后细胞凋亡的改变.结果 S-AbPS和P-AbPS对A549细胞的增殖有抑制作用,且存在时间和剂量相关性.S-AbPS和P-AbPS的48 h抑瘤活性均较同浓度的牛膝多糖强.流式细胞仪分析显示,S-AbPS和P-AbPS均可不同程度地诱导细胞凋亡.结论 S-AbPS和P-AbPS较牛膝多糖能更有效地抑制人肺癌A549细胞的增殖,其机制可能是通过诱导A549细胞凋亡而实现.

  12. Eosinophil Granulocytes Account for Indoleamine 2,3-Dioxygenase-Mediated Immune Escape in Human Non Small Cell Lung Cancer

    Directory of Open Access Journals (Sweden)

    Simonetta Astigiano

    2005-04-01

    Full Text Available Indoleamine 2,3-dioxygenase (IDO, a catabolizing enzyme of tryptophan, is supposed to play a role in tumor immune escape. Its expression in solid tumors has not yet been well elucidated: IDO can be expressed by the tumor cells themselves, or by ill-defined infiltrating cells, possibly depending on tumor type. We have investigated IDO expression in 25 cases of non small cell lung cancer (NSCLC. Using histochemistry and immunohistochemistry, we found that IDO was expressed not by tumor cells, but by normal cells infiltrating the peritumoral stroma. These cells were neither macrophages nor dendritic cells, and were identified as eosinophil granulocytes. The amount of IDO-positive eosinophils varied in different cases, ranging from a few cells to more than 50 per field at x200 magnification. IDO protein in NSCLC was enzymatically active. Therefore, at least in NSCLC cases displaying a large amount of these cells in the inflammatory infiltrate, IDO-positive eosinophils could exert an effective immunosuppressive action. On analyzing the 17 patients with adequate follow-up, a significant relationship was found between the amount of IDO-positive infiltrate and overall survival. This finding suggests that the degree of IDO-positive infiltrate could be a prognostic marker in NSCLC.

  13. Non-small-cell lung cancer-induced immunosuppression by increased human regulatory T cells via Foxp3 promoter demethylation.

    Science.gov (United States)

    Ke, Xing; Zhang, Shuping; Xu, Jian; Liu, Genyan; Zhang, Lixia; Xie, Erfu; Gao, Li; Li, Daqian; Sun, Ruihong; Wang, Fang; Pan, Shiyang

    2016-05-01

    Patients with non-small-cell lung cancer (NSCLC) have immune defects that are poorly understood. Forkhead box protein P3 (Foxp3) is crucial for immunosuppression by CD4(+) regulatory T cells (Tregs). It is not well known how NSCLC induces Foxp3 expression and causes immunosuppression in tumor-bearing patients. Our study found a higher percentage of CD4(+) Tregs in the peripheral blood of NSCLC compared with healthy donors. NSCLC patients showed demethylation of eight CpG sites within the Foxp3 promoter with methylation ratios negatively correlated with CD4(+)CD25(+)Foxp3(+) T levels. Foxp3 expression in CD4(+) Tregs was directly regulated by Foxp3 promoter demethylation and was involved in immunosuppression by NSCLC. To verify the effect of tumor cells on the phenotype and function of CD4(+) Tregs, we established a coculture system using NSCLC cell line and healthy CD4(+) T cells and showed that SPC-A1 induced IL-10 and TGF-β1 secretion by affecting the function of CD4(+) Tregs. The activity of DNA methyltransferases from CD4(+) T was decreased during this process. Furthermore, eight CpG sites within the Foxp3 promoter also appeared to have undergone demethylation. Foxp3 is highly expressed in CD4(+) T cells, and this may be caused by gene promoter demethylation. These induced Tregs are highly immunosuppressive and dramatically inhibit the proliferative activity of naïve CD4(+) T cells. Our study provides one possible mechanism describing Foxp3 promoter demethylation changes by which NSCLC down-regulates immune responses and contributes to tumor progression. Foxp3 represents an important target for NSCLC anti-tumor immunotherapy.

  14. Gracilaria edulis exhibit antiproliferative activity against human lung adenocarcinoma cell line A549 without causing adverse toxic effect in vitro and in vivo.

    Science.gov (United States)

    Sakthivel, Ravi; Muniasamy, Samuthirapandi; Archunan, Govindaraju; Devi, Kasi Pandima

    2016-02-01

    In the present study, the antiproliferative potential of various solvent extracts of Gracilaria edulis (GE) was tested against various cancer cell lines. In the A549 lung cancer cell line model, GE ethyl acetate extract (GEEA) (100 μg mL(-1)) treated group showed the maximum and significant (P < 0.05) growth inhibition at 48 h. The IC50 value was found to be 24.5 ± 19.1 μg mL(-1) at 48 h. Moreover, a low level of LDH release was observed at 48 h at various concentrations of (40, 60, 80 and 100 μg mL(-1)) GEEA extract-treated group compared to a control group. Changes in the cell morphology and echinoid spikes formation were observed at 48 h. Safety evaluation of GEEA in a non-cancerous liver cell line, PBMC and in Wistar rats positively revealed that the extract did not show any adverse toxic effects. The GEEA extract was partially purified by column chromatography and the active fraction was characterized through LC-MS analysis. Furthermore, HPLC and FT-IR analysis of the active fractions confirmed the presence of phytol, a diterpene compound with potent antiproliferative activity, which positively suggests that the red alga G. edulis contains a potent anticancer active principle.

  15. Effect of Paclitaxel-Mesoporous Silica Nanoparticles with a Core-Shell Structure on the Human Lung Cancer Cell Line A549

    Science.gov (United States)

    Wang, Tieliang; Liu, Ying; Wu, Chao

    2017-01-01

    A nanodrug delivery system of paclitaxel-mesoporous silica nanoparticles with a core-shell structure (PAC-csMSN) was used to increase the dissolution of paclitaxel (PAC) and improve its treatment of lung cancer. PAC was loaded into the core-shell mesoporous silica nanoparticles (csMSN) by the adsorption equilibrium method and was in an amorphous state in terms of its mesoporous structure. In vitro and in vivo studies showed that csMSN increased the dissolution rate of PAC and improved its lung absorption. The area under concentration-time curve (AUC) value of PAC-csMSN used for pulmonary delivery in rabbits was 2.678-fold higher than that obtained with the PAC. After continuous administration for 3 days, a lung biopsy showed no signs of inflammation. Cell apoptosis results obtained by flow cytometry indicated that PAC-csMSN was more potent than pure PAC in promoting cell apoptosis. An absorption investigation of PAC-csMSN in A549 cells was carried out by transmission electron microscopy (TEM) and laser scanning confocal microscopy (LSCM). The obtained results indicated that the cellular uptake was time-dependent and csMSN was uptaken into the cytoplasm. All these results demonstrate that csMSN have the potential to achieve pulmonary inhalation administration of poorly water-soluble drugs for the treatment of lung cancer.

  16. Green tea induces annexin-I expression in human lung adenocarcinoma A549 cells: involvement of annexin-I in actin remodeling.

    Science.gov (United States)

    Lu, Qing-Yi; Jin, Yu Sheng; Zhang, Zuo-Feng; Le, Anh D; Heber, David; Li, Frederick P; Dubinett, Steven M; Rao, Jian Yu

    2007-05-01

    Green tea polyphenols exhibit multiple antitumor activities in various in vitro and in vivo tumor models, and the mechanisms of action are not clear. Previously, we found that green tea extract (GTE) regulates actin remodeling in different cell culture systems. Actin remodeling plays an important role in cancer cell morphology, cell adhesion, motility, and invasion. Using proteomic approaches, we found GTE-induced expression of annexin-I, a multifunctional actin binding protein, in these cell lines. In this study, we aimed to further define the functional role of GTE-induced annexin-I expression in actin remodeling, cell adhesion, and motility in lung adenocarcinoma A549 cells. We found that GTE stimulates the expression of annexin-I in a dose-dependent fashion. The GTE-induced annexin-I expression appears to be at the transcription level, and the increased annexin-I expression mediates actin polymerization, resulting in enhanced cell adhesion and decreased motility. Annexin-I specific interference resulted in loss of GTE-induced actin polymerization and cell adhesion, but not motility. In fact, annexin-I specific interference itself inhibited motility even without GTE. Together, annexin-I plays an important role in GTE-induced actin remodeling, and it may serve as a potential molecular target associated with the anticancer activities of green tea.

  17. TGF-β1 downregulates COX-2 expression leading to decrease of PGE2 production in human lung cancer A549 cells, which is involved in fibrotic response to TGF-β1.

    Directory of Open Access Journals (Sweden)

    Erina Takai

    Full Text Available Transforming growth factor-ß1 (TGF-β1 is a multifunctional cytokine that is involved in various pathophysiological processes, including cancer progression and fibrotic disorders. Here, we show that treatment with TGF-β1 (5 ng/mL induced downregulation of cyclooxygenase-2 (COX-2, leading to reduced synthesis of prostaglandin E2 (PGE2, in human lung cancer A549 cells. Treatment of cells with specific inhibitors of COX-2 or PGE2 receptor resulted in growth inhibition, indicating that the COX-2/PGE2 pathway contributes to proliferation in an autocrine manner. TGF-β1 treatment induced growth inhibition, which was attenuated by exogenous PGE2. TGF-β1 is also a potent inducer of epithelial mesenchymal transition (EMT, a phenotype change in which epithelial cells differentiate into fibroblastoid cells. Supplementation with PGE2 or PGE2 receptor EP4 agonist PGE1-alcohol, as compared with EP1/3 agonist sulprostone, inhibited TGF-β1-induced expression of fibronectin and collagen I (extracellular matrix components. Exogenous PGE2 or PGE2 receptor agonists also suppressed actin remodeling induced by TGF-β1. These results suggest that PGE2 has an anti-fibrotic effect. We conclude that TGF-β1-induced downregulation of COX-2/PGE2 signaling is involved in facilitation of fibrotic EMT response in A549 cells.

  18. Irciniastatin A induces potent and sustained activation of extracellular signal-regulated kinase and thereby promotes ectodomain shedding of tumor necrosis factor receptor 1 in human lung carcinoma A549 cells.

    Science.gov (United States)

    Quach, Hue Tu; Hirano, Seiya; Fukuhara, Sayuri; Watanabe, Tsubasa; Kanoh, Naoki; Iwabuchi, Yoshiharu; Usui, Takeo; Kataoka, Takao

    2015-01-01

    Irciniastatin A is a pederin-type marine product that potently inhibits translation. We have recently shown that irciniastatin A induces ectodomain shedding of tumor necrosis factor (TNF) receptor 1 with slower kinetics than other translation inhibitors. In human lung carcinoma A549 cells, irciniastatin A induced a marked and sustained activation of extracellular signal-regulated kinase (ERK) and induced little activation of p38 mitogen-activated protein (MAP) kinase and c-Jun N-terminal kinase (JNK). Moreover, the TNF receptor 1 shedding induced by irciniastatin A was blocked by the MAP kinase/ERK kinase inhibitor U0126, but not by the p38 MAP kinase inhibitor SB203580 or the JNK inhibitor SP600125. Thus unlike other translation inhibitors that trigger ribotoxic stress response, our results show that irciniastatin A is a unique translation inhibitor that induces a potent and sustained activation of the ERK pathway, and thereby promotes the ectodomain shedding of TNF receptor 1 in A549 cells.

  19. TGF-β1 Downregulates COX-2 Expression Leading to Decrease of PGE2 Production in Human Lung Cancer A549 Cells, Which Is Involved in Fibrotic Response to TGF-β1

    Science.gov (United States)

    Takai, Erina; Tsukimoto, Mitsutoshi; Kojima, Shuji

    2013-01-01

    Transforming growth factor-ß1 (TGF-β1) is a multifunctional cytokine that is involved in various pathophysiological processes, including cancer progression and fibrotic disorders. Here, we show that treatment with TGF-β1 (5 ng/mL) induced downregulation of cyclooxygenase-2 (COX-2), leading to reduced synthesis of prostaglandin E2 (PGE2), in human lung cancer A549 cells. Treatment of cells with specific inhibitors of COX-2 or PGE2 receptor resulted in growth inhibition, indicating that the COX-2/PGE2 pathway contributes to proliferation in an autocrine manner. TGF-β1 treatment induced growth inhibition, which was attenuated by exogenous PGE2. TGF-β1 is also a potent inducer of epithelial mesenchymal transition (EMT), a phenotype change in which epithelial cells differentiate into fibroblastoid cells. Supplementation with PGE2 or PGE2 receptor EP4 agonist PGE1-alcohol, as compared with EP1/3 agonist sulprostone, inhibited TGF-β1-induced expression of fibronectin and collagen I (extracellular matrix components). Exogenous PGE2 or PGE2 receptor agonists also suppressed actin remodeling induced by TGF-β1. These results suggest that PGE2 has an anti-fibrotic effect. We conclude that TGF-β1-induced downregulation of COX-2/PGE2 signaling is involved in facilitation of fibrotic EMT response in A549 cells. PMID:24098479

  20. Effect of piperine combined with cisplation on proliferation and MMP-2, VEGF of human lung adenocarcinoma A549 cell%胡椒碱联合顺铂对肺腺癌 A549 细胞生长及 MMP-2、VEGF 的影响

    Institute of Scientific and Technical Information of China (English)

    何含含; 岳红梅; 鲁文强; 罗亚娟

    2015-01-01

    目的:研究胡椒碱与顺铂联合对肺腺癌A549细胞生长和凋亡的作用,以及检测A549细胞基质金属蛋白酶-2(matrix metalloproteinase-2,MMP-2)和血管内皮生长因子(vascular endothelial growth factor,VEGF)的表达水平. 方法:MTT法检测不同质量浓度的胡椒碱、顺铂及两者联合对A549细胞增殖的影响. 流式细胞术分析细胞凋亡. RT-PCR测定各组A549细胞MMP-2 mRNA和VEGF mRNA的表达变化. 结果:胡椒碱与顺铂均可抑制人肺腺癌A549细胞的生长,呈时间—浓度依赖性( P<0.05 );联合组随作用时间延长,对A549细胞的抑制效应增加( P<0.05 ). 联合组较对照组、胡椒碱组、顺铂组的A549细胞凋亡率显著增高( P<0.05 ). 联合组与对照组及单用胡椒碱、顺铂比较, MMP-2 mRNA和VEGF mRNA 的表达明显下调( P<0.05 ). 结论:胡椒碱显著提高顺铂对肺腺癌A549细胞的生长抑制和促进凋亡效应,降低MMP-2、VEGF的表达,从而可能抑制肺癌细胞的迁移.%Objective:To investigate the proliferation and apoptosis of piperine combined with cisplation on A 549 cells and the changes in the expression of MMP-2 and VEGF.Methods: MTT assay was used to detect the proliferation on A549 cells after treatment with series of monotherapy group ( piperine or cisplation alone ) or combination group ( piperine combined with cisplation ) .Flow cytometry was performed to assess the cellular apoptosis.The expression of MMP-2,VEGF mRNA were measured by RT-PCR in each group.Results: Both piperine and cisplation could inhibit the growth of A 549 cells in a time-dose dependent manner ( P<0.05 ) .With the increasing time , the cell proliferation was significantly inhibited in the combination group ( P <0.05 ) .The combination group also showed an obviously higher apoptosis rate than piperine alone , cisplation alone and control group, respectively( P<0.05).In addition, the expression of MMP-2 and VEGF mRNA in combination group were evidently lower than

  1. Experimental studies of the menispermum dauricum on anti-proliferation of human lung cancer cell line A549%蝙蝠葛活性成分对人肺癌A549细胞株抗增殖作用的实验研究

    Institute of Scientific and Technical Information of China (English)

    王永刚; 杨万山; 孙抒

    2010-01-01

    目的 探讨蝙蝠葛活性成分对人肺癌A549细胞株抗增殖作用及其机制.方法 应用MTT法测定蝙蝠葛活性成分对人肺癌A549细胞株的生长抑制作用;通过吖碇橙(AO)/溴化乙啶(EB)染色荧光显微镜观察肿瘤细胞的形态学变化;采用流式细胞仪检测A549细胞的周期分布相;应用免疫细胞化学技术SP法检测药物处理前后增殖细胞核抗原Ki-67、Bcl-2的表达.结果 蝙蝠葛活性成分对人肺癌A549细胞株有明显的抑制生长的作用,且呈现出浓度依赖性;蝙蝠葛活性成分可诱导A549细胞发生细胞周期阻滞;蝙蝠葛活性成分作用后Ki-67和Bcl-2阳性表达率较对照组降低(P<0.01).结论 蝙蝠葛活性成分在体外对人肺癌A549细胞株有显著的抑制增殖作用,可能与下调Ki-67、Bcl-2蛋白表达,细胞周期发生G0/G1期阻滞有关.

  2. CCL21/CCR7 up-regulate vascular endothelial growth factor-D expression via ERK pathway in human non-small cell lung cancer cells.

    Science.gov (United States)

    Sun, Limei; Zhang, Qingfu; Li, Yang; Tang, Na; Qiu, Xueshan

    2015-01-01

    Lymphangiogenesis has received considerable attention and become a new research hotspot of tumor metastasis. Recently, C-C chemokine receptor 7 (CCR7) is known to promote metastasis of non-small cell lung cancer (NSCLC) cells into lymph nodes. In this study, we investigated the relationship between CCL21/CCR7 and the lymphangiogenic factor vascular endothelial growth factor (VEGF)-D in human lung cancer cells and its impact on patients' prognosis. We found that CCL21/CCR7 increase the expression of VEGF-D in NSCLC Cell Lines through induced ERK1/2 and Akt phosphorylation. In addition, our study found that the expression levels of CCR7 and CCL21 were correlated with VEGF-D, lymphatic vessels density (LVD), clinical stages, lymph node metastasis, and patient Survival in 90 human non-small cell lung cancer (NSCLC) specimens. Taken together, our results provide evidence that CCL21/CCR7 induce VEGF-D up-regulation and promote lymphangiogenesis via ERK/Akt pathway in lung cancer.

  3. Multifunctional gold nanocomposites designed for targeted CT/MR/optical trimodal imaging of human non-small cell lung cancer cells

    Science.gov (United States)

    Chen, Jingwen; Sun, Yingqi; Chen, Qian; Wang, Le; Wang, Suhe; Tang, Yun; Shi, Xiangyang; Wang, Han

    2016-07-01

    Multifunctional gold nanocomposites, which were designed as dendrimer-entrapped gold nanoparticles functionalized with gadolinium, cyanine dye (Cy5.5), and folic acid, were synthesized to be used as the first dendrimer-based clinical nanoprobes for targeted X-ray computed tomography/magnetic resonance/optical trimodal imaging in vitro and in vivo of human non-small cell cancer cells.Multifunctional gold nanocomposites, which were designed as dendrimer-entrapped gold nanoparticles functionalized with gadolinium, cyanine dye (Cy5.5), and folic acid, were synthesized to be used as the first dendrimer-based clinical nanoprobes for targeted X-ray computed tomography/magnetic resonance/optical trimodal imaging in vitro and in vivo of human non-small cell cancer cells. Electronic supplementary information (ESI) available: Synthesis and characterization data of the nanoprobes; biocompatibility results; confirmation of the tumor cell uptake of the nanoprobes in vitro and in vivo; biodistribution results in vivo. See DOI: 10.1039/c6nr03143a

  4. Human RNA polymerase II associated factor 1 complex promotes tumorigenesis by activating c-MYC transcription in non-small cell lung cancer

    Energy Technology Data Exchange (ETDEWEB)

    Zhi, Xiuyi [Department of Thoracic Surgery, Xuanwu Hospital, Capital Medical University, Beijing, 100053 (China); Giroux-Leprieur, Etienne [ER2 GRC UPMC04 Theranoscan, Pierre et Marie Curie University, Tenon Hospital, 4 Rue de La Chine, 75020, Paris (France); Respiratory Diseases and Thoracic Oncology Department, Ambroise Pare Hospital – APHP, Versailles Saint Quentin en Yvelines University, 9 Avenue Charles de Gaulle, 92100, Boulogne-Billancourt (France); Wislez, Marie [ER2 GRC UPMC04 Theranoscan, Pierre et Marie Curie University, Tenon Hospital, 4 Rue de La Chine, 75020, Paris (France); Hu, Mu; Zhang, Yi [Department of Thoracic Surgery, Xuanwu Hospital, Capital Medical University, Beijing, 100053 (China); Shi, Huaiyin [Department of Pathology, Chinese PLA General Hospital, Fu-xing Road #28, Beijing, 100853 (China); Du, Kaiqi, E-mail: kaiqidu_zhejiang@163.com [Department of Cardiothoracic Surgery, Chinese People' s Armed Police Force, Zhejiang Corps Hospital, Jiaxing, Zhejiang Province (China); Wang, Lei, E-mail: leiwang_hebei@163.com [Department of Human Anatomy, Hebei Medical University, Hebei Province (China)

    2015-10-02

    Human RNA polymerase II (RNAPII)-associated factor 1 complex (hPAF1C) plays a crucial role in protein-coding gene transcription. Overexpression of hPAF1C has been implicated in the initiation and progression of various human cancers. However, the molecular pathways involved in tumorigenesis through hPAF1C remain to be elucidated. The current study suggested hPAF1C expression as a prognostic biomarker for early stage non-small cell lung cancer (NSCLC) and patients with low hPAF1C expression levels had significantly better overall survival. Furthermore, the expression of hPAF1C was found to be positively correlated with c-MYC expression in patient tumor samples and in cancer cell lines. Mechanistic studies indicated that hPAF1C could promote lung cancer cell proliferation through regulating c-MYC transcription. These results demonstrated the prognostic value of hPAF1C in early-stage NSCLC and the role of hPAF1C in the transcriptional regulation of c-MYC oncogene during NSCLC tumorigenesis. - Highlights: • hPAF1C expression is a prognostic biomarker for early stage non-small cell lung cancer. • The expression of hPAF1C was positively correlated with c-MYC in tumor samples of patients and in several NSCLC cell lines. • hPAF1C could promote lung cancer cell proliferation through regulating c-MYC transcription.

  5. The combined effects of physicochemical properties of size-fractionated ambient particulate matter on in vitro toxicity in human A549 lung epithelial cells

    Directory of Open Access Journals (Sweden)

    Umme S. Akhtar

    2014-01-01

    Full Text Available Epidemiological and toxicological studies have suggested that the health effects associated with exposure to particulate matter (PM are related to the different physicochemical properties of PM. These effects occur through the initiation of differential cellular responses including: the induction of antioxidant defenses, proinflammatory responses, and ultimately cell death. The main objective of this study was to investigate the effects of size-fractionated ambient PM on epithelial cells in relation to their physicochemical properties. Concentrated ambient PM was collected on filters for three size fractions: coarse (aerodynamic diameter [AD] 2.5–10 μm, fine (0.15–2.5 μm, and quasi-ultrafine (<0.2 μm, near a busy street in Toronto, Ontario, Canada. Filters were extracted and analyzed for chemical composition and redox activity. Chemical analyses showed that the coarse, fine, and quasi-ultrafine particles were comprised primarily of metals, water-soluble species, and organic compounds, respectively. The highest redox activity was observed for fine PM. After exposure of A549 cells to PM (10–100 μg/ml for 4 h, activation of antioxidant, proinflammatory and cytotoxic responses were assessed by determining the expression of heme oxygenase (HMOX-1, mRNA, interleukin-8 (IL-8, mRNA, and metabolic activity of the cells, respectively. All three size fractions induced mass-dependent antioxidant, proinflammatory, and cytotoxic responses to different degrees. Quasi-ultrafine PM caused significant induction of HMOX-1 at the lowest exposure dose. Correlation analyses with chemical components suggested that the biological responses correlated mainly with transition metals and organic compounds for coarse and fine PM and with organic compounds for quasi-ultrafine PM. Overall, the observed biological responses appeared to be related to the combined effects of size and chemical composition and thus both of these physicochemical properties should be

  6. Induction of human microsomal prostaglandin E synthase 1 by activated oncogene RhoA GTPase in A549 human epithelial cancer cells

    Energy Technology Data Exchange (ETDEWEB)

    Choi, Hye Jin [Laboratory of Systems Mucosal Biomodulation, Department of Microbiology and Immunology, Pusan National University School of Medicine, Yangsan (Korea, Republic of); Lee, Dong-Hyung [Department of Obstetrics and Gynecology, Medical Research Institute, Pusan National University, Busan (Korea, Republic of); Park, Seong-Hwan; Kim, Juil; Do, Kee Hun [Laboratory of Systems Mucosal Biomodulation, Department of Microbiology and Immunology, Pusan National University School of Medicine, Yangsan (Korea, Republic of); An, Tae Jin; Ahn, Young Sup; Park, Chung Berm [Department of Herbal Crop Research, NIHHS, RDA, Eumseong (Korea, Republic of); Moon, Yuseok, E-mail: moon@pnu.edu [Laboratory of Systems Mucosal Biomodulation, Department of Microbiology and Immunology, Pusan National University School of Medicine, Yangsan (Korea, Republic of); Medical Research Institute and Research Institute for Basic Sciences, Pusan National University, Busan (Korea, Republic of)

    2011-09-30

    Highlights: {yields} As a target of oncogene RhoA-linked signal, a prostaglandin metabolism is assessed. {yields} RhoA activation increases PGE{sub 2} levels and its metabolic enzyme mPGES-1. {yields} RhoA-activated NF-{kappa}B and EGR-1 are positively involved in mPGES-1 induction. -- Abstract: Oncogenic RhoA GTPase has been investigated as a mediator of pro-inflammatory responses and aggressive carcinogenesis. Among the various targets of RhoA-linked signals, pro-inflammatory prostaglandin E{sub 2} (PGE{sub 2}), a major prostaglandin metabolite, was assessed in epithelial cancer cells. RhoA activation increased PGE{sub 2} levels and gene expression of the rate-limiting PGE{sub 2} producing enzymes, cyclooxygenase-2 and microsomal prostaglandin E synthase 1 (mPGES-1). In particular, human mPGES-1 was induced by RhoA via transcriptional activation in control and interleukin (IL)-1{beta}-activated cancer cells. To address the involvement of potent signaling pathways in RhoA-activated mPGES-1 induction, various signaling inhibitors were screened for their effects on mPGES-1 promoter activity. RhoA activation enhanced basal and IL-1{beta}-mediated phosphorylated nuclear factor-{kappa}B and extracellular signal-regulated kinase1/2 proteins, all of which were positively involved in RhoA-induced gene expression of mPGES-1. As one potent down-stream transcription factor of ERK1/2 signals, early growth response gene 1 product also mediated RhoA-induced gene expression of mPGES-1 by enhancing transcriptional activity. Since oncogene-triggered PGE{sub 2} production is a critical modulator of epithelial tumor cells, RhoA-associated mPGES-1 represents a promising chemo-preventive or therapeutic target for epithelial inflammation and its associated cancers.

  7. Autophagy is induced by 3β-O-succinyl-lupeol (LD9-4) in A549 cells via up-regulation of Beclin 1 and down-regulation mTOR pathway.

    Science.gov (United States)

    Hao, Jing; Pei, Ying; Ji, Guiyuan; Li, Weijie; Feng, Shixiu; Qiu, Shengxiang

    2011-11-16

    The purpose of this study is to investigate the antitumor activity of a new derivative of lupeol-3β-O-succinyl-lupeol (LD9-4) and the molecular mechanism underlying cell death in human non-small cell lung cancer A549 cells. The results revealed that LD9-4 inhibited A549 cell proliferation in a time- and dose-dependent manner, with an IC(50) value of 5.78 ± 0.48 μM after cells exposed to LD9-4 for 72 h. Markers indicative of apoptosis (cell cycle arrest, phosphatidylserine externalization and Hoechst33258 staining) were uniformly negative in LD9-4 exposed cells. Interestingly, transmission electron microscope, MDC staining and LC3 level determination all confirmed that autophagy was induced in LD9-4 treated A549 cells. Furthermore, we found that LD9-4-induced autophagy in A549 cells was associated with the increase of intracellular reactive oxygen species and the decrease of phosphorylated mTOR and p70S6K levels. In the meanwhile, both mRNA and protein levels of Beclin 1 were up-regulated in a time-dependent manner. Our data suggest that autophagy is induced by LD9-4 in A549 cells, and the accumulating reactive oxygen species, up-regulation of Beclin 1 and inhibition of the mTOR signaling pathway are involved in this process.

  8. Synergistic effect of 8-Br-cAMP and cisplatin in human pulmonary adenocarcinoma cell A549%8-Br-cAMP联合顺铂作用于肺腺癌细胞A549的初步研究

    Institute of Scientific and Technical Information of China (English)

    陈曦; 刘志勇

    2010-01-01

    目的:观察8-溴-环腺苷酸(8-Br-cAMP)联合顺铂(DDP)对人肺腺癌细胞A549生长及其表皮生长因子受体(EGFR)表达的影响.方法:MTT法测定不同浓度8-Br-cAMP、DDP以及两药联用处理人肺腺癌细胞A549后细胞增殖活性的差异;应用RT-PCR技术检测EGFR mRNA表达水平的变化;Western blot技术检测EGFR蛋白表达水平的变化;利用流式细胞术测定细胞周期的改变.结果:8-Br-cAMP及DDP均能抑制人肺腺癌细胞A549的增殖,并呈浓度及时间依赖性;DDP联合8-Br-cAMP细胞抑制率显著高于相应浓度单药组(P<0.05),两者呈现出相加的抗瘤效果.8-Br-cAMP可下调人肺腺癌A549细胞EGFR mRNA和蛋白的表达.8-Br-cAMP将细胞周期阻滞在G2/M期(P<0.05).结论:8-Br-cAMP、DDP均可抑制A549细胞增殖.8-Br-cAMP可显著提高顺铂的细胞毒作用,增强化疗效果,并将细胞阻滞于G2/M期.8-Br-cAMP可以下调EGFR表达,可一定程度地逆转化癌细胞.

  9. Isothiocyanates induce oxidative stress and suppress the metastasis potential of human non-small cell lung cancer cells

    Directory of Open Access Journals (Sweden)

    Zhou Qinghua

    2010-06-01

    Full Text Available Abstract Background Isothiocyanates are natural compounds found in consumable cruciferous vegetables. They have been shown to inhibit chemical carcinogenesis by a wide variety of chemical carcinogens in animal models. Recent studies have also shown that isothiocyanates have antitumor activity, inhibiting the growth of several types of cultured human cancer cells. Our previous study showed that PEITC inhibited human leukemia cells growth by inducing apoptosis. However, the effect of isothiocyanates on lung cancer cell metastasis has not been studied. In the present study, we investigated the inhibitory effects of BITC and PEITC on metastatic potential of highly metastatic human lung cancer L9981 cells. Methods Cell migration and invasion were measured by wound healing assay and transwell chemotaxis assay. Expression of metastasis-related genes was assessed by quantitative RT-PCR and Western blotting. The mechanisms of action were evaluated by flow cytometry, reporter assay and Western blotting. Results Our data showed that both BITC and PEITC inhibited L9981 cell growth in a dose-dependent manner, the IC50 values were 5.0 and 9.7 μM, respectively. Cell migrations were reduced to 8.1% and 16.5% of control, respectively; and cell invasions were reduced to 2.7% and 7.3% of control, respectively. Metastasis-related genes MMP-2, Twist and β-catenin were also modulated. BITC and PEITC inhibited cell survival signaling molecules Akt and NFκB activation. Moreover, BITC and PEITC increased ROS generation and caused GSH depletion. Pretreatment with NAC blocked BITC and PEITC induced ROS elevation and NFκB inhibition. Conclusion Our results indicated that BITC and PEITC suppress lung cancer cell metastasis potential by modulation of metastasis-related gene expression, inhibition of Akt/NFκB pathway. Induction of oxidative stress may play an important role.

  10. Stable low-level expression of p21WAF1/CIP1 in A549 human bronchogenic carcinoma cell line-derived clones down-regulates E2F1 mRNA and restores cell proliferation control

    Directory of Open Access Journals (Sweden)

    Crawford Erin L

    2006-01-01

    Full Text Available Abstract Background Deregulated cell cycle progression and loss of proliferation control are key properties of malignant cells. In previous studies, an interactive transcript abundance index (ITAI comprising three cell cycle control genes, [MYC × E2F1]/p21 accurately distinguished normal from malignant bronchial epithelial cells (BEC, using a cut-off threshold of 7,000. This cut-off is represented by a line with a slope of 7,000 on a bivariate plot of p21 versus [MYC × E2F1], with malignant BEC above the line and normal BEC below the line. This study was an effort to better quantify, at the transcript abundance level, the difference between normal and malignant BEC. The hypothesis was tested that experimental elevation of p21 in a malignant BEC line would decrease the value of the [MYC × E2F1]/p21 ITAI to a level below this line, resulting in loss of immortality and limited cell population doubling capacity. In order to test the hypothesis, a p21 expression vector was transfected into the A549 human bronchogenic carcinoma cell line, which has low constitutive p21 TA expression relative to normal BEC. Results Following transfection of p21, four A549/p21 clones with stable two-fold up-regulated p21 expression were isolated and expanded. For each clone, the increase in p21 transcript abundance (TA was associated with increased total p21 protein level, more than 5-fold reduction in E2F1 TA, and 10-fold reduction in the [MYC × E2F1]/p21 ITAI to a value below the cut-off threshold. These changes in regulation of cell cycle control genes were associated with restoration of cell proliferation control. Specifically, each transfectant was capable of only 15 population doublings compared with unlimited population doublings for parental A549. This change was associated with an approximate 2-fold increase in population doubling time to 38.4 hours (from 22.3 hrs, resumption of contact-inhibition, and reduced dividing cell fraction as measured by flow

  11. Extracellular HSP70 Activates ERK1/2, NF-kB and Pro-Inflammatory Gene Transcription Through Binding with RAGE in A549 Human Lung Cancer Cells.

    Science.gov (United States)

    Somensi, Nauana; Brum, Pedro Ozorio; de Miranda Ramos, Vitor; Gasparotto, Juciano; Zanotto-Filho, Alfeu; Rostirolla, Diana Carolina; da Silva Morrone, Maurilio; Moreira, José Claudio Fonseca; Pens Gelain, Daniel

    2017-08-22

    Heat shock protein 70 (HSP70) has been recently described with extracellular actions, where it is actively released in inflammatory conditions. Acting as DAMPs (damage associated molecular pattern), extracellular HSP70 (eHSP70) interacts with membrane receptors and activates inflammatory pathways. At this context, the receptor for advanced glycation endproducts (RAGE) emerges as a possible candidate for interaction with eHSP70. RAGE is a pattern-recognition receptor and its expression is increased in several diseases related to a chronic pro-inflammatory state. One of the main consequences of RAGE ligand-binding is the ERK1/2 (extracellular signal-regulated kinases)-dependent activation of NF-kB (nuclear factor kappa B), which leads to expression of TNF-α (tumor necrosis factor alpha) and other cytokines. The purpose of this work is to elucidate if eHSP70 is able to evoke RAGE-dependent signaling using A549 human lung cancer cells, which constitutively express RAGE. Immunoprecipitation and protein proximity assay were utilized to demonstrate the linkage between RAGE and eHSP70. To investigate RAGE relevance on cell response to eHSP70, siRNA was used to knockdown the receptor expression. Signaling pathways activation were evaluated by western blotting, gene reporter luciferase and real time quantitative PCR. Protein eHSP70 shown to be interacting physically with the receptor RAGE in our cell model. Treatment with eHSP70 caused ERK1/2 activation and NF-κB transactivation impaired by RAGE knockdown. Moreover, the stimulation of pro-inflammatory cytokines expression by eHSP70 was inhibited in RAGE-silenced cells. Finally, conditioned medium of eHSP70-treated A549 cells caused differential effects in monocytes cytokine expression when A549 RAGE expression is inhibited. Our results evidence eHSP70 as a novel RAGE agonist capable of influence the cross-talk between cancer and immune system cells. © 2017 The Author(s). Published by S. Karger AG, Basel.

  12. Induction of IL-6 and CCL5 (RANTES in human respiratory epithelial (A549 cells by clinical isolates of respiratory syncytial virus is strain specific

    Directory of Open Access Journals (Sweden)

    Levitz Ruth

    2012-09-01

    Full Text Available Abstract Background Respiratory syncytial virus (RSV is the major respiratory pathogen of infants and young children. During each seasonal epidemic, multiple strains of both subgroup A and B viruses circulate in the community. Like other RNA viruses, RSV genome replication is prone to errors that results in a heterogeneous population of viral strains some of which may possess differences in virulence. We sought to determine whether clinical isolates of RSV differ in their capacity to induce inflammatory cytokines IL-6 and CCL5 (previously known as RANTES [regulated upon activation, normal T-cell expressed and secreted protein], which are known to be induced in vitro and in vivo in response to RSV, during infection of A549 cells. Results Screening of subgroup A and B isolates revealed heterogeneity among strains to induce IL-6 and CCL5. We chose two subgroup B strains, New Haven (NH1067 and NH1125, for further analysis because of their marked differences in cytokine inducing properties and because subgroup B strains, in general, are less genetically heterogeneous as compared to subgroup A strains. At 12 and 24 hours post infection RSV strains, NH1067 and NH1125 differed in their capacity to induce IL-6 by an order of magnitude or more. The concentrations of IL-6 and CCL5 were dependent on the dose of infectious virus and the concentration of these cytokines induced by NH1125 was greater than that of those induced by NH1067 when the multiplicity of infection of NH1067 used was as much as 10-fold higher than that of NH1125. The induction of IL-6 was dependent on viable virus as infection with UV-inactivated virus did not induce IL-6. The difference in IL-6 induction most likely could not be explained by differences in viral replication kinetics. The intracellular level of RSV RNA, as determined by quantitative RT-PCR, was indistinguishable between the 2 strains though the titer of progeny virus produced by NH1125 was greater than that produced by

  13. ETS2 mediated tumor suppressive function and MET oncogene inhibition in human non-small cell lung cancer

    Science.gov (United States)

    Kabbout, Mohamed; Garcia, Melinda M.; Fujimoto, Junya; Liu, Diane D.; Woods, Denise; Chow, Chi-Wan; Mendoza, Gabriela; Momin, Amin A.; James, Brian P.; Solis, Luisa; Behrens, Carmen; Lee, J. Jack

    2013-01-01

    PURPOSE The ETS2 transcription factor is an evolutionarily conserved gene that is deregulated in cancer. We analyzed the transcriptome of lung adenocarcinomas and normal lung tissue by expression profiling and found that ETS2 was significantly down-regulated in adenocarcinomas. In this study, we probed the yet unknown functional role of ETS2 in lung cancer pathogenesis. EXPERIMENTAL DESIGN Lung adenocarcinomas (n=80) and normal lung tissues (n=30) were profiled using the Affymetrix Human Gene 1.0 ST platform. Immunohistochemical (IHC) analysis was performed to determine ETS2 protein expression in NSCLC histological tissue specimens (n=201). Patient clinical outcome, based on ETS2 IHC expression, was statistically assessed using the log-rank and Kaplan-Meier tests. RNA interference and over-expression strategies were employed to assess effects of ETS2 expression on the transcriptome and on various malignant phenotypes. RESULTS ETS2 expression was significantly reduced in lung adenocarcinomas compared to normal lung (precurrence in NSCLC (p=0.009, HR=1.89) and adenocarcinoma (p=0.03, HR=1.86). Moreover, ETS2 was found to significantly inhibit lung cancer cell growth, migration and invasion (p<0.05), and microarray and pathways analysis revealed significant (p<0.001) activation of the HGF pathway following ETS2 knockdown. In addition, ETS2 was found to suppress MET phosphorylation and knockdown of MET expression significantly attenuated (p<0.05) cell invasion mediated by ETS2-specific siRNA. Furthermore, knockdown of ETS2 augmented HGF-induced MET phosphorylation, cell migration and invasion. CONCLUSION(S) Our findings point to a tumor suppressor role for ETS2 in human NSCLC pathogenesis through inhibition of the MET proto-oncogene. PMID:23659968

  14. Yes-associated protein regulates the growth of human non-small cell lung cancer in response to matrix stiffness.

    Science.gov (United States)

    Yuan, Yonggang; Zhong, Weiliang; Ma, Ge; Zhang, Baoxiang; Tian, Hui

    2015-06-01

    The Yes‑associated protein (YAP) transcriptional coactivator is recognized as a crucial regulator of human cancer. However, its involvement in human non‑small cell lung cancer (NSCLC) in response to physical cues remains unclear. In this study, substrates with different rigidity were generated in order to evaluate the role of YAP, and its upstream regulators in the Hippo pathway, in the regulation of growth of an NSCLC cell line within particular environments. It was shown that the expression of the YAP protein in SPCA-1 NSCLC cells was significantly increased when cultured on a stiff substrate compared to a soft substrate. However, the expression of phospho‑YAP protein and large tumor suppressor kinase 1 (LATS1) were markedly decreased after culturing on the stiff substrate. Phosphorylation of YAP by LATS1 leads to cytoplasmic retention of YAP, which inhibits its function as a nuclear transcription coactivator. The study also found that the stiff substrate promoted the growth of NSCLC cells in vitro, and an increase in the transcription levels of Survivin, connective tissue growth factor, amphiregulin and Ki67, as well as a decrease in the expression level of YAP in the cytoplasm, and adecrease in p-YAP. In conclusion, the findings showed that the stiffness of the subcellular matrix altered the behavior of NSCLC cells, and that YAP regulated the growth of NSCLC cells in response to matrix stiffness, thereby suggesting a role for the Hippo‑YAP pathway in the response of NSCLC cell growth to specific microenvironments.

  15. Effects of tangeretin on growth and invasion of human non-small-cell lung cancer cells%桔皮素对人非小细胞肺癌细胞生长及侵袭的影响

    Institute of Scientific and Technical Information of China (English)

    石歆

    2015-01-01

    目的:探讨桔皮素( TGN)对非小细胞肺癌( NSCLC)细胞生长和侵袭的影响及其分子机制。方法:体外培养非小细胞肺癌A549细胞,分别用不同浓度的TGN处理,MTT比色法检测细胞活性,Annexin V-FITC/PI染色及流式细胞术检测细胞凋亡率, Transwell检测细胞侵袭, RT-PCR分析MMP-2和MMP-9的mRNA表达水平, Western blotting检测Ki67、Cyt C、caspase-3、cleaved caspase-3、MMP-2、MMP-9、Akt、p-Akt以及p-PI3K表达水平。结果:桔皮素剂量依赖性地抑制A549细胞增殖( P<0.05),同时伴随有增殖标记分子Ki67表达水平的下调。分析发现,桔皮素诱导细胞中Cyt C、caspase-3和cleaved caspase-3的表达上调( P<0.01),加速A549细胞的凋亡。此外,桔皮素作用后,A549细胞中侵袭相关蛋白MMP-2和MMP-9的表达量下降,且侵袭数目随桔皮素浓度增加而减少。进一步研究表明,桔皮素作用后A549细胞中p-Akt和p-PI3K表达水平降低(P<0.05),阻断PI3K/Akt信号通路后,不同浓度TGN对细胞活性影响没有变化。结论:桔皮素能抑制A549细胞生长及侵袭,促进细胞凋亡,可能通过抑制PI3 K/Akt信号通路的激活起作用。因此,本研究将为非小细胞肺癌的防治提供新的研究方向。%[ ABSTRACT] AIM: To study the influences of tangeretin ( TGN) on the growth and invasion of non-small-cell lung cancer ( NSCLC) cells, and to explore the molecular mechanisms.METHODS: The A549 cells were treated with different concentrations of TGN in vitro.The relative cell activity was determined by MTT assay.The apoptotic rate was an-alyzed by flow cytometry with Annexin V-FITC/PI staining.The number of the invasive cells was measured by Transwell as-say.The mRNA expression of MMP-2 and MMP-9 was detected by RT-PCR, and the protein levels of Ki67, Cyt C, caspase-3, cleaved caspase-3, MMP-2, MMP-9, Akt, p-Akt and p-PI3K were determined by Western

  16. 斑蝥素通过MAPK途径对肺癌A549细胞周期阻滞及其分子机制的研究%Role and molecular mechanism of cantharidin in cell cycle arrest of A549 human lung carcinoma cells through mitogen-activated protein kinase pathway

    Institute of Scientific and Technical Information of China (English)

    张卫东; 赵惠儒; 于秉治; 王晓华; 宗志红; 刘莹

    2006-01-01

    目的:探讨斑蝥素对人肺癌A549细胞的周期阻滞作用及其分子机制.方法:采用MTT法检测斑蝥素对A549细胞的增殖抑制作用;流式细胞仪检测细胞周期;以蛋白印迹法分析斑蝥素作用后细胞周期蛋白B1(cyclinB1)、p34cdc2、phos-p34cdc2、p21、survivin、ERK1/ERK2、phos-ERK1/phos-ERK2等蛋白表达或活性的改变.结果:斑蝥素能抑制A549细胞的增殖;斑蝥素作用于A549细胞后引起G2/M期阻滞;斑蝥素处理后,p34cdc2、phos-p34cdc2表达量没有改变,cyclinB1、survivin蛋白表达量减少,p21蛋白表达量增加.ERK1/ERK2、phos-ERK1/phos-ERK2表达量降低.结论:斑蝥素通过调节cyclinB1、p21、survivin、ERK1/ERK2、phos-ERK1/phos-ERK2等蛋白表达或活性的改变引起G2/M期阻滞.

  17. The effect of ataxia-telangiectasia mutated kinase-dependent hyperphosphorylation of checkpoint kinase-2 on oligodeoxynucleotide 7909 containing CpG motifs-enhanced sensitivity to X-rays in human lung adenocarcinoma A549 cells

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    Liu XQ

    2015-06-01

    Full Text Available Xiaoqun Liu,1,* Xiangdong Liu,2,* Tiankui Qiao,1 Wei Chen,1 Sujuan Yuan1 1Department of Oncology, 2Department of Ophthalmology, Affiliated Jinshan Hospital, Fudan University, Shanghai, People’s Republic of China *These authors contributed equally to this work Objective: The aim of the study reported here was to further investigate the potential effect of ataxia-telangiectasia mutated (ATM kinase-dependent hyperphosphorylation of checkpoint kinase-2 (Chk2 on radiosensitivity enhanced by oligodeoxynucleotide 7909 containing CpG motifs (CpG ODN7909 in human lung adenocarcinoma A549 cells. Methods: In vitro A549 cells were randomly separated into control, CpG, X-ray, CpG+X-ray, ATM kinase-small interfering RNA (siRNA+CpG+X-ray (ATM-siRNA, and Chk2-siRNA+CpG+X-ray (Chk2-siRNA groups. siRNAs were adopted to silence the ATM and Chk2 genes. Expression and phosphorylation of ATM kinase and Chk2 were detected by Western blot assay. Cell colonies were observed under inverted phase-contrast microscopy. Cellular survival curves were fitted using a multi-target single-hitting model. Cell cycle and apoptosis were analyzed by flow cytometry. Results: Expression of ATM kinase and Chk2 was similar among the control, CpG, X-ray, and CpG+X-ray groups. Phosphorylated ATM kinase and Chk2 were significantly increased in the CpG+X-ray group compared with in the X-ray group (t=6.00, P<0.01 and t=3.13, P<0.05, respectively, though these were hardly detected in the control and CpG groups. However, expression of ATM kinase and Chk2 was clearly downregulated in the ATM-siRNA and Chk2-siRNA groups, respectively. Similarly, their phosphorylation levels were also significantly decreased in the ATM-siRNA group (t=14.35, P<0.01 and t=8.46, P<0.01, respectively and a significant decrease in phosphorylated Chk2 was observed in the Chk2-siRNA group (t=7.28, P<0.01 when compared with the CpG+X-ray group. Further, the number of A549 cells at Gap 2/mitotic phase and the apoptosis

  18. Thromboxane A2 receptor-mediated release of matrix metalloproteinase-1 (MMP-1) induces expression of monocyte chemoattractant protein-1 (MCP-1) by activation of protease-activated receptor 2 (PAR2) in A549 human lung adenocarcinoma cells.

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    Li, Xiuling; Tai, Hsin-Hsiung

    2014-08-01

    Matrix metalloproteinases (MMPs) and monocyte chemoattractant protein-1 (MCP-1, CCL2) are known to be upregulated in many tumors. Their roles in tumor invasion and metastasis are being uncovered. How they are related to each other and involved in tumor progression remains to be determined. Earlier it was reported that I-BOP-initiated activation of thromboxane A2 receptor (TP) induced the release of MMP-1, MMP-3, and MMP-9 from lung cancer A549 cells overexpressing TPα (A549-TPα). Herein it was found that MMP-1, but not MMP-3 or MMP-9, induced the expression of MCP-1 in A549 cells. Conditioned medium (CM) from I-BOP activated, MMP-1 siRNA pretreated A549-TPα cells induced greatly attenuated expression of MCP-1 in A549 cells indicating that MMP-1 in the CM contributed significantly to the expression of MCP-1. MMP-1 was shown to activate protease-activated receptor 2 (PAR2) instead of commonly assumed PAR1 to increase the expression of MCP-1 in A549 cells. This conclusion was reached from the following findings: (1) expression of MCP-1 induced by trypsin, a PAR2 agonist, and also PAR2 agonist peptide, was inhibited by a PAR2 antagonist; (2) expression of MCP-1 induced by MMP-1 and by CM from I-BOP activated A549-TPα cells was blocked by a PAR2 antagonist but not by other PAR antagonists; (3) expression of MCP-1 induced by MMP-1 and by CM from I-BOP activated A549-TPα cells was attenuated significantly by pretreatment of cells with PAR2-siRNA. These results suggest that PAR2 is a novel MMP-1 target mediating MMP-1-induced signals in A549 lung cancer cells.

  19. Artesunate induces AIF-dependent apoptosis in A549 cells

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    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.

  20. Determination of in vitro free radical scavenging and antiproliferative effect of Pennisetum alopecuroides on cultured A549 human lung cancer cells

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    Githa Elizabeth Mathew

    2015-01-01

    Conclusions: This is the 1 st time a pharmacological exploration of P. alopecuroides grasses has been conducted. We have shown that P. alopecuroides exhibits good free radical scavenging and strong in vitro cytotoxic activities against human lung cancer cell lines.

  1. Inhibition of TRPC6 reduces non-small cell lung cancer cell proliferation and invasion

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    Lu, Xiao-Yu; Yan, Yan; Zhai, Yu-Jia; Bao, Qing; Doetsch, Paul W.; Deng, Xingming; Thai, Tiffany L.; Alli, Abdel A.; Eaton, Douglas C.; Shen, Bao-Zhong; Ma, He-Ping

    2017-01-01

    Recent studies indicate that the transient receptor potential canonical 6 (TRPC6) channel is highly expressed in several types of cancer cells. However, it remains unclear whether TRPC6 contributes to the malignancy of human non-small cell lung cancer (NSCLC). We used a human NSCLC A549 cell line as a model and found that pharmacological blockade or molecular knockdown of TRPC6 channel inhibited A549 cell proliferation by arresting cell cycle at the S-G2M phase and caused a significant portion of cells detached and rounded-up, but did not induce any types of cell death. Western blot and cell cycle analysis show that the detached round cells at the S-G2M phase expressed more TRPC6 than the still attached polygon cells at the G1 phase. Patch-clamp data also show that TRPC whole-cell currents in the detached cells were significantly higher than in the still attached cells. Inhibition of Ca2+-permeable TRPC6 channels significantly reduced intracellular Ca2+ in A549 cells. Interestingly, either blockade or knockdown of TRPC6 strongly reduced the invasion of this NSCLC cell line and decreased the expression of an adherent protein, fibronectin, and a tight junction protein, zonula occluden protein-1 (ZO-1). These data suggest that TRPC6-mediated elevation of intracellular Ca2+ stimulates NSCLC cell proliferation by promoting cell cycle progression and that inhibition of TRPC6 attenuates cell proliferation and invasion. Therefore, further in vivo studies may lead to a consideration of using a specific TRPC6 blocker as a complement to treat NSCLC. PMID:28030826

  2. Human RNA polymerase II associated factor 1 complex promotes tumorigenesis by activating c-MYC transcription in non-small cell lung cancer.

    Science.gov (United States)

    Zhi, Xiuyi; Giroux-Leprieur, Etienne; Wislez, Marie; Hu, Mu; Zhang, Yi; Shi, Huaiyin; Du, Kaiqi; Wang, Lei

    2015-10-02

    Human RNA polymerase II (RNAPII)-associated factor 1 complex (hPAF1C) plays a crucial role in protein-coding gene transcription. Overexpression of hPAF1C has been implicated in the initiation and progression of various human cancers. However, the molecular pathways involved in tumorigenesis through hPAF1C remain to be elucidated. The current study suggested hPAF1C expression as a prognostic biomarker for early stage non-small cell lung cancer (NSCLC) and patients with low hPAF1C expression levels had significantly better overall survival. Furthermore, the expression of hPAF1C was found to be positively correlated with c-MYC expression in patient tumor samples and in cancer cell lines. Mechanistic studies indicated that hPAF1C could promote lung cancer cell proliferation through regulating c-MYC transcription. These results demonstrated the prognostic value of hPAF1C in early-stage NSCLC and the role of hPAF1C in the transcriptional regulation of c-MYC oncogene during NSCLC tumorigenesis. Copyright © 2015. Published by Elsevier Inc.

  3. Expression of telomerase hTERT in human non-small cell lung cancer and its correlation with c-myc gene

    Institute of Scientific and Technical Information of China (English)

    耿志华; 张敦华; 刘银坤

    2003-01-01

    Objective To investigate the expression of human telomerase catalytic subunit, hTERT, in human non-small cell lung cancer (NSCLC) and its correlations to c-myc gene.Methods hTERT and c-myc mRNA expressions were detected by semiquantitative reverse transcription-polymerase chain reaction (RT-PCR). Statistical correlation analysis was made to estimate whether there was interrelation between them.Results Positive rate of hTERT expression in 51 surgically resected lung cancer specimens was 86.3%, significantly higher than that in adjacent non-neoplastic lung tissues and benign lesions, which were 14.3% and 27.3% respectively. No statistical significance was observed between the frequency of hTERT expression and histologic types, degree of differentiation, TNM stages, tumor size or lymph nodes metastases. Correlation analysis revealed that the expression of c-myc gene was significantly related to that of hTERT (correlation coefficient, r=0.633, P<0.001).Conclusions hTERT may be a useful tumor marker in diagnosing lung cancer. Significant correlation between the expression of hTERT and c-myc mRNA indicates that the activation and up-regulation of hTERT might be conferred by over-expression of c-myc gene.

  4. Differential expression of the RANKL/RANK/OPG system is associated with bone metastasis in human non-small cell lung cancer.

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    Xianbo Peng

    Full Text Available BACKGROUND: Human non-small cell lung cancer (NSCLC patients exhibit a high propensity to develop skeletal metastasis, resulting in excessive osteolytic activity. The RANKL/RANK/OPG system, which plays a pivotal role in bone remodeling by regulating osteoclast formation and activity, is of potential interest in this context. MATERIALS AND METHODS: Reverse transcriptase polymerase chain reaction, western blotting, and immunohistochemical analysis were used to examine the expression of RANKL, RANK, and OPG in human NSCLC cell lines with different metastatic potentials, as well as in 52 primary NSCLC samples and 75 NSCLC bone metastasis samples. In primary NSCLC patients, the expression of these proteins was correlated with clinicopathological parameters. Recombinant human RANKL and transfected RANKL cDNA were added to the PAa cell line to evaluate the promoter action of RANKL during the process of metastasis in vitro and in vivo. RESULTS: Up-regulated RANKL, RANK, and OPG expression and increased RANKL:OPG ratio were detected in NSCLC cell lines and in tumor tissues with bone metastasis, and were correlated with higher metastatic potential. The metastatic potential of NSCLC in vitro and in vivo, including migration and invasion ability, was significantly enhanced by recombinant human RANKL and the transfection of RANKL cDNA, and was impaired after OPG was added. The increased expression of RANKL and OPG correlated with tumor stage, lymph node metastasis, and distant metastasis. CONCLUSIONS: Differential expression of RANKL, RANK, and OPG is associated with the metastatic potential of human NSCLC to skeleton, raising the possibility that the RANKL/RANK/OPG system could be a therapeutic target for the treatment of metastatic NSCLC patients.

  5. 吉西他滨对人肺癌A549细胞株CN-Ⅱ,APE/Ref-1mRNA和蛋白表达的影响%Effects of Gemcitabine on Expression of CN-Ⅱ,APE/Ref-1 mRNA and Protein in Human Lung Cancer Cell Line A549

    Institute of Scientific and Technical Information of China (English)

    宋东; 周曙光; 刘玥; 李晓栋; 王姗姗; 唐小龙

    2009-01-01

    [目的]研究人肺癌A549细胞株在吉西他滨化疗时CN-Ⅱ,APE/Ref-1基因表达的变化,并探讨其在肺癌化疗耐药中所起的作用.[方法]不同浓度吉西他滨0、10、20、40及60 μmol/L作用人肺癌A549细胞株24 h,分别以RT-PCR 及Western blot方法测定用药后CN-Ⅱ和APE/Ref-1的mRNA及蛋白表达情况. [结果]吉西他滨作用人肺癌A549细胞株24 h后,CN-Ⅱ和APE/Ref-1的mRNA及蛋白表达水平均明显上升,并与吉西他滨的浓度呈正相关(CN-Ⅱ RT-PCR:r=0.687,P=0.009;Western blot:r=0.594,P=0.021;APE/Ref-1 RT-PCR:r=0.669,P=0.010;Western blot:r=0.562,P=0.029). [结论]CN-Ⅱ和APE/Ref-1在肺癌化疗时表达明显增强,可能与化疗耐药性的产生有关,并提示针对CN-Ⅱ和APE/Ref-1的靶向干预可能有助于提高肺癌的化疗敏感性.

  6. Proteomic investigation in A549 lung cell line stably infected by HPV16E6/E7 oncogenes.

    Science.gov (United States)

    Ciotti, Marco; Marzano, Valeria; Giuliani, Laura; Nuccetelli, Marzia; D'Aguanno, Simona; Azzimonti, Barbara; Bernardini, Sergio; Perno, Carlo Federico; Urbani, Andrea; Favalli, Cartesio; Federici, Giorgio

    2009-01-01

    Data have accumulated implicating the involvement of oncogenic human papillomaviruses (HPVs) in bronchial carcinogenesis. We recently described the presence of oncogenic HPV transcripts in non-small cell lung cancers. To investigate the role of oncogenic HPVs in lung carcinogenesis. The lung cell line A549 stably infected with HPV16E6, HPV16E7 and HPVE6/E7 constructs was used to investigate the protein profile changes associated with the expression of these oncogenes. Replicated two-dimensional gel electrophoresis gels from uninfected and stably HPV16E6-, E7-, and E6/E7-infected A549 cells were compared for changes in protein profile. Protein identification was achieved by peptide mass fingerprinting by MALDI-TOF-MS and nLC-ESI-Q-TOF-MS/MS peptide ladder sequencing. We identified 17 different polypeptides whose average normalized spot intensity was statistically significant (p < 0.05) and differed by 2-fold. Relationships between differentially expressed proteins and the HPV-induced infection mechanism have been clustered by knowledge-base database functional association network analysis. The impact of Hsp27, annexin III, annexin IV, Gp96 and TPT1 on the cellular response mechanism to HPV infection is presented and discussed. Copyright 2008 S. Karger AG, Basel.

  7. Down-regulation of the DNA-repair endonuclease 8-oxo-guanine DNA glycosylase 1 (hOGG1) by sodium dichromate in cultured human A549 lung carcinoma cells.

    Science.gov (United States)

    Hodges, N J; Chipman, J K

    2002-01-01

    Hexavalent chromium is a genotoxic human pulmonary carcinogen that elevates DNA oxidation, apparently through the generation of reactive DNA-damaging intermediates including Cr(V), Cr(IV) and reactive oxygen species. We tested the hypothesis that elevation of DNA oxidation may also be through inhibition of the expression of the repair glycosylase for 8-oxo deoxyguanine (hOGG1) in cultured A549 human lung epithelial cells. Treatment with sodium dichromate (0-100 microM, 16 h) resulted in a concentration-dependent decrease in the levels of OGG1 mRNA as measured by both RT-PCR and RNase protection assay. Sodium dichromate at 25 microM and above gave a marked reduction of OGG1 mRNA expression which was not seen at 1 microM and below. No effect on the expression of the apurinic endonuclease hAPE or the house-keeping gene GAPDH was observed at any of the concentrations of sodium dichromate investigated. Treatment of cells with the pro-oxidant H(2)O(2) (0-200 microM) for 16 h had no detectable effect on the levels of OGG1 mRNA or protein expression suggesting that the effect of sodium dichromate is not mediated by H(2)O(2). Western blotting demonstrated that sodium dichromate (100 microM; 16 h and >25 microM; 28 h) markedly reduced levels of OGG1 protein in nuclear cell extracts. Additionally, treatment of cells with sodium dichromate (>25 microM, 28 h) resulted in a concentration-dependent decrease in the ability of nuclear extracts to nick a synthetic oligonucleotide containing 8-oxo deoxyguanine (8-oxo dG). We conclude that the elevation of 8-oxo dG levels observed in A549 cells treated with sodium dichromate may be, at least in part, due to a reduced capacity to repair endogenous and hexavalent chromium-induced 8-oxo dG.

  8. Irradiation and various cytotoxic drugs enhance tyrosine phosphorylation and {beta}{sub 1}-integrin clustering in human A549 lung cancer cells in a substratum-dependent manner in vitro

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    Cordes, N.; Beinke, C.; Beuningen, D. van [Inst. of Radiobiology, German Armed Forces, Munich (Germany); Plasswilm, L. [Dept. of Radiation Oncology, Univ. Hospital Basel (Swaziland)

    2004-03-01

    Background and purpose: interactions of cells with a substratum, especially extracellular matrix proteins, initiate clustering of integrin receptors in the cell membrane. This process represents the initial step for the activation of signaling pathways regulating survival, proliferation, differentiation, adhesion, and migration, and could, furthermore, be important for cellular resistance-mediating mechanisms against radiation or cytotoxic drugs. The lack of data elucidating the impact of irradiation or cytotoxic drugs on this important phenomenon led to this study on human A549 lung cancer cells in vitro. Material and methods: the human lung carcinoma cell line A549 grown on polystyrene or fibronectin (FN) was irradiated with 0-8 Gy or treated with cisplatin (0.1-50 {mu}M), paclitaxel (0.1-50 nM), or mitomycin (0.1-50 {mu}M). Colony formation assays, immunofluorescence staining in combination with activation of integrin clustering using anti-{beta}{sub 1}-integrin antibodies (K20), and Western blotting for tyrosine phosphorylation under treatment of cells with the IC{sub 50} for irradiation (2 Gy; IC{sub 50} = 2.2 Gy), cisplatin (2 {mu}M), paclitaxel (5 nM), or mitomycin (7 {mu}M) were performed. Results: attachment of cells to FN resulted in a significantly reduced radio- and chemosensitivity compared to polystyrene. The clustering of {beta}{sub 1}-integrins examined by immunofluorescence staining was only stimulated by irradiation, cisplatin, paclitaxel, or mitomycin in case of cell attachment to FN. By contrast, tyrosine phosphorylation, as one of the major events following {beta}{sub 1}-integrin clustering, showed a 3.7-fold, FN-related enhancement, and treatment of cells with the IC{sub 50} of radiation, cisplatin, paclitaxel, or mitomycin showed a substratum-dependent induction. Conclusion: for the first time, a strong influence of irradiation and a variety of cytotoxic drugs on the clustering of {beta}{sub 1}-integrins could be shown. This event is a

  9. HGF induces EMT in non-small-cell lung cancer through the hBVR pathway.

    Science.gov (United States)

    Liu, Fang; Song, Shasha; Yi, Zhi; Zhang, Min; Li, Jiali; Yang, Fang; Yin, Hongtao; Yu, Xiufeng; Guan, Chao; Liu, Ying; Liu, Zizhen; Wang, Jing; Zhu, Daling

    2017-09-15

    The epithelial-to-mesenchymal transition (EMT) is a crucial event during non-small-cell lung cancer (NSCLC) invasion and metastasis. However, the mechanisms involved in NSCLC EMT have not been fully clarified. Hepatocyte growth factor (HGF) and human biliverdin reductase (hBVR) are reported to contribute to EMT in several diseases. Here, we show that compared with transforming growth factor beta (TGF-β), fibroblast growth factor (FGF), and epidermal growth factor (EGF), HGF is an important cell factor for EMT in NSCLC cell lines A549 and H460. Met protein, HGF receptors, and hBVR were found to be highly expressed and positively correlated with EMT in NSCLC tissue sections. In addition, HGF and hBVR induced a decrease in epithelial protein marker expression and an increase in mesenchymal protein marker expression as well as increased cellular migration and invasion, indicating that both HGF and hBVR mediate EMT in A549 and H460 cell lines. Furthermore, HGF-induced EMT and migration and invasion in both cell lines was inhibited by si-hBVR. Taken together, our data show that HGF induces EMT in NSCLC through the hBVR pathway. Copyright © 2017. Published by Elsevier B.V.

  10. Imaging of non-small cell lung cancers with a monoclonal antibody, KC-4G3, which recognizes a human milk fat globule antigen

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    Dienhart, D.G.; Schmelter, R.F.; Lear, J.L.; Miller, G.J.; Glenn, S.D.; Bloedow, D.C.; Kasliwal, R.; Moran, P.; Seligman, P.; Murphy, J.R. (Univ. of Colorado Cancer Center, Denver (USA))

    1990-11-01

    To determine the role of lung cancer tumor imaging with monoclonal antibodies directed against high molecular weight human milk fat globule antigens, we administered i.v. 111In-KC-4G3 to 24 patients with advanced non-small cell lung cancer. One mg of 111In-KC-4G3 was mixed with 0, 9, 49, 99, or 499 mg of unlabeled KC-4G3 and infused i.v. over 1 to 5 h. The mean 111In-KC-4G3 radiochemical purity was greater than 97% and the resultant immunoreactivity averaged 62%. Successful imaging of cancer sites was accomplished in 92% of 24 patients, and 57% of 91 total lesions were visualized. Successful localization of tumor sites related to size (P less than 0.001), with 81% of lesions greater than 3.0 cm in diameter, 50% of lesions 1.5 to 3 cm, and 6% of lesions less than 1.5 cm successfully imaging, and to location (P less than 0.05), with 69% of pulmonary lesions, 80% of soft tissue lesions, and only 32% of bone metastases being visualized. Nonspecific reticulo-endothelial uptake of radioactivity was a major problem. Approximately 35% of 111In was chelated to serum transferrin by 24 and 48 h after infusion. The mean t 1/2 beta for plasma radioisotope and immunoreactive KC-4G3 was 29 and 27 h, respectively. There was no correlation between total infused antibody dose and imaging success or between total dose and effect on 111In and KC-4G3 kinetics. Circulating free KC-4 antigen was measurable in all but one patient before study. Tumor biopsy following infusion could demonstrate antibody presence but not saturable antigen binding. We conclude that (a) 111In-KC-4G3 demonstrates successful tumor localization in non-small cell lung cancers bearing generally high expression of its antigen and (b) further investigations to diminish nonspecific radioactivity for imaging and utilization of high dose radiolabeled antibody for therapeutic intent are warranted.

  11. Usp9x- and Noxa-mediated Mcl-1 downregulation contributes to pemetrexed-induced apoptosis in human non-small-cell lung cancer cells.

    Science.gov (United States)

    Yan, J; Zhong, N; Liu, G; Chen, K; Liu, X; Su, L; Singhal, S

    2014-07-03

    Pemetrexed, a folate antimetabolite, combined with cisplatin is used as a first-line therapy for malignant pleural mesothelioma (MPM) and locally advanced or metastatic non-small-cell lung cancer (NSCLC). Pemetrexed arrests cell cycle by inhibiting three enzymes in purine and pyrimidine synthesis that are necessary for DNA synthesis. Pemetrexed also promotes apoptosis in target cells, but little is known about its mechanism in cancer cells. We have previously shown that pemetrexed can result in endoplasmic reticulum (ER) stress, and it can lead to downstream apoptosis. In this study, we further elucidate this mechanism. Our data show that pemetrexed increases Noxa expression through activating transcription factor 4 (ATF4) and activating transcription factor 3 (ATF3) upregulation. Furthermore, pemetrexed induces apoptosis by activating the Noxa-Usp9x-Mcl-1 pathway. Inhibition of Noxa by small interfering RNA (siRNA) promotes Usp9x (ubiquitin-specific peptidase 9, X-linked) expression. Moreover, downregulation of the deubiquitinase Usp9x by pemetrexed results in downstream reduction of myeloid cell leukemia 1 (Mcl-1) expression. Mechanistically, Noxa upregulation likely reduces the availability of Usp9x to Mcl-1, thereby promoting its ubiquitination and degradation, leading to the apoptosis of neoplastic cells. Thus, our findings demonstrate that Noxa-Usp9x-Mcl-1 axis may contribute to pemetrexed-induced apoptosis in human lung cancer cells.

  12. Fisetin induces apoptosis and endoplasmic reticulum stress in human non-small cell lung cancer through inhibition of the MAPK signaling pathway.

    Science.gov (United States)

    Kang, Kyoung Ah; Piao, Mei Jing; Madduma Hewage, Susara Ruwan Kumara; Ryu, Yea Seong; Oh, Min Chang; Kwon, Taeg Kyu; Chae, Sungwook; Hyun, Jin Won

    2016-07-01

    Fisetin (3,3',4',7-tetrahydroxyflavone), a dietary flavonoid compound, is currently being investigated for its anticancer effect in various cancer models, including lung cancer. Recent studies show that fisetin induces cell growth inhibition and apoptosis in the human non-small cell lung cancer line NCI-H460. In this study, we investigated whether fisetin can induce endoplasmic reticulum (ER) stress-mediated apoptosis in NCI-H460 cells. Fisetin induced mitochondrial reactive oxygen species (ROS) and characteristic signs of ER stress: ER staining; mitochondrial Ca(2+) overload; expression of ER stress-related proteins; glucose-regulated protein (GRP)-78, phosphorylation of protein kinase RNA (PKR)-like endoplasmic reticulum kinase (PERK) and phosphorylation of eukaryotic initiation factor-2 α subunit; cleavage of activating transcription factor-6; phosphorylation of inositol-requiring kinase-1 and splicing of X-box transcription factor-1; induction of C/EBP homologous protein and cleaved caspase-12. siRNA-mediated knockdown of CHOP and ATF-6 attenuated fisetin-induced apoptotic cell death. In addition, fisetin induced phosphorylation of ERK, JNK, and p38 MAPK. Moreover, silencing of the MAPK signaling pathway prevented apoptotic cell death. In summary, our results indicate that, in NCI-H460 cells, fisetin induces apoptosis and ER stress that is mediated by induction of the MAPK signaling pathway.

  13. Synergistic cytotoxicity of ampelopsin sodium and carboplatin in human non-small cell lung cancer cell line SPC-A1 by G1 cell cycle arrested.

    Science.gov (United States)

    Lu, Li; Yang, Li-Ning; Wang, Xue-Xi; Song, Chun-Li; Qin, Hong; Wu, Yong-Jie

    2017-02-01

    To evaluate the cytotoxic effects of ampelopsin sodium (Amp-Na) and carboplatin (CBP) used alone or in combination on human non-small cell lung cancer (NSCLC) cells SPC-A1 in vitro and its related mechanism. Cytotoxic effects were assessed by 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays. The synergistic effects of the drugs were calculated with coefficient of drug interaction (CDI). Cell cycle was determined by flow cytometry (FCM). The levels of p53, p21, cyclinE, cyclinD1, and phosphorylated cyclin-dependent kinase-2 (p-CDK2) were evaluated by Western blot. Amp-Na (6.25-200 μg/mL) and CBP (3.13-100 μg/mL) alone exhibited prominent cytotoxic activity in a concentration-dependent manner on SPC-A1 cells with 50% inhibitive concentration values of 57.07±14.46 and 34.97±6.30 μg/mL, respectively. Drug combinations were associated with significantly higher cytotoxic effects than each drug alone (PSPC-A1. These results suggest that Amp-Na may be applied to enhance the anticancer action of CBP.

  14. Siamese crocodile bile induces apoptosis in NCI-H1299 human non-small cell lung cancer cells via a mitochondria-mediated intrinsic pathway and inhibits tumorigenesis.

    Science.gov (United States)

    Tian, Ling; Deng, Yi-Tao; Dong, Xin; Fan, Jia-Yi; Li, Hua-Liang; Ding, Yu-Mei; Peng, Wei-Xi; Chen, Qing-Xi; Shen, Dong-Yan

    2017-04-01

    Non-small-cell lung cancer (NSCLC) is a widespread and particularly aggressive form of cancer. Patients with NSCLC and early metastases typically have poor prognosis, highlighting the critical need for additional drugs to improve disease outcome following surgical resection. The present study aimed to determine if Siamese crocodile bile (SCB) had an anti‑cancer effect on NCI‑H1299 human NSCLC cells. The inhibitory mechanism of SCB was examined in cell culture and nude mice. In vitro experimental results revealed that SCB inhibited the proliferation and colony‑forming ability of NCI‑H1299 cells by arresting cell cycle and inducing apoptosis. The loss of the mitochondrial membrane potential and the release of cytochrome c indicated that SCB treatment may lead to mitochondrial dysfunction in NCI‑H1299 cells. At the molecular level, SCB altered the ratio of protein expression of Bax/Bcl‑2 and activated associated caspases, suggesting that intrinsic pathway involvement in the SCB‑induced apoptosis of NCI‑H1299 cells. In the in vivo experiments, intraperitoneal injection of SCB for 4 weeks inhibited xenograft tumor growth by 46.8% without observable toxicity in nude mice. Immunohistochemistry analysis of proliferating cell nuclear antigen and vascular endothelial growth factor also revealed that SCB inhibited cell proliferation and metastasis in NSCLC xenograft tumors. Overall, SCB exerted an anti-cancer effect on NCI‑H1299 human NSCLC cells in vitro and in vivo and may have therapeutic potential for the treatment of human NSCLC.

  15. MiR-107 and MiR-185 can induce cell cycle arrest in human non small cell lung cancer cell lines.

    Directory of Open Access Journals (Sweden)

    Yukari Takahashi

    Full Text Available BACKGROUND: MicroRNAs (miRNAs are short single stranded noncoding RNAs that suppress gene expression through either translational repression or degradation of target mRNAs. The annealing between messenger RNAs and 5' seed region of miRNAs is believed to be essential for the specific suppression of target gene expression. One miRNA can have several hundred different targets in a cell. Rapidly accumulating evidence suggests that many miRNAs are involved in cell cycle regulation and consequentially play critical roles in carcinogenesis. METHODOLOGY/PRINCIPAL FINDINGS: Introduction of synthetic miR-107 or miR-185 suppressed growth of the human non-small cell lung cancer cell lines. Flow cytometry analysis revealed these miRNAs induce a G1 cell cycle arrest in H1299 cells and the suppression of cell cycle progression is stronger than that by Let-7 miRNA. By the gene expression analyses with oligonucleotide microarrays, we find hundreds of genes are affected by transfection of these miRNAs. Using miRNA-target prediction analyses and the array data, we listed up a set of likely targets of miR-107 and miR-185 for G1 cell cycle arrest and validate a subset of them using real-time RT-PCR and immunoblotting for CDK6. CONCLUSIONS/SIGNIFICANCE: We identified new cell cycle regulating miRNAs, miR-107 and miR-185, localized in frequently altered chromosomal regions in human lung cancers. Especially for miR-107, a large number of down-regulated genes are annotated with the gene ontology term 'cell cycle'. Our results suggest that these miRNAs may contribute to regulate cell cycle in human malignant tumors.

  16. Targeting Hsp90 with small molecule inhibitors induces the over-expression of the anti-apoptotic molecule, survivin, in human A549, HONE-1 and HT-29 cancer cells

    Directory of Open Access Journals (Sweden)

    Lyu Kevin W

    2010-04-01

    Full Text Available Abstract Background Survivin is a dual functioning protein. It inhibits the apoptosis of cancer cells by inhibiting caspases, and also promotes cancer cell growth by stabilizing microtubules during mitosis. Since the molecular chaperone Hsp90 binds and stabilizes survivin, it is widely believed that down-regulation of survivin is one of the important therapeutic functions of Hsp90 inhibitors such as the phase III clinically trialed compound 17-AAG. However, Hsp90 interferes with a number of molecules that up-regulate the intracellular level of survivin, raising the question that clinical use of Hsp90 inhibitors may indirectly induce survivin expression and subsequently enhance cancer anti-drug responses. The purpose of this study is to determine whether targeting Hsp90 can alter survivin expression differently in different cancer cell lines and to explore possible mechanisms that cause the alteration in survivin expression. Results Here, we demonstrated that Hsp90 inhibitors, geldanamycin and 17-AAG, induced the over-expression of survivin in three different human cancer cell lines as shown by Western blotting. Increased survivin mRNA transcripts were observed in 17-AAG and geldanamycin-treated HT-29 and HONE-1 cancer cells. Interestingly, real-time PCR and translation inhibition studies revealed that survivin was over-expressed partially through the up-regulation of protein translation instead of gene transcription in A549 cancer cells. In addition, 17-AAG-treated A549, HONE-1 and HT-29 cells showed reduced proteasomal activity while inhibition of 26S proteasome activity further increased the amount of survivin protein in cells. At the functional level, down-regulation of survivin by siRNA further increased the drug sensitivity to 17-AAG in the tested cancer cell lines. Conclusions We showed for the first time that down-regulation of survivin is not a definite therapeutic function of Hsp90 inhibitors. Instead, targeting Hsp90 with small

  17. Flavonoids from Gynostemma pentaphyllum exhibit differential induction of cell cycle arrest in H460 and A549 cancer cells.

    Science.gov (United States)

    Tsui, Ko-Chung; Chiang, Tzu-Hsuan; Wang, Jinn-Shyan; Lin, Li-Ju; Chao, Wei-Chih; Chen, Bing-Huei; Lu, Jyh-Feng

    2014-10-31

    Flavonoids, containing mainly kaempferol rhamnohexoside derivatives, were extracted from Gynostemma pentaphyllum (G. pentaphyllum) and their potential growth inhibition effects against H460 non-small cell lung cancer cells was explored and compared to that on A549 cells. The extracted flavonoids were found to exhibit antiproliferation effects against H460 cells (IC50 = 50.2 μg/mL), although the IC50 of H460 is 2.5-fold that of A549 cells (IC50 = 19.8 μg/mL). Further investigation revealed that H460 cells are more susceptible to kaempferol than A549, whereas A549 cell growth is better inhibited by kaempferol rhamnohexoside derivatives as compared with H460. In addition, flavonoids from G. pentaphyllum induced cell cycle arrest at both S and G2/M phases with concurrent modulated expression of the cellular proteins cyclin A, B, p53 and p21 in A549 cells, but not H460. On the contrary, apoptosis and concomitant alteration in balance of BCL-2 and BAX expression as well as activation of caspase-3 were equally affected between both cells by flavonoid treatment. These observations strongly suggest the growth inhibition discrepancy between H460 and A549 following flavonoid treatment can be attributed to the lack of cell cycle arrest in H460 cells and the differences between H460 and A549 cells may serve as contrasting models for further mechanistic investigations.

  18. Flavonoids from Gynostemma pentaphyllum Exhibit Differential Induction of Cell Cycle Arrest in H460 and A549 Cancer Cells

    Directory of Open Access Journals (Sweden)

    Ko-Chung Tsui

    2014-10-01

    Full Text Available Flavonoids, containing mainly kaempferol rhamnohexoside derivatives, were extracted from Gynostemma pentaphyllum (G. pentaphyllum and their potential growth inhibition effects against H460 non-small cell lung cancer cells was explored and compared to that on A549 cells. The extracted flavonoids were found to exhibit antiproliferation effects against H460 cells (IC50 = 50.2 μg/mL, although the IC50 of H460 is 2.5-fold that of A549 cells (IC50 = 19.8 μg/mL. Further investigation revealed that H460 cells are more susceptible to kaempferol than A549, whereas A549 cell growth is better inhibited by kaempferol rhamnohexoside derivatives as compared with H460. In addition, flavonoids from G. pentaphyllum induced cell cycle arrest at both S and G2/M phases with concurrent modulated expression of the cellular proteins cyclin A, B, p53 and p21 in A549 cells, but not H460. On the contrary, apoptosis and concomitant alteration in balance of BCL-2 and BAX expression as well as activation of caspase-3 were equally affected between both cells by flavonoid treatment. These observations strongly suggest the growth inhibition discrepancy between H460 and A549 following flavonoid treatment can be attributed to the lack of cell cycle arrest in H460 cells and the differences between H460 and A549 cells may serve as contrasting models for further mechanistic investigations.

  19. Lithium-Acetate-Mediated Biginelli One-Pot Multicomponent Synthesis under Solvent-Free Conditions and Cytotoxic Activity against the Human Lung Cancer Cell Line A549 and Breast Cancer Cell Line MCF7

    Directory of Open Access Journals (Sweden)

    Harshita Sachdeva

    2012-01-01

    Full Text Available Various Biginelli compounds (dihydropyrimidinones have been synthesized efficiently and in high yields under mild, solvent-free, and eco-friendly conditions in a one-pot reaction of 1,3-dicarbonyl compounds, aldehydes, and urea/thiourea/acetyl thiourea using lithium-acetate as a novel catalyst without the addition of any proton source. Comparative catalytic efficiency of lithium-acetate and polyphosphoric acid to catalyze Biginelli condensation is also studied under neat conditions. The reaction is carried out in the absence of any solvent and represents an improvement of the classical Biginelli protocol and an advantage in comparison with FeCl3·6H2O, NiCl2·6H2O and CoCl2·6H2O that were used with HCl as a cocatalyst. Compared to classical Biginelli reaction conditions, the present method has advantages of good yields, short reaction times, and experimental simplicity. The obtained products have been identified by spectral (1H NMR and IR data and their melting points. The prepared compounds are evaluated for anticancer activity against two human cancer cell lines (lung cancer cell line A549 and breast cancer cell line MCF7.

  20. Comparative Cytotoxicity of Glycyrrhiza glabra Roots from Different Geographical Origins Against Immortal Human Keratinocyte (HaCaT), Lung Adenocarcinoma (A549) and Liver Carcinoma (HepG2) Cells.

    Science.gov (United States)

    Basar, Norazah; Oridupa, Olayinka Ayotunde; Ritchie, Kenneth J; Nahar, Lutfun; Osman, Nashwa Mostafa M; Stafford, Angela; Kushiev, Habibjon; Kan, Asuman; Sarker, Satyajit D

    2015-06-01

    Glycyrrhiza glabra L. (Fabaceae), commonly known as 'liquorice', is a well-known medicinal plant. Roots of this plant have long been used as a sweetening and flavouring agent in food and pharmaceutical products, and also as a traditional remedy for cough, upper and lower respiratory ailments, kidney stones, hepatitis C, skin disorder, cardiovascular diseases, diabetes, gastrointestinal ulcers and stomach ache. Previous pharmacological and clinical studies have revealed its antitussive, antiinflammatory, antiviral, antimicrobial, antioxidant, immunomodulatory, hepatoprotective and cardioprotective properties. While glycyrrhizin, a sweet-tasting triterpene saponin, is the principal bioactive compound, several bioactive flavonoids and isoflavonoids are also present in the roots of this plant. In the present study, the cytotoxicity of the methanol extracts of nine samples of the roots of G. glabra, collected from various geographical origins, was assessed against immortal human keratinocyte (HaCaT), lung adenocarcinoma (A549) and liver carcinoma (HepG2) cell lines using the in vitro 3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyl tetrazoliumbromide cell toxicity/viability assay. Considerable variations in levels of cytotoxicity were observed among various samples of G. glabra.

  1. 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.

    Science.gov (United States)

    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.

  2. Pharmacokinetic correlation between experimental and clinical effects on human non-small cell lung cancers of cis-diammineglycolatoplatinum (254-S) and cis-diamminedichloroplatinum.

    Science.gov (United States)

    Koenuma, M; Kasai, H; Uchida, N; Wada, T; Hattori, M; Oguma, T; Totani, T; Inaba, M

    1995-01-01

    We attempted to correlate the in vitro and in vivo antitumor activities of cis-diammineglycolatoplatinum (254-S), a novel platinum complex, and cis-diamminedichloro-platinum (CDDP) against the established culture cell lines and xenografts of human non-small cell lung cancer (NSCLC) with their clinical effects, based on the previous finding that the cytotoxicity of CDDP depends on the area under the curve (AUC). The concentration of 254-S and CDDP inhibiting the in vitro growth of 4 cultured NSCLC lines by 50% (IC50) was 0.82-7.8 and 0.53-4.2 micrograms/ml, respectively, showing a similar level. Of the 4 cell lines, only the most sensitive line, RERF-LC-AI, showed an IC50 close to a specific concentration (0.50 for 254-S and 0.32 micrograms/ml for CDDP) that reproduces in vitro the clinical AUCfree (24.8 and 5.34 micrograms-hr/ml) of the respective drugs. We treated 6 lines of human NSCLC xenografts implanted in nude mice with 254-S and CDDP at a particular dose (13.2 and 3.7 mg/kg) that is equivalent to the clinical doses with respect to the plasma AUCfree. 254-S and CDDP exhibited significant antitumor effects on 2 and 1 of the 6 lines, respectively. These in vitro and in vivo findings were considered to be relatively well correlated with the reported clinical response rates of 15-19% for 254-S and 14-15% for CDDP.

  3. Breviscapine suppresses the growth of non-small cell lung cancer by enhancing microRNA-7 expression

    Indian Academy of Sciences (India)

    JIAN ZENG; SHUNV CAI

    2017-03-01

    Breviscapine (BVP) has previously been shown to inhibit the proliferation of hepatocellular carcinoma cells.However, little is known about the effects of BVP on non-small cell lung cancer (NSCLC) growth. Here, we aimedto study the effects of BVP on human NSCLC growth. We employed A549, NCL-H460 and A549 cells transfectedwith microRNA-7 (miR-7) mimic and inhibitor to investigate the effect of BVP on cell proliferation, apoptosis andapoptosis-associated molecules. The results showed that BVP significantly reduced the growth of A549 and NCLH460cells in a concentration-dependent and time-dependent manner, accompanied by a significant elevation ofapoptosis. Additionally, the present study also confirmed that BVP-treated A549 cells showed increased levels of Baxand microRNA-7 (miR-7) and a decreased level of Bcl-2. The up-regulation of miR-7 enhanced the BVP sensitivity ofNSCLC cells by suppressing cell proliferation and promoting cell apoptosis, while the inhibition of miR-7 reversedthe anti-proliferative pro-apoptotic effects of BVP. Pre-treatment with miR-7 mimics enhanced the BVP-mediateddown-regulation of Bax/Bcl-2 in NSCLC cells, while pre-treatment with the miR-7 inhibitor blocked the BVPmediateddown-regulation of Bax/Bcl. Taken together, these results confirm that BVP effectively inhibits NSCLCproliferation and that miR-7, as a novel target, is likely involved in BVP-induced growth suppression and theapoptosis of NSCLC cells.

  4. [Grape seed proanthocyanidins inhibits the invasion and migration of A549 lung cancer cells].

    Science.gov (United States)

    Zhou, Yehan; Ye, Xiufeng; Shi, Yao; Wang, Ke; Wan, Dan

    2016-02-01

    To explore the effect of grape seed proanthocyanidins (GSPs) on the invasion and migration of A549 lung cancer cells and the underlying mechanism. Trypan blue dye exclusion assay was used to determine the cytotoxic effect of varying doses of GSPs on the BEAS-2B normal human pulmonary epithelial cells. After treated with 0, 10, 20, 40, 80 μg/mL GSP, the proliferation of A549 cells was detected by MTT assay; the invasion and migration of A549 cells were determined by Transwell(TM) assay and scratch wound assay, respectively. The levels of epithelial growth factor receptor (EGFR), E-cadherin, N-cadherin in A549 cells treated with GSPs were detected by Western blotting. (0-40) μg/mL GSPs had no significant toxic effect on BEAS-2B cells, while 80 μg/mL GSPs had significant cytotoxicity to BEAS-2B cells. The proliferation of A549 cells was significantly inhibited within limited dosage in a dose-dependent manner, and the abilities of invasion and migration of A549 cells were also inhibited. Western blotting showed that the expression of EGFR and N-cadherin decreased, while E-cadherin increased after GSPs treatment. GSPs could inhibit the abilities of proliferation, invasion and migration of A549 cells, which might be related to the dow-regulation of EGFR and N-cadherin and the up-regulation of E-cadherin.

  5. Fibroblasts weaken the anti-tumor effect of gefitinib on co-cultured non-small cell lung cancer cells

    Institute of Scientific and Technical Information of China (English)

    Yong Xiao; Wang Peiqin; Jiang Tao; Yu Wenchen; Shang Yan; Han Yiping; Zhang Pingping

    2014-01-01

    Background Non-small cell lung cancer (NSCLC) is the most common lung malignancy worldwide.The metastatic potential of NSCLC cells has been shown to be associated with the tumor microenvironment,which consists of tumor cells,stroma,blood vessels,immune infiltrates and the extracellular matrix.Fibroblasts can produce numerous extraceilular matrix molecules and growth factors.Gefitinib has been evaluated as a first-line treatment in selected patients,and it has shown favorable efficacy especially in NSCLC,but it is not effective for everyone.Methods In this study,we examined the antitumor activity of gefitinib on lung fibroblasts co-cultured of lung cancer cells.A series of co-culture experiments that employed cell counting kit-8 (CCK8),transwells,real-time polymerase chain reaction (RT-PCR) and Western blotting with HFL-1 fibroblasts and A549 human lung carcinoma cells were performed to learn more about tumor cell proliferation,migration and invasion; and to determine any change of epithelial mesenchymal transition (EMT)-associated tumor markers vimentin,matrix metallopro-teinase 2 (MMP2) and chemotaxis cytokines receptor 4 (CXCR4) mRNA levels.Results A549 cell proliferation in the presence of HFL-1 cells was not significantly increased compared with A549 cells alone,but A549 cell spheroid body formation was increased after co-culture,and treatment with gefitinib increased further.Our study also revealed that fibroblasts attenuated the lung cancer cell inhibition ratio of migration and invasion after gefitinib treatment in vitro.To further study this mechanism,RT-PCR analysis showed that vimentin,MMP2 and CXCR4 mRNA levels were more highly expressed in the lung cancer cells after co-culture,but did not obviously decrease compared with the control cells following gefitinib treatment.This suggests the mechanism by which fibroblasts attenuate gefitinib-induced expression of EMT-associated tumor markers.Finally,our results demonstrated that co-culture with A549 lung

  6. Characterization of the effects of cyclooxygenase-2 inhibition in the regulation of apoptosis in human small and non-small cell lung cancer cell lines.

    LENUS (Irish Health Repository)

    Alam, Mahmood

    2012-02-03

    BACKGROUND: Cyclooxygenase-2 enzyme (COX-2) is overexpressed in human non-small cell lung cancer (NSCLC) but is not expressed in small cell lung cancer. Selective COX-2 inhibitors have been shown to induce apoptosis in NSCLC cells, an effect which is associated with the regulation of intracellular MAP kinase (MAPK) signal pathways. Our aims were to characterize the effects of COX-2 inhibition by rofecoxib on apoptosis in human NSCLC and small cell lung cancer cell lines. METHODS: The human NSCLC cell line NCI-H2126 and small cell lung cancer cell line DMS-79 were used. Constitutive COX-2 protein levels were first determined by Western blot test. Levels of apoptosis were evaluated by using propidium iodide staining on FACScan analysis after incubation of NCI-H2126 and DMS-79 with p38 MAPK inhibitor SB202190 (25 ?microM), NF-kappaB inhibitor SN50 (75 microg\\/mL), and rofecoxib at 100 and 250 microM. All statistical analysis was performed by analysis of variance. RESULTS: Western blot test confirmed the presence of COX-2 enzyme in NCI-H2126 and absence in DMS-79. Interestingly, rofecoxib treatment demonstrated a dose-dependent increase in apoptosis in both cell lines. Given this finding, the effect of rofecoxib on NF-kappaB and p38 MAPK pathways was also examined. Apoptosis in both cell lines was unaltered by SN50, either alone or in combination with rofecoxib. A similar phenomenon was observed in NCI-H2126 cells treated with SB202190, either alone or in combination with rofecoxib. In contrast, p38 MAPK inhibition greatly upregulated DMS-79 apoptosis in a manner that was unaltered by the addition of rofecoxib. CONCLUSIONS: Rofecoxib led to a dose-dependent increase in apoptosis in both tumor cell lines. This effect occurred independently of COX-2, NF-kappaB, and p38 MAPK pathways in DMS-79 cells. As such, rofecoxib must act on alternative pathways to regulate apoptosis in human small cell lung cancer cells.

  7. Overexpression of collagen triple helix repeat containing 1 (CTHRC1) is associated with tumour aggressiveness and poor prognosis in human non-small cell lung cancer.

    Science.gov (United States)

    Ke, Zunfu; He, Weiling; Lai, Yuanhui; Guo, Xuefeng; Chen, Sharon; Li, Shuhua; Wang, Yuefeng; Wang, Liantang

    2014-10-15

    Collagen triple helix repeat-containing 1 (CTHRC1), a novel oncogene, was identified to be aberrantly overexpressed in several malignant tumors. However, the expression profile of CTHRC1 and its clinical significance in non-small cell lung cancer (NSCLC) remain unknown. In this study, we showed that CTHRC1 was evidently overexpressed in human NSCLC tissues and NSCLC cell lines at the protein and mRNA level. Ectopic up-regulation of CTHRC1 in cancer cells resulted in elevated invasive and proliferative abilities, which were attenuated by the specific CTHRC1 siRNA. The biological effect of CTHRC1 on metastasis and proliferation was mediated by the activation of the Wnt/β-catenin pathway. Furthermore, CTHRC1 immunoreactivity was evidently overexpressed in paraffin-embedded NSCLC tissues (212/292, 72.60%) in comparison to corresponding adjacent non-cancerous tissues (6/66, 9.09%) (p<0.001). Clinicopathologic analysis showed that CTHRC1 expression was significantly correlated with differentiation degree (p<0.001), clinical stage (p<0.001), T classification (p<0.001), lymph node metastasis (p=0.013) and distant metastasis (p<0.001). Kaplan-Meier analysis revealed that patients with high CTHRC1 expression had poorer overall survival rates than those with low CTHRC1 expression. Multivariate analysis indicated that CTHRC1 expression was an independent prognostic factor for the overall survival of NSCLC patients. Collectively, CTHRC1 plays important roles in NSCLC progression, and the evaluation of CTHRC1 expression could serve as a potential marker for metastasis progression and prognosis in NSCLC patients.

  8. Identification of microRNA profiles in docetaxel-resistant human non-small cell lung carcinoma cells (SPC-A1).

    Science.gov (United States)

    Rui, Wang; Bing, Feng; Hai-Zhu, Song; Wei, De; Long-Bang, Chen

    2010-01-01

    Docetaxel has been used as first-line chemotherapy in advanced non-small cell lung carcinoma (NSCLC), but further extensive and effective application is prevented by drug resistance. MicroRNAs (miRNAs) have recently been identified as important posttranscriptional regulators, which are involved in various biological processes. The aim of this study was to identify microRNA expression profiles involved in the development of docetaxel resistance in NSCLC. Here, microarray chip technology was employed to identify miRNA expression profiles in docetaxel-resistant human NSCLC cell line (SPC-A1/docetaxel). Then, the changes of miRNAs expression (>2-fold compared with control SPC-A1 cell line) were testified by quantitative real-time RT-PCR (qRT-PCR) assay. Furthermore, the potential target genes regulated by selected miRNAs were analysed by various target prediction tools. The expression of a total of 52 miRNAs showed significant difference between SPC-A1/docetaxel cells and control SPC-A1 cells (P SPC-A1/docetaxel cells, while the expression of other three miRNAs (miR-192, 424 and 98) was significantly up-regulated in SPC-A1/docetaxel cells (P SPC-A1/docetaxel and SPC-A1 cells were further confirmed by qRT-PCR and Western blot. Taken together, the identification of microRNA expression profiles in docetaxel-resistant NSCLC cells could provide a better understanding of mechanisms involved in drug sensitivity or resistance, which would be helpful to develop novel strategies for targeted therapies in chemorefractive NSCLC patients.

  9. rmhTNF-αCombined with Cisplatin Inhibits Proliferation of A549 Cell Line In Vitro

    Institute of Scientific and Technical Information of China (English)

    Le-min Xia; Yi-yang Zhou

    2014-01-01

    Objective To explore the inhibitory effect of recombinant mutant human tumor necrosis factor-α(rmhTNF-α) in combination with cisplatin on human lung adenocarcinoma cell line A549. Methods Human lung adenocarcinoma cell line A549 was treated with varying concentrations of rmhTNF-α (0.38, 0.75, 1.50, 6.00 and 12.00 IU/ml) or cisplatin (3.91, 7.81, 15.63, 31.25 and 62.50 μg/ml) for 24 hours. Viable cell number was analyzed by using crystal violet staining. The inhibitory rates of A549 cells growth by the two drugs were calculated. For analyzing whether there was a synergistic effect of rmhTNF-α with cisplatin, A549 cells were treated with 0.75 IU/ml rmhTNF-αand increased concentrations of cisplatin. Results rmhTNF-αor cisplatin inhibited the growth of A549 cell lines in a dose-dependent manner. The inhibitory effect of rmhTNF-αcombined with cisplatin was significantly greater than cisplatin alone at the same concentration (all P Conclusion rmhTNF-αcombined with cisplatin might have synergistic inhibitory effect on human lung adenocarcinoma cell line A549.

  10. Phospholipid flippase associates with cisplatin resistance in plasma membrane of lung adenocarcinoma A549 cells

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    The fusion of the liposomes containing N-(7-nitro-2, 1, 3-benzoxadiazol-4-yl)-i ,2-hexadecanoylSn-glycero-3-1abeled phosphatidylethanolamine (NBD-PE) with A549 and A549/DDP cells was performed, and the activity of the phospholipid flippase in the plasma membrane of the cells was measured by fluorescence intensity change of NBDPE in the outer membrane. When A549 or A549/DDP cells containing N BD-PE were incubated at 37 C for 0, 30, 60 and 90 min, the fluorescence intensities in the outer membrane of the cells were 0%, 1.4%, 2.9% and 7.8% for A59cells, and 0%, 10.5 %, 15. 5 % and 18.3 % for A549/DDP cells respectively, demonstrating that the phospholipid flippase was distributed in the plasma membrane of As49 cells, but its activity in the drug-resistant A549/DDP cells was much higher than that in the A549 cells. When the A549/DDP cells were incubated with a multidrug resistance reverse agent, verapamil, for 60 min at 37C, the results showed that the NBD-PE in outer membrane decreased by 25.0% compared with the control's. Furthermore, when A549/DDP cells were incubated with 25 μmol/L cisplatin, which is a specific anticancer drug, the flippase activity decreased by 31.6%, and it further decreased with the increase of cisplatin concentration, suggesting that phospholipid flippase in the membrane might be related to the cisplatin-resistance of human lung adenocarcinoma cancer cells.

  11. Comparison of the uptakes of Tc-99m MIBI and Tc-99m tetrofosmin in A549, an MRP-expressing cancer cell, in vitro and in vivo

    Energy Technology Data Exchange (ETDEWEB)

    Yoo, Jeong Ah; Jeong, Shin Young; Seo, Myung Rang; Bae, Jin Ho; Ahn, Byeong Cheol; Lee, Kyu Bo; Lee, Jae Tae [School of Medicine, Kyungpook National Univ., Daegu (Korea, Republic of); Choi, Sang Woon; Lee, Byung Ho [Korea Institute of Chemical Technology, Daejon (Korea, Republic of)

    2003-12-01

    Uptakes of Tc-99m MIBI (MIBI) and Tc-99m tetrofosmin (tetrofosmin) in human non-small cell lung cancer A549, multidrug-resistance associated protein (MRP) expressing cell, were investigated in vitro and in vivo. Western blot analysis and immunohistochemistry were used for detetion of MRP in A549 cells with anti-MRPr1 antibody. Cellular uptakes of two tracers were evaluated at 100 {mu}M of verapamil (Vrp), 50 {mu}M of cyclosporin A (CsA) and 25 {mu}M of butoxysulfoximide (BSO) after incubation with MIBI and tetrofosmin for 30 and 60 min at 37.deg.C, using single cell suspensions at 1x10{sup 6} cells/ml. Radioactivities in supernatants and pellets were measured with gamma well counter. A549 cells were inoculated in each flanks of 24 nude mice. Group 1 (Gr1) and Gr3 mice were treated with only MIBI or tetrofosmin, and Gr2 and Gr4 mice were treated with 70mg/kg of CsA i.p. for 1 hour before injection of 370KBq of MIBI or tetrofosmin. Mice were sacrificed at 10, 60 and 240 min. Radioactivities of organs and tumors were expressed as percentage injected dose per gram of tissue (%ID/gm). Western blot analysis of the A549 cells detected expression of MRPr1 (190 kDa) and immunohistochemical staining of tumor tissue for MRPr1 revealed brownish staining in cell membrane but not P-gp. Upon incubating A549 cells for 60 min with MIBI and tetrofosmin, cellular uptake of MIBI was higher than that of tetrofosmin. Coincubation with modulators resulted in an increase in cellular uptakes of MIBI and tetrofosmin. Coincubation with modulators resulted in an increase in cellular uptakes of MIBI and tetorfosmin. Percentage increase of MIBI was higher than that of tetrofosmin with Vrp by 623% and 427%, CsA by 753% and 629% and BSO by 219% and 140%, respectively. There was no significant difference in tumoral uptakes of MIBI and tetrofosmin between Gr1 and Gr3. Percentage increases in MIBI (114% at 10 min, 257% at 60 min, 396% at 240 min) and tetrofosmin uptake (110% at 10 min, 205% at 60

  12. Ski prevents TGF-β-induced EMT and cell invasion by repressing SMAD-dependent signaling in non-small cell lung cancer.

    Science.gov (United States)

    Yang, Haiping; Zhan, Lei; Yang, Tianjie; Wang, Longqiang; Li, Chang; Zhao, Jun; Lei, Zhe; Li, Xiangdong; Zhang, Hong-Tao

    2015-07-01

    Epithelial-mesenchymal transition (EMT) is a key event in cancer metastasis, which confers cancer cells with increased motility and invasiveness, and EMT is characterized by loss of epithelial marker E-cadherin and gain of mesenchymal marker N-cadherin. Transforming growth factor-β (TGF-β) signaling is a crucial inducer of EMT in various types of cancer. Ski is an important negative regulator of TGF-β signaling, which interacts with SMADs to repress TGF-β signaling activity. Although there is accumulating evidence that Ski functions as a promoter or suppressor in human types of cancer, the molecular mechanisms by which Ski affects TGF-β-induced EMT and invasion in non-small cell lung cancer (NSCLC) are not largely elucidated. In the present study, we investigated the mechanistic role of Ski in NSCLC metastasis. Ski was significantly reduced in metastatic NSCLC cells or tissues when compared with non-metastatic NSCLC cells or tissues. Moreover, following TGF-β stimulation Ski-silenced A549 cells had more significant features of EMT and a higher invasive activity when compared with A549 cells overexpressing Ski. Mechanistically, Ski-silenced and overexpressed A549 cells showed an increase and a reduction in the SMAD3 phosphorylation level, respectively. This was supported by plasminogen activator inhibitor-1 (PAI-1) promoter activity obtained in Ski-silenced and overexpressed A549 cells. However, after treatment of SIS3 (inhibitor of SMAD3 phosphorylation) followed by TGF-β1 stimulation, we did not observe any effect of Ski on TGF-β-induced EMT, and invasion in Ski-silenced and overexpressed A549 cells. In conclusion, our findings suggest that Ski represses TGF-β-induced EMT and invasion by inhibiting SMAD-dependent signaling in NSCLC.

  13. Synergistic activity of vorinostat combined with gefitinib but not with sorafenib in mutant KRAS human non-small cell lung cancers and hepatocarcinoma

    Directory of Open Access Journals (Sweden)

    Jeannot V

    2016-11-01

    apoptosis. The sorafenib and vorinostat combination sustained the IGF-1R-, AKT-, and mitogen-activated protein kinase-dependent signaling pathways. These results showed that there was synergistic cytotoxicity when vorinostat was combined with gefitinib for both lung adenocarcinoma and hepatocarcinoma with mutant KRAS in vitro and in vivo but that the combination of vorinostat with sorafenib did not show any benefit. These findings highlight the important role of the IGF-1R/AKT pathway in the resistance to targeted therapies and support the use of histone deacetylase inhibitors in combination with EGFR-tyrosine kinase inhibitors, especially for treating patients with mutant KRAS resistant to other treatments. Keywords: targeted therapy, combined treatments, non-small cell lung cancer, hepatocarcinoma

  14. A proteomic study on human lung adenocarcinoma cell line A549 by 2-DE and MALDI-TOF-MS%人肺腺癌细胞系A549细胞双向电泳-飞行时间质谱研究

    Institute of Scientific and Technical Information of China (English)

    杨拴盈; 田应选; 南岩东; 卜丽娜; 霍树芬; 阮禹松

    2007-01-01

    目的 初步分析人肺腺癌细胞系A549细胞蛋白质表达情况,从蛋白组水平探讨肺腺癌发病的分子机制.方法 应用2-DE技术对人肺腺癌细胞系A549细胞总蛋白进行分离,获得蛋白质表达谱;应用基质辅助激光解吸电离飞行时间质谱(MALDI-TOF-MS)结合生物信息学进行蛋白质鉴定.结果 3块胶平均蛋白斑点数为1 138±49,平均匹配点数为986±32,匹配率为85.8%.随机选取背景清晰、分辨清楚、蛋白含量较高的20个斑点进行MALDI-TOF-MS分析,共获得了18个肽质量指纹图谱(PMF).将PMFs质量数据通过Aldente软件查询SWISS-PROT数据库,根据匹配片段及氨基酸序列覆盖率等,初步鉴定出15种蛋白质.根据功能,这些蛋白质可分为:①基本代谢相关的酶类:果糖2-磷酸醛缩酶、视网醛脱氢酶1(RALDH1)、吡多醛激酶;②细胞骨架类:蛋白细胞角蛋白8(CK8)、β-2链微管蛋白;③应激相关蛋白:蛋白二硫化物异构酶A3(PDIA3);④ 信号转导分子:膜联蛋白1(ANX1)、膜联蛋白4(ANX4);⑤分子伴侣:热休克蛋白60(HSP60)、热休克蛋白β-1、抑制素(PHB);⑥转录及翻译相关蛋白:不均一性核糖核蛋白H(hnRNP H);⑦杂类:Rgr protein、NPM、PCBP1.结论 应用2-DE/MALDI-MS方法获得了较为理想的人肺腺癌细胞系A549细胞2-DE蛋白质表达谱,初步鉴定了15种蛋白质.

  15. Gleevec (STI-571 inhibits lung cancer cell growth (A549 and potentiates the cisplatin effect in vitro

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    Turner Steven

    2003-01-01

    Full Text Available Abstract Background Gleevec (aka STI571, Imatinib is a recently FDA approved anti-tumor drug for chronic myelogenous leukemia. Gleevec binds specifically to BCR-ABL tyrosine kinase and inhibit the tyrosine kinase activity. It cross-reacts with another two important membrane tyrosine kinase receptors, c-kit and PDGF receptors. We sought to investigate if Gleevec has a potential role in treatment of non-small cell lung cancer. Results We have shown that Gleevec alone can inhibit the A549 lung cancer cell growth in dose-dependent manner, and the optimal concentration of Gleevec inhibition of A549 cell growth is at the range of 2–3 μM (IC50. We have also shown that A549 cells are resistant to cisplatin treatment (IC50 64 μM. Addition of Gleevec to the A549 cells treated with cisplatin resulted in a synergistic cell killing effect, suggesting that Gleevec can potentiate the effect of cisplatin on A549 cells. We also showed that the A549 lung cancer cells expresses the platelet derived growth factor receptor α, and the inhibitory effects of Gleevec on A549 cells is likely mediated through inhibition of PDGFR α phosphorylation. We further tested 33 lung cancer patients' tumor specimens to see the frequency of PDGFR-α expression by tissue micro-arrays and immunohistochemistry. We found that 16 of the 18 squamous carcinomas (89%, 11 of the 11 adenocarcinomas (100%, and 4 of the 4 small cell lung cancers (100% expressed PDGFR-α. Conclusion These results suggest a potential role of Gleevec as adjuvant therapeutic agent for treatment of non-small cell lung cancer.

  16. Expressions and Significances of PRL-3 and RhoC in A549 Cell

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    Ping ZHANG

    2010-12-01

    Full Text Available Background and objective The expression of phosphatase of regenerating liver-3 (PRL-3 is correlated with Ras homologue C (RhoC in non-small cell lung cancer (NSCLC, suggesting that they have interactions. The aim of this study is to investigate the functions of PRL-3 and RhoC in the migration of A549 cell and the potential mechanism of PRL-3 and RhoC in carcinogenesis and cancer development. Methods PRL-3Ab and RhoCAb were used to block the functions of PRL-3 and RhoC respectively. Wound healing assay was applied to detect the migration of A549 cell and the expression levels of PRL-3 and RhoC were detected by RT-PCR. Results The migration of A549 cell decreased after blockage of PRL-3 and RhoC. The expression of RhoC decreased when PRL-3 was blocked without any changes on the expression of PRL-3. Conclusion PRL-3, RhoC could increase cell migration in A549 cells.

  17. Inhibition of epidermal growth factor receptor expression by RNA interference in A549 cells

    Institute of Scientific and Technical Information of China (English)

    MinZHANG; XinZHANG; Chun-xueBAI; JieCHEN; MinQWEI

    2004-01-01

    AIM: To investigate the biological features of A549 cells in which epidermal growth factor (EGF) receptors expression were suppressed by RNA interference (RNAi). METHODS: A549 cells were transfected using short small interfering RNAs (siRNAs) formulated with Lipofectamine 2000. The EGF receptor numbers were determined by Western blotting and flowcytometry. The antiproliferative effects of sequence specific double stranded RNA (dsRNA) were assessed using cell count, colony assay and scratch assay. The chemosensitivity of transfected cells to cisplatin was measured by MTT. RESULTS: Sequence specific dsRNA-EGFR down-regulated EGF receptor expression dramatically. Compared with the control group, dsRNA-EGFR reduced the cell number by 85.0 %, decreased the colonies by 63.3 %, inhibited the migration by 87.2 %, and increased the sensitivity of A549 to cisplatin by four-fold. CONCLUSION: Sequence specific dsRNA-EGFR were capable of suppressing EGF receptor expression, hence significantly inhibiting cellular proliferation and motility, and enhancing chemosensitivity of A549 cells to cisplatin. The successful application of dsRNA-EGFR for inhibition of proliferation in EGF receptor overexpressing cells can help extend the list of available therapeutic modalities in the treatment of non-small-cell lung carcinoma (NSCLC).

  18. Enhanced Replication of Hepatitis E Virus Strain 47832c in an A549-Derived Subclonal Cell Line

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    Schemmerer, Mathias; Apelt, Silke; Trojnar, Eva; Ulrich, Rainer G.; Wenzel, Jürgen J.; Johne, Reimar

    2016-01-01

    Hepatitis E virus (HEV) is a human pathogen with increasing importance. The lack of efficient cell culture systems hampers systematic studies on its replication cycle, virus neutralization and inactivation. Here, several cell lines were inoculated with the HEV genotype 3c strain 47832c, previously isolated from a chronically infected transplant patient. At 14 days after inoculation the highest HEV genome copy numbers were found in A549 cells, followed by PLC/PRF/5 cells, whereas HepG2/C3A, Huh-7 Lunet BLR and MRC-5 cells only weakly supported virus replication. Inoculation of A549-derived subclone cell lines resulted in most cases in reduced HEV replication. However, the subclone A549/D3 was susceptible to lower virus concentrations and resulted in higher virus yields as compared to parental A549 cells. Transcriptome analysis indicated a downregulation of genes for carcinoembryonic antigen-related cell adhesion molecules (CEACAM) 5 and 6, and an upregulation of the syndecan 2 (SDC2) gene in A549/D3 cells compared to A549 cells. However, treatment of A549/D3 cells or A549 cells with CEACAM- or syndecan 2-specific antisera did not influence HEV replication. The results show that cells supporting more efficient HEV replication can be selected from the A549 cell line. The specific mechanisms responsible for the enhanced replication remain unknown. PMID:27690085

  19. NPRL2 sensitizes human non-small cell lung cancer (NSCLC cells to cisplatin treatment by regulating key components in the DNA repair pathway.

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    Gitanjali Jayachandran

    Full Text Available NPRL2, one of the tumor suppressor genes residing in a 120-kb homozygous deletion region of human chromosome band 3p21.3, has a high degree of amino acid sequence homology with the nitrogen permease regulator 2 (NPR2 yeast gene, and mutations of NPRL2 in yeast cells are associated with resistance to cisplatin-mediated cell killing. Previously, we showed that restoration of NPRL2 in NPRL2-negative and cisplatin-resistant cells resensitize lung cancer cells to cisplatin treatment in vitro and in vivo. In this study, we show that sensitization of non-small cell lung cancer (NSCLC cells to cisplatin by NPRL2 is accomplished through the regulation of key components in the DNA-damage checkpoint pathway. NPRL2 can phosphorylate ataxia telangiectasia mutated (ATM kinase activated by cisplatin and promote downstream gamma-H2AX formation in vitro and in vivo, which occurs during apoptosis concurrently with the initial appearance of high-molecular-weight DNA fragments. Moreover, this combination treatment results in higher Chk1 and Chk2 kinase activity than does treatment with cisplatin alone and can activate Chk2 in pleural metastases tumor xenograft in mice. Activated Chk1 and Chk2 increase the expression of cell cycle checkpoint proteins, including Cdc25A and Cdc25C, leading to higher levels of G2/M arrest in tumor cells treated with NPRL2 and cisplatin than in tumor cells treated with cisplatin only. Our results therefore suggest that ectopic expression of NPRL2 activates the DNA damage checkpoint pathway in cisplatin-resistant and NPRL2-negative cells; hence, the combination of NPRL2 and cisplatin can resensitize cisplatin nonresponders to cisplatin treatment through the activation of the DNA damage checkpoint pathway, leading to cell arrest in the G2/M phase and induction of apoptosis. The direct implication of this study is that combination treatment with NPRL2 and cisplatin may overcome cisplatin resistance and enhance therapeutic efficacy.

  20. Leptin promotes metastasis by inducing an epithelial-mesenchymal transition in A549 lung cancer cells.

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    Feng, Helin; Liu, Qingyi; Zhang, Ning; Zheng, Lihua; Sang, Meixiang; Feng, Jiangang; Zhang, Jinming; Wu, Xiangyun; Shan, Baoen

    2013-01-01

    Leptin, an adipocyte-derived cytokine associated with obesity, has been reported to participate in carcinogenesis. Epithelial-mesenchymal transition (EMT) is also considered as a key event in tumor metastasis. The aim of this study is to investigate the mechanism of leptin in the promotion of EMT leading to metastasis in A549 lung cancer cells. We investigated the effect of leptin on migration of A549 cells using wound healing and transwell assays. The incidence of EMT in A549 cells was examined by real-time PCR and immunofluorescence staining. The expression of TGF-β in A549 cells was detected by real-time PCR, and blocking of TGF-β in A549 cells was achieved by siRNA techniques. Additional work was performed using 100 patient samples, which included samples from 50 patients diagnosed with lung cancer and an additional 50 patients diagnosed with lung cancer with metastatic bone lesions. Leptin expression was measured using immunohistochemistry techniques. We demonstrated that leptin can effectively enhance the metastasis of human lung cancer A549 cell line using both wound healing and transwell assays. We also found the incidence of EMT in A549 cells after leptin exposure. Furthermore, we detected the expression of TGF-β in A549 cells, which had been reported to play an important role in inducing EMT. We showed that leptin can significantly upregulate TGF-β at both the mRNA and protein levels in A549 cells. Using siRNA to block the expression of TGF-β in A549 cells, we confirmed the role of TGF-β in the promotion of metastasis and induction of EMT. Furthermore, we found that in patient samples leptin was present at higher levels in samples associated with diagnosis of lung cancer bone metastases tissue than lung cancer tissue. Our results indicated that leptin promoted the metastasis of A549 human lung cancer cell lines by inducing EMT in a TGF-β-dependent manner.

  1. The influence of ciprofloxacin on viability of A549, HepG2, A375.S2, B16 and C6 cell lines in vitro.

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    Kloskowski, Tomasz; Gurtowska, Natalia; Nowak, Monika; Joachimiak, Romana; Bajek, Anna; Olkowska, Joanna; Drewa, Tomasz

    2011-01-01

    Ciprofloxacin is a chemotherapeutic agent mainly used in the treatment of the pulmonary and urinary tract infections but is also known for its anticancer properties. The aim of these study was to check the anticancer effect of ciprofloxacin on selected five cell lines. Human non-small cell lung cancer line A549, human hepatocellular carcinoma line HepG2, human and mouse melanoma lines (A375.S2 and B16) and rat glioblastoma line C6 were used for evaluation of cytotoxic properties of ciprofloxacin (in concentration range: 10-1000 microg/mL). Viability was established using trypan blue assay and MTT. Ciprofloxacin induced morphological changes and decreased viability of A549 cells in a concentration and time dependent manner. In case of A375.S2 and B16 cell lines, cytotoxicyty of ciprofloxacin was observed but we were not able to eradicate all cells from A375.S2 and B16 cultures. HepG2 line was sensitive to ciprofloxacin, but this effect was independent from concentration and incubation time. The C6 cells were insensitive to ciprofloxacin. Our results showed that ciprofloxacin can be potentially used for the experimental adjunctive therapy of lung cancer.

  2. Rab27A regulates exosome secretion from lung adenocarcinoma cells A549: involvement of EPI64.

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    Li, Wenhai; Hu, Yunsheng; Jiang, Tao; Han, Yong; Han, Guoliang; Chen, Jiakuan; Li, Xiaofei

    2014-11-01

    Exosomes are small membrane vesicles secreted into the extracellular compartment by exocytosis. The unique composition of exosomes can be transported to other cells which allow cells to exert biological functions at distant sites. However, in lung cancer, the regulation of exosome secretion was poorly understood. In this study, we employed human lung adenocarcinoma A549 cells to determine the exosome secretion and involved regulation mechanism. We found that Rab27A was expressed in A549 cells and the reduction of Rab27A by Rab27A-specific shRNA could significantly decrease the secretion of exosome by A549 cells. EPI64, a candidate GAP that is specific for Rab27, was also detected in A549 cells. By pull-down assay, we found that EPI64 participated in the exosome secretion of A549 cells by acting as a specific GAP for Rab27A, not Rab27B. Overexpression of EPI64 enhanced exosome secretion. Taken together, in A549 cells, EPI64 could regulate the exosome secretion by functioning as a GAP specific for Rab27A.

  3. Toward chelerythrine optimization: Analogues designed by molecular simplification exhibit selective growth inhibition in non-small-cell lung cancer cells.

    Science.gov (United States)

    Yang, Rosania; Tavares, Maurício T; Teixeira, Sarah F; Azevedo, Ricardo A; C Pietro, Diego; Fernandes, Thais B; Ferreira, Adilson K; Trossini, Gustavo H G; Barbuto, José A M; Parise-Filho, Roberto

    2016-10-01

    A series of novel chelerythrine analogues was designed and synthesized. Antitumor activity was evaluated against A549, NCI-H1299, NCI-H292, and NCI-H460 non-small-cell lung cancer (NSCLC) cell lines in vitro. The selectivity of the most active analogues and chelerythrine was also evaluated, and we compared their cytotoxicity in NSCLC cells and non-tumorigenic cell lines, including human umbilical vein endothelial cells (HUVECs) and LL24 human lung fibroblasts. In silico studies were performed to establish structure-activity relationships between chelerythrine and the analogues. The results showed that analogue compound 3f induced significant dose-dependent G0/G1 cell cycle arrest in A549 and NCI-H1299 cells. Theoretical studies indicated that the molecular arrangement and electron characteristics of compound 3f were closely related to the profile of chelerythrine, supporting its activity. The present study presents a new and simplified chelerythrinoid scaffold with enhanced selectivity against NSCLC tumor cells for further optimization.

  4. Role of gambogic acid and NaI131 in A549/DDP cells

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    Huang, Jing; Zhu, Xiaoli; Wang, Huan; Han, Shuhua; Liu, Lu; Xie, Yan; Chen, Daozhen; Zhang, Qiang; Zhang, Li; Hu, Yue

    2017-01-01

    Resistance to platinum in tumor tissue is a considerable barrier against effective lung cancer treatment. Radionuclide therapy is the primary adjuvant treatment, however, the toxic side effects limit its dosage in the clinical setting. Therefore, the present study aimed to determine whether an NaI131 radiosensitizer could help reduce the toxic side effects of radionuclide therapy. In vitro experiments were conducted to determine whether NaI131 can inhibit platinum resistance in A549/DDP cells, which are cisplatin-resistant non-small cell lung cancer cells, and whether gambogic acid (GA) is an effective NaI131 radiosensitizer. Cell proliferation following drug intervention was analyzed using MTT and isobolographic analysis. Apoptosis was assessed by flow cytometry. In addition, the mechanisms of drug intervention were analyzed by measuring the expression of P-glycoprotein (P-gP), B cell lymphoma 2 (Bcl-2), Bcl2-associated X protein (Bax) and P53 using western blot analysis and immunocytochemistry. According to isobolographic analysis, a low concentration of NaI131 combined with GA had a synergistic effect on the inhibition of A549/DDP cell proliferation, which was consistent with an increased rate of apoptosis. Furthermore, the overexpression of Bax, and the downregulation of P-gP, P53 and Bcl-2 observed demonstrated the potential mechanism(s) of NaI131 and GA intervention. NaI131 may induce apoptosis in A549/DDP cells by regulating apoptosis-related proteins. A low concentration combination of NaI131 and GA was able to significantly inhibit A549/DDP cell proliferation and induce cell apoptosis. Thus, the two drugs appear to have a synergistic effect on apoptosis of A549/DDP cells. PMID:28123519

  5. Open reading frame 3 of genotype 1 hepatitis E virus inhibits nuclear factor-κappa B signaling induced by tumor necrosis factor-α in human A549 lung epithelial cells.

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    Xu, Jian; Wu, Fan; Tian, Deying; Wang, Jingjing; Zheng, Zizheng; Xia, Ningshao

    2014-01-01

    Hepatitis E virus (HEV) is one of the primary causative agents of acute hepatitis, and represents a major cause of severe public health problems in developing countries. The pathogenesis of HEV is not well characterized, however, primarily due to the lack of well-defined cell and animal models. Here, we investigated the effects of genotype 1 HEV open reading frame 3 (ORF3) on TNF-α-induced nucleus factor-κappa B (NF-κB) signaling. Human lung epithelial cells (A549) were transiently transfected with ORF3 containing plasmids. These cells were then stimulated with TNF-α and the nucleus translocation of the p65 NF-κB subunit was assessed using western blot and laser confocal microscopy. DNA-binding activity of p65 was also examined using electrophoretic mobility shift assay (EMSA), and the suppression of NF-κB target genes were detected using real-time RT-PCR and ELISA. These results enabled us to identify the decreased phosphorylation levels of IKBα. We focused on the gene of negative regulation of NF-κB, represented by TNF-α-induced protein 3 (TNFAIP3, also known as A20). Reducing the levels of A20 with siRNAs significantly enhances luciferase activation of NF-κB. Furthermore, HEV ORF3 regulated A20 primarily via activating transcription factor 6 (ATF6), involved in unfolded protein response (UPR), resulting in the degradation or inactivation of the receptor interacting protein 1 (RIP1), a major upstream activator of IKB kinase compounds (IKKs). Consequently, the phosphorylation of IKBα and the nucleus translocation of p65 are blocked, which contributes to diminished NF-κB DNA-binding activation and NF-κB-dependent gene expression. The findings suggest that genotype 1 HEV, through ORF3, may transiently activate NF-κB through UPR in early stage, and subsequently inhibit TNF-α-induced NF-κB signaling in late phase so as to create a favorable virus replication environment.

  6. Open reading frame 3 of genotype 1 hepatitis E virus inhibits nuclear factor-κappa B signaling induced by tumor necrosis factor-α in human A549 lung epithelial cells.

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    Jian Xu

    Full Text Available Hepatitis E virus (HEV is one of the primary causative agents of acute hepatitis, and represents a major cause of severe public health problems in developing countries. The pathogenesis of HEV is not well characterized, however, primarily due to the lack of well-defined cell and animal models. Here, we investigated the effects of genotype 1 HEV open reading frame 3 (ORF3 on TNF-α-induced nucleus factor-κappa B (NF-κB signaling. Human lung epithelial cells (A549 were transiently transfected with ORF3 containing plasmids. These cells were then stimulated with TNF-α and the nucleus translocation of the p65 NF-κB subunit was assessed using western blot and laser confocal microscopy. DNA-binding activity of p65 was also examined using electrophoretic mobility shift assay (EMSA, and the suppression of NF-κB target genes were detected using real-time RT-PCR and ELISA. These results enabled us to identify the decreased phosphorylation levels of IKBα. We focused on the gene of negative regulation of NF-κB, represented by TNF-α-induced protein 3 (TNFAIP3, also known as A20. Reducing the levels of A20 with siRNAs significantly enhances luciferase activation of NF-κB. Furthermore, HEV ORF3 regulated A20 primarily via activating transcription factor 6 (ATF6, involved in unfolded protein response (UPR, resulting in the degradation or inactivation of the receptor interacting protein 1 (RIP1, a major upstream activator of IKB kinase compounds (IKKs. Consequently, the phosphorylation of IKBα and the nucleus translocation of p65 are blocked, which contributes to diminished NF-κB DNA-binding activation and NF-κB-dependent gene expression. The findings suggest that genotype 1 HEV, through ORF3, may transiently activate NF-κB through UPR in early stage, and subsequently inhibit TNF-α-induced NF-κB signaling in late phase so as to create a favorable virus replication environment.

  7. 龙葵生物碱体外抑制肿瘤细胞增殖作用的实验研究%Study of Antineoplastic Effects In Vitro of Solanine Extract on Human Lung Cancer A549 Cells

    Institute of Scientific and Technical Information of China (English)

    黄越燕; 朱琦峰; 周燕; 张莹楠

    2012-01-01

    目的:研究龙葵生物碱提取物对人肺癌A549细胞株增殖的抑制作用.方法:采用盐酸-乙醇混合溶剂加热回流法从龙葵中提取分离生物碱.以MTT法考察龙葵生物碱不同浓度对人肺癌A549细胞株增殖的抑制作用,采用倒置显微镜观察药物对肿瘤细胞株形态的影响.结果:龙葵生物碱提取物对人肺癌A549细胞株具有显著的细胞增殖抑制作用,且呈剂量依赖关系,并可使肿瘤细胞形态发生显著变化.结论:龙葵生物碱具有对肺癌细胞的抑制作用.

  8. Identification and significance of differential proteins in A549 cells transfected with HLCDG1

    Institute of Scientific and Technical Information of China (English)

    ZOU Fei-yan; HU Wei; YU Yan-hui; OUYANG Yong-mei; XIE Hai-long; ZENG Ping-yao; CHEN Zhu-chu; LI Feng; XIAO Zhi-qiang; FENG Xue-ping; ZHANG Peng-fei; YANG Hai-yan

    2005-01-01

    HLCDG1, which locates in chromosome 5q33, is a novel gene cloned recently. The HLCDG1 expression was significantly down regulated in the primary lung carcinoma. It was previously studied that HLCDG1 acted like a tumor suppressor gene. In this paper, proteomics studies were performed to analyze the proteomic expression patterns in the HLCDG1-transfected human lung carcinoma cell line (A549-HLCDG1) and in the control vector-transfecred human lung carcinoma cell line (A549-vector). Employing two dimensional gel eleetrophoresis (2DE), the global pattern of protein expressions in A549-HLCDG1 human lung adenocarcinoma cell line expressing stably HL-CDG1 gene were compared with those of control A549-vector cell line to generate a differential protein expression catalog. Forty-two differentially expressed proteins were screened. Thirteen differential proteins were identified by matrix-assisted laser desorption/ionization time of flight mass spectrometry (MALDI-TOF-MS), which were 6 upregulated (MSH5, MOD, MDH precursor, ETFβ, Prxd Ⅵ and JM23) and 7 downregulated (PLC-δ1, hnRNPA2,hnRNPB1, TIM, TCTP, nm23H-1 and PrxdⅤ) proteins in A549-HLCDG1 cells compared to control A549-vector cells. The above identified proteins were involved in energy metabolism, transcription regulation, antioxidation,cell cycle, metastasis, DNA methylation and mismatch repair. Therefore, these differential expression proteins by HLCDG1 transfection may play some important roles for investigation of the biochemical basis of growth suppression of HLCDG1 gene in lung carcinoma cells A549. Further understanding of this data base may provide valuable resources for the developing novel diagnostic markers and therapeutic targets of lung cancer.

  9. Inhibition of autophagy enhances heat-induced apoptosis in human non-small cell lung cancer cells through ER stress pathways.

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    Xie, Wen-Yue; Zhou, Xiang-Dong; Yang, Juan; Chen, Ling-Xiu; Ran, Dan-Hua

    2016-10-01

    The occurrence and mechanisms of autophagy induced by heat stress are not well known in lung cancer cells. Here, we have demonstrated that heat stress induces autophagy in A549 and NCI-H460 cells through morphological and biochemical analyses. The inhibition of autophagy by chloroquine, 3-methyladenine and Beclin 1 siRNA enhanced heat-induced apoptosis. Moreover, the combination of chloroquine and heat stress inhibited tumor growth and enhanced apoptosis in vivo experiments. In addition, heat-induced autophagy involved the ER stress pathway (PERK- or IRE1-dependent). Further, heat treatment led to the increased phosphorylation of AMPK and the decreased phosphorylation of mTOR in vitro and in vivo. Knockdown of GRP78 inhibited the AMPK-mTOR pathway, and the AMPK inhibitor compound C decreased heat-induced autophagy, suggesting that activation of ER stress was involved in autophagy induction and promotion of the AMPK-mTOR pathway. In conclusion, our data suggested that the heat treatment of lung cancer cells triggered protective autophagy, as mediated by ER stress. Thus, inhibition of autophagy can be a promising strategy to enhance hyperthermia in the treatment of lung cancer patients.

  10. Synergistic effect of phenformin in non-small cell lung cancer (NSCLC) ionizing radiation treatment.

    Science.gov (United States)

    Wang, Jia; Xia, Shi'an; Zhu, Zhizhen

    2015-03-01

    Biguanides, used for anti-diabetic drugs, bring more attention in cancer research for their beneficial effects. Phenformin is more potent than metformin. However its potential application as a anti-cancer regent is far behind metformin. In order to investigate any beneficial effect of combination of Phenformin and radiotherapy, non-small cell lung cancer cell lines A549 and H1299 were exposure under different dose of ionizing radiation with or without Phenformin. Results indicated Phenformin showed synergistic effect and could induce more cancer cell apoptosis and inhibition of tumor growth compared with ionizing radiation alone. Furthermore, this synergistic effect may be through different pathway according to cancer cell genotype background. Our results showed Phenformin induced AMPK activation in A549 but not H1299. However, Phenformin activated eIF2α in both cell lines. Our findings implicated Phenformin may be used as radiosensitizer for non-small cell lung cancer therapy.

  11. Downregulated TIPE2 is associated with poor prognosis and promotes cell proliferation in non-small cell lung cancer

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    Li, Yuexia [Intensive Care Unit, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan 450052 (China); Li, Xiaohui [Department of Cardiovascular Surgery, Henan Provincial People’s Hospital, Zhengzhou, Henan 450003 (China); Liu, Gang; Sun, Rongqing; Wang, Lirui [Intensive Care Unit, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan 450052 (China); Wang, Jing, E-mail: jing_wang1980@163.com [Department of Respiratory Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan 450052 (China); Wang, Hongmin, E-mail: hmwangzz@126.com [Department of Respiratory Medicine, The First Affiliated Hospital of Zhengzhou University, Zhengzhou, Henan 450052 (China)

    2015-01-30

    Highlights: • TIPE2 is down-regulated in NSCLC tissues. • TIPE2 inhibits NSCLC cell proliferation, colony formation and invasion. • TIPE2 reduces the anti-apoptotic Bcl-XL protein and mesenchymal marker N-cadherin expression. - Abstract: The present study aims to investigate the expression pattern of TIPE2 protein and its clinical significance in human non-small cell lung cancer (NSCLC). We investigated the expression levels of TIPE2 in 96 NSCLC tumor samples by immunohistochemistry and then analyzed its clinical significance. Furthermore, the role of TIPE2 on the biological properties of the NSCLC cell line H1299 and A549 was experimentally tested in vitro and in vivo. We found that the expression level of TIPE2 was significantly higher in normal lung tissues compared with NSCLC tissues (P < 0.001), and TIPE2 downregulation was significantly correlated with advanced TNM stage (P = 0.006). TIPE2 expression was lower in lung cancer cell lines than normal bronchial cell line HBE. Transfection of TIPE2 plasmid was performed in H1299 and A549 cells. TIPE2 overexpression inhibited lung cancer cell proliferation, colony formation and cell invasive in vitro, and prevented lung tumor growth in vivo. In addition, TIPE2 transfection reduced the anti-apoptotic Bcl-XL protein and mesenchymal marker N-cadherin expression. Taken together, our results demonstrate that TIPE2 might serve as a tumor suppressor in NSCLC progression.

  12. Green tea inhibits cycolooxygenase-2 in non-small cell lung cancer cells through the induction of Annexin-1.

    Science.gov (United States)

    Lu, Qing-Yi; Jin, Yusheng; Mao, Jenny T; Zhang, Zuo-Feng; Heber, David; Dubinett, Steven M; Rao, Jianyu

    2012-11-02

    Elevated cyclooygenase-2 (COX-2) expression is frequently observed in human non-small cell lung cancer (NSCLC) and associated with poor prognosis, indicating critical involvement of the inflammatory pathway in lung carcinogenesis. Recently, we found that green tea extract (GTE) induced Annexin-1 (ANX1) in the lung adenocarcinoma A549 cells. ANX1 is a glucocorticoid-inducible 37kDa protein involved in a wide range biological function and is an important anti-inflammatory mediator. The present study further examines the interplay between the expressions and production of ANX1, COX-2, phospholipase A(2) (cPLA(2)) and prostaglandin E(2) (PGE(2)) following the treatment of NSCLC cell lines with GTE. We found that GTE induced ANX1 and inhibited COX-2 expression in lung cancer A549, H157 and H460 cell lines. Addition of pro-inflammatory cytokine IL-1β diminished GTE-induced ANX1. Silence of ANX1 in cells abrogates the inhibitory activity on COX-2, indicating that the anti-inflammatory activity of GTE is mediated at least partially by the up-regulation of ANX1. However, differential pattern of inhibitory effects of ANX1 on cPLA(2) expression was observed among various cell types, suggesting that the anti-inflammatory activity mediated by ANX1 is cell type specific. Our study may provide a new mechanism of GTE on the prevention of lung cancer and other diseases related to inflammation.

  13. Mechanism of Wnt/β-catenin signaling pathway in enhanced malignant phenotype of non-small cell lung cancer induced by anti-angiogenesis therapy

    Institute of Scientific and Technical Information of China (English)

    Xiao-Xue Zhang; Ling-Ling Zhang; Huan-Lian Yang; Xiu-Wen Wang

    2016-01-01

    Objective: To study the mechanism of Wnt/β-catenin signaling pathway in the enhanced malignant phenotype of A549 cells of human non-small cell lung cancer induced by the anti-angiogenesis therapy. Methods: The siRNA technique was employed to inhibit the expression of vascular endothelial growth factor (VEGF) in A549 cells and simulate the clinical course of anti-angiogenesis therapy. Real-time PCR and western-blot were used to study the change in the expression of Wnt/β-catenin signaling molecules at the mRNA and protein level respectively, as well as the effect on the epithelial mesenchymal transition in A549 cells. The proliferation and invasion abilities of tumor cells were detected to discuss the mechanism of Wnt/β-catenin signaling pathway in the enhanced malignant phenotype of non-small cell lung cancer induced by the anti-angiogenesis therapy. Results: The specific siRNA could significantly inhibit the expression of VEGF in cells to simulate the anti-angiogenesis therapy. Under the action of 50nM VEGF siRNA, the proliferation ability of A549 significantly increased (P<0.05). After being treated with VEGF siRNA, the invasion ability of cells increased. Twenty-four hours after the transcription of 50 nM siRNA into cells, the number of cells that come through the membrane was 278.3 ± 12.9. Compared with the Ctrl siRNA group, when VEGF was inhibited, the expression ofβ-catenin and Cyclin D1 increased by 86% and 55% respectively. Meanwhile, the expression of E-cadherin decreased, while the one of vimentin increased. Conclusions: siRNA can significantly inhibit the expression of VEGF. For the anti-angiogenesis therapy, the inhibited expression of VEGF can activate the Wnt/β-catenin signaling pathway to cause the epithelial mesenchymal transition and then the enhanced malignant phenotype of non-small cell lung cancer.

  14. Elevated pressure, a novel cancer therapeutic tool for sensitizing cisplatin-mediated apoptosis in A549

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    Oh, Sangnam [Cellular and Developmental Biology, Division of Biomedical Science, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of); Department of Preventive Medicine and Medical Research Center for Environmental Toxico-Genomics and Proteomics, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of); Kim, Yanghee [Department of Preventive Medicine and Medical Research Center for Environmental Toxico-Genomics and Proteomics, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of); Kim, Joonhee [Cellular and Developmental Biology, Division of Biomedical Science, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of); Department of Preventive Medicine and Medical Research Center for Environmental Toxico-Genomics and Proteomics, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of); Kwon, Daeho [Department of Preventive Medicine and Medical Research Center for Environmental Toxico-Genomics and Proteomics, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of); Lee, Eunil, E-mail: eunil@korea.ac.kr [Cellular and Developmental Biology, Division of Biomedical Science, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of); Department of Preventive Medicine and Medical Research Center for Environmental Toxico-Genomics and Proteomics, College of Medicine, Korea University, Seoul 136-705 (Korea, Republic of)

    2010-08-13

    Research highlights: {yields} Sensitized apoptosis in cancer cells stimulated by EP precondition with p53 dependence. {yields} EP attenuates several CDDP-resistance mechanisms. {yields} No harmful effect of EP on normal fibroblasts. -- Abstract: Intensive cancer therapy strategies have thus far focused on sensitizing cancer cells to anticancer drug-mediated apoptosis to overcome drug resistance, and this strategy has led to more effective cancer therapeutics. Cisplatin (cis-diamminedichloroplatinum(II), CDDP) is an effective anticancer drug used to treat many types of cancer, including non-small cell lung carcinoma (NSCLC), and can be used in combination with various chemicals to enhance cancer cell apoptosis. Here, we introduce the use of elevated pressure (EP) in combination with CDDP for cancer treatment and explore the effects of EP on CDDP-mediated apoptosis in NSCLC cells. Our findings demonstrate that preconditioning NSCLC cells with EP sensitizes cells for CDDP-induced apoptosis. Enhanced apoptosis was dependent on p53 and HO-1 expression, and was associated with increased DNA damage and down-regulation of genes involved in nucleotide excision repair. The transcriptional levels of transporter proteins indicated that the mechanism by which EP-induced CDDP sensitization was intracellular drug accumulation. The protein levels of some antioxidants, such as hemeoxygenase-1 (HO-1), glutathione (GSH) and glutathione peroxidase (Gpx), were decreased in A549 cells exposed to EP via the down-regulation of the transcription factor nuclear factor (erythroid-derived 2)-like 2 (Nrf-2). Furthermore, normal human fibroblasts were resistant to EP treatment, with no elevated DNA damage or apoptosis. Collectively, these data show that administration of EP is a potential adjuvant tool for CDDP-based chemosensitivity of lung cancer cells that may reduce drug resistance.

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

    Science.gov (United States)

    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

  16. Cathepsin L upregulation-induced EMT phenotype is associated with the acquisition of cisplatin or paclitaxel resistance in A549 cells.

    Science.gov (United States)

    Han, Mei-Ling; Zhao, Yi-Fan; Tan, Cai-Hong; Xiong, Ya-Jie; Wang, Wen-Juan; Wu, Feng; Fei, Yao; Wang, Long; Liang, Zhong-Qin

    2016-12-01

    Cathepsin L (CTSL), a lysosomal acid cysteine protease, is known to play important roles in tumor metastasis and chemotherapy resistance. In this study we investigated the molecular mechanisms underlying the regulation of chemoresistance by CTSL in human lung cancer cells. Human lung cancer A549 cells, A549/PTX (paclitaxel-resistant) cells and A549/DDP (cisplatin-resistant) cells were tested. The resistance to cisplatin or paclitaxel was detected using MTT and the colony-formation assays. Actin remodeling was observed with FITC-Phalloidin fluorescent staining or immunofluorescence. A wound-healing assay or Transwell assay was used to assess the migration or invasion ability. The expression of CTSL and epithelial and mesenchymal markers was analyzed with Western blotting and immunofluorescence. The expression of EMT-associated transcription factors was measured with Western blotting or q-PCR. BALB/c nude mice were implanted subcutaneously with A549 cells overexpressing CTSL, and the mice were administered paclitaxel (10, 15 mg/kg, ip) every 3 d for 5 times. Cisplatin or paclitaxel treatment (10-80 ng/mL) induced CTSL expression in A549 cells. CTSL levels were much higher in A549/PTX and A549/DDP cells than in A549 cells. Silencing of CTSL reversed the chemoresistance in A549/DDP and A549/TAX cells, whereas overexpression of CTSL attenuated the sensitivity of A549 cells to cisplatin or paclitaxel. Furthermore, A549/DDP and A549/TAX cells underwent morphological and cytoskeletal changes with increased cell invasion and migration abilities, accompanied by decreased expression of epithelial markers (E-cadherin and cytokeratin-18) and increased expression of mesenchymal markers (N-cadherin and vimentin), as well as upregulation of EMT-associated transcription factors Snail, Slug, ZEB1 and ZEB2. Silencing of CTSL reversed EMT in A549/DDP and A549/TAX cells; In contrast, overexpression of CTSL induced EMT in A549 cells. In xenograft nude mouse model, the mice implanted

  17. A Potent Derivative of Indolizino[6,7-b]Indole for Treatment of Human Non-Small Cell Lung Cancer Cells.

    Science.gov (United States)

    Chen, Chi-Wei; Wu, Ming-Hsi; Chen, Yi-Fan; Yen, Tsai-Yi; Lin, Yi-Wen; Chao, Shu-Hsin; Tala, Satishkumar; Tsai, Tung-Hu; Su, Tsann-Long; Lee, Te-Chang

    2016-04-01

    The therapeutic effect in non-small cell lung cancer (NSCLC) patients is limited because of intrinsic and acquired resistance. Thus, an unmet need exists for the development of new drugs to improve the therapeutic efficacy in NSCLC patients. In this study, the novel small molecule indolizino[6,7-b]indole derivative BO-1978 was selected to evaluate its therapeutic effects on NSCLC and its preclinical toxicity in animal models. An in vitro cytotoxicity assay revealed that BO-1978 significantly suppressed the growth of various NSCLC cell lines with or without mutations in epidermal growth factor receptor (EGFR). Mechanistically, we demonstrated that BO-1978 exhibited multiple modes of action, including inhibition of topoisomerase I/II and induction of DNA cross-linking. Treatment of NSCLC cells with BO-1978 caused DNA damage, disturbed cell cycle progression, and triggered apoptotic cell death. Furthermore, BO-1978 significantly suppressed the growth of EGFR wild-type and mutant NSCLC tumors in xenograft tumor and orthotopic lung tumor models with negligible body weight loss. The combination of BO-1978 with gefitinib further suppressed EGFR mutant NSCLC cell growth in xenograft tumor and orthotopic lung tumor models. Preclinical toxicity studies showed that BO-1978 administration did not cause apparent toxicity in mice. Based on its significant therapeutic efficacy and low drug toxicity, BO-1978 is a potential therapeutic agent for treatment of NSCLC.

  18. Change of CMTM7 expression, a potential tumor suppressor, is associated with poor clinical outcome in human non-small cell lung cancer

    Institute of Scientific and Technical Information of China (English)

    LIU Qiang; SU Yu; JIANG Guan-chao; ZHOU Zu-li; LIU Bao-cai; BU Liang; YANG Fan

    2013-01-01

    Background CKLF-like MARVEL transmembrane domain-containing 7 (CMTM7) located at 3p22.3,is a frequent deletion site and a tumor suppressor gene (TSG) locus in many cancer,which suggests CMTM7 may be a potential TSG.The aim of this study was to investigate the correlations of CMTM7 expression and survival rate in patients with non-small-cell lung cancer (NSCLC).Methods Surgical specimens of 180 cases with pathologically confirmed NSCLC were grouped into 18 tissue microarray slides.CMTM7 expression in these specimens were detected by immunohistochemistry staining and representative cases were confirmed by Western blotting.Univariate and multivariate analyses were performed to identify the association of CMTM7 expression with pathological features and survival of patients with NSCLC.Results A total of 78.9% of the 180 patients had variations of CMTM7 protein expression,either up-regulated or downregulated.Univariate analysis showed that the patients' survival rate after surgery was highly correlated with CMTM7 expression (P=0.0091).In addition,prognostic factors were examined by multivariate Cox regression analysis,and results suggested that CMTM7 expression was a unique prognostic factor in NSCLC survival.Conclusions The CMTM7 expression may be related to survival of patients with NSCLC and a unique prognostic factor.CMTM7 may play an important role in NSCLC development.

  19. Synergistic and attenuated effect of HSS in combination treatment with docetaxel plus cisplatin in human non-small-cell lung SPC-A-1 tumor xenograft.

    Science.gov (United States)

    Jia, Yuping; Zhou, Dongshun; Jia, Qingwen; Ying, Yong; Chen, Shuntai

    2016-04-01

    Platinum based combination regimens are first-line treatment option in treatment of non-small cell lung cancer (NSCLC) but the clinical utility has been limited because of their toxicities. Many reports indicated that patients with tumors can benefit from adjuvant chemotherapy drugs. The aim of this study was to confirm adjuvant chemotherapy of HSS with docetaxel plus cisplatin (DP) against NSCLC by evaluating antitumor activity and attenuated effect. In vivo SPC-A-1 xenograft model was established to evaluate antitumor activity and toxicity of HSS along or combination with DP. Evaluation indexes include the relative tumor proliferation rate, tumor growth inhibition rate, body weight, food consumption, hematological and biochemical analysis. HSS treatment showed inhibited tumor growth and increased tumor inhibition of DP treatment at doses of 250 mg/kg and 500 mg/kg. No significant toxicity was found in HSS-treated mice, but significant toxicity was found in DP-treated mice. HSS combination with DP could reduce toxicity of DP treatment by improving body weight and food consumption, and increasing the number of WBC and PLT, decreasing the level of ALT, AST and BUN. HSS combined with DP treatment has additive effect which contributes to enhance tumor growth inhibition of DP treatment and attenuated effect which contributes to reduce toxicity of DP treatment. These findings indicate potential benefit for use of HSS adjuvant chemotherapy for NSCLC treatment. Copyright © 2016. Published by Elsevier Masson SAS.

  20. Lung cancer - non-small cell

    Science.gov (United States)

    Cancer - lung - non-small cell; Non-small cell lung cancer; NSCLC; Adenocarcinoma - lung; Squamous cell carcinoma - lung ... Smoking causes most cases (around 90%) of lung cancer. The risk depends on the number of cigarettes ...

  1. Re-188 Enhances the Inhibitory Effect of Bevacizumab in Non-Small-Cell Lung Cancer

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    Jie Xiao

    2016-09-01

    Full Text Available The malignant behaviors of solid tumors such as growth, infiltration and metastasis are mainly nourished by tumor neovascularization. Thus, anti-angiogenic therapy is key to controlling tumor progression. Bevacizumab, a humanized anti-vascular endothelial growth factor (VEGF antibody, plus chemotherapy or biological therapy can prolong survival for cancer patients, but treatment-related mortality is a concern. To improve inhibitory effect and decrease side-effects on non-small-cell lung cancer (NSCLC, we used Re-188, which is a β emitting radionuclide, directly labeled with bevacizumab for radioimmunotherapy in a human A549 tumor model. Cytotoxic assay data showed that, after 188ReO4− or 188Re-bevacizumab at different concentration for 4 and 24 h, a time- and radioactivity does-dependent reduction in cell viability occurred. Also, an apoptosis assay conformed great apoptosis in the 188Re-bevacizumab group compared with controls and other treatment groups. In vivo, tumor volumes in the 188Re-bevacizumab (11.1 MBq/mice group were not reduced but growth was delayed compared with other groups. Thus, 188Re-bevacizumab enhanced the therapeutic effect of bevacizumab, suggesting a potential therapeutic strategy for NSCLC treatment.

  2. Arsenic trioxide exerts synergistic effects with cisplatin on non-small cell lung cancer cells via apoptosis induction

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    Zhang Yawei

    2009-08-01

    Full Text Available Abstract Background Despite multidisciplinary treatment, lung cancer remains a highly lethal disease due to poor response to chemotherapy. The identification of therapeutic agents with synergistic effects with traditional drugs is an alternative for lung cancer therapy. In this study, the synergistic effects of arsenic trioxide (As2O3 with cisplatin (DDP on A549 and H460 non-small cell lung cancer (NSCLC cells were explored. Methods A549 and H460 human lung cancer cells were treated with As2O3 and/or DDP. Cell growth curves, cell proliferation, cell cycle, and apoptosis of human cancer cell lines were determined by the 3-(4,5-dimethylthiahiazo (-z-y1-3,5-di-phenytetrazoliumromide (MTT method, clonogenic assay, and flow cytometry (FCM. Apoptosis was further assessed by TUNEL staining. Cell cycle and apoptosis related protein p21, cyclin D1, Bcl-2, bax, clusterin, and caspase-3 were detected by western blot. Results MTT and clonogenic assay showed As2O3 within 10-2 μM to 10 μM exerted inhibition on the proliferation of NSCLC cells, and 2.5 μM As2O3 exerted synergistic inhibition on proliferation with 3 μg/ml DDP. The combination indices (CI for A549 and H460 were 0.5 and 0.6, respectively, as confirmed by the synergism of As2O3 with DDP. FCM showed As2O3 did not affect the cell cycle. The G0/G1 fraction ranged from 57% to 62% for controlled A549 cells and cells treated with As2O3 and/or DDP. The G0/G1 fraction ranged from 37% to 42% for controlled H460 cells and cells treated with As2O3 and/or DDP. FCM and TUNEL staining illustrated that the combination of As2O3 and DDP provoked synergistic effects on apoptosis induction based on the analysis of the apoptosis index. Western blotting revealed that the expression of cell cycle related protein p21 and cyclin D1 were not affected by the treatments, whereas apoptosis related protein bax, Bcl-2, and clusterin were significantly regulated by As2O3 and/or DDP treatments compared with controls. The

  3. Influence of Berberine on Cisplatin Antineoplastic Effect in A549 Cells

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    Guojun JIANG

    2015-08-01

    Full Text Available Background and objective Cisplatin is a standard first-line chemotherapeutic agents for treating advanced non-small cell lung cancer. Unfortunately, the clinical application cisplatin is restricted because it induces serious adverse reaction. The aim of this study is to investigate the influence and probable mechanism of berberine on cisplatin antineoplastic effect on lung cancer A549 cells. Methods The total Cx43 protein amount, localization of Cx43 on cell membrane, and gap junction function were observed after the A549 cells were treated with berberine. The influence of berberine on the antitumor action of cisplatin was detected by standard colony-forming assay. Protein kinase C (PKC protein, which regulates the gap junction, was subsequently determined. Results Berberine did not affect cell survival at concentrations of 0 μM to 10 μM in the A549 cells. The gap junction function between the cells was enhanced through increased Cx43 protein expression and localization of Cx43 on the membrane after berberine treatment. The intercellular dye coupling through gap junction increased when the cells exposed to 0.1 μM, 1 μM, 10 μM berberine [33.3% (P=0.002,3, 67.0% (P<0.001, 160.0% (P<0.001] compared withcontrols. This effect was associated with the PKC activity. The cisplatin-induced inhibition of colony growth was enhanced when berberine was combined with cisplatin. Conclusion Berberine can obviously increase the antitumor effect of cisplatin by enhancing the function of the gap junction possibly in A549 cells.

  4. Roles of PI3K/Akt and c-Jun signaling pathways in human papillomavirus type 16 oncoprotein-induced HIF-1α, VEGF, and IL-8 expression and in vitro angiogenesis in non-small cell lung cancer cells.

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    Erying Zhang

    Full Text Available Human papillomavirus (HPV-16 infection may be related to non-smoking associated lung cancer. Our previous studies have found that HPV-16 oncoproteins promoted angiogenesis via enhancing hypoxia-inducible factor-1α (HIF-1α, vascular endothelial growth factor (VEGF, and interleukin-8 (IL-8 expression in non-small cell lung cancer (NSCLC cells. In this study, we further investigated the roles of PI3K/Akt and c-Jun signaling pathways in it.Human NSCLC cell lines, A549 and NCI-H460, were stably transfected with pEGFP-16 E6 or E7 plasmids. Western blotting was performed to analyze the expression of HIF-1α, p-Akt, p-P70S6K, p-P85S6K, p-mTOR, p-JNK, and p-c-Jun proteins. VEGF and IL-8 protein secretion and mRNA levels were determined by ELISA and Real-time PCR, respectively. The in vitro angiogenesis was observed by human umbilical vein endothelial cells (HUVECs tube formation assay. Co-immunoprecipitation was performed to analyze the interaction between c-Jun and HIF-1α.HPV-16 E6 and E7 oncoproteins promoted the activation of Akt, P70S6K, P85S6K, mTOR, JNK, and c-Jun. LY294002, a PI3K inhibitor, inhibited HPV-16 oncoprotein-induced activation of Akt, P70S6K, and P85S6K, expression of HIF-1α, VEGF, and IL-8, and in vitro angiogenesis. c-Jun knockdown by specific siRNA abolished HPV-16 oncoprotein-induced HIF-1α, VEGF, and IL-8 expression and in vitro angiogenesis. Additionally, HPV-16 oncoproteins promoted HIF-1α protein stability via blocking proteasome degradation pathway, but c-Jun knockdown abrogated this effect. Furthermore, HPV-16 oncoproteins increased the quantity of c-Jun binding to HIF-1α.PI3K/Akt signaling pathway and c-Jun are involved in HPV-16 oncoprotein-induced HIF-1α, VEGF, and IL-8 expression and in vitro angiogenesis. Moreover, HPV-16 oncoproteins promoted HIF-1α protein stability possibly through enhancing the interaction between c-Jun and HIF-1α, thus making a contribution to angiogenesis in NSCLC cells.

  5. Autophagy Accompanied with Bisdemethoxycurcumin-induced Apoptosis in Non-small Cell Lung Cancer Cells

    Institute of Scientific and Technical Information of China (English)

    XU Jin Hong; YANG He Ping; ZHOU Xiang Dong; WANG Hai Jing; GONG Liang; TANG Chun Lan

    2015-01-01

    Objective To investigate the effects of bisdemethoxycurcumin (BDMC) on non-small cell lung cancer (NSCLC) cell line, A549, and the highly metastatic lung cancer 95D cells. Methods CCK-8 assay was used to assess the effect of BDMC on cytotoxicity. Flow cytometry was used to evaluate apoptosis. Western blot analysis, electron microscopy, and quantification of GFP-LC3 punctuates were used to test the effect of BDMC on autophagy and apoptosis of lung cancer cells. Results BDMC inhibited the viability of NSCLC cells, but had no cytotoxic effects on lung small airway epithelial cells (SAECs). The apoptotic cell death induced by BDMC was accompanied with the induction of autophagy in NSCLC cells. Blockage of autophagy by the autophagy inhibitor 3-methyladenine (3-MA) repressed the growth inhibitory effects and induction of apoptosis by BDMC. In addition, BDMC treatment significantly decreased smoothened (SMO) and the transcription factor glioma-associated oncogene 1 (Gli1) expression. Furthermore, depletion of Gli1 by siRNA and cyclopamine (a specific SMO inhibitor) induced autophagy. Conclusion Aberrant activation of Hedgehog (Hh) signaling has been implicated in several human cancers, including lung cancers. The present findings provide direct evidence that BDMC-induced autophagy plays a pro-death role in NSCLC, in part, by inhibiting Hedgehog signaling.

  6. 采用RNA干扰技术抑制A549细胞表皮生长因子受体表达%Inhibition of epidermal growth factor receptor expression by RNA interference in A549 cells

    Institute of Scientific and Technical Information of China (English)

    张敏; Min ZHANG; 张新; Xin ZHANG; Chun-xue BAI; 白春学; Jie CHEN; 陈杰; MinQ WEI; Wei MQ

    2004-01-01

    AIM: To investigate the biological features of A549 cells in which epidermal growth factor (EGF) receptors expression were suppressed by RNA interference (RNAi). METHODS: A549 cells were transfected using short small interfering RNAs (siRNAs) formulated with Lipofectamine 2000. The EGF receptor numbers were determined by Western blotting and flowcytometry. The antiproliferative effects of sequence specific double stranded RNA(dsRNA) were assessed using cell count, colony assay and scratch assay. The chemosensitivity of transfected cells to cisplatin was measured by MTT. RESULTS: Sequence specific dsRNA-EGFR down-regulated EGF receptor expression dramatically. Compared with the control group, dsRNA-EGFR reduced the cell number by 85.0 %,decreased the colonies by 63.3 %, inhibited the migration by 87.2 %, and increased the sensitivity of A549 to cisplatin by four-fold. CONCLUSION: Sequence specific dsRNA-EGFR were capable of suppressing EGF receptor expression, hence significantly inhibiting cellular proliferation and motility, and enhancing chemosensitivity of A549 cells to cisplatin. The successful application of dsRNA-EGFR for inhibition of proliferation in EGF receptor overexpressing cells can help extend the list of available therapeutic modalities in the treatment of non-small-cell lung carcinoma (NSCLC).

  7. Effect of antisense transfecting of monocarboxylate transporter gene on biological characteristics of lung adenocarcinoma A549 cells

    Institute of Scientific and Technical Information of China (English)

    ZHANG Gui-zhi; HUANG Gui-jun; GUO Xian-jian; QIAN Gui-sheng

    2002-01-01

    Objective: To study the influence of transfecting antisense expression vector of the first subtype of the monocarboxylate transporter (MCT1) gene into lung cancer cells on pHi regulation, lactate transportation and cell growth, Methods: MCT1 antisense gene recombinant vector was introduced into human lung cancer cell line A549 by electroporation. The transfected A549 cells resistant to G418 were selected. Positive clones were examined by using PCR. The changes of intracellular pH and lactate were examined with spectrophotometric method. Cell growth was studied with cell growth curve. Results: Intracellular pH and lactate were remarkably decreased in the cells transfected pLXSN-MCT1 in comparison with A549 cells without transfection (P<0. 001). The growth of A549 cells transfected pLXSN-MCT1 was also inhibited remarkably. Conclusion: MCT1 gene may play an important role in pHi regulation, lactate transportation and cell growth in tumor cells.

  8. Green tea polyphenol EGCG reverse cisplatin resistance of A549/DDP cell line through candidate genes demethylation.

    Science.gov (United States)

    Zhang, Youwei; Wang, Xiang; Han, Liang; Zhou, Yizhou; Sun, Sanyuan

    2015-02-01

    Epigallocatechin-3-gallate (EGCG), the major polyphenol in green tea, has been extensively studied as a potential demethylating agent. Our hypothesis is that EGCG could resensitize non-small-cell lung cancer (NSCLC) cells to cisplatin (DDP) through candidate genes demethylation. The A549/DDP cell line was established by continuous exposure of A549 cells to increasing concentrations of DDP. MTT, colony formation assay, flow cytometric analysis, Hoechst staining, real time-PCR, quantitative methylation-specific PCR and in vivo experiments were performed in this study. EGCG+DDP treatment significantly caused proliferation inhibition, cell cycle arrest in G1 phase, increase of apoptosis in A549/DDP cells, along with inhibition of DNA methyltransferase (DNMT) activity and histone deacetylase (HDAC) activity, reversal of hypermethylated status and downregulated expression of GAS1, TIMP4, ICAM1 and WISP2 gene in A549/DDP cells. Furthermore, pre-treatment with EGCG followed by DDP caused significant tumor inhibition in vivo. Methylation levels of GAS1, TIMP4, ICAM1 and WISP2 were decreased and their expression levels were increased in EGCG-treatment groups, but only combinatorial treatment group caused growth inhibition. In conclusion, we identified EGCG pretreatment resensitized cells to DDP, along with the demethylation and restoration of expression of candidate genes.

  9. Resveratrol Inhibited Non-small Cell Lung Cancer Through Inhibiting STAT-3 Signaling.

    Science.gov (United States)

    Li, Xin; Wang, Dan; Zhao, Qing Chun; Shi, Tao; Chen, Jun

    2016-11-01

    Resveratrol has demonstrated many beneficial effects against cancers; however, the mechanism remains unclear. Non-small cell lung cancer accounts for 80% of lung cancers. The present study was designed to observe the effects and related mechanisms of resveratrol on non-small cell lung cancer in in vitro A549 cells. The anticancer effects of resveratrol were analyzed on cell viability, migration and invasion, proliferation and apoptosis. Cell viability was determined by sulphorhodamine B assays. Cell proliferation and apoptosis were determined by flow cytometry and migration and invasion by transwell chamber analysis. Expression of STAT-3 was examined by real-time polymerase chain reaction and western blot. Overexpressing vector of STAT-3 was also constructed and transfected into A549 cells to observe the effects of resveratrol on STAT-3 signaling. The results showed that resveratrol displayed a dose-dependent and time-dependent cytotoxicity action on A549 cell viability. Resveratrol also inhibited proliferation, migration and invasion and promoted apoptosis in a time-dependent manner from 0-72 hours. Further study showed that resveratrol inhibited the messenger RNA and protein expression of STAT-3, and overexpressed STAT-3 abolished the effects of resveratrol on proliferation, apoptosis, migration and invasion totally or in part. These results suggest that the anticancer effects of resveratrol are mediated by STAT-3 signaling. Copyright © 2016 Southern Society for Clinical Investigation. Published by Elsevier Inc. All rights reserved.

  10. Infection of A549 human type II epithelial cells with Mycobacterium tuberculosis induces changes in mitochondrial morphology, distribution and mass that are dependent on the early secreted antigen, ESAT-6.

    Science.gov (United States)

    Fine-Coulson, Kari; Giguère, Steeve; Quinn, Frederick D; Reaves, Barbara J

    2015-10-01

    Pulmonary infection by Mycobacterium tuberculosis (Mtb) involves the invasion of alveolar epithelial cells (AECs). We used Mitotracker Red(®) to assess changes in mitochondrial morphology/distribution and mass from 6 to 48 h post infection (hpi) by confocal microscopy and flow cytometry in Mtb-infected A549 type II AECs. During early infection there was no effect on mitochondrial morphology, however, by 48 hpi mitochondria appeared fragmented and concentrated around the nucleus. In flow cytometry experiments, the median fluorescence intensity (MFI) decreased by 44% at 48 hpi; double-labelling using antibodies to the integral membrane protein COXIV revealed that these changes were due to a decrease in mitochondrial mass. These changes did not occur with the apathogenic strain, Mycobacterium bovis BCG. ESAT-6 is a virulence factor present in Mtb Erdman but lacking in M. bovis BCG. We performed similar experiments using Mtb Erdman, an ESAT-6 deletion mutant and its complement. MFI decreased at 48 hpi in the parent and complemented strains versus uninfected controls by 52% and 36% respectively; no decrease was detected in the deletion mutant. These results indicate an involvement of ESAT-6 in the perturbation of mitochondria induced by virulent Mtb in AECs and suggest mitophagy may play a role in the infection process.

  11. Functional EpoR pathway utilization is not detected in primary tumor cells isolated from human breast, non-small cell lung, colorectal, and ovarian tumor tissues.

    Directory of Open Access Journals (Sweden)

    Scott D Patterson

    Full Text Available Several clinical trials in oncology have reported increased mortality or disease progression associated with erythropoiesis-stimulating agents. One hypothesis proposes that erythropoiesis-stimulating agents directly stimulate tumor proliferation and/or survival through cell-surface receptors. To test this hypothesis and examine if human tumors utilize the erythropoietin receptor pathway, the response of tumor cells to human recombinant erythropoietin was investigated in disaggregated tumor cells obtained from 186 patients with colorectal, breast, lung, ovarian, head and neck, and other tumors. A cocktail of well characterized tumor growth factors (EGF, HGF, and IGF-1 were analyzed in parallel as a positive control to determine whether freshly-isolated tumor cells were able to respond to growth factor activation ex vivo. Exposing tumor cells to the growth factor cocktail resulted in stimulation of survival and proliferation pathways as measured by an increase in phosphorylation of the downstream signaling proteins AKT and ERK. In contrast, no activation by human recombinant erythropoietin was observed in isolated tumor cells. Though tumor samples exhibited a broad range of cell-surface expression of EGFR, c-Met, and IGF-1R, no cell-surface erythropoietin receptor was detected in tumor cells from the 186 tumors examined (by flow cytometry or Western blot. Erythropoiesis-stimulating agents did not act directly upon isolated tumor cells to stimulate pathways known to promote proliferation or survival of human tumor cells isolated from primary and metastatic tumor tissues.

  12. Intracellular and Extracellular Cytokines in A549 Cells and THP1 Cells Exposed to Cigarette Smoke.

    Science.gov (United States)

    Holownia, A; Wielgat, P; Rysiak, E; Braszko, J J

    Cigarette smoke (CS) activates inflammatory cells and increases cytokine levels producing local and systemic inflammation. To assess changes in intracellular and extracellular cytokine levels we used human epithelial (A549 cells) and monocyte (THP-1) cell lines grown for 24 h in cigarette smoke-conditioned media. Cytokines were assessed using immunostaining/flow cytometry and ELISA assay. In THP1cells, grown in CS-conditioned media, the intracellular interleukins IL-1β, IL-6, and IL-10 increased by more than tenfold, while less significant increases were found in A549 cells. IL-1α and IL-1β, but not IL-6 or IL-10, were increased in the culture media, while IL-2 was raised by about fivefold only in the culture medium of A549 cells. IL-4, IL-6, IL-8, IL-10, IL-12, and tumor necrosis factor alpha were undetectable, while only a slight increase was observed in extracellular IL-17A (by about 60 %) in the medium of A549 cells and by about 115 % in the medium of THP1 cells. The interferon gamma (IFNγ) was increased by about eightfold, but only in the medium of THP1 cells grown with CS. We conclude that IL-1 and INFγ are the key cytokines responsible for pro-inflammatory signaling in epithelial cells and monocytes, respectively, exposed to cigarette smoke.

  13. [Effect of two different acellular lung matrices on α-SMA expression in A549 cells].

    Science.gov (United States)

    Chen, C; Wang, Z Y; Weng, J; Wang, Z B; Mei, J; Du, X H; Wang, L

    2017-01-24

    Objective: To explore the effect of acellular normal and fibrotic lung matrices on alpha smooth muscle actin (α-SMA) expression in human lung adenocarcinoma cell line A549. Methods: Twenty adult SD rats were randomly divided into normal group and idiopathic pulmonary fibrosis(IPF)group (n=10 each). The pulmonary fibrosis was induced by Bleomycin. Normal and fibrotic decellularized lungs were made, then sections with 500 μm thick were cut by a standard Vibratome. None scaffold was set as control group. A549 cells were seeded dropwise into different slices (normal and fibrotic scaffolds), and cultured for one week in vitro. The expression of α-SMA was measured by immunofluorescence staining and quantitative real time polymerase chain reaction (qRT-PCR). Results: In control group, the expression of α-SMA protein was positive in A549 cells by immunofluorescence staining. However, it expressed weakly both in normal and fibrotic scaffold group, and the fluorescence intensity in fibrotic scaffold group was significant lower than that in normal group (PSMA mRNA in normal and fibrotic scaffold group were (0.70±0.11) and (0.55±0.12), which were significant lower than that of control group (1.28±0.21) (PSMA mRNA in fibrotic scaffold group was decreased compared to that in normal scaffold group (PSMA in human lung adenocarcinoma cell line A549. It may inhibit the movement of A549 cells in acellular normal and fibrotic lung matrices, especially in acellular fibrotic lung scaffold.

  14. A novel aminothiazole KY-05009 with potential to inhibit Traf2- and Nck-interacting kinase (TNIK attenuates TGF-β1-mediated epithelial-to-mesenchymal transition in human lung adenocarcinoma A549 cells.

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    Jiyeon Kim

    Full Text Available Transforming growth factor (TGF-β triggers the epithelial-to-mesenchymal transition (EMT of cancer cells via well-orchestrated crosstalk between Smad and non-Smad signaling pathways, including Wnt/β-catenin. Since EMT-induced motility and invasion play a critical role in cancer metastasis, EMT-related molecules are emerging as novel targets of anti-cancer therapies. Traf2- and Nck-interacting kinase (TNIK has recently been considered as a first-in-class anti-cancer target molecule to regulate Wnt signaling pathway, but pharmacologic inhibition of its EMT activity has not yet been studied. Here, using 5-(4-methylbenzamido-2-(phenylaminothiazole-4-carboxamide (KY-05009 with TNIK-inhibitory activity, its efficacy to inhibit EMT in cancer cells was validated. The molecular docking/binding study revealed the binding of KY-05009 in the hinge region of TNIK, and the inhibitory activity of KY-05009 against TNIK was confirmed by an ATP competition assay (Ki, 100 nM. In A549 cells, KY-05009 significantly and strongly inhibited the TGF-β-activated EMT through the attenuation of Smad and non-Smad signaling pathways, including the Wnt, NF-κB, FAK-Src-paxillin-related focal adhesion, and MAP kinases (ERK and JNK signaling pathways. Continuing efforts to identify and validate potential therapeutic targets associated with EMT, such as TNIK, provide new and improved therapies for treating and/or preventing EMT-based disorders, such as cancer metastasis and fibrosis.

  15. MiR-122 Induces Radiosensitization in Non-Small Cell Lung Cancer Cell Line

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    Debin Ma

    2015-09-01

    Full Text Available MiR-122 is a novel tumor suppresser and its expression induces cell cycle arrest, or apoptosis, and inhibits cell proliferation in multiple cancer cells, including non-small cell lung cancer (NSCLC cells. Radioresistance of cancer cell leads to the major drawback of radiotherapy for NSCLC and the induction of radiosensitization could be a useful strategy to fix this problem. The present work investigates the function of miR-122 in inducing radiosensitization in A549 cell, a type of NSCLC cells. MiR-122 induces the radiosensitization of A549 cells. MiR-122 also boosts the inhibitory activity of ionizing radiation (IR on cancer cell anchor-independent growth and invasion. Moreover, miR-122 reduced the expression of its targeted genes related to tumor-survival or cellular stress response. These results indicate that miR-122 would be a novel strategy for NSCLC radiation-therapy.

  16. Cancer-associated fibroblasts promote non-small cell lung cancer cell invasion by upregulation of glucose-regulated protein 78 (GRP78) expression in an integrated bionic microfluidic device.

    Science.gov (United States)

    Yu, Ting; Guo, Zhe; Fan, Hui; Song, Jing; Liu, Yuanbin; Gao, Zhancheng; Wang, Qi

    2016-05-01

    The tumor microenvironment is comprised of cancer cells and various stromal cells and their respective cellular components. Cancer-associated fibroblasts (CAFs), a major part of the stromal cells, are a key determinant in tumor progression, while glucose-regulated protein (GRP)78 is overexpressed in many human cancers and is involved in tumor invasion and metastasis. This study developed a microfluidic-based three dimension (3D) co-culture device to mimic an in vitro tumor microenvironment in order to investigate tumor cell invasion in real-time. This bionic chip provided significant information regarding the role of GRP78, which may be stimulated by CAFs, to promote non-small cell lung cancer cell invasion in vitro. The data showed that CAF induced migration of NSCLC A549 and SPCA-1 cells in this three-dimensional invasion microdevice, which is confirmed by using the traditional Transwell system. Furthermore, CAF induced GRP78 expression in A549 and SPCA-1 cells to facilitate NSCLC cell migration and invasion, whereas knockdown of GRP78 expression blocked A549 and SPCA-1 cell migration and invasion capacity. In conclusion, these data indicated that CAFs might promote NSCLC cell invasion by up-regulation of GRP78 expression and this bionic chip microdevice is a robust platform to assess the interaction of cancer and stromal cells in tumor environment study.

  17. CD40-CD40 ligand (CD154) engagement is required but not sufficient for modulating MHC class I, ICAM-1 and Fas expression and proliferation of human non-small cell lung tumors.

    Science.gov (United States)

    Yamada, M; Shiroko, T; Kawaguchi, Y; Sugiyama, Y; Egilmez, N K; Chen, F A; Bankert, R B

    2001-05-15

    To determine the possible functional significance of CD40 expression on human non-small cell lung carcinomas and to assess the potential of CD40 as a therapeutic target, 18 lung tumor cell lines were established from biopsy tissues and were monitored for phenotypic changes on the cell surface and alterations in tumor cell proliferation after the ligation of CD40 with a trimeric fusion protein complex of CD40 ligand (CD40Lt). CD40 cross-linking resulted in up to a 6-fold increase in the surface expression of major histocompatibility complex (MHC) class I, Fas and intracellular adhesion molecule (ICAM)-1 in a subset of tumors expressing the highest levels of CD40. Suppression of tumor proliferation was seen after the ligation of CD40 on CD40Lt-responsive cell lines. The suppression was dose dependent, reversible and resulted from a delay of the tumor cells entering S-phase. No change in the cell phenotype or in proliferation were observed in CD40-negative tumors or in tumors expressing moderate-to-low levels of CD40 after incubation with CD40Lt. CD40-negative tumors transfected with the CD40 gene expressed high levels of CD40 on their surface, but were also unresponsive to CD40Lt cross-linking of CD40. Our data establish that CD40 is required (but not sufficient) for transducing a signal that results in phenotypic changes in human lung tumors and suppression in their proliferation. We conclude that CD40 on non-small cell lung tumors may represent a potential therapeutic target, but only on a subset of the CD40+ tumors.

  18. Downregulation of Cyclophilin A by siRNA diminishes non-small cell lung cancer cell growth and metastasis via the regulation of matrix metallopeptidase 9

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    Qian Zhe

    2012-10-01

    Full Text Available Abstract Background Cyclophilin A (CypA is a cytosolic protein possessing peptidyl-prolyl isomerase activity that was recently reported to be overexpressed in several cancers. Here, we explored the biology and molecular mechanism of CypA in non-small cell lung cancer (NSCLC. Methods The expression of CypA in human NSCLC cell lines was detected by real-time reverse transcription PCR. The RNA interference-mediated knockdown of CypA was established in two NSCLC cell lines (95C and A549. 239836 CypA inhibitor was also used to suppress CypA activity. Tumorigenesis was assessed based on cellular proliferation, colony formation assays, and anchorage-independent growth assays; metastasis was assessed based on wound healing and transwell assays. Results Suppression of CypA expression inhibited the cell growth and colony formation of A549 and 95C cells. CypA knockdown resulted in the inhibition of cell motility and invasion. Significantly, we show for the first time that CypA increased NSCLC cell invasion by regulating the activity of secreted matrix metallopeptidase 9 (MMP9. Likewise, suppression of CypA with 239836 CypA inhibitor decreased cell proliferation and MMP9 activity. Conclusions The suppression of CypA expression was correlated with decreased NSCLC cell tumorigenesis and metastasis.

  19. Down-regulated βIII-tubulin Expression Can Reverse Paclitaxel Resistance in A549/Taxol Cells Lines

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    Yinling ZHUO

    2014-08-01

    Full Text Available Background and objective Chemotherapy drug resistance is the primary causes of death in patients with pulmonary carcinoma which make tumor recurrence or metastasis. β-tubulin is the main cell targets of anti-microtubule drug. Increased expression of βIII-tubulin has been implicated in non-small cell lung cancer (NSCLC cell lines. To explore the relationship among the expression level of βIII-tubulin and the sensitivity of A549/Taxolcell lines to Taxol and cell cycles and cell apoptosis by RNA interference-mediated inhibition of βIII-tubulin in A549/Taxol cells. Methods Three pairs of siRNA targetd βIII-tubulin were designed and prepared, which were transfected into A549/Taxol cells using LipofectamineTM 2000. We detected the expression of βIII-tubulin mRNA using Real-time fluorescence qRT-PCR. Tedhen we selected the most efficient siRNA by the expression of βIII-tubulin mRNA in transfected group. βIII-tubulin protein level were mesured by Western blot. The taxol sensitivity in transfected group were evaluated by MTT assay. And the cell apoptosis and cell cycles were determined by flow cytometry. Results βIII-tubulin mRNA levels in A549/Taxol cells were significantly decreased in transfected grop by Real-time qRT-PCR than control groups. And βIII-tubulin siRNA-1 sequence showed the highest transfection efficiency, which was (87.73±4.87% (P<0.01; Western blot results showed that the expressional level of BIII tublin protein was significantly down-reulated in the transfectant cells than thant in the control cells. By MTT assay, we showed that the inhibition ratio of Taxol to A549/Taxol cells transfeced was higher than that of control group (51.77±4.60% (P<0.01. The early apoptosis rate of A549/Taxol cells in transfected group were significantly higher than that of control group (P<0.01; G2-M content in taxol group obviously increased than untreated samples by the cell cycle (P<0.05. Conclusion βIII-tubulin down-regulated significantly

  20. In vitro cytotoxic effects of PM{sub 2.5} from the city of Abidjan (Ivory Coast) on human A549 lung cells; Effets cytotoxiques in vitro des PM{sub 2,} {sub 5} de la ville d'Abidjan (Cote-d'Ivoire) sur des cellules pulmonaires humaines

    Energy Technology Data Exchange (ETDEWEB)

    Kouassi, Kouakou-Serge [Universite Lille Nord de France, Lille (France); Unite de Chimie Environnementale et Interactions sur le Vivant, EA 4492 MREI, Universite du Littoral Cote d' Opale, Dunkerque (France); Universite Cocody-Abidjan et Institut Pasteur, Abidjan (Cote d' Ivoire); Billet, Sylvain; Garcon, Guillaume; Verdin, Anthony; Courcot, Dominique; Shirali, Pirouz [Universite Lille Nord de France, Lille (France); Unite de Chimie Environnementale et Interactions sur le Vivant, EA 4492 MREI, Universite du Littoral Cote d' Opale, Dunkerque (France); Diouf, Amadou [Laboratoire de Toxicologie, Faculte de Medecine Pharmacologie Odontologie, Universite Cheikh Anta Diop, Dakar (Senegal); Cazier, Fabrice [Universite Lille Nord de France, Lille (France); Centre Commun de Mesures, MREI 1, Universite du Littoral Cote d' Opale, Dunkerque (France); Djaman, Joseph [Universite Cocody-Abidjan et Institut Pasteur, Abidjan (Cote d' Ivoire)

    2012-01-15

    Epidemiological studies associate air pollution, especially particulate, increased morbidity and mortality from respiratory and cardiovascular origin . Africa, which has an urbanization rate among the highest in the world, is particularly exposed. The 'Initiative on the air quality in Sub-Saharan Africa' showed the importance of atmospheric concentrations of certain pollutants such as nitrogen oxides, sulfur dioxide and particulate matter (PM{sub 10}). Like the great capitals of Africa, Abidjan, economic capital and most industrialized city of Ivory Coast is facing an air pollution from industrial-urban and health consequences for its population of nearly 6 million inhabitants. To better understand the mechanisms of action resulting from pulmonary exposure to particulate atmospheric aerosols, we proposed: (i) to collect atmospheric particles (PM{sub 2.5}) using high volume cascade impaction in the District of Abidjan in three influences (rural, urban or industrial), (ii) to determine their main physicochemical, (iii) assess their cytotoxicity and their role in the induction of oxidative damage in a model of human lung cells (A549) in culture. The chemical composition of the atmospheric particles revealed their heterogeneity, and many inorganic (e.g. Al, Ca, Fe, Mn, Zn, Ni, Cr, Cu, Pb, Mg) and organic compounds (e.g. paraffins) were quantified at the three sites. Their effect concentrations (EC) to 10 and 50% on the A549 were as follows: influence rural: EC{sub 10} = 5.91 {mu}g/cm{sup 2} and EC{sub 50} 29.55 {mu}g/cm{sup 2}, urban influence: EC{sub 10} = 5 .45 {mu}g/cm{sup 2} and EC{sub 50} = 27.23 {mu}g/cm{sup 2}, and industrial influence: EC{sub 10} = 6.86 {mu}g/cm{sup 2} and EC{sub 50} = 34.29 {mu}g/cm{sup 2}. Exposure of A549 cells to Abidjan city's PM samples for 24, 48 or 72 hours to their EC{sub 10} or EC{sub 50} induced oxidative damage, as demonstrated by the formation of malon-dialdehyde, changes in enzyme activity of superoxide dismutase

  1. Overexpression of the lung cancer-prognostic miR-146b microRNAs has a minimal and negative effect on the malignant phenotype of A549 lung cancer cells.

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    Santosh Kumar Patnaik

    Full Text Available INTRODUCTION: Expression levels of miR-146b-5p and -3p microRNAs in human non-small cell lung cancer (NSCLC are associated with recurrence of the disease after surgery. To understand this, the effect of miR-146b overexpression was studied in A549 human lung cancer cells. METHODS: A549 cells, engineered with lentiviruses to overexpress the human pre-miR-146b precursor microRNA, were examined for proliferation, colony formation on plastic surface and in soft agar, migration and invasiveness in cell culture and in vivo in mice, chemosensitivity to cisplatin and doxorubicin, and global gene expression. miR-146b expressions were assessed in microdissected stroma and epithelia of human NSCLC tumors. Association of miR-146b-5p and -3p expression in early stage NSCLC with recurrence was analyzed. PRINCIPAL FINDINGS: A549 pre-miR-146b-overexpressors had 3-8-fold higher levels of both miR-146b microRNAs than control cells. Overexpression did not alter cellular proliferation, chemosensitivity, migration, or invasiveness; affected only 0.3% of the mRNA transcriptome; and, reduced the ability to form colonies in vitro by 25%. In human NSCLC tumors, expression of both miR-146b microRNAs was 7-10-fold higher in stroma than in cancerous epithelia, and higher miR-146b-5p but lower -3p levels were predictive of recurrence. CONCLUSIONS: Only a minimal effect of pre-miR-146b overexpression on the malignant phenotype was seen in A549 cells. This could be because of opposing effects of miR-146b-5p and -3p overexpression as suggested by the conflicting recurrence-predictive values of the two microRNAs, or because miR-146b expression changes in non-cancerous stroma and not cancerous epithelia of tumors are responsible for the prognostic value of miR-146b.

  2. Estimations of chick embryo chorioallantoic membrane transplantation model of non-small cell lung cancer%非小细胞肺癌鸡胚绒毛膜移植瘤模型的建立

    Institute of Scientific and Technical Information of China (English)

    侯欣喜; 李萍; 罗殿中; 陈罡

    2015-01-01

    Objective To establish chick embryo chorioallantoic membrane(CAM) transplantatioin model of non-small cell lung cancer (NSCLC). Methods A549 lung cancer cells in gradient increasing concentrations were engrafted in CAM. Photographs of orthoyopic transplantation tumors were taken by anatomy microscope. The volume of orthoyopic transplantation tumors was measured and the tissue sections were observed. Results A549 lung cancer cells with (0.6-2.5)×106 cells could form transplantation tumors in CAM. The morphology of the transplantation tumors was in consistence with human lung cancer. Conclusion The CAM transplantation model of NSCLC is successful established. The model can provide a new technology foundation for further research on NSCLC.%目的:建立非小细胞肺癌的鸡胚绒毛膜(CAM)模型。方法以梯度递增的肺癌细胞株A549接种鸡胚,并对生长出的移植瘤行原位解剖学显微镜摄影,体积测量,切片显微观察。结果细胞数为(0.6~2.5)×106的A549细胞株可在CAM上形成移植瘤,该移植瘤与人体肺癌的形态学一致。结论成功建立非小细胞肺癌鸡胚绒毛膜移植瘤模型,为非小细胞肺癌的研究打开一个新的技术平台。

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

    Science.gov (United States)

    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.

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

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    Sun, Lixin; Zhang, Shuang; Jiang, Zhenzhou; Huang, Xin; Wang, Tao; Huang, Xiao; Li, Han [Jiangsu Center for Drug Screening, China Pharmaceutical University, Nanjing 210009 (China); Zhang, Luyong, E-mail: lyzhang@cpu.edu.cn [Jiangsu Center for Drug Screening, China Pharmaceutical University, Nanjing 210009 (China)

    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. 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.

  5. Upregulation of the Chemokine Receptor CCR7 expression by HIF-1αand HIF-2α in non-small cell lung cancer

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    Yang LI

    2008-10-01

    Full Text Available Background and objective CCR7 is closely related with the lymph node metastasis of non-small cell lung cancer. The objective of this work is to investigate the expressions of chemokine receptor CCR7, hypoxiainducible factor 1α (HIF-1α and hypoxia inducible factor 2α (HIF-2α protein in non small cell lung cancer and the relationships of their expression, and to study the mechanism of CCR7 upregulation in NSCLC. Methods T he levels of expressions of CCR7, HIF-1α and HIF-2α protein were detected in 94 specimens of human primary non small cell lung cancer by immunohistochemical S-P method. Human lung adenocarcinoma cell line A549 cells were transfected by lipofection with HIF-1α siRNA、HIF-2α siRNA, the change of CCR7 was observed by RT-PCR and immunofluorescence staining. Correlations between the expression of CCR7 and HIF-1α, HIF-2α were respectively analyzed. Results Immunohistochemistry showed that CCR7 was distributed in cytoplasm and/or membrane of tumor cells, HIF-1α, HIF-2α was distributed in nucleus and/or cytoplasm of tumor cells. The levels of expressions of CCR7, HIF-1α and HIF-2α protein were found to be 75.53% (71/94, 54.25% (51/ 94 and 70.21% (66/94 in non small celllung cancer, respectively. the levels of expression of CCR7 protein were closely related to the clinical stages (P 0.05. Furthermore, A significant correlation were found among CCR7, Hif-1α and HIF-2α (r =0.272, P <0.01 (r=0.225, P <0.05. In addition, the expression of CCR7 mRNA and protein levels were decreased in the transfected specificHIF-1α, HIF-2αsiRNA group (P <0.05. Conclusion CCR7 expression is significantly associated with non small cell lung cancer invasion and metastasis. The upregulation of CCR7 is regulated by HIF-1α and HIF-2α in non small cell lung cancer.

  6. BOK displays cell death-independent tumor suppressor activity in non-small-cell lung carcinoma.

    Science.gov (United States)

    Moravcikova, Erika; Krepela, Evzen; Donnenberg, Vera S; Donnenberg, Albert D; Benkova, Kamila; Rabachini, Tatiana; Fernandez-Marrero, Yuniel; Bachmann, Daniel; Kaufmann, Thomas

    2017-11-15

    As the genomic region containing the Bcl-2-related ovarian killer (BOK) locus is frequently deleted in certain human cancers, BOK is hypothesized to have a tumor suppressor function. In the present study, we analyzed primary non-small-cell lung carcinoma (NSCLC) tumors and matched lung tissues from 102 surgically treated patients. We show that BOK protein levels are significantly downregulated in NSCLC tumors as compared to lung tissues (p < 0.001). In particular, we found BOK downregulation in NSCLC tumors of grades two (p = 0.004, n = 35) and three (p = 0.031, n = 39) as well as in tumors with metastases to hilar (pN1) (p = 0.047, n = 31) and mediastinal/subcarinal lymph nodes (pN2) (p = 0.021, n = 18) as opposed to grade one tumors (p = 0.688, n = 7) and tumors without lymph node metastases (p = 0.112, n = 51). Importantly, in lymph node-positive patients, BOK expression greater than the median value was associated with longer survival (p = 0.002, Mantel test). Using in vitro approaches, we provide evidence that BOK overexpression is inefficient in inducing apoptosis but that it inhibits TGFβ-induced migration and epithelial-to-mesenchymal transition (EMT) in lung adenocarcinoma-derived A549 cells. We have identified epigenetic mechanisms, in particular BOK promoter methylation, as an important means to silence BOK expression in NSCLC cells. Taken together, our data point toward a novel mechanism by which BOK acts as a tumor suppressor in NSCLC by inhibiting EMT. Consequently, the restoration of BOK levels in low-BOK-expressing tumors might favor the overall survival of NSCLC patients. © 2017 UICC.

  7. Erlotinib pretreatment improves photodynamic therapy of non-small cell lung carcinoma xenografts via multiple mechanisms

    Science.gov (United States)

    Gallagher-Colombo, Shannon M.; Miller, Joann; Cengel, Keith A.; Putt, Mary E.; Vinogradov, Sergei A.; Busch, Theresa M.

    2015-01-01

    Aberrant expression of the epidermal growth factor receptor (EGFR) is a common characteristic of many cancers including non-small cell lung carcinoma (NSCLC), head and neck squamous cell carcinoma, and ovarian cancer. While EGFR is currently a favorite molecular target for treatment of these cancers, inhibition of the receptor with small molecule inhibitors (i.e.- erlotinib) or monoclonal antibodies (i.e.- cetuximab) does not provide long-term therapeutic benefit as standalone treatment. Interestingly, we have found that addition of erlotinib to photodynamic therapy (PDT) can improve treatment response in typically erlotinib-resistant NSCLC tumor xenografts. Ninety-day complete response rates of 63% are achieved when erlotinib is administered in three doses before PDT of H460 human tumor xenografts, compared to 16% after PDT-alone. Similar benefit is found when erlotinib is added to PDT of A549 NCSLC xenografts. Improved response is accompanied by increased vascular shutdown, and erlotinib increases the in vitro cytotoxicity of PDT to endothelial cells. Tumor uptake of the photosensitizer (benzoporphyrin derivative monoacid ring A; BPD) is increased by the in vivo administration of erlotinib; nevertheless, this elevation of BPD levels only partially accounts for the benefit of erlotinib to PDT. Thus, pretreatment with erlotinib augments multiple mechanisms of PDT effect that collectively lead to large improvements in therapeutic efficacy. These data demonstrate that short-duration administration of erlotinib before PDT can greatly improve the responsiveness of even erlotinib-resistant tumors to treatment. Results will inform clinical investigation of EGFR-targeting therapeutics in conjunction with PDT. PMID:26054596

  8. Erlotinib Pretreatment Improves Photodynamic Therapy of Non-Small Cell Lung Carcinoma Xenografts via Multiple Mechanisms.

    Science.gov (United States)

    Gallagher-Colombo, Shannon M; Miller, Joann; Cengel, Keith A; Putt, Mary E; Vinogradov, Sergei A; Busch, Theresa M

    2015-08-01

    Aberrant expression of the epidermal growth factor receptor (EGFR) is a common characteristic of many cancers, including non-small cell lung carcinoma (NSCLC), head and neck squamous cell carcinoma, and ovarian cancer. Although EGFR is currently a favorite molecular target for the treatment of these cancers, inhibition of the receptor with small-molecule inhibitors (i.e., erlotinib) or monoclonal antibodies (i.e., cetuximab) does not provide long-term therapeutic benefit as standalone treatment. Interestingly, we have found that addition of erlotinib to photodynamic therapy (PDT) can improve treatment response in typically erlotinib-resistant NSCLC tumor xenografts. Ninety-day complete response rates of 63% are achieved when erlotinib is administered in three doses before PDT of H460 human tumor xenografts, compared with 16% after PDT-alone. Similar benefit is found when erlotinib is added to PDT of A549 NCSLC xenografts. Improved response is accompanied by increased vascular shutdown, and erlotinib increases the in vitro cytotoxicity of PDT to endothelial cells. Tumor uptake of the photosensitizer (benzoporphyrin derivative monoacid ring A; BPD) is increased by the in vivo administration of erlotinib; nevertheless, this elevation of BPD levels only partially accounts for the benefit of erlotinib to PDT. Thus, pretreatment with erlotinib augments multiple mechanisms of PDT effect that collectively lead to large improvements in therapeutic efficacy. These data demonstrate that short-duration administration of erlotinib before PDT can greatly improve the responsiveness of even erlotinib-resistant tumors to treatment. Results will inform clinical investigation of EGFR-targeting therapeutics in conjunction with PDT.

  9. Co-expression of ILT4/HLA-G in human non-small cell lung cancer correlates with poor prognosis and ILT4-HLA-G interaction activates ERK signaling.

    Science.gov (United States)

    Zhang, Yanwen; Zhao, Jianqiang; Qiu, Lijun; Zhang, Pei; Li, Juan; Yang, Dong; Wei, Xiaojuan; Han, Yali; Nie, Siyue; Sun, Yuping

    2016-08-01

    Non-small cell lung cancer (NSCLC) is the most common malignant tumor in the world, of which prognosis is generally poor due to insufficient mechanistic understanding. To explore the molecular pathogenesis of NSCLC, the co-expression of immunoglobulin-like transcript 4 (ILT4) and its ligand human leukocyte antigen G (HLA-G) in NSCLC tissues and cells were investigated. Here, we detected the expression of ILT4 and HLA-G in 81 tumor specimens from primary NSCLC patients, and we found that co-expression of ILT4/HLA-G was significantly associated with regional lymph node involvement, advanced stages, and the overall survival of patients. In NSCLC cell lines, HLA-G expression increased/decreased accordingly when ILT4 was up-/down-regulated, and ILT4 expression increased in a concentration-dependent manner via the stimulation of HLA-G fusion protein. Interestingly, HLA-G fusion protein could also up-regulate the phospho-ERK1/2 expression, which means the activation of extracellular signal-regulated kinase (ERK) signaling. All in all, our results indicate that the ILT4-HLA-G interaction might play an important role in NSCLC progression. Identification of ILT4 and HLA-G expression may provide an indicator to predict prognosis and guide prevention and treatment of NSCLC.

  10. Cyclin B1和survivin在非小细胞肺癌中的表达和意义%The expression and significance of cyclin B1 and survivin in human non-small cell lung cancer

    Institute of Scientific and Technical Information of China (English)

    Guosheng Li; Xuhan Liu; Daorong Zhang; Dong Liu; Zhiyong Li

    2011-01-01

    Objective: We studied the expression of cyclin B1 and survivin in human non-small cell lung cancer (NSCLC), and the relationship between such expression and clinicopathological features of NSCLC. Methods: One hundred cases of tissue specimen including NSCLC, neighboring noncancerous tissue and normal lung tissue were collected at random. These specimens were detected by immunohistochemical methods. Results: The expression of cyclin B1 and survivin showed significant difference (P 0.05) in NSCLC. Statistical significance was marked between different clinical stages of NSCLC and the expression of cyclin B1 and survivin (P < 0.05). Conclusion: The overexpression of cyclin B1 and survivin was found in NSCLC. The expression of cyclin B1 and survivin might be up-regulated during an early step of tumorigenesis and during the development of NSCLC. The progression of cell cycle could be efficiently connected with the control of apoptosis by the interrelations between the overexpression of cyclin B1 and that of survivin in NSCLC during the G2/M phase. The overexpression of cyclin B1 and survivin might be used as marker in showing the dividing and proliferating ability, and the inhibiting apoptosis ability (lengthening cell lifespan) of NSCLC. Moreover, the overexpression of cyclin B1 and survivin was associated with the clinic stages of NSCLC.

  11. MicroRNA-1228(*) inhibit apoptosis in A549 cells exposed to fine particulate matter.

    Science.gov (United States)

    Li, Xiaobo; Ding, Zhen; Zhang, Chengcheng; Zhang, Xin; Meng, Qingtao; Wu, Shenshen; Wang, Shizhi; Yin, Lihong; Pu, Yuepu; Chen, Rui

    2016-05-01

    Studies have reported associations between fine particulate matter (PM2.5) and respiratory disorders; however, the underlying mechanism is not completely clear owing to the complex components of PM2.5. microRNAs (miRNAs) demonstrate tremendous regulation to target genes, which are sensitive to exogenous stimulation, and facilitate the integrative understood of biological responses. Here, significantly modulated miRNA were profiled by miRNA microarray, coupled with bioinformatic analysis; the potential biological function of modulated miRNA were predicted and subsequently validated by cell-based assays. Downregulation of miR-1228-5p (miR-1228(*)) expression in human A549 cells were associated with PM2.5-induced cellular apoptosis through a mitochondria-dependent pathway. Further, overexpression of miR-1228(*) rescued the cellular damages induced by PM2.5. Thus, our results demonstrate that PM2.5-induced A549 apoptosis is initiated by mitochondrial dysfunction and miR-1228(*) could protect A549 cells against apoptosis. The involved pathways and target genes might be used for future mechanistic studies.

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

    Science.gov (United States)

    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.

  13. Clozapine Induces Autophagic Cell Death in Non-Small Cell Lung Cancer Cells

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    Yu-Chun Yin

    2015-02-01

    Full Text Available Background/Aims: Previous studies have shown that patients with schizophrenia have a lower incidence of cancer than the general population, and several antipsychotics have been demonstrated to have cytotoxic effects on cancer cells. However, the mechanisms underlying these results remain unclear. The present study aimed to investigate the effect of clozapine, which is often used to treat patients with refractory schizophrenia, on the growth of non-small cell lung carcinoma cell lines and to examine whether autophagy contributes to its effects. Methods: A549 and H1299 cells were treated with clozapine, and cell cytotoxicity, cell cycle and autophagy were then assessed. The autophagy inhibitor bafilomycin A1 and siRNA-targeted Atg7 were used to determine the role of autophagy in the effect of clozapine. Results: Clozapine inhibited A549 and H1299 proliferation and increased p21 and p27 expression levels, leading to cell cycle arrest. Clozapine also induced a high level of autophagy, but not apoptosis, in both cell lines, and the growth inhibitory effect of clozapine was blunted by treatment with the autophagy inhibitor bafilomycin A1 or with an siRNA targeting atg7. Conclusions: Clozapine inhibits cell proliferation by inducing autophagic cell death in two non-small cell lung carcinoma cell lines. These findings may provide insights into the relationship between clozapine use and the lower incidence of lung cancer among patients with schizophrenia.

  14. Dammarane-type saponins from heat-processed Gynostemma pentaphyllum show fortified activity against A549 cells.

    Science.gov (United States)

    Piao, Xiang-Lan; Wu, Qian; Yang, Jing; Park, Seo Young; Chen, Dao-Jin; Liu, Hui-Min

    2013-07-01

    An ethanol extract from heat-processed Gynostemma pentaphyllum showed more potent cytotoxic activity against human lung adenocarcinoma A549 cells than that of raw G. pentaphyllum. Four constituents were isolated from heat-processed G. pentaphyllum using resin HP-20, silica gel and reversed ODS column chromatography. They were identified by mass and NMR spectra as damulin A and damulin B, gypenoside L and gypenoside LI, respectively. To evaluate the efficacy of these four constituents, the MTT cytotoxicity assay was performed using A549 cells. Based on the structure of these four constituents, the results indicate that the hydroxyl group in C-2 and double bond in C20(21) and C20(22) positions are of importance in inhibition of A549 cell proliferation.

  15. MicroRNA-944 Affects Cell Growth by Targeting EPHA7 in Non-Small Cell Lung Cancer

    Science.gov (United States)

    Liu, Minxia; Zhou, Kecheng; Cao, Yi

    2016-01-01

    MicroRNAs (miRNAs) have critical roles in lung tumorigenesis and development. To determine aberrantly expressed miRNAs involved in non-small cell lung cancer (NSCLC) and investigate pathophysiological functions and mechanisms, we firstly carried out small RNA deep sequencing in NSCLC cell lines (EPLC-32M1, A549 and 801D) and a human immortalized cell line 16HBE, we then studied miRNA function by cell proliferation and apoptosis. cDNA microarray, luciferase reporter assay and miRNA transfection were used to investigate interaction between the miRNA and target gene. miR-944 was significantly down-regulated in NSCLC and had many putative targets. Moreover, the forced expression of miR-944 significantly inhibited the proliferation of NSCLC cells in vitro. By integrating mRNA expression data and miR-944-target prediction, we disclosed that EPHA7 was a potential target of miR-944, which was further verified by luciferase reporter assay and microRNA transfection. Our data indicated that miR-944 targets EPHA7 in NSCLC and regulates NSCLC cell proliferation, which may offer a new mechanism underlying the development and progression of NSCLC. PMID:27681722

  16. Thiazolidinediones enhance vascular endothelial growth factor expression and induce cell growth inhibition in non-small-cell lung cancer cells

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    Yoshizaki Yumiko

    2010-03-01

    Full Text Available Abstract Background It is known that thiazolidinediones are involved in regulating the expression of various genes, including the vascular endothelial growth factor (VEGF gene via peroxisome proliferator-activated receptor γ (PPARγ; VEGF is a prognostic biomarker for non-small-cell lung cancer (NSCLC. Methods In this study, we investigated the effects of troglitazone and ciglitazone on the mRNA expression of VEGF and its receptors in human NSCLC cell lines, RERF-LC-AI, SK-MES-1, PC-14, and A549. These mRNA expressions were evaluated by quantitative real-time reverse transcription-polymerase chain reaction (RT-PCR analysis. We also studied the effect of Je-11, a VEGF inhibitor, on the growth of these cells. Results In NSCLC cells, thiazolidinediones increased the mRNA expression of VEGF and neuropilin-1, but not that of other receptors such as fms-like tyrosine kinase and kinase insert domain receptor-1. Furthermore, the PPARγ antagonist GW9662 completely reversed this thiazolidinedione-induced increase in VEGF expression. Furthermore, the addition of VEGF inhibitors into the culture medium resulted in the reversal of thiazolidinedione-induced growth inhibition. Conclusions Our results indicated that thiazolidinediones enhance VEGF and neuropilin-1 expression and induce the inhibition of cell growth. We propose the existence of a pathway for arresting cell growth that involves the interaction of thiazolidinedione-induced VEGF and neuropilin-1 in NSCLC.

  17. COPD promotes migration of A549 lung cancer cells: the role of chemokine CCL21

    Directory of Open Access Journals (Sweden)

    Kuźnar-Kamińska B

    2016-05-01

    Full Text Available Barbara Kuźnar-Kamińska,1 Justyna Mikuła-Pietrasik,2 Patrycja Sosińska,2 Krzysztof Książek,2 Halina Batura-Gabryel1 1Department of Pulmonology, Allergology and Respiratory Oncology, 2Department of Pathophysiology, Poznań University of Medical Sciences, Poznań, Poland Abstract: Patients with COPD develop lung cancer more frequently than healthy smokers. At the same time, molecular mediators promoting various aspects of cancer cell progression are still elusive. In this report, we examined whether COPD can be coupled with increased migration of non-small-cell lung cancer cells A549 and, if so, whether this effect may be related to altered production and activity of chemokines CCL21, CXCL5, and CXCL12. The study showed that the migration of A549 cells through the polycarbonate membrane and basement membrane extract toward a chemotactic gradient elicited by serum from patients with COPD was markedly higher as compared with serum from healthy donors. The concentration of CCL21 and CXCL12, but not CXCL5, in serum from patients with COPD was also increased. Experiments in which CCL21- and CXCL12-dependent signaling was blocked revealed that increased migration of the cancer cells upon treatment with serum from patients with COPD was mediated exclusively by CCL21. Collectively, our results indicate that COPD may contribute to the progression of lung cancer via CCL21-dependent intensification of cancer cell migration. Keywords: chemokines, COPD, lung cancer, migration

  18. IL-6 promotes growth and epithelial-mesenchymal transition of CD133+ cells of non-small cell lung cancer.

    Science.gov (United States)

    Lee, Soo Ok; Yang, Xiaodong; Duan, Shanzhou; Tsai, Ying; Strojny, Laura R; Keng, Peter; Chen, Yuhchyau

    2016-02-09

    We examined IL-6 effects on growth, epithelial-mesenchymal transition (EMT) process, and metastatic ability of CD133+ and CD133- cell subpopulations isolated from three non-small cell lung cancer (NSCLC) cell lines: A549, H157, and H1299. We developed IL-6 knocked-down and scramble (sc) control cells of A549 and H157 cell lines by lentiviral infection system, isolated CD133+ and CD133- sub-populations, and investigated the IL-6 role in self-renewal/growth of these cells. IL-6 showed either an inhibitory or lack of effect in modulating growth of CD133- cells depending on intracellular IL-6 levels, but there was higher self-renewal ability of IL-6 expressing CD133+ cells than IL-6 knocked down cells, confirming the promoter role of IL-6 in CD133+ cells growth. We then examined tumor growth of xenografts developed from CD133+ cells of A549IL-6si vs. A549sc cell lines. Consistently, there was retarded growth of tumors developed from A549IL-6si, CD133+ cells compared to tumors originating from A549sc, CD133+ cells. The effect of IL-6 in promoting CD133+ self-renewal was due to hedgehog (Hhg) and Erk signaling pathway activation and higher Bcl-2/Bcl-xL expression. We also investigated whether IL-6 regulates the EMT process of CD133- and CD133+ cells differently. Expression of the EMT/metastasis-associated molecules in IL-6 expressing cells was higher than in IL-6 knocked down cells. Together, we demonstrated dual roles of IL-6 in regulating growth of CD133- and CD133+ subpopulations of lung cancer cells and significant regulation of IL-6 on EMT/metastasis increase in CD133+ cells, not in CD133- cells.

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

    Science.gov (United States)

    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.

  20. Ophiopogonin B induces apoptosis, mitotic catastrophe and autophagy in A549 cells.

    Science.gov (United States)

    Chen, Meijuan; Guo, Yuanyuan; Zhao, Ruolin; Wang, Xiaoxia; Jiang, Miao; Fu, Haian; Zhang, Xu

    2016-07-01

    Ophiopogonin B (OP-B), a saponin compound isolated from Radix Ophiopogon japonicus, was verified to inhibit cell proliferation in numerous non-small cell lung cancer (NSCLC) cells in our previous study. However, the precise mechanisms of action have remained unclear. In the present study, we mainly investigated the effects of OP-B on adenocarcinoma A549 cells to further elaborate the underlying mechanisms of OP-B in different NSCLC cell lines. Detection by high content screening (HCS) and TUNEL assay verified that OP-B induced apoptosis in this cell line, while detection of Caspase-3, Bcl-2 and Bax showed that OP-B induced cell death was caspase and mitochondrial independent. Further experiments showed that OP-B induced cell cycle arrest in the S and G2/M phases by inhibiting the expression of Myt1 and phosphorylation of Histone H3 (Ser10), which resulted in mitotic catastrophe in the cells. Transmission electron microscopy (TEM) observation of cell micro-morphology combined with detection of Atgs by western blot analysis showed that OP-B induced autophagy in this cell line. Autophagy inhibition by the lysosome inhibitor CQ or Beclin1-siRNA knockdown both attenuated cell viability, demonstrated that autophagy also being the vital reason resulted in cell death. More importantly, the xenograft model using A549 cells provided further evidence of the inhibition of OP-B on tumor proliferation. Immunohistochemistry detection of LC3 and Tunel assay both verified that high dose of OP-B (75 mg/kg) induced autophagy and apoptosis in vivo, and western blot detection of p-Histone H3 (Ser10), Survivin and XIAP further indicated the molecular mechanism of OP-B in vivo. As our findings revealed, multiple types of cell death overlapped in OP-B treated A549 cells, it displayed multitarget characteristics of the compounds extracted from the Chinese herbal, which may be used as candidate anticancer medicine in clinic.

  1. Acid-sensing ion channels contribute to the effect of extracellular acidosis on proliferation and migration of A549 cells.

    Science.gov (United States)

    Wu, Yu; Gao, Bo; Xiong, Qiu-Ju; Wang, Yu-Chan; Huang, Da-Ke; Wu, Wen-Ning

    2017-06-01

    Acid-sensing ion channels, a proton-gated cation channel, can be activated by low extracellular pH and involved in pathogenesis of some tumors such as glioma and breast cancer. However, the role of acid-sensing ion channels in the growth of lung cancer cell is unclear. In this study, we investigated the expression of acid-sensing ion channels in human lung cancer cell line A549 and their possible role in proliferation and migration of A549 cells. The results show that acid-sensing ion channel 1, acid-sensing ion channel 2, and acid-sensing ion channel 3 are expressed in A549 cells at the messenger RNA and protein levels, and acid-sensing ion channel-like currents were elicited by extracellular acid stimuli. Moreover, we found that acidic extracellular medium or overexpressing acid-sensing ion channel 1a promotes proliferation and migration of A549 cells. In addition psalmotoxin 1, a specific acid-sensing ion channel 1a inhibitor, or acid-sensing ion channel 1a knockdown can abolish the effect of acid stimuli on A549 cells. In addition, acid-sensing ion channels mediate increase of [Ca(2+)]i induced by low extracellular pH in A549 cells. All these results indicate that acid-sensing ion channel-calcium signal mediate lung cancer cell proliferation and migration induced by extracellular acidosis, and acid-sensing ion channels may serve as a prognostic marker and a therapeutic target for lung cancer.

  2. Adenovirus vector infection of non-small-cell lung cancer cells is a trigger for multi-drug resistance mediated by P-glycoprotein

    Energy Technology Data Exchange (ETDEWEB)

    Tomono, Takumi [Laboratory of Clinical Pharmacokinetics, Graduate School of Pharmaceutical Sciences, Takasaki University of Health and Welfare, 60 Nakaorui-machi, Takasaki-shi, Gunma 370-0033 (Japan); Kajita, Masahiro [Laboratory of Molecular Pharmaceutics and Technology, Faculty of Pharmacy, Takasaki University of Health and Welfare, 60 Nakaorui-machi, Takasaki-shi, Gunma 370-0033 (Japan); Yano, Kentaro [Laboratory of Biopharmaceutics, Faculty of Pharmacy, Takasaki University of Health and Welfare, 60 Nakaorui-machi, Takasaki-shi, Gunma 370-0033 (Japan); Ogihara, Takuo, E-mail: togihara@takasaki-u.ac.jp [Laboratory of Clinical Pharmacokinetics, Graduate School of Pharmaceutical Sciences, Takasaki University of Health and Welfare, 60 Nakaorui-machi, Takasaki-shi, Gunma 370-0033 (Japan)

    2016-08-05

    P-glycoprotein (P-gp) is an ATP-binding cassette protein involved in cancer multi-drug resistance (MDR). It has been reported that infection with some bacteria and viruses induces changes in the activities of various drug-metabolizing enzymes and transporters, including P-gp. Although human adenoviruses (Ad) cause the common cold, the effect of Ad infection on MDR in cancer has not been established. In this study, we investigated whether Ad infection is a cause of MDR in A549, H441 and HCC827 non-small-cell lung cancer (NSCLC) cell lines, using an Ad vector system. We found that Ad vector infection of NSCLC cell lines induced P-gp mRNA expression, and the extent of induction was dependent on the number of Ad vector virus particles and the infection time. Heat-treated Ad vector, which is not infectious, did not alter P-gp mRNA expression. Uptake experiments with doxorubicin (DOX), a P-gp substrate, revealed that DOX accumulation was significantly decreased in Ad vector-infected A549 cells. The decrease of DOX uptake was blocked by verapamil, a P-gp inhibitor. Our results indicated that Ad vector infection of NSCLC cells caused MDR mediated by P-gp overexpression. The Ad vector genome sequence is similar to that of human Ad, and therefore human Ad infection of lung cancer patients may lead to chemoresistance in the clinical environment. -- Highlights: •Adenovirus vector infection induced P-gp mRNA expression in three NSCLC cell lines. •Adenovirus vector infection enhanced P-gp-mediated doxorubicin efflux from the cells. •The increase of P-gp was not mediated by nuclear receptors (PXR, CAR) or COX-2.

  3. 下调βIII-tubulin逆转肺腺癌A549/Taxol细胞株紫杉醇耐药%Down-regulatedβIII-tubulin Expression Can Reverse Paclitaxel Resistance in A549/Taxol Cells Lines

    Institute of Scientific and Technical Information of China (English)

    禚银玲; 郭其森

    2014-01-01

    Background and objective Chemotherapy drug resistance is the primary causes of death in patients with pulmonary carcinoma which make tumor recurrence or metastasis.β-tubulin is the main cell targets of anti-microtubule drug. Increased expression ofβIII-tubulin has been implicated in non-small cell lung cancer (NSCLC) cell lines. To explore the relationship among the expression level ofβIII-tubulin and the sensitivity of A549/Taxolcell lines to Taxol and cell cycles and cell apoptosis by RNA interference-mediated inhibition ofβIII-tubulin in A549/Taxol cells. Methods hTree pairs of siRNA targetdβIII-tubulin were designed and prepared, which were transfected into A549/Taxol cells using LipofectamineTM 2000. We detected the expression ofβIII-tubulin mRNA using Real-time lfuorescence qRT-PCR. Tedhen we selected the most eff-cient siRNA by the expression ofβIII-tubulin mRNA in transfected group.βIII-tubulin protein level were mesured by Western blot. hTe taxol sensitivity in transfected group were evaluated by MTT assay. And the cell apoptosis and cell cycles were deter-mined by lfow cytometry. Results βIII-tubulin mRNA levels in A549/Taxol cells were signiifcantly decreased in transfected grop by Real-time qRT-PCR than control groups. AndβIII-tubulin siRNA-1 sequence showed the highest transfection eff-ciency, which was (87.73±4.87)%(P<0.01);Western blot results showed that the expressional level of BIII tublin protein was signiifcantly down-reulated in the transfectant cells than thant in the control cells. By MTT assay, we showed that the inhibition ratio of Taxol to A549/Taxol cells transfeced was higher than that of control group (51.77±4.60)%(P<0.01). hTe early apopto-sis rate of A549/Taxol cells in transfected group were signiifcantly higher than that of control group (P<0.01);G2-M content in taxol group obviously increased than untreated samples by the cell cycle (P<0.05). Conclusion βIII-tubulin down-regulated signiifcantly sensitized NSCLC A549

  4. Matrine reduces proliferation of human lung cancer cells by inducing apoptosis and changing miRNA expression profiles.

    Science.gov (United States)

    Liu, Yong-Qi; Li, Yi; Qin, Jie; Wang, Qian; She, Ya-Li; Luo, Ya-Li; He, Jian-Xin; Li, Jing-Ya; Xie, Xiao-Dong

    2014-01-01

    Matrine, a main active component extracted from dry roots of Sophora flavecens , has been reported to exert antitumor effects on A549 human non-small lung cancer cells, but its mechanisms of action remain unclear. To determine effects of matrine on proliferation of A549 cells and assess possible mechanisms, MTT assays were employed to detect cytotoxicity, along with o flow cytometric analysis of DNA content of nuclei of cells following staining with propidium iodide to analyze cell cycle distribution. Western blotting was performed to determined expression levels of Bax, Bcl-2, VEGF and HDAC1, while a microarray was used to assessed changes of miRNA profiles. In the MTT assay, matrine suppressed growth of human lung cancer cell A549 in a dose- and time- dependent manner at doses of 0.25-2.5 mg/ml for 24h, 48h or 72h. Matrine induced cell cycle arrest in G0/G1 phase and decreased the G2/M phase, while down-regulating the expression of Bcl2 protein, leading to a reduction in the Bcl-2/Bax ratio. In addition, matrine down regulated the expression level of VEGF and HDAC1 of A549 cells. Microarray analysis demonstrated that matrine altered the expression level of miRNAs compared with untreated control A549 cells. In conclusion, matrine could inhibit proliferation of A549 cells, providing useful information for understanding anticancer mechanisms.

  5. Construction of a CD147 Lentiviral Expression Vector and Establishment of Its Stably Transfected A549 Cell Line

    Directory of Open Access Journals (Sweden)

    Shaoxing YANG

    2012-12-01

    Full Text Available Background and objective CD147, a type of transmembrane glycoprotein embedded on the surface of tumor cells, can promote tumor invasion and metastasis. This aim of this study is to construct a CD147 lentiviral expression vector, establish its stably transfected A549 cell line, and observe the effect of CD147 on MMP-9 proliferation as well as on the invasive ability of human lung adenocarcinoma cells. Methods Full-length CD147 gene was amplified by real-time polymerase chain reaction (RT-PCR, inserted into a pEGFP vector to construct pEGFP-CD147 and pEGFP vectors, and then transfected into 293FT cells to precede the lentivirus equipment package. Subsequently, we collected the lentivirus venom to infect the A549 cells and establish a stable, overexpressed cell line named A549-CD147. The mRNA expression of MMP-9 was examined by RT-PCR. The proliferation and invasive ability of the human lung cancer cells before and after transfection were examined by the CCK-8 and Transwell methods. Results A CD147 lentiviral expression vector (pEGFP-CD147 was successfully constructed by restrictive enzyme digestion and plasmid sequencing. RT-PCR and Western blot analyses revealed increased mRNA and protein expression of CD147 gene in cells transfected with pEGFP-CD147 compared with the control groups. Therefore, the A549-CD147 cell line was successfully established through the experiment. The mRNA expression of MMP-9 also significantly increased after the upregulation of CD147 expression. Meanwhile, CCK-8 and Transwell assays indicated that the proliferation and invasive ability significantly increased in the A549-CD147 cells. Conclusion A lentiviral CD147 expression vector and its A549 cell line (A549-CD14 were successfully constructed. CD147 overexpression upregulated the protein expression of MMP-9, and strengthened the proliferation and invasive ability of human lung adenocarcinoma cells.

  6. Dual roles of extracellular signal-regulated kinase (ERK) in quinoline compound BPIQ-induced apoptosis and anti-migration of human non-small cell lung cancer cells.

    Science.gov (United States)

    Fong, Yao; Wu, Chang-Yi; Chang, Kuo-Feng; Chen, Bing-Hung; Chou, Wan-Ju; Tseng, Chih-Hua; Chen, Yen-Chun; Wang, Hui-Min David; Chen, Yeh-Long; Chiu, Chien-Chih

    2017-01-01

    2,9-Bis[2-(pyrrolidin-1-yl)ethoxy]-6-{4-[2-(pyrrolidin-1-yl)ethoxy] phenyl}-11H-indeno[1,2-c]quinoline-11-one (BPIQ), is a synthetic quinoline analog. A previous study showed the anti-cancer potential of BPIQ through modulating mitochondrial-mediated apoptosis. However, the effect of BPIQ on cell migration, an index of cancer metastasis, has not yet been examined. Furthermore, among signal pathways involved in stresses, the members of the mitogen-activated protein kinase (MAPK) family are crucial for regulating the survival and migration of cells. In this study, the aim was to explore further the role of MAPK members, including JNK, p38 and extracellular signal-regulated kinase (ERK) in BPIQ-induced apoptosis and anti-migration of human non-small cell lung cancer (NSCLC) cells. Western Blot assay was performed for detecting the activation of MAPK members in NSCLC H1299 cells following BPIQ administration. Cellular proliferation was determined using a trypan blue exclusion assay. Cellular apoptosis was detected using flow cytometer-based Annexin V/propidium iodide dual staining. Cellular migration was determined using wound-healing assay and Boyden's chamber assay. Zymography assay was performed for examining MMP-2 and -9 activities. The assessment of MAPK inhibition was performed for further validating the role of JNK, p38, and ERK in BPIQ-induced growth inhibition, apoptosis, and migration of NSCLC cells. Western Blot assay showed that BPIQ treatment upregulates the phosphorylated levels of both MAPK proteins JNK and ERK. However, only ERK inhibitor rescues BPIQ-induced growth inhibition of NSCLC H1299 cells. The results of Annexin V assay further confirmed the pro-apoptotic role of ERK in BPIQ-induced cell death of H1299 cells. The results of wound healing and Boyden chamber assays showed that sub-IC50 (sub-lethal) concentrations of BPIQ cause a significant inhibition of migration in H1299 cells accompanied with downregulating the activity of MMP-2 and -9, the

  7. Anti-Tumor Effect of Heat Shock Protein 70-Peptide Complexes on A-549 Cells

    Institute of Scientific and Technical Information of China (English)

    2007-01-01

    Objective: To investigate the anti-tumor immunity in vitro of heat shock protein 70-peptide complexes (HSP70-PC) from human lung cancer tissue. Methods: HSP70-PC was purified from lung tumor tissues and corresponding non-tumor lung samples with the methods of ADP-affinity chromatography, DEAE ion-exchange chromatography and Western-blot. The activation and proliferation of PBMC induced by different HSP70-PC and tumor cytotoxic reactivity to A549 cells in vitro were measured by the MTT cell proliferation assay. Results: The purified HSP70-PC had a very high purity found by SDS-PAGE and Western-blot. Human lymphocytes were sensitized efficiently by HSP70 preparation purified from lung cancer tissues and a definite cytotoxicity to A-549 cells was observed. There was significant difference with HSP70-PC purified from lung cancer, compared with the control group (P<0.001). Conclusion: High purity of HSP70-PC could be achieved from tumor tissues in this study. HSP70-PC purified from human tumor tissues can induce anti-tumor immunity in vitro mainly implemented by eliciting CTL immunity.

  8. Apoptosis inducing ability of silver decorated highly reduced graphene oxide nanocomposites in A549 lung cancer.

    Science.gov (United States)

    Khan, Merajuddin; Khan, Mujeeb; Al-Marri, Abdulhadi H; Al-Warthan, Abdulrahman; Alkhathlan, Hamad Z; Siddiqui, Mohammed Rafiq H; Nayak, Vadithe Lakshma; Kamal, Ahmed; Adil, Syed F

    2016-01-01

    Recently, graphene and graphene-based materials have been increasingly used for various biological applications due to their extraordinary physicochemical properties. Here, we demonstrate the anticancer properties and apoptosis-inducing ability of silver doped highly reduced graphene oxide nanocomposites synthesized by employing green approach. These nano composites (PGE-HRG-Ag) were synthesized by using Pulicaria glutinosa extract (PGE) as a reducing agent and were evaluated for their anticancer properties against various human cancer cell lines with tamoxifen as the reference drug. A correlation between the amount of Ag nanoparticles on the surface of highly reduced graphene oxide (HRG) and the anticancer activity of nanocomposite was observed, wherein an increase in the concentration of Ag nanoparticles on the surface of HRG led to the enhanced anticancer activity of the nanocomposite. The nanocomposite PGE-HRG-Ag-2 exhibited more potent cytotoxicity than standard drug in A549 cells, a human lung cancer cell line. A detailed investigation was undertaken and Fluorescence activated cell sorting (FACS) analysis demonstrated that the nanocomposite PGE-HRG-Ag-2 showed G0/G1 phase cell cycle arrest and induced apoptosis in A549 cells. Studies such as, measurement of mitochondrial membrane potential, generation of reactive oxygen species (ROS) and Annexin V-FITC staining assay suggested that this compound induced apoptosis in human lung cancer cells.

  9. TRAIL-coated lipid-nanoparticles overcome resistance to soluble recombinant TRAIL in non-small cell lung cancer cells

    Science.gov (United States)

    De Miguel, Diego; Gallego-Lleyda, Ana; María Ayuso, José; Erviti-Ardanaz, Sandra; Pazo-Cid, Roberto; del Agua, Celia; José Fernández, Luis; Ochoa, Ignacio; Anel, Alberto; Martinez-Lostao, Luis

    2016-05-01

    Purpose. Non-small cell lung cancer (NSCLC) is one the types of cancer with higher prevalence and mortality. Apo2-Ligand/TRAIL is a TNF family member able to induce apoptosis in tumor cells but not in normal cells. It has been tested in clinical trials against different types of human cancer including NSCLC. However, results of clinical trials have shown a limited efficacy of TRAIL-based therapies. Recently we have demonstrated that artificial lipid nanoparticles coated with bioactive Apo2L/TRAIL (LUV-TRAIL) greatly improved TRAIL cytotoxic ability being capable of killing chemoresistant hematological cancer cells. In the present work we have extended the study to NSCLC. Methods/patients. LUV-TRAIL-induced cytotoxicity was assessed on different NSCLC cell lines with different sensitivity to soluble TRAIL and on primary human tumor cells from three patients suffering from NSCLC cancer. We also tested LUV-TRAIL-cytotoxic ability in combination with several anti-tumor agents. Results. LUV-TRAIL exhibited a greater cytotoxic effect compared to soluble TRAIL both in A549 cells and primary human NSCLC cells. LUV-TRAIL-induced cell death was dependent on caspase-8 and caspase-3 activation. Moreover, combination of LUV-TRAIL with other anti-tumor agents such as flavopiridol, and SNS-032 clearly enhanced LUV-TRAIL-induced cytotoxicity against NSCLC cancer cells. Conclusion. The novel formulation of TRAIL based on displaying it on the surface of lipid nanoparticles greatly increases its anti-tumor activity and has clinical potential in cancer treatment.

  10. Intratumoral injection of taxol in vivo suppresses A549 tumor showing cytoplasmic vacuolization.

    Science.gov (United States)

    Wang, Chaoyang; Chen, Tongsheng

    2012-04-01

    Based on our recent in vitro studies, this report was designed to explore the mechanism by which high concentration of taxol (70 µM) induced paraptosis-like cell death in human lung carcinoma (A549) cells, and to evaluate the therapeutic efficacy of taxol using A549 tumor-bearing mice in vivo. Exposure of cells to taxol induced time-dependent cytotoxicity and cytoplasmic vacuolization without the involvement of Bax, Bak, Mcl-1, Bcl-XL, and caspase-3. Although taxol treatment induced activating transcription factor 6 (ATF6) cleavage indicative of endoplasmic reticulum (ER) stress, silencing ATF6 by shATF6 did not prevent taxol-induced both cytotoxcity and cytoplasmic vacuolization, suggesting that taxol-induced cytoplasmic vacuolization and cell death were not due to ER stress. Moreover, taxol-treated cells did not show DNA fragmentation and loss of mitochondrial membrane potential, the typical characteristics of apoptosis. In addition, taxol-induced cytoplasmic vacuolization did not show the cellular lysis, the characteristics of oncosis, and positive of β-galactosidase, the characteristic of senescence, indicating that taxol induced paraptosis-like cell death is neither oncosis nor senescence. Moreover, our in vivo data showed that intratumoral injection of taxol (50 mg/kg) in A549 tumor xenograft mice on day 1 and day 19 potently suppressed tumor growth showing significant ER vacuolization without toxicity. In conclusion, high concentration of taxol exhibits a significant anticancer activity by inducing paraptosis-like cell death in vitro and in vivo, without significant toxicity, suggesting a promising therapeutic strategy for apoptosis-resistance cancer by inducing ER vacuolization.

  11. JAM-C promotes lymphangiogenesis and nodal metastasis in non-small cell lung cancer.

    Science.gov (United States)

    Hao, SongNan; Yang, YanMei; Liu, Yan; Yang, ShuCai; Wang, Geng; Xiao, JianBing; Liu, HuiDong

    2014-06-01

    This study aims to investigate lymphatic metastasis-related genes in non-small cell lung carcinomas (NSCLC). NSCLC tissue was analyzed for expression of junctional adhesion molecule-C (JAM-C) protein. Our data revealed novel associations between JAM-C overexpression in primary tumors and lymphatic microvessel density (LMVD), lymph node metastasis, and poorer overall survival and recurrence-free survival. We used the highly metastatic human lung adenocarcinoma cell line Anip973 and its parental line AGZY83-a, which has a low metastatic capacity, in vivo and vitro. We found that JAM-C played an important role in different metastasis capacity of lymph node. JAM-C affected tumor growth, LNM, JAM-C, VEGF-C, vasculature, and ERK1/2 phosphorylation (p-ERK1/2). β1 integrin was involved in lymph node metastasis. Moreover, JAM-C knockdown in highly metastatic Anip973 decreased cell migration in scratch-wound assays. The JAM-C knockdown in Anip973 cells and JAM-C cDNA in AGZY83-a cells regulated the vascular endothelial growth factor C (VEGF-C) expression. Immunofluorescence showed that blocked VEGF-C expression in JAM-C shRNA Anip973 cells were restored after JAM-C treatment. JAM-C-induced VEGF-C in JAM-C cDNA AGZY83-a cells was also effectively inhibited by treatment with an antibody specifically against JAM-C. Use of media from Anip973 cells, AGZY83-a, and A549cells lung cancer cells that overexpressed or downregulated JAM-C was demonstrated to affect activity of VEGF-C-induced β1 integrin subunit or ERK activity in human dermal lymphatic endothelial cells (HDLEC) treated with VEGF-C or inhibitory antibody to JAM-C. Overall, these results indicate that JAM-C could mediate metastasis as it contributes to VEGF-C expression in cancer cells. JAM-C affects β1and ERK activation in HDLEC, thus promoting lymphangiogenesis and nodal metastasis. Our findings indicate that JAM-C may be a therapeutic target for preventing and treating lymphatic metastases.

  12. DNA Repair Genes ERCC1 and BRCA1 Expression in Non-Small Cell Lung Cancer Chemotherapy Drug Resistance.

    Science.gov (United States)

    Wang, Shuai; Liu, Feng; Zhu, Jingyan; Chen, Peng; Liu, Hongxing; Liu, Qi; Han, Junqing

    2016-06-12

    BACKGROUND Surgery combined with chemotherapy is an important therapy for non-small cell lung cancer (NSCLC). However, chemotherapy drug resistance seriously hinders the curative effect. Studies show that DNA repair genes ERCC1 and BRCA1 are associated with NSCLC chemotherapy, but their expression and mechanism in NSCLC chemotherapy drug-resistant cells has not been elucidated. MATERIAL AND METHODS NSCLC cell line A549 and drug resistance cell line A549/DDP were cultured. Real-time PCR and Western blot analyses were used to detect ERCC1 and BRCA1 mRNA expression. A549/DDP cells were randomly divided into 3 groups: the control group; the siRNA-negative control group (scramble group); and the siRNA ERCC1 and BRCA1siRNA transfection group. Real-time PCR and Western blot analyses were used to determine ERCC1 and BRCA1 mRNA and protein expression. MTT was used to detect cell proliferation activity. Caspase 3 activity was tested by use of a kit. Western blot analysis was performed to detect PI3K, AKT, phosphorylated PI3K, and phosphorylated AKT protein expression. RESULTS ERCC1 and BRCA1 were overexpressed in A549/DDP compared with A549 (P<0.05). ERCC1 and BRCA1siRNA transfection can significantly reduce ERCC1 and BRCA1 mRNA and protein expression (P<0.05). Downregulating ERCC1 and BRCA1 expression obviously inhibited cell proliferation and increased caspase 3 activity (P<0.05). Downregulating ERCC1 and BRCA1 significantly decreased PI3K and AKT phosphorylation levels (P<0.05). CONCLUSIONS ERCC1 and BRCA1 were overexpressed in NSCLC drug-resistant cells, and they regulated lung cancer occurrence and development through the phosphorylating PI3K/AKT signaling pathway.

  13. Development and in vitro evaluation of core-shell type lipid-polymer hybrid nanoparticles for the delivery of erlotinib in non-small cell lung cancer.

    Science.gov (United States)

    Mandal, Bivash; Mittal, Nivesh K; Balabathula, Pavan; Thoma, Laura A; Wood, George C

    2016-01-01

    Core-shell type lipid-polymer hybrid nanoparticles (CSLPHNPs) have emerged as a multifunctional drug delivery platform. The delivery system combines mechanical advantages of polymeric core and biomimetic advantages of the phospholipid shell into a single platform. We report the development of CSLPHNPs composed of the lipid monolayer shell and the biodegradable polymeric core for the delivery of erlotinib, an anticancer drug, clinically used to treat non-small cell lung cancer (NSCLC). Erlotinib loaded CSLPHNPs were prepared by previously reported single-step sonication method using polycaprolactone (PCL) as the biodegradable polymeric core and phospholipid-shell composed of hydrogenated soy phosphatidylcholine (HSPC) and 1,2-distearoyl-sn-glycero-3-phosphoethanolamine-N-[methoxy(polyethylene glycol)-2000 (DSPE-PEG2000). Erlotinib loaded CSLPHNPs were characterized for physicochemical properties including mean particle size, polydispersity index (PDI), zeta potential, morphology, thermal and infrared spectral analysis, drug loading, in vitro drug release, in vitro serum stability, and storage stability. The effect of critical formulation and process variables on two critical quality attributes (mean particle size and drug entrapment efficiency) of erlotinib loaded CSLPHNPs was studied and optimized. In addition, in vitro cellular uptake, luminescent cell viability assay and colony formation assay were performed to evaluate efficacy of erlotinib loaded CSLPHNPs in A549 cells, a human lung adenocarcinoma cell line. Optimized erlotinib loaded CSLPHNPs were prepared with mean particle size of about 170nm, PDI<0.2, drug entrapment efficiency of about 66% with good serum and storage stability. The evaluation of in vitro cellular efficacy results indicated enhanced uptake and efficacy of erlotinib loaded CSLPHNPs compared to erlotinib solution in A549 cells. Therefore, CSLPHNPs could be a potential delivery system for erlotinib in the therapy of NSCLC.

  14. AZD5438, an Inhibitor of Cdk1, 2, and 9, Enhances the Radiosensitivity of Non-Small Cell Lung Carcinoma Cells

    Energy Technology Data Exchange (ETDEWEB)

    Raghavan, Pavithra; Tumati, Vasu; Yu Lan [Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, Texas (United States); Chan, Norman [Departments of Medical Biophysics and Radiation Oncology, Princess Margaret Hospital, University Health Network, University of Toronto, Ontario (Canada); Tomimatsu, Nozomi [Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, Texas (United States); Burma, Sandeep [Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, Texas (United States); Simmons Comprehensive Cancer Center, Dallas, Texas (United States); Bristow, Robert G. [Departments of Medical Biophysics and Radiation Oncology, Princess Margaret Hospital, University Health Network, University of Toronto, Ontario (Canada); Saha, Debabrata, E-mail: debabrata.saha@utsouthwestern.edu [Department of Radiation Oncology, University of Texas Southwestern Medical Center, Dallas, Texas (United States); Simmons Comprehensive Cancer Center, Dallas, Texas (United States)

    2012-11-15

    Purpose: Radiation therapy (RT) is one of the primary modalities for treatment of non-small cell lung cancer (NSCLC). However, due to the intrinsic radiation resistance of these tumors, many patients experience RT failure, which leads to considerable tumor progression including regional lymph node and distant metastasis. This preclinical study evaluated the efficacy of a new-generation cyclin-dependent kinase (Cdk) inhibitor, AZD5438, as a radiosensitizer in several NSCLC models that are specifically resistant to conventional fractionated RT. Methods and Materials: The combined effect of ionizing radiation and AZD5438, a highly specific inhibitor of Cdk1, 2, and 9, was determined in vitro by surviving fraction, cell cycle distribution, apoptosis, DNA double-strand break (DSB) repair, and homologous recombination (HR) assays in 3 NSCLC cell lines (A549, H1299, and H460). For in vivo studies, human xenograft animal models in athymic nude mice were used. Results: Treatment of NSCLC cells with AZD5438 significantly augmented cellular radiosensitivity (dose enhancement ratio rangeing from 1.4 to 1.75). The degree of radiosensitization by AZD5438 was greater in radioresistant cell lines (A549 and H1299). Radiosensitivity was enhanced specifically through inhibition of Cdk1, prolonged G{sub 2}-M arrest, inhibition of HR, delayed DNA DSB repair, and increased apoptosis. Combined treatment with AZD5438 and irradiation also enhanced tumor growth delay, with an enhancement factor ranging from 1.2-1.7. Conclusions: This study supports the evaluation of newer generation Cdk inhibitors, such as AZD5438, as potent radiosensitizers in NSCLC models, especially in tumors that demonstrate variable intrinsic radiation responses.

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

    Science.gov (United States)

    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

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

    Science.gov (United States)

    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.

  17. CCDC106 promotes non-small cell lung cancer cell proliferation.

    Science.gov (United States)

    Zhang, Xiupeng; Zheng, Qin; Wang, Chen; Zhou, Haijing; Jiang, Guiyang; Miao, Yuan; Zhang, Yong; Liu, Yang; Li, Qingchang; Qiu, Xueshan; Wang, Enhua

    2017-04-18

    Coiled-coil domain containing (CCDC) family members enhance tumor cell proliferation, and high CCDC protein levels correlate with unfavorable prognoses. Limited research demonstrated that CCDC106 may promote the degradation of p53/TP53 protein and inhibit its transactivity. The present study demonstrated that CCDC106 expression correlates with advanced TNM stage (P = 0.008), positive regional lymph node metastasis (P CCDC106-low and CCDC106-high expressing cell lines, respectively. CCDC106 overexpression promoted A549 cell proliferation and xenograft tumor growth in nude mice, while siRNA-mediated CCDC106 knockdown inhibited H1299 cell proliferation. CCDC106 promoted AKT phosphorylation and upregulated the cell cycle-regulating proteins Cyclin A2 and Cyclin B1. Cell proliferation promoted by CCDC106 via Cyclin A2 and Cyclin B1 was rescued by treatment with the AKT inhibitor, LY294002. Our studies revealed that CCDC106 is associated with non-small cell lung cancer progression and unfavorable prognosis. CCDC106 enhanced Cyclin A2 and Cyclin B1 expression and promoted A549 and H1299 cell proliferation, which depended on AKT signaling. These results suggest that CCDC106 may be a novel target for lung cancer treatment.

  18. 地塞米松诱导人肺腺癌A549细胞对紫杉醇耐药性及Bcl-xL基因表达的影响%Influence of lung adenocarcinoma A549 cells induced by dexamethasone on the drug resistance of PTX and expression of Bcl-xL gene

    Institute of Scientific and Technical Information of China (English)

    康马飞; 李林凤

    2014-01-01

    Objective To observe the changes of the drug resistance of lung adenocarcinoma A549 cells and the expres-sion of mRNA and protein of Bcl-xL in human lung adenocarcinoma A549 cell after dexamethasone(DEX) pretreatment in dif-ferent concentrations and intervention with paclitaxel(PTX),and to explore the molecular mechanism of DEX-induced A549 cell to the drug resistance of PTX. Methods The cell viability rate of lung adenocarcinoma A549 cells was determined by MTT assay after treatment of different concentrations of PTX,and to screen the half inhibitory concentration(IC50) of PTX. The cell viability rate of A549 cells which pretreated with different concentrations of DEX and different concentrations of PTX was determined by MTT assay,and the expression level of mRNA and protein of Bcl-xL in the A549 cells were determined by reverse transcriptase-poly merase chain reaction(RT-PCR) and Western blotting. Results After being pretreated with DEX in different concentrations, A549 cells were induced to resist to PTX,and the rate of resistance increased gradually with the increasing concentrations of DEX and the expression level of Bcl-xL gene and protein also increased gradually with the increasing of DEX concentrations. Conclu-sions DEX can induce resistance to PTX in lung adenocarcinoma A549 cells ,whose mechanism might be involved in increase of Bcl-xL gene(anti-apoptosis gene) in DEX-induced lung adenocarcinoma A549 cells.%目的:观察紫杉醇(PTX)干预不同浓度地塞米松(DEX)预处理人肺腺癌A549细胞后的耐药情况和Bcl-xL基因及蛋白表达变化,探讨DEX诱导A549细胞对PTX产生耐药的分子机制。方法采用四甲基偶氮唑蓝比色法(MTT法)测定不同浓度PTX作用于A549细胞后的细胞存活率,筛选出PTX的半数抑制浓度(IC50);用不同浓度DEX预处理A549细胞后,再给予不同浓度PTX作用于A549细胞,用MTT法测定细胞存活率,逆转录-聚合酶链反应和蛋白质印迹法

  19. Inhibition Mechanism of Novel Pyrazolo[1,5-a]pyrazin-4(5H)-one Derivatives Against Proliferation of A549 and H322 Cancer Cells

    Institute of Scientific and Technical Information of China (English)

    Jin-hui Shao; Gui-hua Feng

    2015-01-01

    Objective To explore the inhibition mechanism and safety of pyrazolo[1,5-a]pyrazin-4(5H)-one derivatives against proliferation of human lung cancer A549 cells, H322 cells, and human umbilical vein endothelial cell (HUVEC). Methods Cells were treated with 40μmol/L of the ppo3a, ppo3b, ppo3i, and 0.1% DMSO (control) for 48 hours, respectively. Apoptosis was determined by Hoechst 33258 staining assay in H322 and A549 cells. Cell cycle distribution was determined by flow cytometry analysis in A549 cell. LC3-II, p53, and heat shock protein (HSP) 70 protein levels were detected by Western blotting in A549 cells treated with ppo3b for 48 hours. The morphology and viability of HUVEC were observed by inverted microscope and sulforhodamine B (SRB) assay. Results Ppo3a, ppo3b, and ppo3i significantly induced apoptosis in H322 and A549 cells. A strong G1-phase arrest was concomitant with the growth inhibitory effect on A549 cells. Ppo3b effectively elevated the p53 protein level, but significantly reduced the HSP70 protein level. There were no significantly inhibitory effect on the morphology and viability of HUVEC when treated with ppo3a, ppo3b, and ppo3i. Conclusions ppo3a, ppo3b, and ppo3i could inhibit H322 proliferation through apoptosis and inhibit A549 through apoptosis and G1-phase arrest. The protein p53 and HSP70 might involve in the inhibition effects. These derivatives might be a clue to find effective and safe drug for lung cancers.

  20. 沉默COX-2抑制A549细胞的恶性增殖%COX-2 silencing inhibits cell proliferation in A549 cell

    Institute of Scientific and Technical Information of China (English)

    Weiying Li; Wentao Yue; Lina Zhang; Xiaoting Zhao; Li Ma; Xuehui Yang; Chunyan Zhang; Yue Wang; Meng Gu

    2011-01-01

    Objective: The aim of this study was to explore the effects on malignant proliferation of A549 cell by silencing cyclooxygenase (COX)-2. Methods: In the present study, we constructed three siRNA vectors producing small interference RNA. The siRNA vectors and the vacant vectors were transfected into A549 cell with lipofectamine respectively and the transfected cell strains were constructed. The change of COX-2 expression levels was examined by Western blot and RT-PCR. The effects on the proliferation of lung cancer cells were studied by cell growth curve, clonogenic assay and xenograft assays. Results: The siRNA expression vectors produced marked effects in A549 cell but the inhibited effects were different. The effect of psi-10 was best and the mRNA and protein levels of COX-2 reduced 61.2% and 56.2% respectively in A549-si10 cell in contrast to the control.The growth of A549 cell slowed and the colony formation rate reduced after silencing COX-2. In xenograft assays, the growth speeds of tumor became slow and the numbers of tumor reduced after silencing COX-2. Conclusion: The si10 target of COX-2 has the best silencing effect in A549 cell and the best inhibition effect on malignant proliferation of A549 cell in vivo and in vitro.

  1. Suppression of SCARA5 by Snail1 is essential for EMT-associated cell migration of A549 cells

    OpenAIRE

    Liu, J.; Hu, G; Chen, D.; Gong, A-Y; Soori, G. S.; Dobleman, T J; Chen, X-M

    2013-01-01

    Accumulating evidence indicates that epithelial-to-mesenchymal transition (EMT) might be a key event for cancer progression. The upregulation of Snail1, one of the most extensively studied EMT regulators, has been implicated in cancer metastasis, but the underlying mechanisms remain unclear. This study aims to identify that Snail1 targets regulating EMT-associated cancer cell migration. Human lung carcinoma A549 cells were treated with transforming growth factor beta 1 (TGF-β1), and EMT-assoc...

  2. Treatment Options by Stage (Non-Small Cell Lung Cancer)

    Science.gov (United States)

    ... Lung Cancer Screening Research Non-Small Cell Lung Cancer Treatment (PDQ®)–Patient Version General Information About Non-Small ... clinical trials before, during, or after starting their cancer treatment. Some clinical trials only include patients who have ...

  3. Treatment Option Overview (Non-Small Cell Lung Cancer)

    Science.gov (United States)

    ... Lung Cancer Screening Research Non-Small Cell Lung Cancer Treatment (PDQ®)–Patient Version General Information About Non-Small ... clinical trials before, during, or after starting their cancer treatment. Some clinical trials only include patients who have ...

  4. Non-small-cell lung cancer cell lines A549 and NCI-H460 express hypoxanthine guanine phosphoribosyltransferase on the plasma membrane

    OpenAIRE

    Townsend MH; Anderson MD; Weagel EG; Velazquez EJ; Weber KS; Robison RA; O'Neill KL

    2017-01-01

    Michelle H Townsend, Michael D Anderson, Evita G Weagel, Edwin J Velazquez, K Scott Weber, Richard A Robison, Kim L O’Neill Department of Microbiology and Molecular Biology, Brigham Young University, Provo, UT, USA Abstract: In both males and females, lung cancer is one of the most lethal cancers worldwide and accounts for >30% of cancer-related deaths. Despite advances in biomarker analysis and tumor characterization, there remains a need to find suitable biomarker antigen...

  5. Effect of phosphorylation and methylation on the function of the p16INK4a protein in non-small cell lung cancer A549 cells

    OpenAIRE

    Ma, Wen-Long; Wang, Lin; LIU, LING-XIA; Wang, Xiu-Li

    2015-01-01

    The p16INK4a protein (p16) has been reported to be a tumor suppressor gene that suppresses the proliferation of cells through the direct inhibition of cell cycle progression. Accordingly, p16 is a potential target for cancer gene therapy. In the present study, the arginine 22, 131 and 138 residues of p16 were found to be methylation sites, as the mutation of these arginine residues to lysine resulted in the hypomethylation of p16. Furthermore, the protein arginine methyltransferases (PRMTs), ...

  6. Targeted therapy in non-small cell lung cancer

    Institute of Scientific and Technical Information of China (English)

    Shou-Ching Tang

    2004-01-01

    @@ 1 Introduction Recent progress in molecular biology has enabled us to better understand the molecular mechanism underlying pathogenesis of human malignancy including lung cancer. Sequencing of human genome has identified many oncogenes and tumor suppressor genes,giving us a better understanding of the molecular events leading to the formation, progression, metastasis, and the development of drug resistance in human lung cancer. In addition, many signal transduction pathways have been discovered that play important roles in lung cancer. Novel strategy of anti-cancer drug development now involves the identification and development of targeted therapy that interrupts one or more than one pathways or cross-talk among different signal transduction pathways. In addition, efforts are underway that combine the traditional cytotoxic (non-targeted) agents with the biological (targeted) therapy to increase the response rate and survival in patients with lung cancer, especially advanced non-small cell lung cancer (NSCLC).

  7. Curcumin Inhibits Non-Small Cell Lung Cancer Cells Metastasis through the Adiponectin/NF-κb/MMPs Signaling Pathway

    Science.gov (United States)

    Tsai, Jong-Rung; Liu, Po-Len; Chen, Yung-Hsiang; Chou, Shah-Hwa; Cheng, Yu-Jen; Hwang, Jhi-Jhu; Chong, Inn-Wen

    2015-01-01

    Adipose tissue is now considered as an endocrine organ involved in metabolic and inflammatory reactions. Adiponectin, a 244–amino acid peptide hormone, is associated with insulin resistance and carcinogenesis. Curcumin (diferuloylmethane) is the principal curcuminoid of the popular Indian spice, turmeric. Curcumin possesses antitumor effects, including the inhibition of neovascularization and regulation of cell cycle and apoptosis. However, the effects of adiponectin and curcumin on non-small cell lung cancer (NSCLC) remain unclear. In this study, we evaluated the expression of adiponectin in paired tumors and normal lung tissues from 77 patients with NSCLC using real-time polymerase chain reaction, western blotting, and immunohistochemistry. Kaplan–Meier survival analysis showed that patients with low adiponectin expression ratio (1) (p = 0.015). Curcumin inhibited the migratory and invasive ability of A549 cells via the inhibition of adiponectin expression by blocking the adiponectin receptor 1. Curcumin treatment also inhibited the in vivo tumor growth of A549 cells and adiponectin expression. These results suggest that adiponectin can be a prognostic indicator of NSCLC. The effect of curcumin in decreasing the migratory and invasive ability of A549 cells by inhibiting adiponectin expression is probably mediated through NF-κB/MMP pathways. Curcumin could be an important potential adjuvant therapeutic agent for lung cancer in the future. PMID:26656720

  8. Effects of a protein kinase C inhibitor combined with cisplatin on non-small cell lung cancer%蛋白激酶C抑制剂联合顺铂治疗非小细胞肺癌的实验研究

    Institute of Scientific and Technical Information of China (English)

    高志强; 韩宝惠; 沙慧芳; 史振余; 杨晓华; 冯久贤

    2010-01-01

    Objective To study the effects and possible mechanisms of the protein kinase C (PKC) inhibitor chelerythrine chloride (CH),combined with cisplatin (DDP) on human non-small cell lung cancer.Methods The effect of CH,DDP and the combination on proliferation and apoptosis of human lung cancer cell line A549 were evaluated by MTT assay and flow cytometry respectively.The inhibitory effects of CH and DDP on neoplasia were verified on subcutaneous implanted tumor of nude mice.Implanted tumor models were constructed in nude mice using human lung adenocarcinoma cell line A549.Twenty-four BALB/ c nude mice with implanted tumors were divided into 4 groups randomly:group CH,group DDP,group CH + DDP,and normal saline group (group NS),each with 5 mice.CH,DDP or NS were intraperitoneally injected into nude mice for 3 weeks (DDP or NS was injected once a week,CH was injected twice a week).Results CH inhibited A549 cell proliferation in a concentration-dependent pattern.When CH and DDP were combined,the inhibitory effect was enhanced in a synergistic or additive pattern.Both CH and DDP significantly increased the apoptosis of A549 cells,and this action was remarkably increased when DDP was combined with CH.CH and DDP inhibited the growth of subcutaneous implanted tumor in nude mice and the inhibitory rate of group CH + DDP (80.5%) was significantly higher than that of group CH (72.4%) or group DDP(64.3%) (t=11.34,P < 0.01).Conclusion CH combined with DDP shows significantly synergistic anti-tumor effects on non-small cell lung cancer cell line A549 and subcutaneous implanted tumor in nude mice,possibly by enhancement of growth inhibition and apoptosis induction on tumor cells.%目的 初步探讨蛋白激酶C(PKC)抑制剂白屈菜红碱(CH)与化疗药物顺铂联合应用在非小细胞肺癌(NSCLC)治疗中的作用及其可能的机制.方法 应用四甲基偶氮唑蓝试验及流式细胞术分别检测CH、顺铂以及CH与顺铂联用对人NSCLC细胞株A549增殖和凋

  9. Overview of KRAS-Driven Genetically Engineered Mouse Models of Non-Small Cell Lung Cancer.

    Science.gov (United States)

    Sheridan, Clare; Downward, Julian

    2015-01-01

    KRAS, the most frequently mutated oncogene in non-small cell lung cancer, has been utilized extensively to model human lung adenocarcinomas. The results from such studies have enhanced considerably an understanding of the relationship between KRAS and the development of lung cancer. Detailed in this overview are the features of various KRAS-driven genetically engineered mouse models (GEMMs) of non-small cell lung cancer, their utilization, and the potential of these models for the study of lung cancer biology.

  10. High throughput determination of TGFβ1/SMAD3 targets in A549 lung epithelial cells.

    Directory of Open Access Journals (Sweden)

    Yingze Zhang

    Full Text Available BACKGROUND: Transforming growth factor beta 1 (TGFβ1 plays a major role in many lung diseases including lung cancer, pulmonary hypertension, and pulmonary fibrosis. TGFβ1 activates a signal transduction cascade that results in the transcriptional regulation of genes in the nucleus, primarily through the DNA-binding transcription factor SMAD3. The objective of this study is to identify genome-wide scale map of SMAD3 binding targets and the molecular pathways and networks affected by the TGFβ1/SMAD3 signaling in lung epithelial cells. METHODOLOGY: We combined chromatin immunoprecipitation with human promoter region microarrays (ChIP-on-chip along with gene expression microarrays to study global transcriptional regulation of the TGFβ1/SMAD3 pathway in human A549 alveolar epithelial cells. The molecular pathways and networks associated with TGFβ1/SMAD3 signaling were identified using computational approaches. Validation of selected target gene expression and direct binding of SMAD3 to promoters were performed by quantitative real time RT-PCR and electrophoretic mobility shift assay on A549 and human primary lung epithelial cells. RESULTS AND CONCLUSIONS: Known TGFβ1 target genes such as SERPINE1, SMAD6, SMAD7, TGFB1 and LTBP3, were found in both ChIP-on-chip and gene expression analyses as well as some previously unrecognized targets such as FOXA2. SMAD3 binding of FOXA2 promoter and changed expression were confirmed. Computational approaches combining ChIP-on-chip and gene expression microarray revealed multiple target molecular pathways affected by the TGFβ1/SMAD3 signaling. Identification of global targets and molecular pathways and networks associated with TGFβ1/SMAD3 signaling allow for a better understanding of the mechanisms that determine epithelial cell phenotypes in fibrogenesis and carcinogenesis as does the discovery of the direct effect of TGFβ1 on FOXA2.

  11. Apoptosis inducing ability of silver decorated highly reduced graphene oxide nanocomposites in A549 lung cancer

    Directory of Open Access Journals (Sweden)

    Khan M

    2016-03-01

    Full Text Available Merajuddin Khan,1 Mujeeb Khan,1 Abdulhadi H Al-Marri,1 Abdulrahman Al-Warthan,1 Hamad Z Alkhathlan,1 Mohammed Rafiq H Siddiqui,1 Vadithe Lakshma Nayak,2 Ahmed Kamal,2 Syed F Adil1 1Department of Chemistry, College of Science, King Saud University, Riyadh, Kingdom of Saudi Arabia; 2Department of Medicinal Chemistry and Pharmacology, CSIR – Indian Institute of Chemical Technology, Hyderabad, India Abstract: Recently, graphene and graphene-based materials have been increasingly used for various biological applications due to their extraordinary physicochemical properties. Here, we demonstrate the anticancer properties and apoptosis-inducing ability of silver doped highly reduced graphene oxide nanocomposites synthesized by employing green approach. These nano­composites (PGE-HRG-Ag were synthesized by using Pulicaria glutinosa extract (PGE as a reducing agent and were evaluated for their anticancer properties against various human cancer cell lines with tamoxifen as the reference drug. A correlation between the amount of Ag nanoparticles on the surface of highly reduced graphene oxide (HRG and the anticancer activity of nanocomposite was observed, wherein an increase in the concentration of Ag nanoparticles on the surface of HRG led to the enhanced anticancer activity of the nanocomposite. The nanocomposite PGE-HRG-Ag-2 exhibited more potent cytotoxicity than standard drug in A549 cells, a human lung cancer cell line. A detailed investigation was undertaken and Fluorescence activated cell sorting (FACS analysis demonstrated that the nanocomposite PGE-HRG-Ag-2 showed G0/G1 phase cell cycle arrest and induced apoptosis in A549 cells. Studies such as, measurement of mitochondrial membrane potential, generation of reactive oxygen species (ROS and Annexin V-FITC staining assay suggested that this compound induced apoptosis in human lung cancer cells. Keywords: plant extract, graphene/silver nanocomposites, anticancer, apoptosis

  12. Mechanisms for inhibition effects of polypeptide extract from scorpion venom (PESV) on proliferation of A549 cell lines in vitro%蝎毒多肽提取物对A549细胞增殖抑制作用机制研究

    Institute of Scientific and Technical Information of China (English)

    王晓慧; 王兆朋; 张月英; 贾青; 王朝霞; 张捷; 张维东

    2012-01-01

    目的:观察蝎毒多肽提取物( PESV)对非小细胞肺癌细胞株A549的增殖抑制作用及可能的作用机制.方法:采用噻唑蓝(MTT)法观察不同浓度PESV对A549细胞生长与增殖的影响,下游实验将对数生长期的A549细胞分为阴性对照组、PESV低、中、高剂量组,应用流式细胞术、免疫细胞化学法、Western blot法检测PESV干预后细胞周期及VEGF,HIF-1α和PTEN蛋白表达的变化.结果:MTT结果显示,PESV在一定浓度范围内对A549细胞的增殖活性有明显抑制作用(P<0.01).流式细胞法、免疫细胞化学法及Western blot法结果显示,PESV干预后能使A549细胞阻滞于G0/G1期,并显著下调HIF-1α,VEGF表达,上调PTEN表达.结论:PESV能够抑制A549细胞的增殖,其作用机制可能与影响血管生成因子VEGF,HIF-1α和PTEN的表达而直接抑制细胞增殖、阻滞细胞周期和抑制血管生成有关.%Objective: To investigate the mechanisms for inhibition effects of PESV on proliferation of non-small cell lung cancer cell line A549. Method: MTT was used to observe cell growth and proliferation of A549 at different concentrations of PESV. Flow eytometry(FCM)was applied to analyze cell cycle distribution. Immunocytochemistry and western blot assay was recruited to detect the expression of VEGF,HIF-lα,PTEN after the intervention of PESV. Result: A549 cells may he arrested mainly in G0/G1 phase and cell proliferation was significantly inhibited ( P < 0.01) after PESV intervention in a certain range of concentration. PESV can significantly reduce the expression of HIF-lα,VEGF and increase the expression of PTEN. Conclusion: PESV can block cell cycle and inhibit angiogenesis directly to inhibit cell proliferation of non-small cell lung cancer cell line A549 mainly through reducing the expression of HIF-lα.VEGF and increasing the expression of PTEN.

  13. [Ru(pipe)(dppb)(bipy)]PF6: A novel ruthenium complex that effectively inhibits ERK activation and cyclin D1 expression in A549 cells.

    Science.gov (United States)

    Ferreira-Silva, Guilherme A; Ortega, Marina M; Banionis, Marco A; Garavelli, Graciana Y; Martins, Felipe T; Dias, Julia S M; Viegas, Cláudio; Oliveira, Jaqueline C de; Nascimento, Fabio B do; Doriguetto, Antonio C; Barbosa, Marilia I F; Ionta, Marisa

    2017-10-01

    Lung cancer is the most frequent type of cancer worldwide. In Brazil, only 14% of the patients diagnosed with lung cancer survived 5years in the last decades. Although improvements in the therapeutic approach, it is relevant to identify new chemotherapeutic agents. In this framework, ruthenium metal compounds emerge as a promising alternative to platinum-based compounds once they displayed lower cytotoxicity and more selectivity for tumor cells. The present study aimed to evaluate the antitumor potential of innovative ruthenium(II) complex, [Ru(pipe)(dppb)(bipy)]PF6 (PIPE) on A549 cells, which is derived from non-small cell lung cancer. Results demonstrated that PIPE effectively reduced the viability and proliferation rate of A549 cells. When PIPE was used at 9μM there was increase in G0/G1 cell population with concomitant reduction in frequency of cells in S-phase, indicating cell cycle arrest in G1/S transition. Antiproliferative activity of PIPE was associated to its ability of reducing cyclin D1 expression and ERK phosphorylation levels. Cytotoxic activity of PIPE on A549 cells was observed when PIPE was used at 18μM, which was associated to its ability of inducing apoptosis by intrinsic pathway. Taken together, the data demonstrated that PIPE is a promising antitumor agent and further in vivo studies should be performed. Copyright © 2017 Elsevier Ltd. All rights reserved.

  14. Silencing ATM by siRNA enhances the radiosensitization effect of CpG ODN 7909 on lung cancer A549 cells%siRNA沉默ATM增强CpG ODN 7909对肺癌A549细胞的放射增敏作用

    Institute of Scientific and Technical Information of China (English)

    袁素娟; 乔田奎; 刘小群; 陈伟

    2013-01-01

    目的:研究siRNA沉默毛细血管扩张—共济失调突变(atxia-telangiectasia mutated,ATM)基因的表达增强胞嘧啶鸟嘌呤二核苷酸寡脱氧核苷酸(cytosine-phophate-guanine oligodeoxynucleotide,CpG ODN) 7909对人非小细胞肺癌A549细胞的放射增敏作用.方法:将ATM-siRNA转染至A549细胞中,Western blotting检测A549细胞中ATM蛋白的表达.A549细胞随机分为6组:对照组、CpG组、X射线(IR)组、CpG+ IR组、ATM-siRNA+ CpG+ IR组和NC-siRNA+ CpG+ IR组,克隆形成分析法检测各组细胞克隆形成率,Graphpad prism 5.0软件进行单击多靶模型和L-Q线性模型拟合辐射后A549细胞的生存曲线,以D0、Dq、N、α/β及SF2等参数分析A549细胞辐射损伤修复能力,流式细胞术检测A549细胞的凋亡.结果:ATM-siRNA转染可明显抑制A549细胞中ATM蛋白的表达(P<0.01).X射线可剂量依赖性抑制A549细胞的克隆形成能力(P<0.05);且CpG+ IR组A549细胞的克隆形成能力进一步降低(P<0.01);ATM-siRNA转染后,CpG处理的A549细胞克隆形成能力再度降低[10 Gy时,(0.05±0.00)%vs(0.71±0.00)%,P<0.01].辐射损伤剂量生存曲线结果显示,ATM-siRNA转染后,ATM-siRNA+ CpG+ IR组较CpG+ IR组A549细胞的α/β值明显增大(1.48 vs0.97,P<0.05),对放射损伤修复能力明显减弱.CpG+ IR组较IR组细胞凋亡率显著升高[(9.18±0.16)%vs(6.56±0.33)%,P<0.01];ATM-siRNA+ CpG+ IR组A549细胞凋亡率进一步升高[(10.45 ±0.40)% vs (9.18±0.16)%,P<0.05].结论:siRNA沉默ATM的表达可增强CpGODN 7909对A549细胞的放射增敏作用,ATM可作为肺癌治疗的潜在靶点.%Objective:To explore the potentiation of silencing atxia-telangiectasia mutated (ATM) gene expression in the radiosensitization effect of cytosine-phophate-guanine oligodeoxynucleotide (CpG ODN) 7909 on non-small cell lung cancer A549 cell line.Methods:ATM-siRNA was transfected into A549 cells,and the expression of ATM protein in A549 cells was

  15. Cetuximab in non-small-cell lung cancer.

    Science.gov (United States)

    Carillio, Guido; Montanino, Agnese; Costanzo, Raffaele; Sandomenico, Claudia; Piccirillo, Maria Carmela; Di Maio, Massimo; Daniele, Gennaro; Giordano, Pasqualina; Bryce, Jane; Normanno, Nicola; Rocco, Gaetano; Perrone, Francesco; Morabito, Alessandro

    2012-02-01

    Cetuximab is a chimeric human-mouse anti-EGF receptor monoclonal antibody. In Phase I studies, no dose-limiting toxicities were observed with cetuximab as a single agent or combined with chemotherapy; pharmacokinetic and pharmacodynamic analyses supported 250 mg/m(2) weekly administration. Skin toxicity, diarrhea and fatigue were the most common toxicities. The positive results obtained in Phase II trials in patients with advanced non-small-cell lung cancer prompted two randomized Phase III trials evaluating cetuximab in addition to first-line chemotherapy. Both trials showed a small benefit in overall survival for the experimental treatment, which was considered insufficient by the EMA for marketing approval. However, a subgroup analysis of the FLEX Phase III trial recently demonstrated a larger survival benefit from the experimental treatment in patients with high immunohistochemical EGF receptor expression. This finding, if confirmed prospectively, could represent a new opportunity for positioning cetuximab into the standard treatment of advanced non-small-cell lung carcinoma.

  16. Zinc supplementation induces apoptosis and enhances antitumor efficacy of docetaxel in non-small-cell lung cancer

    Directory of Open Access Journals (Sweden)

    Kocdor H

    2015-07-01

    Full Text Available Hilal Kocdor,1,2 Halil Ates,1 Suleyman Aydin,3 Ruksan Cehreli,1 Firat Soyarat,2 Pinar Kemanli,2 Duygu Harmanci,2 Hakan Cengiz,2 Mehmet Ali Kocdor4 1Institute of Oncology, Dokuz Eylul University, 2Department of Molecular Medicine, Institute of Health Sciences, Dokuz Eylul University, Izmir Turkey; 3Department of Biochemistry, Firat University School of Medicine, Elazig, 4Department of Surgery, School of Medicine, Dokuz Eylul University, Izmir, Turkey Background: Exposure to exogenous zinc results in increased apoptosis, growth inhibition, and altered oxidative stress in cancer cells. Previous studies also suggested that zinc sensitizes some cancer cells to cytotoxic agents depending on the p53 status. Therefore, zinc supplementation may show anticancer efficacy solely and may increase docetaxel-induced cytotoxicity in non-small-cell lung cancer cells.Methods: Here, we report the effects of several concentrations of zinc combined with docetaxel on p53-wild-type (A549 and p53-null (H1299 cells. We evaluated cellular viability, apoptosis, and cell cycle progression as well as oxidative stress parameters, including superoxide dismutase, glutathione peroxidase, and malondialdehyde levels.Results: Zinc reduced the viability of A549 cells and increased the apoptotic response in both cell lines in a dose-dependent manner. Zinc also amplified the docetaxel effects and reduced its inhibitory concentration 50 (IC50 values. The superoxide dismutase levels increased in all treatment groups; however, glutathione peroxidase was slightly increased in the combination treatments. Zinc also caused malondialdehyde elevations at 50 µM and 100 µM.Conclusion: Zinc has anticancer efficacy against non-small-cell lung cancer cells in the presence of functionally active p53 and enhances docetaxel efficacy in both p53-wild-type and p53-deficient cancer cells. Keywords: lung cancer, zinc, docetaxel, A549, H1299

  17. Molecular mechanism underlying the antiproliferative effect of suppressor of cytokine signaling-1 in non-small-cell lung cancer cells.

    Science.gov (United States)

    Shimada, Kazuki; Serada, Satoshi; Fujimoto, Minoru; Nomura, Shintaro; Nakatsuka, Rie; Harada, Emi; Iwahori, Kota; Tachibana, Isao; Takahashi, Tsuyoshi; Kumanogoh, Atsushi; Kishimoto, Tadamitsu; Naka, Tetsuji

    2013-11-01

    Lung cancer (LC) is the major cause of death by cancer and the number of LC patients is increasing worldwide. This study investigated the therapeutic potential of gene delivery using suppressor of cytokine signaling 1 (SOCS-1), an endogenous inhibitor of intracellular signaling pathways, for the treatment of LC. To examine the antitumor effect of SOCS-1 overexpression on non-small-cell lung cancer (NSCLC) cells, NSCLC cells (A549, LU65, and PC9) were infected with adenovirus-expressing SOCS-1 vector. The cell proliferation assay showed that A549 and LU65, but not PC9, were sensitive to SOCS-1 gene-mediated suppression of cell growth. Although JAK inhibitor I could also inhibit proliferation of A549 and LU65 cells, SOCS-1 gene delivery appeared to be more potent as SOCS-1 could suppress focal adhesion kinase and epidermal growth factor receptor, as well as the JAK/STAT3 signaling pathway. Enhanced phosphorylation of the p53 protein was detected by means of phospho-kinase array in SOCS-1 overexpressed A549 cells compared with control cells, whereas no phosphorylation of p53 was observed when JAK inhibitor I was used. Furthermore, treatment with adenoviral vector AdSOCS-1 in vivo significantly suppressed NSCLC proliferation in a xenograft model. These results suggest that the overexpression of SOCS-1 gene is effective for antitumor therapy by suppressing the JAK/STAT, focal adhesion kinase, and epidermal growth factor receptor signaling pathways and enhancing p53-mediated antitumor activity in NSCLC.

  18. A study on the allelic deletion and mutation of FHIT gene in human non-small cell lung cancer%人非小细胞肺癌中FHIT等位基因缺失和突变的研究

    Institute of Scientific and Technical Information of China (English)

    周清华; 陈军; 覃扬; 孙芝琳; 刘伦旭; 孙泽芳; 车国卫; 李潞; 秦建军; 宫友陵

    2001-01-01

    Objective To explore the role of the allelic deletion and mutation of FHIT gene on the carcinogenesis and development of lung cancer. Methods The allelic alterations of FHIT gene and microsatellites D3S1300, D3S1312,D3S1313 were detected in 35 cancer samples of NSCLC, their corresponding normal tissues, and 4 lung cancer cell lines, and 10 lung tissues of benign pulmonary lesions as control by PCR-SSCP and DNA sequence. Results Loss of heterozygosity (LOH) affecting at least one locus of FHIT gene was observed in 22 out of 35 tumors, with a LOH rate of 62.86%. LOH of FHIT gene in squamous cell carcinoma (88.24%) was significantly higher than that in adenocarcinoma (38.89%) (P<0.01). The LOH rate of FHIT gene in smoking patients (76.19%) was also significantly higher than