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Sample records for a549 cell line

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

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    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.

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

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    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.

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

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    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.

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

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    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

  5. MicroRNA-126 inhibits the proliferation of lung cancer cell line A549

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    Xun Yang; Bei-Bei Chen; Ming-Hua Zhang; Xin-Rong Wang

    2015-01-01

    Objective:To study the role of microRNA-126 in the development of lung cancer.Methods:The biological function of microRNA-126 was detected using EdU assay and CCK-8 assay;the target gene of microRNA-126 was analyzed using real time RT-PCR and Western blot assay.Results: In A549 cell line, overexpression of microRNA-126 inhibits the proliferation rate; VEGF is the target gene of microRNA-126; microRNA-126 exerts its function via regulating VEGF protein level.Conclusions: microRNA-126 inhibits the proliferation in A549 cell line.

  6. 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

  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

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    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. Different maspin functions in the lung adenocarcinoma A549 and SPC-A1 cell lines.

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    Zhou, Jun; Hualong, Qin; Zhou, Peng; Guo, Feng

    2015-11-01

    Mammary serine protease inhibitor (maspin) is a tumor suppressor gene that is silenced in the majority of cancer cells during metastatic progression by transcriptional and epigenetic mechanisms. The function of maspin in non‑small cell lung cancer cells (NSCLC) has not been clearly defined. In the present study, the expression of maspin in NSCLC cell lines, in particular, the adenocarcinoma cell lines, was heterogeneous. While the expression levels of maspin in PC‑9 and H460 cell lines were intact, the expression of maspin in the A549 and SPC‑A1 cells was hardly detected. Ectopic expression of maspin in A549 cells carrying the K‑ras gene point mutation significantly inhibited cell migration and invasion abilities, which was associated with downregulated expression of matrix metalloproteinase‑2 and integrin β1. Ectopic expression of maspin in SPC‑A1 cells harboring the wild‑type K‑ras gene predominantly affected cell growth via targeting the AKT signaling molecules. Maspin functions differently in lung adenocarcinoma cells, possibly due to the varied molecular characteristics.

  9. In vitro anticancer activity of fucoidan from Turbinaria conoides against A549 cell lines.

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    Marudhupandi, Thangapandi; Ajith Kumar, Thipramalai Thankappan; Lakshmanasenthil, Shanmugaasokan; Suja, Gunasekaran; Vinothkumar, Thirumalairaj

    2015-01-01

    The present study was conducted to evaluate the anticancer activity of fucoidan isolated from brown seaweed Turbinaria conoides. Extracted fucoidan contained 53 ± 0.69% of fucose and 38 ± 0.42% of sulphate, respectively. Functional groups and structural characteristics of the fucoidan were analyzed by FT-IR and NMR. In vitro anticancer effect was studied on A549 cell line. Fucoidan inhibited the growth of cancer cells in a dose-dependent manner and potent anticancer activities were 24.9-73.5% in the concentrations of 31.25-500 μg/ml. The CTC50 value against the cancer cell was found to be 45 μg/ml and the CTC50 value of normal Vero cell line is 325 μg/ml. This study suggests that the fucoidan from T. conoides could be significantly improved if the active component is further purified and tested for further investigation in various cancer cell lines.

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

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    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.

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

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    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.

  12. Construction of a CD147 Lentiviral Expression Vector and Establishment of Its Stably Transfected A549 Cell Line

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    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.

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

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    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.

  14. 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

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

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    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.

  16. 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

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    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

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

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    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.

  18. The mRNA and protein expression of folylpolyglutamate synthetase in methotrexate enantiomer-resistant A549 cell lines%信息动态

    Institute of Scientific and Technical Information of China (English)

    2011-01-01

    Objective To study the expression of folylpolyglutamate synthetase ( FPGS ) in methotrexate ( MTX ) enantiomer-resistant A549 cell lines [ L-( + )-MTX and D-( - )-MTX ]. Methods The expression of FPGS on genetic and protein level was determined by FQ-PCR and Western blot in lung cancer A549 cells, and MTX enantiomer-resistant A549 cells [ L-( + )-MTX and D-( - )-MTX ], with the concentration of drug resistance was 15 μmol/L. Results The genetic expression level of FPGS was ( 0.80 ± 0. 09 ) and ( 2. 04 ± 0. 34 ) folds in L-( + )- MTX/A549 cells and D-( - )-MTX/A549 cells compared with lung cancer A549 cells, there was statistical difference between two groups ( P < 0.05 ). The protein expression level of FPGS was ( 0. 85 ± 0. 12 ) and( 1.62 ± 0. 24 ) folds in L-( + )-MTX/A549 cells and D-( - )-MTX/A549 cells compared with lung cancer A549 cells,there was statistical difference ( P < 0. 05 ). Conclusion The expression level of FPGS on genetic and protein level in drug resistant cells have been changed, and significant difference in two enantiomer-resistant cells are appeared.

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

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    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. 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.

  1. Cytotoxic, Antiproliferative and Apoptotic Effects of New Benzimidazole Derivatives on A549 Lung Carcinoma and C6 Glioma Cell Lines.

    Science.gov (United States)

    Yurttas, Leyla; Demirayak, Seref; Ciftci, Gulsen Akalın

    2015-01-01

    Benzimidazole ring is a versatile structure which has been extensively utilized in medicinal chemistry. Since we are working on 1,2-disubstutited benzimidazoles, we have reported new antitumor active derivatives. As a continuation to our previous work, we have synthesized a new series of 1-(2-aryl-2-oxoethyl)-2-[(N,Ndimethylamino/pyrrolidinyl/piperidinyl)thiocarbamoyl] benzimidazole derivatives. Anticancer activity of the compounds was evaluated using MTT assay, BrdU assay and flow cytometric analysis on A549 human lung carcinoma and C6 rat glioma cell lines. Compounds bearing dimethylamino moiety exhibited higher antitumor activity.

  2. Direct electric current treatment modifies mitochondrial function and lipid body content in the A549 cancer cell line.

    Science.gov (United States)

    Holandino, Carla; Teixeira, Cesar Augusto Antunes; de Oliveira, Felipe Alves Gomes; Barbosa, Gleyce Moreno; Siqueira, Camila Monteiro; Messeder, Douglas Jardim; de Aguiar, Fernanda Silva; da Veiga, Venicio Feo; Girard-Dias, Wendell; Miranda, Kildare; Galina, Antonio; Capella, Marcia Alves Marques; Morales, Marcelo Marcos

    2016-10-01

    Electrochemical therapy (EChT) entails treatment of solid tumors with direct electric current (DC). This work evaluated the specific effects of anodic flow generated by DC on biochemical and metabolic features of the A549 human lung cancer cell line. Apoptosis was evaluated on the basis of caspase-3 activity and mitochondrial transmembrane potential dissipation. Cell morphology was analyzed using transmission electron microscopy, and lipid droplets were studied through morphometric analysis and X-ray qualitative elemental microanalysis. High-resolution respirometry was used to assess mitochondrial respiratory parameters. Results indicated A549 viability decreased in a dose-dependent manner with a prominent drop between 18 and 24h after treatment (p<0.001), together with a two-fold increase in caspase-3 activity. AF-treatment induced a significantly increase (p<0.01) in the cell number with disrupted mitochondrial transmembrane potential. Furthermore, treated cells demonstrated important ultrastructural mitochondria damage and a three-fold increase in the cytoplasmic lipid bodies' number, quantified by morphometrical analyses. Conversely, 24h after treatment, the cells presented a two-fold increase of residual oxygen consumption, accounting for 45.3% of basal oxygen consumption. These results show remarkable alterations promoted by anodic flow on human lung cancer cells which are possibly involved with the antitumoral effects of EChT.

  3. 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.

  4. Cytotoxic and pro-apoptotic activities of leaf extract of Croton bonplandianus Baill. against lung cancer cell line A549.

    Science.gov (United States)

    Bhavana, J; Kalaivani, M K; Sumathy, A

    2016-06-01

    The acetone extract (AcE) of the Croton bonplandianus Baill., an exotic weed of the Euphorbiaceae family was studied for cytotoxicity, apoptosis, cell cycle arrest in A549 cell line and antioxidant capacities using MTT assay, acridine orange/ethidium bromide (AO/EB staining), cell cycle analysis and DPPH radical scavenging assay respectively. Based on the cytotoxic activity, the extract was tested for the apoptotic effect using AO/EB and Hoechst 33258 staining. The apoptosis was characterized by chromatin condensation and DNA fragmentation. Further, to determine the stage of cell death, cell cycle analysis was performed by flow cytometry and AcE was found to arrest G2/M phase in a dose dependent manner. The number of cells in G2/M phase increases with concurrent accumulation of cells in sub G₀/G₁phase indicates the induction of apoptosis at G2M phase. The free radical scavenging activity of the AcE against DPPH was considerably significant. The cytotoxic, apoptotic and antioxidant effect of the AcE could be well correlated with the presence of potent free radical scavenging secondary metabolites such as phenols (43 ± 0.05 µg/mL), flavonoids (3.5 ± 0.07 µg/mL) and tannin (0.36 ± 0.1 µg/mL). Our study has shown that A549 cells were more sensitive to AcE with an IC₅₀ of 15.68 ± 0.006 µg/mL compared to the standard drug 2.20 ± 0.008 µg/mL (cisplatin). The results suggest that Croton bonplandianus could serve as a potential source of alternative therapeutic agent for treating cancer. Further research is required to isolate the active principle compound and determination of its anticancer property.

  5. 盐霉素对人肺腺癌A549细胞株生物学特性的影响%Effects of Salinomycin on the biological characteristics of lung adenocarcinoma cell line A549 cells

    Institute of Scientific and Technical Information of China (English)

    石俊杰; 王哲; 李阳; 张翼翔; 刘晶; 顾春东

    2013-01-01

    Objective To investigate the effects of Salinomycin on the biological characteristics of lung adenocarcinoma cell line A549 cells.Methods The scratch test,Transwell assay and cell counting kit-8 assay were applied to study the influence of Salinomycin on biological characteristics of lung adenocarcinoma A549 cells.Flow cytometry was used to analyze the proportion of side population cells.Western blotting was used to detect the expression of Oct4 protein in A549 cells treated with Salinomycin.Results In blank control group,30 mg/L Salinomycin group and 60 mg/L Salinomycin group,migration distance was 70,40 and 10 μm,the number of invasive cells was (32.3 ± 2.5),(21.4 ± 1.8) and (1.3 ± 0.3)/HP,and cell survival rate was 98%,74% and 50%,respectively.The proportion of side population cells in 60 mg/L Salinomycin group,30 mg/L Salinomycin group,blank control group and reserpine blocking group was 3.4%,10.5%,12.5% and 0.7% respectively.Western blotting revealed that the Oct4 expression was decreased with increasing Salinomycin.Conclusion Salinomycin has an important impact on the biological characteristics of lung adenocarcinoma cell line A549.Salinomycin might become a new effective drug for the treatment of lung adenocarcinoma.%目的 观察盐霉素对人肺腺癌A549细胞株生物学特性的影响.方法 无血清培养A549干细胞,进行划痕、Transwell和细胞计数实验,运用流式细胞仪检测细胞侧群比例,采用Westernblot检测盐霉素对转录因子Oct4表达的影响.结果 空白对照组、30 mg/L、60 mg/L盐霉素组迁移距离分别为70、40、10μm,侵袭细胞数目分别为(32.3±2.5)、(21.4±1.8)、(1.3±0.3)个/HP,细胞存活率分别为98%、74%、50%;60 mg/L组、30 mg/L组、空白对照组、利血平阻断组侧群比例分别为3.4%、10.5%、12.5%、0.7%.Western blot示随着盐霉素浓度增加,Oct4的表达量下降.结论 盐霉素对人肺腺癌细胞株A549生物学特性具有重要影响.

  6. Purification and characterization of protease enzyme from actinomycetes and its cytotoxic effect on cancer cell line (A549)

    Institute of Scientific and Technical Information of China (English)

    C Balachandran; V Duraipandiyan; S Ignacimuthu

    2012-01-01

    Objective: To isolate active actinomycetes from soil samples of Northern Himalayas and study their culture characterization, protease production and cytotoxic effects on cancer cell line (A549). Methods: Forty six strains of actinomycetes were isolated from the soil collected from Northern Himalayas, India. Isolation of actinomycetes was performed by serial dilution plate technique. Forty six isolated actinomycetes cultures were grown in ISP 2 medium to study the morphology and biochemical characteristics. Isolated strains were studied for protease enzyme production in skim milk agar medium with solubilising capacity. Seven isolates were studied for melanin pigmentation and different NaCl concentration. Effects of environmental conditions influencing protease enzyme production of seven isolated strains were also studied at different pH, temperature and metal ions (β-mercaptoethanol, dithiothreitol, iodoacetamide, MgSO4, CaCl2 and EDTA). The seven isolates were also studied for lytic enzyme activity using different bacteria and yeast such as Pseudomonas aeruginosa (P. aeruginosa), Enterococcus feacalis (E. feacalis), Escherishia coli (E. coli), Candida albicans (C. albicans), Bacillus subtilis (B. subtilis), Klebsiella pneumonia (K. pneumonia) and Staphylococcus aureus (S. aureus). Results: Isolates ERIA-31 and ERIA-33 produced more protease enzyme activity in modified nutrient agar media compared to other actinomycetes cultures. ERIA-31 and ERIA-33 were tested for cytotoxic effect in human adenocarcinoma cancer cell line (A549). IC50 for ERIA-31 was 57.04 μg/mL and IC50 for ERIA-33 was 55.07 μg/mL. Conclusion: Actinomycete being a protease producing bacteria has the potential for use in industrial purpose, pharmaceuticals, cytotoxic agent and its proteolytic activity. Isolates of ERIA-31 and ERIA-33 produced significant amount of protease enzymes.

  7. 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

  8. 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.

  9. 三种缝线材料对人肺腺癌细胞A549增殖和细胞周期的影响%Effect of three suture lines on the proliferation and cell cycle of lung adenocarcinoma cell A549 in vitro

    Institute of Scientific and Technical Information of China (English)

    Lianhua Ye; Yunchao Huang; Qilin Jin; Feng Hua; Guangqiang Zhao

    2011-01-01

    Objective: The interaction of cell and medical biomaterial is one of the significant factors to affect clinical application of medical biomaterial. This research is to investigate three of suture lines how to affect the proliferation and cell cycle of lung adenocarcinoma cell A549 in vitro. Methods: Three of suture lines were respectively cultivated with lung adenocarcinoma cell A549, after of 72 hours, we detected absorptions of each group by MTT method in order to reflect the proliferation of lung adenocarcinoma cell A549, and also examined percentage of G1 period cells and S period cells of each group by flow cytometry. Results: Different of suture lines had different effects on the proliferation and cell cycle of lung adenocarcinoma cell A549 (P < 0.05). The effect of absorbent suture line was the strongest on the proliferation and cell cycle of lung adenocarcinoma cell A549, the effect of chorda serica chirurgicalis was medium, and the effect of slide wire was poor. Different length of each suture line had different effects on the proliferation and cell cycle of lung adenocarcinoma cell A549 (P < 0.05).Conclusion: Three of suture line materials have different effects on the proliferation and cell cycle of lung adenocarcinoma cell A549, with dose-effect relationship.

  10. 流感病毒NS1蛋白稳定表达的A549细胞系建立%Establishment of A549 Cell Line Stably Expressing NS1 Protein of Influenza Virus

    Institute of Scientific and Technical Information of China (English)

    李志辉; 曾琳姣; 王慧煜; 梅琳; 刘永飞; 韩雪清

    2013-01-01

    NS1 of influenza A virus is a key multifunctional protein that plays various roles in regulating viral replication mechanisms, disease pathogenesis. In order to establish stable A549 cell line expressing NS1 protein of influenza A Virus, NSl cDNA was obtained by RT-PCR using 2009 A(H1N1) influenza virus total RNA as template. The fragment was cloned in the pMD19-T vector, then the fragment was obtained by BamHI and NdeI digestion, and ligated with pCMV-HA. Linearized pCMV-HA-NS1 and neo were transfect-ed into A549 cells. The stable expressing NSl protein cell line was screened by G418. DNA, RNA, protein levels of NS1 were detected in A549 cells by PCR, RT-PCR and Western blot, the location of the NSl protein in cells was observed by immunofluorescence. The result indicated that NS1 protein was stable expressed in A549 cell line, suggesting that NSl stable expression A549 cell line was successfully constructed, and the NS1 protein is located in nucleus. This stable cell line can be used for further study of biological functions of NS1.%A型流感病毒的NSl(Nonstructurol 1 protein,NSl)蛋白是病毒复制、毒力等的重要调节蛋白.运用RT-PCR方法扩增A/Beijing/501/2009 (H1N1)流感病毒NS1基因,克隆至真核表达载体pCMV-HA,用Lipofectamine 2000将线性化pCMV-HA-NS1与neo基因共同转染A549细胞,通过G418筛选获得阳性重组细胞,并采用PCR、RT-PCR、Western blot技术检测重组细胞中NS1蛋白的表达,通过免疫荧光技术观察NS1蛋白在细胞中的定位.PCR、RT-PCR检测显示NS1基因成功整合进入细胞基因组,并转录为mRNA;Western blot检测显示重组细胞系稳定表达NS1蛋白,免疫荧光显示NS1蛋白定位于细胞核内.表明通过G418筛选,成功构建稳定表达NS1蛋白的重组A549-HA-NS1细胞系,且NS1蛋白定位于细胞核内,为进一步研究NS1蛋白的生物学功能奠定基础.

  11. Impact of siRNA targeting pirh2 on proliferation and cell cycle control of the lung adenocarcinoma cell line A549

    Institute of Scientific and Technical Information of China (English)

    SU Yuan; ZHU Liping; JIN Yang; ZHANG Xiaoju; ZHOU Qiong; BAI Ming

    2007-01-01

    The aim of this study was to investigate the role of pirh2(p53-induced RING-H2)protein in the proliferation,apoptosis and cell cycle control of the lung cancer cell line A549.Pirh2 expression was detected by immunofluorescence,Western blot analysis and real-time polymerase chain reaction(PCR).Cell proliferation was assessed by cell counting kit-8(CCK-8).Cell cycle control and apoptosis were analyzed by flow cytometry.The results showed that pirh2 was expressed in the cytoplasm ofA549 cells.The inhibition of pirh2 expression by siRNA(psiRNA-pirh2)resulted in reduced cell proliferation and increased apoptosis.In addition,the number of G0/G1 phase cells was increased but G2/M cells were not affected significantly.Taken together,the inhibition of pirh2 expression in the lung adenocarcinoma cell line A549 resulted in reduced tumor cell growth via the inhibition of cell proliferation,the activation of apoptosis and the interruption of cell cycle transition.

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

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    Khaghanzadeh Narges

    2012-10-01

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

  13. Umbelliprenin is Cytotoxic against QU-DB Large Cell Lung Cancer Cell Line but Anti-Proliferative against A549 Adenocarcinoma Cells

    Directory of Open Access Journals (Sweden)

    Abbas Ghaderi

    2012-10-01

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

  14. 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

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

    Science.gov (United States)

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

    2016-01-01

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

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

    Directory of Open Access Journals (Sweden)

    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.

  18. Combination therapy in A549 cells

    Energy Technology Data Exchange (ETDEWEB)

    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.

  19. 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.

  20. 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.

  1. Polyurethane dispersion containing quaternized ammonium groups: An efficient nanosize gene delivery carrier for A549 cancer cell line transfection.

    Science.gov (United States)

    Yousefpour Marzbali, Mahsa; Yari Khosroushahi, Ahmad; Movassaghpour, AliAkbar; Yeganeh, Hamid

    2016-01-25

    A novel polyurethane containing cationic ammonium groups (QPU) was synthesized and used as vector for gene therapy and cancer gene targeting. The synthesized QPU was characterized by Fourier transform infrared and nuclear magnetic resonance spectroscopy methods. An agarose gel retardation electrophoresis assay was conducted to verify the complete complex formation between QPU and pDNA. The particles size and zeta potential of neat polymers, plasmid DNA, polymers/DNA polyplexes were determined by the dynamic light scattering technique. The polyplexes cytotoxicity was determined using [3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide] assay and its transfection efficiency was examined qualitatively by fluorescent microscopy and quantitatively by flow cytometery methods. The gel retardation assay, particle size and zeta potential measurements were confirmed that the synthesized cationic polymer could condense DNA efficiently in the physiologic condition. QPU polyplexes showed a significantly lower cytotoxicity compared to Polyfect polyplexes in the examined human cancerous (A549) or normal cells (KDR). Based on our findings, the transfection efficiency by QPU was 2.2 fold higher than Polyfect in the A549 cells whereas in the KDR cells, the cell transfection by Polyfect was 18.1 fold higher than QPU. Due to low cytotoxicity for normal cells and high transfection efficiency in cancer cells, the potential applicability of designed QPU as a non-viral gene carrier for targeting of cancer gene therapy was confirmed.

  2. 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.

  3. 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.

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

    Science.gov (United States)

    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.

  5. 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.

  6. 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.

  7. [Down-regulated βIII-tubulin expression can reverse paclitaxel resistance in A549/taxol cells lines].

    Science.gov (United States)

    Zhuo, Yinling; Guo, Qisen

    2014-08-20

    背景与目的 化疗耐药导致肿瘤很快复发和/或转移,是目前肺癌死亡的主要原因之一。β-tubulin是抗微管药物的主要细胞靶点。已有的研究证明:βIII-tubulin高表达与非小细胞肺癌(non-small cell lung cancer, NSCLC)耐药有关。利用RNA干扰技术沉默耐紫杉醇A549细胞(A549/Taxol)中βIII-tubulin基因表达,探讨靶基因下调后对化疗药物紫杉醇的敏感性的变化以及细胞周期和细胞凋亡情况。方法 构建靶向βIII-tubulin的siRNA,以脂质体为载体介导βIII-tubulin siRNA转染A549/Taxol细胞,利用qRT-PCR检测细胞内βIII-tubulin mRNA的变化情况,并筛选出最佳干扰序列;Western blot法检测A549/Taxol细胞内βIII-tubulin蛋白表达的变化;MTT法检测转染后细胞株对紫杉醇敏感性的变化;流式细胞仪检测细胞周期和细胞凋亡的变化。结果 实时荧光qRT-PCR法显示转染后细胞株靶基因水平较对照组降低,其中βIII-tubulin siRNA-1序列抑制率最高为(87.73±4.87)%(P<0.01);Western blot显示转染后靶蛋白水平较对照组明显降低;MTT法表明紫杉醇处理转染后细胞株的细胞抑制率较对照组明显增加(51.77±4.60)%(P<0.01);细胞凋亡显示βIII-tubulin siRNA+Taxol组细胞早期凋亡率较对照组明显增加(P<0.01),两者的差异有统计学意义;细胞周期检测结果显示紫杉醇处理组的G2/M期细胞百分率高于对照组,且转染后紫杉醇处理组的细胞晚期凋亡率较对照组增加。结论 βIII-tubulin表达下调明显提高A549/Taxol细胞株对Taxol的敏感性。

  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. Evaluation of the cytotoxicity and the genotoxicity induced by {alpha} radiation in an A549 cell line

    Energy Technology Data Exchange (ETDEWEB)

    Belchior, Ana, E-mail: anabelchior@itn.pt [Instituto Tecnologico e Nuclear, Estrada Nacional no 10, 2686-953 Sacavem (Portugal) and Universidade de Lisboa, Instituto de Biofisica e Engenharia Biomedica, Campo Grande, 1749-016 Lisboa (Portugal); Gil, Octavia Monteiro [Instituto Tecnologico e Nuclear, Estrada Nacional no 10, 2686-953 Sacavem (Portugal); Almeida, Pedro [Universidade de Lisboa, Instituto de Biofisica e Engenharia Biomedica, Campo Grande, 1749-016 Lisboa (Portugal); Vaz, Pedro [Instituto Tecnologico e Nuclear, Estrada Nacional no 10, 2686-953 Sacavem (Portugal)

    2011-09-15

    Exposure to radon and its progenies represents one of the greatest risks of ionizing radiation from natural sources. Nowadays, these risks are assessed by the extrapolation of biological effects observed from epidemiological data. In the present study, we made a dose response curve, to evaluate the in vitro response of A549 human lung cells to {alpha}-radiation resulting from the decay of a {sup 210}Po source, evaluated by the cytokinesis blocked micronuclei assay. The clonogenic assay was used to measure the survival cell fraction. As expected, the results revealed an increase of cellular damage with increased doses made evident from the increased number of micronuclei (MN) per binucleated cell (BN). Besides this study involving the biological effects induced by direct irradiation, and due to the fact that radiation-induced genomic instability is thought to be an early event in radiation carcinogenesis, we analyzed the genomic instability in early and delayed untargeted effects, by using the medium transfer technique. The obtained results show that unirradiated cells exposed to irradiated medium reveal a higher cellular damage in earlier effects when compared to the delayed effects. The obtained results may provide clues for the biodosimetric determination of radon dose to airway cells at cumulative exposures.

  10. 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.

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

    Science.gov (United States)

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

    2015-10-19

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

  12. 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.

  13. Oxidative damage to DNA and repair induced by Norwegian wood smoke particles in human A549 and THP-1 cell lines

    DEFF Research Database (Denmark)

    Danielsen, Pernille Høgh; Loft, Steffen; Kocbach, Anette;

    2009-01-01

    particulate matter (WSPM), authentic traffic-generated particles, mineral PM and standard reference material (SRM2975) of diesel exhaust particles in human A549 lung epithelial and THP-1 monocytic cell lines. DNA damage was measured as strand breaks (SB) and formamidopyrimidine DNA glycosylase (FPG) sites......Genotoxic effects of traffic-generated particulate matter (PM) are well described, whereas little data are available on PM from combustion of biomass and wood, which contributes substantially to air pollution world wide. The aim of this study was to compare the genotoxicity of wood smoke...

  14. Culture phases, cytotoxicity and protein expressions of agarose hydrogel induced Sp2/0, A549, MCF-7 cell line 3D cultures.

    Science.gov (United States)

    Ravi, Maddaly; Kaviya, S R; Paramesh, V

    2016-05-01

    Advancements in cell cultures are occurring at a rapid pace, an important direction is culturing cells in 3D conditions. We demonstrate the usefulness of agarose hydrogels in obtaining 3 dimensional aggregates of three cell lines, A549, MCF-7 and Sp2/0. The differences in culture phases, susceptibility to cisplatin-induced cytotoxicity are studied. Also, the 3D aggregates of the three cell lines were reverted into 2D cultures and the protein profile differences among the 2D, 3D and revert cultures were studied. The analysis of protein profile differences using UniProt data base further augment the usefulness of agarose hydrogels for obtaining 3D cell cultures.

  15. Induction of Apoptotic Effects of Antiproliferative Protein from the Seeds of Borreria hispida on Lung Cancer (A549 and Cervical Cancer (HeLa Cell Lines

    Directory of Open Access Journals (Sweden)

    S. Rupachandra

    2014-01-01

    Full Text Available A 35 KDa protein referred to as F3 was purified from the seeds of Borreria hispida by precipitation with 80% ammonium sulphate and gel filtration on Sephadex G-100 column. RP-HPLC analysis of protein fraction (F3 on an analytical C-18 column produced a single peak, detected at 220 nm. F3 showed an apparent molecular weight of 35 KDa by SDS PAGE and MALDI-TOF-MS analyses. Peptide mass fingerprinting analysis of F3 showed the closest homology with the sequence of 1-aminocyclopropane-1-carboxylate deaminase of Pyrococcus horikoshii. The protein (F3 exhibited significant cytotoxic activity against lung (A549 and cervical (HeLa cancer cells in a dose-dependent manner at concentrations ranging from 10 µg to 1000 µg/mL, as revealed by the MTT assay. Cell cycle analysis revealed the increased growth of sub-G0 population in both cell lines exposed to a concentration of 1000 µg/mL of protein fraction F3 as examined from flow cytometry. This is the first report of a protein from the seeds of Borreria hispida with antiproliferative and apoptotic activity in lung (A549 and cervical (HeLa cancer cells.

  16. Cytotoxicity study of Piper nigrum seed mediated synthesized SnO2 nanoparticles towards colorectal (HCT116) and lung cancer (A549) cell lines.

    Science.gov (United States)

    Tammina, Sai Kumar; Mandal, Badal Kumar; Ranjan, Shivendu; Dasgupta, Nandita

    2017-01-01

    Different sized tetragonal tin oxide nanoparticles (SnO2 NPs) were synthesized using Piper nigrum seed extract at three different calcination temperatures (300, 500, 900°C) and these nanoparticles (NPs) were characterized by transmission electron microscopy (TEM), X-ray diffraction (XRD), dynamic light scattering (DLS) and Fourier transform infrared spectrophotometry (FT-IR). The optical properties were studied using UV-Vis and photoluminescence (PL) spectrophotometers. The generation of reactive oxygen species (ROS) was monitored by using a fluorescence spectrophotometer and fluorescence microscope. The cytotoxicity of the synthesized SnO2 NPs was checked against the colorectal (HCT116) and lung (A549) cancer cell lines and the study results show that SnO2 NPs were toxic against cancer cell lines depending on their size and dose. IC50 values of SnO2 NPs having average particle sizes of 8.85±3.5, 12.76±3.9 and 29.29±10.9nm are 165, 174 and 208μgL(-1) against HCT116, while these values are 135, 157 and 187μgL(-1) against A549 carcinoma cell lines, respectively. The generated ROS were responsible for the cytotoxicity of SnO2 NPs to the studied cancer cells and smaller size NPs generated more ROS and hence showed higher cytotoxicity over larger size NPs. The results of this study suggest that the synthesized stable nanoparticles could be a potent therapeutic agent towards cancerous cell lines.

  17. 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.

  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. Forkhead box protein A1 inhibits the expression of uncoupling protein 2 in hydrogen peroxide-induced A549 cell line.

    Science.gov (United States)

    Song, Lan; Xu, Zhaojun; Li, Ling; Hu, Mei; Cheng, Lijuan; Chen, Lingli; Zhang, Bo

    2014-01-01

    Forkhead box protein A1 (FoxA1) is a transcription factor that is involved in embryonic development and cell differentiation. In this study, we show that hydrogen peroxide (H2O2) treatment upregulated expression of FoxA1 and UCP2 in the A549 cell line. Overexpression of FoxA1 by full-length complementary DNA reduced UCP2 expression, while silencing of FoxA1 expression by small interfering RNA significantly increased UCP2 levels. FoxA1 binds to a site from -919 to -913 bp relative to the UCP2 transcription start site. The overexpression of FoxA1 promoted the DNA binding activity and attenuated the transcription of UCP2 promoter as shown by electromobility shift, chromatin immunoprecipitation assays, and luciferase reporter assay. These data indicate an important role of FoxA1 in regulating expression of UCP2.

  20. 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.

  1. Artesunate induces AIF-dependent apoptosis in A549 cells

    Science.gov (United States)

    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.

  2. Evaluation of Synergetic Anticancer Activity of Berberine and Curcumin on Different Models of A549, Hep-G2, MCF-7, Jurkat, and K562 Cell Lines

    Directory of Open Access Journals (Sweden)

    Acharya Balakrishna

    2015-01-01

    Full Text Available Ayurvedic system of medicine is using Berberis aristata and Curcuma longa herbs to treat different diseases including cancer. The study was performed to evaluate the synergetic anticancer activity of Berberine and Curcumin by estimating the inhibition of the cell proliferation by cytotoxicity assay using MTT method on specified human cell lines (A549, Hep-G2, MCF-7, Jurkat, and K562. All the cells were harvested from the culture and seeded in the 96-well assay plates at seeding density of 2.0 × 104 cells/well and were incubated for 24 hours. Test items Berberine with Curcumin (1 : 1, Curcumin 95% pure, and Berberine 95% pure were exposed at the concentrations of 1.25, 0.001, and 0.5 mg/mL, respectively, and incubated for a period of 48 hours followed by dispensing MTT solution (5 mg/mL. The cells were incubated at 37 ± 1°C for 4 hours followed by addition of DMSO for dissolving the formazan crystals and absorbance was read at 570 nm. Separate wells were prepared for positive control, controls (only medium with cells, and blank (only medium. The results had proven the synergetic anticancer activity of Berberine with Curcumin inducing cell death greater percentage of >77% when compared to pure curcumin with <54% and pure Berberine with <45% on average on all cell line models.

  3. Reversal effect of toremifene (TOR) on A549 /cDDP lung cancer cell line with resistance to cisplatin%托瑞米芬逆转肺癌耐药细胞株A549/cDDP耐药性的研究

    Institute of Scientific and Technical Information of China (English)

    刘利则; 夏莉; 刘玉侠; 段北野

    2012-01-01

    目的:探讨托瑞米芬(TOR)对耐顺铂(cDDP)细胞株A549的逆转作用,为临床应用提供实验数据.方法:用不同浓度的托瑞米芬单独及与cDDP联合与耐药细胞A549共同培养,通过MTT法和流式细胞仪法检测其对A549/cDDP的逆转及增敏效果.结果:经不同浓度的托瑞米芬单独及与cDDP联合与耐药细胞A549共同培养后,单独TOR(5 μmol/L、10 μmol/L)对A549/cDDP细胞的增殖均无明显影响,各组间数据无明显差异(P>0.05).当cDDP与TOR联合作用时无论TOR终浓度5 μmol/L或10 μmol/L,均能明显增加cDDP对A549/cDDP细胞的敏感性(P<0.05,P<0.001).其IC50值分别为39.06 μmol/L和30.64 μmol/L,逆转倍数分别为2.05倍和2.65倍.cDDP+TOR终浓度5 μmol/L与cDDP+TOR终浓度10 μmol/L之间除了在cDDP浓度为200 μmol/L时两者有差异(P<0.05)外其它均无明显差异.结论:托瑞米芬与DDP联合应用可以提高A549/cDDP的逆转及治疗效果.%Objective: To investigate the reversal effect of toremifene (TOR) on A549/cDDP lung cancer cell line, which resistance to cisplatin, and provide the experimental data for clinical application. Methods: A549 cell line was cultured with different concentrations of toremifene, with or without cisplatin. Sensitive effect of toremifene on A549/cDDP was measured by MTT assay. The cell apoptotic activity was determined with Annexin V/PI staining by flow cytometry. Results:Using TOR (5 μmol / L, 10 μmol / L) alone had no significant effect on proliferation of A549/cDDP cell line (P > 0. 05) , but TOR combined with cDDP (the final concentration of TOR was 5 (μmol/Lor 10 μmol/L) significantly increased the sensitive effect of A549/cDDP cells to cDDP (P<0.05, P< 0. 001) . The value of IC50 elevated up to 39. 06 μmol/L and 30. 64 μmol/L, and the fold of reversal effect was 2. 05 and 2. 65 times, respectively. In addition to 200 μmol/L of cDDP, there was no significant differences between 5 μmol/L and 10 μmol/L of cDDP combined

  4. 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

  5. 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.

  6. 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.

  7. 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.

  8. 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.

  9. 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.

  10. 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.

  11. Effect of siRNA-mediated silencing Bmi-1 gene expression on the proliferation of lung cancer cell line A549 in vitro and in vivo%Bmi-1-siRNA对肺腺癌A549细胞体内外增殖能力的影响

    Institute of Scientific and Technical Information of China (English)

    郑翔宇; 朱杰; 王艺芳; 刘纯青; 刘奔; 杨春辉; 刘丹丹; 孟秀香

    2013-01-01

    背景与目的:原癌基因Bmi-1是多梳基因家族中的一员,能调节正常干细胞和肿瘤干细胞的自我更新能力。近年来发现其在多种恶性肿瘤中表达上调。本文旨在观察Bmi-1基因沉默对肺腺癌A549细胞体内外增殖的影响,并初步探讨其机制。方法:根据本实验室设计的4条针对Bmi-1的小干扰RNA(siRNA)序列,选择一条已经证实最有效的序列作为靶序列和一条随机序列作为阴性对照,构建重组逆转录病毒siRNA表达载体并将其转染入A549细胞中;应用RT-PCR和蛋白质印迹法(Western blot)检测对Bmi-1基因的沉默效果;应用MTT比色法、台盼蓝拒染法及平板克隆形成实验检测Bmi-1-siRNA对A549细胞体外增殖的影响;利用流式细胞仪分析各组细胞的细胞周期;通过裸鼠腋窝皮下接种各组细胞,观察Bmi-1-siRNA对A549细胞在裸鼠体内的致瘤能力的影响;Western blot检测PTEN、p-AKT、cyclin D1、P21、P27蛋白表达。结果:Bmi-1-siRNA有效地沉默了Bmi-1基因mRNA和蛋白的表达;沉默Bmi-1基因的表达能够抑制A549细胞的体内外增殖能力,使干扰组细胞的细胞周期阻滞于G1期;沉默Bmi-1基因的表达后,干扰组细胞中PTEN、P21、P27蛋白增加,p-AKT、cyclin D1蛋白表达降低。结论:Bmi-1-siRNA通过使细胞周期阻滞于G1期来抑制肺腺癌A549的体内外增殖能力,这种抑制作用涉及cyclin D1和p-AKT表达下降以及P21/P27和PTEN的表达上调。%Background and purpose:The pro-oncogene Bmi-1 is a member of the polycomb-group family, can regulation of the proliferation and self-renewal of normal and tumor stem cells. In recent years, Bmi-1 has been found that it is overexpressed in varieties of human malignant tumors. The study aimed to observe the effects of Bmi-1-siRNA on the growth capacity of lung cancer cell line A549 in vivo and in vivo, and explore its mechanism. Methods:The most effective one as a target

  12. Radiation-induced p53 protein response in the A549 cell line is culture growth-phase dependent

    Energy Technology Data Exchange (ETDEWEB)

    Johnson, N.F.; Gurule, D.M.; Carpenter, T.R.

    1995-12-01

    One role of the p53 tumor suppressor protein has been recently revealed. Kastan, M.B. reported that p53 protein accumulates in cells exposed to ionizing radiation. The accumulation of p53 protein is in response to DNA damage, most importantly double-strand breaks, that results from exposure to ionizing radiation. The rise in cellular p53 levels is necessary for an arrest in the G{sub 1} phase of the cell cycle to provide additional time for DNA repair. The p53 response has also been demonstrated to enhance PCNA-dependent repair. p53 is thus an important regulator of the cellular response to DNA-damaging radiation. From this data, it can be concluded that the magnitude of the p53 response is not dependent on the phase of culture growth.

  13. Effects of epigallocatechin gallate on cisplatin-induced cytotoxicity in HEK293 and A549 cell lines%EGCG对顺铂损伤HEK293细胞及杀伤A549细胞的影响

    Institute of Scientific and Technical Information of China (English)

    连燕娜; 郭淑琴; 周绮云; 高兆兰; 王海; 高丽萍

    2016-01-01

    目的:探讨表儿茶素没食子酸酯(EGCG)对顺铂(DDP)致人胚肾HEK293细胞损伤及对DDP杀伤人肺癌A549细胞的影响.方法:体外培养HEK293细胞和A549细胞,分为对照组、DDP组和DDP+EGCG组,DDP组和DDP+EGCG组同时建立DDP损伤模型,MTT法检测EGCG和/或DDP对HEK293细胞和A549细胞存活率的影响.结果:EGCG对HEK293细胞的IC50值为61.6 mg/L.当EGCG浓度<40 mg/L时,对DDP诱导的HEK293细胞损伤没有显著性影响,EGCG浓度≥40 mg/L时,可显著性增强DDP对HEK293细胞的损伤作用.EGCG对A549细胞的IC50值为33.6 mg/L.当EGCG浓度≥32 mg/L时,可显著增强DDP对A549细胞的杀伤作用.结论:EGCG对DDP所致HEK293细胞损伤无保护作用,但EGCG对癌细胞毒性作用大于其对正常细胞的毒性,且当EGCG与DDP同时使用时可以加重A549细胞损伤.

  14. Induction and repair of DNA cross-links induced by sulfur mustard in the A-549 cell line followed by a comet assay.

    Science.gov (United States)

    Jost, Petr; Svobodova, Hana; Stetina, Rudolf

    2015-07-25

    Sulfur mustard is a highly toxic chemical warfare agent with devastating impact on intoxicated tissues. DNA cross-links are probably the most toxic DNA lesions induced in the cell by sulfur mustard. The comet assay is a very sensitive method for measuring DNA damage. In the present study using the A-549 lung cell line, the comet assay protocol was optimized for indirect detection of DNA cross-links induced by sulfur mustard. The method is based on the additional treatment of the assayed cells containing cross-links with the chemical mutagen, styrene oxide. Alkali-labile adducts of styrene oxide cause DNA breaks leading to the formation of comets. A significant dose-dependent reduction of DNA migration of the comet's tail was found after exposing cells to sulfur mustard, indicative of the amount of sulfur mustard induced cross-links. The remarkable decrease of % tail DNA could be observed as early as 5min following exposure to sulfur mustard and the maximal effect was found after 30min, when DNA migration was reduced to the minimum. Sulfur mustard preincubated in culture medium without cells lost its ability to induce cross-links and had a half-life of about 15min. Pre-incubation longer than 30min does not lead to a significant increase in cross-links when applied to cells. However, the amount of cross-links is decreased during further incubation due to repair. The current modification of the comet assay provides a useful tool for detecting DNA cross-links induced by sulfur mustard and could be used for detection of other DNA cross-linking agents such as chemotherapeutic drugs.

  15. 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.

  16. 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.

  17. 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

  18. 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

    Directory of Open Access Journals (Sweden)

    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.

  19. CD147慢病毒表达载体的构建及稳定转染A549细胞系的建立%Construction of a CD147 Lentiviral Expression Vector and Establishment of Its Stably Transfected A549 Cell Line

    Institute of Scientific and Technical Information of China (English)

    杨绍兴; 汤传昊; 王思涵; 宋三泰; 刘晓晴

    2012-01-01

    背景与目的 CD147是一类位于肿瘤细胞膜表面的跨膜糖蛋白,可促进肿瘤的浸润和转移.本研究拟构建CD147慢病毒表达载体,建立稳定过表达CD147的人肺腺癌A549细胞系,观察过表达CD147后对MMP-9及细胞增殖、侵袭能力的影响.方法 RT-PCR扩增CD147基因全长序列,将序列插入pEGFP载体,构建pEGFP-CD147慢病毒表达载体,随后转入293FT细胞中进行慢病毒包装,用获得的慢病毒毒液感染人肺腺癌细胞系A549,建立稳定过表达CD147的A549细胞系.Real-time PCR检测MMP-9的变化情况,CCK-8及Transwell法检测人肺腺癌细胞增殖、侵袭能力的变化.结果 经限制性内切酶鉴定及测序分析,成功构建了pEGFP-CD 147慢病毒表达载体质粒.Real-time PCR和Western blot检测显示,与对照组相比,转染pEGFP-CD147慢病毒表达载体组的细胞,CD147的表达在mRNA和蛋白两个水平均增高,成功建立了A549-CD147细胞系.上调CD147的表达后,MMP-9的mRNA表达水平明显升高.同时,A549-CD147细胞增殖和侵袭能力明显增加(P<0.05).结论 成功构建CD147慢病毒表达载体和A549-CD147细胞系,过表达CD 147可上调MMP-9的表达,增强人肺腺癌细胞的增殖和侵袭能力.%Background and objective CD 147, a type of transmembrane glycopiotein 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 CD 147 on MMP-9 proliferation as well as on the invasive ability of human lung adenocarcinoma cells. Methods Full-length CD 147 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

  20. 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...

  1. 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

    Directory of Open Access Journals (Sweden)

    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.

  2. Biochemical synthesis of silver nanoprticles using filamentous fungi Penicillium decumbens (MTCC-2494) and its efficacy against A-549 lung cancer cell line.

    Science.gov (United States)

    Majeed, Shahnaz; Abdullah, Mohd Syafiq Bin; Dash, Gouri Kumar; Ansari, Mohammed Tahir; Nanda, Anima

    2016-08-01

    Biosynthesis of silver and other metallic nanoparticles is one of the emerging research area in the field of science and technology due to their potentiality, especially in the field of nano-biotechnology and biomedical sciences in order to develop nanomedicine. In our present study, Penicillium decumbens (MTCC-2494) was brought from Institute of Microbial Technology (IMTECH) Chandigarh and employed for extracellular biological synthesis of silver nanoparticles. Ag-NPs formation was appeared with a dark brown color inside the conical flask. Characterization of Ag-NPs were done by UV-Spectrophotometric analysis which showed absorption peak at 430 nm determines the presence of nanoparticles, Fourier transform infrared (FT-IR) spectroscopic analysis, showed amines and amides are the possible proteins involved in the stabilization of nanoparticles as capping agent. Atomic force Microscopy (AFM) confirmed the particle are spherical, size was around 30 to 60 nm and also the roughness of nanoparticles. Field emission scanning electron microscopy (FE-SEM) showed the topology of the nanoparticles and were spherical in shape. The biosynthesis process was found fast, ecofriendly and cost effective. Nano-silver particle was found to have a broad antimicrobial activity and also it showed good enhancement of antimicrobial activity of Carbenicillin, Piperacillin, Cefixime, Amoxicillin, Ofloxacin and Sparfloxacin in a synergistic mode. These Ag-NPs showed good anti-cancer activity at 80 μg·mL(-1)upon 24 hours of incubation and toxicity increases upon 48 hours of incubation against A-549 human lung cancer cell line and the synergistic formulation of the antibiotic with the synthesized nanoparticles was found more effective against the pathogenic bacteria studied.

  3. 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.

  4. 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.

  5. 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...

  6. Influence of Ciglitazone on A549 Cells Growth in vitro and in vivo and Mechanism

    Institute of Scientific and Technical Information of China (English)

    2006-01-01

    The effect and mechanism of the ciglitazone on lung cancer cells A549 growth in vitro and in vivo were studied. Various concentrations of ciglitazone were added to the cultured A549 line, and the proliferation and differentiation of A549 cells were examined by MTT and cytometry analysis. A549 cells (1 × 106/mouse) were inoculated subcutaneously into 20 nude mice, which were randomly divided into two groups: the control group, the ciglitazone treated group. The weights of subcutaneous tumors were measured. The expression of cyclin D1 and P21 in the lung was detected by immohistochemistry and Western blot respectively. The results showed that the proliferation of A549 was inhibited significantly by ciglitazone in a dose- and time-dependent manner. There were more cells arrested in G1/G0 phase and the expression of PPARγ was markedly upregulated in ciglitazone-treated group. Direct injection of ciglitazone into A549-induced tumors could suppress tumor growth in nude mice and the growth inhibitory rate was 36 %. The expression of cyclin D1 was decreased and P21 increased significantly in ciglitazone-treated group as compared with control group. It was concluded that ciglitazone could inhibit A549 proliferation dose-dependently and time-dependently and induce differentiation, which might be related to the modulation of cell cycle interfered by PPARγ.

  7. 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

  8. 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

  9. 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.

  10. 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.

  11. 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.

  12. 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.

  13. 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.

  14. 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

  15. Study of the photodynamic effect on the A549 cell line by atomic force microscopy and the influence of green tea extract on the production of reactive oxygen species.

    Science.gov (United States)

    Tomankova, Katerina; Kolarova, Hana; Bajgar, Robert; Jirova, Dagmar; Kejlova, Kristina; Mosinger, Jiri

    2009-08-01

    We studied the morphology of the A549 cell line (human lung carcinoma cells) before and after photodynamic therapy (PDT) by atomic force microscopy. PDT was induced by an efficient light-emitting diode source with total light dose of 15 J cm(-2) in the presence of the sensitizer zinc-5,10,15,20-tetrakis(4-sulfonatophenyl)porphyrine. In the presence of molecular oxygen, light activation of the photosensitizer, which accumulates in cancer cells, leads to the local production of reactive oxygen species (ROS). This is one of several reasons leading to cell death, and in some cases we could observe signs of apoptosis. We detected the kinetics of ROS production to be dependent on the presence of green tea extract.

  16. 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.

  17. Transfection of gene Livin α/β into A549 cells and separate effect on the cell growth

    Institute of Scientific and Technical Information of China (English)

    SUN Jian-guo; LIAO Rong-xia; CHEN Zheng-tang; WANG Zhi-xin; ZHANG Qing; HU Yi-de; WANG Dong-lin

    2005-01-01

    Objective:To express two Livin isoforms (Livin α & β genes) with transfection techniques in A549 cell line respectively in order to observe their effect on growth of cell line. Methods:Two eukaryotic expression vectors of Livin, pcDNA3.1-Livin α & β, were transfected into A549 cell line by electroporation. Then G418-resistant clones were screened. RT-PCR, Northern blot and immunofluorescence cytochemistry were used to detect Livin α & β expression level in the transfected cells. Finally, observation of cell morphology, growth curve assay and colony formation analysis were performed to explore the effect of Livin on growth of the cells. Results:Livin α & β were expressed in transfected A549 cells, and induced a faster cell growth, shorter doubling time and stronger cell colony forming ability, yet had no morphology change.Conclusion:Both isoforms can accelerate the growth of A549 cells, indicating a close relationship between Livin expression and the genesis and development of lung cancer. The expression of Livin α & β in A549 cells provides basis for further study of their different biological functions of anti-apoptosis and of their role in lung cancer cell resistance to radiotherapy and chemotherapy.

  18. X线对肺癌细胞株A549 XRCC2和XRCC3表达水平的影响%Effect of X-ray on expression levels of XRCC2 and XRCC3 in lung cancer cell line A549

    Institute of Scientific and Technical Information of China (English)

    史卫林; 李坚; 陈萍; 戴春华

    2009-01-01

    目的:研究X线对肺癌细胞X线修复交叉互补基因2((X-ray repair cross complementing gene 2,XRCC2))与XRCC3表达水平的影响,探讨DNA同源重组修复机制在肺癌放疗过程中的作用.方法:以噻唑蓝还原法(MTT)检测X线对肺腺癌细胞株A549抑制率的影响,实时荧光定量RT-PCR技术检测X线处理肺癌细胞(人肺腺癌细胞株A549)后XRCC2和XRCC3 mR-NA的表达水平.结果:肺癌细胞抑制率多数情况下随X线照射时间的延长及照射剂量的增大,细胞增殖抑制率增加,呈照射时间依赖性(P<0.05)和剂量依赖性(P<0.05),除了16Gy组与32Gy组比较无统计学意义(P=0.211).X线照射后肺癌细胞XRCC2与XRCC3 mRNA的表达水平均先增高后降低,在照射后48 h表达水平达高峰(P<0.05),且随着照射剂量的增大,XRCC2与XR-CC3 mRNA的表达水平也随之增加(P<0.05).结论:X线照射可引起肺癌细胞XRCC2与XRCC3 mRNA表达水平的明显改变,表明DNA同源重组修复机制可能在肺癌放疗耐受中起了重要的作用.%Objective:To study the effect of X-ray on expression levels of X-ray repair cross-complementing gene2(XRCC2)and XRCC3 in lung cancer cells,and explore the effect of DNA homologous recombination repair mechanism in radiotherapy of lung cancer. Methods:Cell inhibition ratio was measured using MTT assay. The expression levels of XRCC2 mRNA and XRCC3 mRNA in lung cancer cell line A549 were measured by RFQ-PCR assay. Results:The rate of proliferation inhibiting of lung cancer cells increased in line with the prolong radiation time of X-ray (P < 0.05) and the increase of radiation dose(P < 0.05) ,but there was no difference between the groups of 16 Gy and 32 Gy (P=0.211). The expression levels of XRCC2 and XRCC3 mRNA in these lung cancer cells increased significantly after treated with X-ray,and then decreased. The expression levels of XRCC2 and XRCC3 mRNA peaked at 48 h after X-ray treatment (P < 0.05). The expression levels of XRCC2 and XRCC3 m

  19. Isolation of eastern equine encephalitis virus in A549 and MRC-5 cell cultures.

    Science.gov (United States)

    Sotomayor, E A; Josephson, S L

    1999-07-01

    Eastern equine encephalitis (EEE) has been diagnosed either serologically or by virus isolation. Until now, the recovery of EEE virus has been delegated to reference laboratories with the expertise and resources needed to amplify the virus in a susceptible vertebrate host and/or to isolate and identify the virus in cell culture. We report a case in which EEE virus was recovered directly from a patient's cerebrospinal fluid in A549 and MRC-5 cell cultures. Many clinical virology laboratories routinely use these cells to recover adenovirus, herpes simplex virus, and enterovirus. To the best of our knowledge, this is the first report of isolation of EEE virus in A549 cell culture. This report demonstrates the possibility of recovery of EEE virus in cell culture without the necessity of bioamplification or maintaining unusual cell lines.

  20. Inhibition of epithelial-mesenchymal transition in A549 cell by transfected Napsin A

    Institute of Scientific and Technical Information of China (English)

    ZHENG Jin-xu; GUAN Shu-hong; XU Qing; LIU Ji-zhu; SONG Ping

    2012-01-01

    Background Epithelial-mesenchymal transition is a cellular process characterized by the loss of cell adhesion,inhibition of E-cadherin expression,and increased cell mobility.Cells without Napsin A are susceptible to transition.Further studies are required to investigate whether this transition can be reversed by restoration of Napsin A.Methods A Napsin A expression vector PLJM1-Napsin A plasmid was constructed and then transfected into the epithelial cell line A549 by lentivirus transfection to obtain A549-PLJM1-Napsin A cell line.Cell proliferation was assayed by 3-[4,5-dimethylthiazol-2-yl]-2,5 diphenyltetrazolium bromide and cell cycle was measured by flow cytometry.The E-cadherin,type I collagen,and focal adhesion kinase mRNA level was detected by reverse transcription-polymerase chain reaction.The Napsin A,E-cadherin,type I collagen,and focal adhesion kinase protein level in A549 cells was detected by Westen blotting.Results Transforming growth factor-β1 induced epithelial-mesenchymal transition in A549 cells,as demonstrated by significant reduction of E-cadherin mRNA and protein levels (P <0.01) as well as up-regulation of type I collagen (P <0.01 ).Transfection of Napsin A in A549 cells can partially block the transforming growth factor-β1-regulated expression of E-cadherin and type I collagen (P <0.01).In addition,transforming growth factor-β1-induced cell proliferation was inhibited by Napsin A (P <0.01).Further study demonstrated that Napsin A caused Go/G1 arrest and inhibited the expression of focal adhesion kinase (P <0.01),a key protein in the integrin signaling pathway,in the in vitro epithelial-mesenchymal transition model.Conclusions Sustained Napsin A expression in A549 cells can inhibit the transforming growth factor-β1-induced epithelial-mesenchymal transition.This may be due to the Napsin A-mediated inhibition of focal adhesion kinase expression and integrin signaling pathway.

  1. 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.

  2. Protection of A549 cells against the toxic effects of sulphur mustard by hexamethylenetetramine.

    Science.gov (United States)

    Lindsay, C D; Hambrook, J L

    1997-02-01

    The A549 cell line was used as a model of the deep lung to study the toxicity and mechanism of action of sulphur mustard (HD), using the neutral red (NR) dye retention and gentian violet (GV) assays as indices of cell viability. It was found that exposure to concentrations in excess of 40 microM HD resulted in a rapid onset of toxicity. Exposure to 1000 microM HD reduced viability in A549 cell cultures to 61% after 2 h (control cultures = 100%), whereas exposure to 40 microM HD did not result in deleterious effects until 26 h at which point viability fell to only 84% (NR assay). Agarose gel electrophoresis of cell cultures exposed to 40 and 1000 microM HD and harvested at 4.5, 19 and 43 h after exposure to HD, indicated that cell death was due to necrosis, despite the observation that at the higher concentration of HD cells displayed many of the features common to cells undergoing apoptotic death. The ability of hexamethylenetetramine (HMT) to protect A549 cells against the effects of an LC50 challenge dose of HD was assessed using the GV and NR assays. It was found that HMT (15 mM) could protect cells against the effects of HD though HMT had to be present at the time of HD challenge. Cultures treated with HD only were 49% viable at 48 h after HD challenge, compared to 101% for protected cultures (NR assay) and 58% and 91% for unprotected and protected cultures respectively using the GV assay. Morphological observations of GV and NR stained cultures confirmed these findings. HMT concentrations of 2.5 to 25 mM were used. Maximal protection against the toxic effects of HD (LC50) was found at 10 to 25 mM HMT. Over this concentration range, HMT did not exert any toxic effects on A549 cells. Pretreatment of A549 cultures with HMT followed by its removal prior to HD challenge had no protective effect. Similarly, treating cultures with HD followed by addition of HMT did not increase the viability of the cultures, even if the HMT was added immediately after HD exposure

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

    Directory of Open Access Journals (Sweden)

    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. A water soluble vitamin B12-ReI fluorescent conjugate for cell uptake screens: use in the confirmation of cubilin in the lung cancer line A549.

    Science.gov (United States)

    Vortherms, Anthony R; Kahkoska, Anna R; Rabideau, Amy E; Zubieta, Jon; Andersen, Louise Lund; Madsen, Mette; Doyle, Robert P

    2011-09-21

    A water soluble vitamin B(12)-rhenium conjugate was synthesized and used in concert with intrinsic factor to screen for cubilin receptor-mediated uptake in lung cancer cells. Internalization of the conjugate demonstrated that it could be used to rapidly screen for the cubilin receptor in living cells, subsequently confirmed with Western blotting and RT-PCR.

  5. In vitro toxicity of cationic micelles and liposomes in cultured human hepatocyte (HepG2) and lung epithelial (A549) cell lines

    DEFF Research Database (Denmark)

    Roursgaard, Martin; Knudsen, Kristina Bram; Northeved, Helle;

    2016-01-01

    The aim of this study was to compare the effects of cationic micelle and liposome drug delivery systems on liver and lung cells in a toxicological in vitro screening model, with observations on cytotoxicity and genotoxicity. A screening battery was established for assessment of a broad range...

  6. 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

    of etheno-adducts or bulky DNA adducts. Furthermore, mRNA expression of the proinflammatory genes monocyte chemoattractant protein-1, interleukin-8, and tumor necrosis factor-R as well as the oxidative stress gene heme oxygenase-1 was upregulated in the THP-1 cells especially by WSPM and ambient PM sampled...

  7. SOCS3 inhibiting migration of A549 cells correlates with PYK2 signaling in vitro

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    Zhang Qingfu

    2008-05-01

    Full Text Available Abstract Background Suppressor of cytokine signaling 3 (SOCS3 is considered to inhibit cytokine responses and play a negative role in migration of various cells. Proline-rich tyrosine kinase 2 (PYK2 is a non-receptor kinase and has been found crucial to cell motility. However, little is known about whether SOCS3 could regulate PYK2 pro-migratory function in lung cancer. Methods The methylation status of SOCS3 was investigated in HBE and A549 cell lines by methylation-specific PCR. A549 cells were either treated with a demethylation agent 5-aza-2'-deoxycytidine or transfected with three SOCS3 mutants with various functional domains deleted. Besides, cells were pretreated with a proteasome inhibitor β-lactacystin where indicated. The effects of SOCS3 up-regulation on PYK2 expression, PYK2 and ERK1/2 phosphorylations were assessed by western blot using indicated antibodies. RT-PCR was used to estimate PYK2 mRNA levels. Transwell experiments were performed to evaluate cell migration. Results SOCS3 expression was found impaired in A549 cells and higher PYK2 activity was correlated with enhanced cell migration. We identified that SOCS3 was aberrantly methylated in the exon 2, and 5-aza-2'-deoxycytidine restored SOCS3 expression. Reactivation of SOCS3 attenuated PYK2 expression and phosphorylation, cell migration was inhibited as well. Transfection studies indicated that exogenous SOCS3 interacted with PYK2, and both the Src homology 2 (SH2 and the kinase inhibitory region (KIR domains of SOCS3 contributed to PYK2 binding. Furthermore, SOCS3 was found to inhibit PYK2-associated ERK1/2 activity in A549 cells. SOCS3 possibly promoted degradation of PYK2 in a SOCS-box-dependent manner and interfered with PYK2-related signaling events, such as cell migration. Conclusion These data indicate that SOCS3 negatively regulates cell motility and decreased SOCS3 induced by methylation may confer a migration advantage to A549 cells. These results also suggest a

  8. 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.

  9. Salvianolic acid A positively regulates PTEN protein level and inhibits growth of A549 lung cancer cells

    Science.gov (United States)

    BI, LEI; CHEN, JIANPING; YUAN, XIAOJING; JIANG, ZEQUN; CHEN, WEIPING

    2013-01-01

    Salvianolic acid A (Sal A) is an effective compound extracted from Salvia miltiorrhiza which has been used in the treatment of various diseases. Preliminary data indicate that Sal A treatment has a specific anti-lung cancer effect. However, the manner in which Sal A regulates cancer growth remains unknown. In this study, the A549 lung cancer cell line and its response to Sal A treatment was examined. Results showed that Sal A treatment significantly decreased A549 cell growth, promoted partial apoptosis and increased mitochondrial membrane permeability. Western blot analysis showed that Sal A upregulated the phosphatase and tensin homolog (PTEN) protein level, while consistently downregulating Akt phosphorylation. These results indicate that Sal A negatively mediates A549 lung cancer cell line growth or apoptosis, most likely by positively regulating PTEN protein level. PMID:24648921

  10. The Effect of dcEFs on migration behavior of A549 cells and Integrin beta1 expression

    Directory of Open Access Journals (Sweden)

    Yunjie WANG

    2008-04-01

    Full Text Available Background and objective The effect of direct-current electric fields (dcEFs on cells attracted extensive attention. Moreover the metastasis and its potential are considered to be related to dcEFs. The aim is to study the effect of dcEFs on migration behavior of A549 cells, Integrin ?1 and its signal pathways. Methods According to exposure to 5 V/cm dcEFs or not and the time of exposure, the A549 cells were divided into 4 groups. Images were taken per 5 min within 2 h to recode the migration of the cells. The data of results were analyzed statistically. Results Most of A549cells exposed to the dcEFs aligned and elongated perpendicularly to the electric field lines and migrated to the cathode continually during 2 h. On the contrary, cells unexposed to dcEFs showed slightly random movements. Immunofluorescence showed that Integrin ?1 on plasma membrane polarized to the cathode of the dcEFs. Western blot showed that Integrin beta1 downstream signal pathways p-FAK and p-ERK were overexpressed in the dcEFs. Conclusion A549 cells have a galvanotatic feature of cathodal directed migration while exposed to the dcEFs. The polarization of Integrin beta1 and the promotion of its downstream signal pathways may play an important roles in the galvanotaxis of A549 cells.

  11. NMR studies of the relationship between the changes of membrane lipids and the cisplatin-resistance of A549/DDP cells

    Directory of Open Access Journals (Sweden)

    Huang Youguo

    2003-04-01

    Full Text Available Abstract Changes of membrane lipids in cisplatin-sensitive A549 and cisplatin-resistant A549/DDP cells during the apoptotic process induced by a clinical dose of cisplatin (30 μM were detected by 1H and 31P-NMR spectroscopy and by membrane fluidity measurement. The apoptotic phenotypes of the two cell lines were monitored with flow cytometry. The assays of apoptosis showed that significant apoptotic characteristics of the A549 cells were induced when the cells were cultured for 24 hours after treatment with cisplatin, while no apoptotic characteristic could be detected for the resistant A549/DDP cells even after 48 hours. The results of 1H-NMR spectroscopy demonstrated that the CH2/CH3 and Glu/Ct ratios of the membrane of A549 cells increased significantly, but those in A549/DDP cell membranes decreased. In addition, the Chol/CH3 and Eth/Ct ratios decreased for the former but increased for the latter cells under the same conditions. 31P-NMR spectroscopy indicated levels of phosphomonoesters (PME and ATP decreased in A549 but increased in A549/DDP cells after being treated with cisplatin. These results were supported with the data obtained from 1H-NMR measurements. The results clearly indicated that components and properties of membrane phospholipids of the two cell lines were significantly different during the apoptotic process when they were treated with a clinical dose of cisplatin. Plasma membrane fluidity changes during cisplatin treatment as detected with the fluorescence probe TMA-DPH also indicate marked difference between the two cell lines. We provided evidence that there are significant differences in plasma membrane changes during treatment of cisplatin sensitive A549 and resistant A549/DDP cells.

  12. 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...

  13. 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.

  14. Influence of suppressor gene p16 on retinoic acid inducing cancer cell A549 differentiation

    Institute of Scientific and Technical Information of China (English)

    2001-01-01

    Objective To investigate the role of suppressor gene p16 in the process of differential regulation of retinoic acid (RA) on the A549 lung cancer cells.Methods Tumor suppressor gene p16 was transferred into A549 cells and the cells were treated with all-trans retinoic acid (ATR) at the dosage of 5×10-6 mol/L for 4 d. After that, the proliferation and differentiation of A549 cells were examined by growth curve and cytometry analysis, the change of lung lineage-specific marker MUC1 was tested by immunohistochemical staining. Meanwhile, Western blot was used to observe the change of p16 protein expression in A549 cells treated with ATRA.Results ATRA could obviously inhibit the growth and induce the differentiation of A549 Cells that were transferred with p16 gene. There were more cells arrested in G1/G0 phase and the expression of MUG1 was markedly down-regulated than in control cells. The expression of p16 protein was up-regulated in A549 cells treated with ATRA.Conclusion Suppressor gene p16 could enhance the effects of RA and proliferated suppression and differential induction of A549 cells.

  15. 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.

  16. 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.

  17. 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.

  18. 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.

  19. DNA damage response signaling in lung adenocarcinoma A549 cells following gamma and carbon beam irradiation

    Energy Technology Data Exchange (ETDEWEB)

    Ghosh, Somnath [Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Trombay, Mumbai 400 085 (India); Narang, Himanshi, E-mail: himinarang@gmail.com [Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Trombay, Mumbai 400 085 (India); Sarma, Asitikantha [Radiation Biology Laboratory, Inter University Accelerator Centre, Aruna Asaf Ali Marg, New Delhi 110 067 (India); Krishna, Malini [Radiation Biology and Health Sciences Division, Bhabha Atomic Research Centre, Trombay, Mumbai 400 085 (India)

    2011-11-01

    Carbon beams (5.16 MeV/u, LET = 290 keV/{mu}m) are high linear energy transfer (LET) radiation characterized by higher relative biological effectiveness than low LET radiation. The aim of the current study was to determine the signaling differences between {gamma}-rays and carbon ion-irradiation. A549 cells were irradiated with 1 Gy carbon or {gamma}-rays. Carbon beam was found to be three times more cytotoxic than {gamma}-rays despite the fact that the numbers of {gamma}-H2AX foci were same. Percentage of cells showing ATM/ATR foci were more with {gamma}-rays however number of foci per cell were more in case of carbon irradiation. Large BRCA1 foci were found in all carbon irradiated cells unlike {gamma}-rays irradiated cells and prosurvival ERK pathway was activated after {gamma}-rays irradiation but not carbon. The noteworthy finding of this study is the early phase apoptosis induction by carbon ions. In the present study in A549 lung adenocarcinoma, authors conclude that despite activation of same repair molecules such as ATM and BRCA1, differences in low and high LET damage responses might be due to their distinct macromolecular complexes rather than their individual activation and the activation of cytoplasmic pathways such as ERK, whether it applies to all the cell lines need to be further explored.

  20. 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.

  1. 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.

  2. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Science.gov (United States)

    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.

  5. 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.

  6. CD147 siRNA 对肺腺癌细胞 A549生长转移的影响%Effect of CD147 siRNA on Cell Proliferation and Invasion of Lung Adenocarcinoma Cell Line A5 4 9

    Institute of Scientific and Technical Information of China (English)

    陈娟; 彭毅强; 梁伟军; 杨红忠

    2013-01-01

    [Objective]To explore the influence of CD147 expression in lung adenocarcinoma cell line A549 on cell proliferation and invasion.[Methods]The siRNA CD147 was transfected into human lung adenocarci-noma cell line A549 mediated by Lipofectamine TM2000.After transfection for 48h,real-time fluorescent quantitive PCR(qRT-PCR)and Western blot were used to assess the inhibition effect.The influence of CD147 siRNA on cell proliferation was determined by MTT method.The effect of CD147 siRNA on cell invasive abil-ity was detected by Transwell method.The expression of MMP-9 was detected by qRT-PCR and Western blot.[Results]After transfection,the expression of CD147 in lung adenocarcinoma cell line A549 significantly decreased,and cell proliferation and invasive ability reduced.After the expression of CD147 decreased,the ex-pression of MMP-9 mRNA and protein markedly decreased.There were significant differences(P <0.05).[Conclusion]The decreasing of CD147 expression can inhibit cell proliferation and invasion of lung adenocari-noma,which may be related to the down-regulation of MMP-9 expression.%[目的]探讨肺腺癌细胞 A549中 CD147的表达对细胞增殖和侵袭能力的影响。[方法]采用脂质体 LipofectamineTM 2000介导 CD147 siRNA 转染人肺腺癌细胞 A549,转染48 h 后采用实时荧光定量 PCR (qRT-PCR)和 Western blot 方法评价干扰效果,MTT 法检测 CD147 siRNA 转染对细胞增殖的影响;Tran-swell 法检测 CD147 siRNA 转染对细胞侵袭能力的影响;qRT-PCR 和 Western blot 方法检测 MMP-9的表达。[结果]转染后肺腺癌细胞系 A549内 CD147的表达显著降低,肺腺癌细胞系 A549的增殖减慢及侵袭能力降低,且 CD147表达降低后 MMP-9的 mRNA 和蛋白表达均显著降低,其差异均有统计学意义(P <0.05)。[结论]CD147表达降低后能抑制肺腺癌细胞的增殖和侵袭能力,可能与其表达降低后使 MMP-9表达降低相关。

  7. MicroRNA-490-3p inhibits proliferation of A549 lung cancer cells by targeting CCND1

    Energy Technology Data Exchange (ETDEWEB)

    Gu, Haihua; Yang, Tao; Fu, Shaozi; Chen, Xiaofan; Guo, Lei; Ni, Yiming, E-mail: ni_yiming@hotmail.com

    2014-01-31

    Highlights: • We examined the level of miR-490-3p in A549 lung cancer cells compared with normal bronchial epithelial cell line. • We are the first to show the function of miR-490-3p in A549 lung cancer cells. • We demonstrate CCND1 may be one of the targets of miR-490-3p. - Abstract: MicroRNAs (miRNAs) are small non-coding RNAs that negatively regulate the translation of messenger RNAs by binding their 3′-untranslated region (3′UTR). In this study, we found that miR-490-3p is significantly down-regulated in A549 lung cancer cells compared with the normal bronchial epithelial cell line. To better characterize the role of miR-490-3p in A549 cells, we performed a gain-of-function analysis by transfecting the A549 cells with chemically synthesized miR-490-3P mimics. Overexpression of miR-490-3P evidently inhibits cell proliferation via G1-phase arrest. We also found that forced expression of miR-490-3P decreased both mRNA and protein levels of CCND1, which plays a key role in G1/S phase transition. In addition, the dual-luciferase reporter assays indicated that miR-490-3P directly targets CCND1 through binding its 3′UTR. These findings indicated miR-490-3P could be a potential suppressor of cellular proliferation.

  8. Cabazitaxel-induced autophagy via the PI3K/Akt/mTOR pathway contributes to A549 cell death

    Science.gov (United States)

    Huo, Ruichao; Wang, Lili; Liu, Peijuan; Zhao, Yong; Zhang, Caiqin; Bai, Bing; Liu, Xueying; Shi, Changhong; Wei, Sanhua; Zhang, Hai

    2016-01-01

    Cabazitaxel has been used to treat castration-resistant prostate cancer since its approval by the US Food and Drug Administration in 2010. However, whether cabazitaxel may inhibit the proliferation of other tissue-derived cancer cells, and its underlying mechanism, remains unknown. In the present study, the A549 lung adenocarcinoma cancer cell line was exposed to cabazitaxel, in order to investigate its cytotoxic effect and determine the underlying mechanism. The results demonstrated that cabazitaxel was able to induce autophagy in A549 cells, as evidenced by the formation of autophagosomes, upregulated LC3-II expression and increased LC3 puncta. Cabazitaxel-induced autophagy had a cytotoxic effect on A549 cells, as evidenced by the induction of cell death and cell cycle arrest at G2/M phase, which was independent of the apoptotic pathway. Furthermore, transfection with Beclin1 small interfering RNA and treatment with the autophagy inhibitor 3-methyladenine protected cells from cabazitaxel-induced cell death, thus confirming that cabazitaxel-induced autophagy contributed to A549 cell death. In addition, cabazitaxel targeted the phosphoinositide 3-kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway to induce autophagy, as indicated by reduced phosphorylation of Akt and mTOR. In conclusion, the present study demonstrated that cabazitaxel exerts a cytotoxic effect on A549 cells by acting on the PI3K/Akt/mTOR pathway to promote autophagic cell death. This result supports the potential use of cabazitaxel as a chemotherapeutic agent for the treatment of lung cancer. PMID:27572899

  9. Cabazitaxel-induced autophagy via the PI3K/Akt/mTOR pathway contributes to A549 cell death.

    Science.gov (United States)

    Huo, Ruichao; Wang, Lili; Liu, Peijuan; Zhao, Yong; Zhang, Caiqin; Bai, Bing; Liu, Xueying; Shi, Changhong; Wei, Sanhua; Zhang, Hai

    2016-10-01

    Cabazitaxel has been used to treat castration-resistant prostate cancer since its approval by the US Food and Drug Administration in 2010. However, whether cabazitaxel may inhibit the proliferation of other tissue‑derived cancer cells, and its underlying mechanism, remains unknown. In the present study, the A549 lung adenocarcinoma cancer cell line was exposed to cabazitaxel, in order to investigate its cytotoxic effect and determine the underlying mechanism. The results demonstrated that cabazitaxel was able to induce autophagy in A549 cells, as evidenced by the formation of autophagosomes, upregulated LC3‑II expression and increased LC3 puncta. Cabazitaxel‑induced autophagy had a cytotoxic effect on A549 cells, as evidenced by the induction of cell death and cell cycle arrest at G2/M phase, which was independent of the apoptotic pathway. Furthermore, transfection with Beclin1 small interfering RNA and treatment with the autophagy inhibitor 3‑methyladenine protected cells from cabazitaxel‑induced cell death, thus confirming that cabazitaxel‑induced autophagy contributed to A549 cell death. In addition, cabazitaxel targeted the phosphoinositide 3‑kinase (PI3K)/Akt/mammalian target of rapamycin (mTOR) pathway to induce autophagy, as indicated by reduced phosphorylation of Akt and mTOR. In conclusion, the present study demonstrated that cabazitaxel exerts a cytotoxic effect on A549 cells by acting on the PI3K/Akt/mTOR pathway to promote autophagic cell death. This result supports the potential use of cabazitaxel as a chemotherapeutic agent for the treatment of lung cancer.

  10. 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.

  11. 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.

  12. Effect of antisense RNA targeting polo-like kinase 1 on cell cycle and proliferation in A549 cells

    Institute of Scientific and Technical Information of China (English)

    周琼; 白明; 苏远

    2004-01-01

    Background Expression of polo-like kinase 1 (Plk1) is elevated in lung cancer and has been proposed as having prognostic value and related to resistance to chemotherapy and radiation. In addition, Plk1 has several functions in mitotic progression. In this study, the authors investigated the effect of Plk1 depletion on cell cycle progression and proliferation in A549 cells, a lung cancer cell line.Methods A recombinant plasmid containing antisense RNA targeting Plk1 (pcDNA3-Plk1) was transfected into A549 cells. Reverse transcription-polymerase chain reaction and Western blot were used to examine Plk1 gene expression. Cell proliferation was evaluated by direct cell counting and bromodeoxyuridine (BrdU) labelling. Cell cycle and apoptosis were examined by flow cytometry. Expression of α-tubulin was detected by immunofluorescence, and the inhibition rate (IR) by chemotherapeutic agents was determined by 3-(4,5-dimethylthiazol-2-yl) 2,5-diphenyl-tetrazolium bromide assay.Results After transfection into A549 cells, pcDNA3-Plk1 reduced Plk1 mRNA by 46.75% for 24 hours and by 61.84% for 48 hours. Plk1 protein was significantly decreased simultaneously (P<0.05). Abnormal morphological changes of cells and growth inhibition were observed in pcDNA3-Plk1 transfected groups. The BrdU labelling index was 25.59% 48 hours after transfection, which was significantly lower than that of the control groups (P<0.05). Forty-eight hours after transfection, there showed absence of microtubule polymerization and spindle abnormalities in staining for α-tubulin. A549 cells showed a strong G2/M arrest and apoptosis 72 hours post transfection. IR of vinorelbine in pcDNA3-Plk1 transfected groups was significantly higher than that of the other groups (P<0.05, respectively).Conclusions Plk1 depletion interferes with spindle formation, induces cell cycle arrest and apoptosis, and consequently inhibits cell proliferation in A549 cells. Moreover, it sensitizes lung cancer cells to chemotherapy.

  13. ERK1/2 activation modulates pyocyanin-induced toxicity in A549 respiratory epithelial cells.

    Science.gov (United States)

    Forbes, Amanda; Davey, Andrew K; Perkins, Anthony V; Grant, Gary D; McFarland, Amelia J; McDermott, Catherine M; Anoopkumar-Dukie, Shailendra

    2014-02-01

    Pyocyanin (PCN), a virulence factor produced by Pseudomonas aeruginosa, has many damaging effects on mammalian cells. Several lines of evidence suggest that this damage is primarily mediated by its ability to generate oxidative stress. However mechanisms underlying PCN-induced oxidative injury remain unclear. Although oxidative stress and subsequent MAPK signaling has been shown to modulate cell death in other models, its role in PCN-induced cytotoxicity remains unknown. Therefore the aim of this study was to investigate the role of redox-sensitive MAPK in PCN-induced toxicity in A549 cells. Here we show that PCN (50μM) rapidly increased ERK1/2 phosphorylation after 5min. Pre-treatment of A549 cells with the MEK1/2 inhibitor U0126 (10μM) decreased PCN-induced ERK1/2 phosphorylation and protected cells against apoptosis and cell injury suggesting a role for ERK signalling. In contrast, JNK and p38 MAPK phosphorylation remained unchanged following exposure to PCN and pretreatment with either the JNK or p38 MAPK inhibitors (10μM SP600125 and 10μM SB203580, respectively) did not afford protection against PCN toxicity. This would suggest that PCN-induced cytotoxicity appears to occur independently of JNK and p38 MAPK signaling pathways. Finally, although we confirm that oxidative stress contributes to PCN-induced toxicity, our data suggest the contribution of oxidative stress is independent of ERK1/2 signaling. These findings may provide insight for novel targeted therapies to reduce PCN-mediated lung injury in patients with chronic P. aeruginosa respiratory infections.

  14. 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.

  15. Effects of hypoxia on the expression of CCR7 and proliferation, invasiveness of A549 cells

    Directory of Open Access Journals (Sweden)

    Yang LI

    2008-10-01

    Full Text Available Background and objective It has been proven that hypoxia could promote tumor cells invasion and metastasis by different mechanisms, but the relationship between hypoxia and CCR7 have not been reported. The aim of this investigate is to evaluate the effects of hypoxia on the expression of CCR7 and the invasiveness of lung adenocarcinoma A549 cells. Methods A549 cells were incubated at either normoxia (37 ℃, 5%CO2, 21%O2 or hypoxia(37 ℃, 5%CO2, 1%O2 condition for 4 h,12 h, 24 h. The expressions of CCR7 mRNA and protein levels were observed by RT-PCR and Western blotting; Cells invasiveness was measured by matrigel invasion assay. Results RT-PCR and Western blotting showed that the expression of CCR7 was detected in lung adenocarcinoma A549 cells, CCR7 mRNA and protein expression level were increased with culture time along either in normoxia or hypoxia condition; Furthermore,compared with normoxia group, the CCR7 mRNA and protein expression level in hypoxia group was increased (P <0.01.The results of Transwell invasion showed that The number of invasive cells was significantly increased in hypoxia group(t =0.006, P <0.01 and A549 cells invasive ability was inhibited after add anti-CCR7 Ab to culture medium (t =0.09, P <0.01. Conclusion The results suggest that hypoxia plays an important role in the augmentation of the CCR7 expression and invasiveness of A549 cells. Invasion of A549 cells in hypoxia condition correlated with CCR7 expression level.

  16. Effect of Juglone in qinglongyi on cell cycle status and apoptosis in A-549 cells

    Institute of Scientific and Technical Information of China (English)

    ZOU Xiang; KONG Ling-sheng; JI Yu-bin

    2008-01-01

    Objective To explore the inhibition of juglone in Qinglongyi on A-549 cells in vitro. Methods MTT assay was used. Laser confocal scanning microscope was used to observe apoptotic morphology.Changes of cell cycle are studied by flow cytometry analysis. Results MTT assay showed that juglone had a marked growth inhibition in A-549 cells and the IC50 is respectively 3.4×10-5 mol·L-1, 1.8×10-5 mol·L-1 and 2.6×10-6 mol·L-1 after treatment for 24, 48 and 72 h by juglone. Through Laser confocal scanning microscope, we can see that juglone can induce the apoptosis. Cell cycle changes are analyzed by flow cytometry with cells at G1 phase significantly less than those of control and ceils at G2 phase significantly more than those of control. Conclusions It suggests that juglone could apoptosis of A-549 cells with the cell cycle arrest on G2 phase in distinct dose-dependent manner.

  17. 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.

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

    Energy Technology Data Exchange (ETDEWEB)

    Brown, David M., E-mail: d.brown@hw.ac.uk [Heriot-Watt University, Nanosafety Research Group, School of Life Sciences (United Kingdom); Varet, Julia, E-mail: julia.varet@IOM-world.org [Institute of Occupational Medicine (United Kingdom); Johnston, Helinor, E-mail: h.johnston@hw.ac.uk; Chrystie, Alison; Stone, Vicki, E-mail: v.stone@hw.ac.uk [Heriot-Watt University, Nanosafety Research Group, School of Life Sciences (United Kingdom)

    2015-10-15

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

  19. 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.

  20. 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).

  1. 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.

  2. 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.

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

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    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.

  4. 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.

  5. 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

  6. PARTICULATE MATTER (PM) INHIBITS NEUROTROPHIN RELEASE FROM A549 CELLS

    Science.gov (United States)

    Several investigations have linked PM exposure to the exacerbation of allergic lung diseases. Many PM effects are mediated by cells within the lung including the airway epithelium, eosinophils, and lymphocytes. These cells also produce neurotophins such as NGF and/or express neur...

  7. 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.

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

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    Chang HB

    2015-08-01

    Full Text Available Hong-Bin Chang,1 Bing-Huei Chen1,21Department of Food Science, 2Graduate Institute of Medicine, Fu Jen Catholic University, Taipei, TaiwanAbstract: 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.Keywords: curcuminoid extract, curcuminoid nanoemulsion, Curcuma longa Linnaeus, lung cancer cell, cell cycle, apoptosis mechanism

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

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    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.

  10. 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.

  11. 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.

  12. Part II-mechanism of adaptation: A549 cells adapt to high concentration of nitric oxide through bypass of cell cycle checkpoints.

    Science.gov (United States)

    Aqil, Madeeha; Deliu, Zane; Elseth, Kim M; Shen, Grace; Xue, Jiaping; Radosevich, James A

    2014-03-01

    Previous work has shown enhanced survival capacity in high nitric oxide (HNO)-adapted tumor cells. In Part I of this series of manuscripts, we have shown that A549-HNO cells demonstrate an improved growth profile under UV and X-ray radiation treatment. These cells exhibit increased expression of proteins involved in DNA damage recognition and repair pathway, both the non-homologous end joining pathway and homologous recombination. These include Ku80, DNA-PK, XLF ligase and MRN complex proteins. Further, the A549-HNO cells show high levels of ATM, ATR, Chk1 and Chk2, and phospho-p53. Activation of these molecules may lead to cell cycle arrest and apoptosis due to DNA damage. This is observed in parent A549 cells in response to NO donor treatment; however, the A549-HNO cells proliferate and inhibit apoptosis. Cell cycle analysis showed slowed progression through S phase which will allow time for DNA repair. Thus, to better understand the increased growth rate in A549-HNO when compared to the parent cell line A549, we studied molecular mechanisms involved in cell cycle regulation in A549-HNO cells. During the initial time period of NO donor treatment, we observe high levels of cyclin/Cdk complexes involved in regulating various stages of the cell cycle. This would lead to bypass of G1-S and G2-M checkpoints. The HNO cells also show much higher expression of Cdc25A. Cdc25A activates Cdk molecules involved in different phases of the cell cycle. In addition, there is enhanced phosphorylation of the Rb protein in HNO cells. This leads to inactivation of Rb/E2F checkpoint regulating G1-S transition. This may lead to faster progression in S phase. Thus, all of these perturbations in HNO cells lead to accelerated cell cycle progression and a higher growth rate. We also assessed expression of cell cycle inhibitors in HNO cells. Interestingly, the HNO cells show a significant decline in p21CIP1 at initial time points, but with prolonged exposure, the levels were much higher

  13. 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.

  14. 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.

  15. Mycelial Extract of Phellinus linteus Induces Cell Death in A549 Lung Cancer Cells and Elevation of Nitric Oxide in Raw 264.7 Macrophage Cells.

    Science.gov (United States)

    Lee, Jong-Jin; Kwon, Ho-Kyun; Lee, Dong-Soo; Lee, Seung-Woo; Lee, Kye-Kwan; Kim, Kyu-Joong; Kim, Jong-Lae

    2006-09-01

    In the present study, in order to investigate the anti-proliferative phenomenon of PLME, the effects of mycelial extract of Phellinus linteus (PLME) on the growth of human lung carcinoma cell line A549 was examined. We studied on the effects of PLME on the release of nitric oxide (NO) in mouse macrophage Raw 264.7 cells. Treatment of PLME to A549 cells resulted in the growth inhibition, morphological change and induction of apoptotic cell death in a dose-dependent manner as measured by MTT assay. We found that PLME stimulated a dose-dependent increase in NO production. These findings suggest that PLME enhances the anti-tumoral activity of macrophage and may be a potential therapeutic agent for the control of human lung carcinoma cells.

  16. PM10-biogenic fraction drives the seasonal variation of proinflammatory response in A549 cells.

    Science.gov (United States)

    Camatini, Marina; Corvaja, Viviana; Pezzolato, Eleonora; Mantecca, Paride; Gualtieri, Maurizio

    2012-02-01

    PM10 was collected in a Milan urban site, representative of the city air quality, during winter and summer 2006. Mean daily PM10 concentration was 48 μg m(-3) during summer and 148 μg m(-3) during winter. Particles collected on Teflon filters were chemically characterized and the endotoxin content determined by the LAL test. PM10-induced cell toxicity, assessed with MTT and LDH methods, and proinflammatory potential, monitored by IL-6 and IL-8 cytokines release, were investigated on the human alveolar epithelial cell line A549 exposed to increasing doses of PM. Besides untreated cells, exposure to inert carbon particles (2-12 μm) was also used as additional control. Both cell toxicity and proinflammatory potency resulted to be higher for summer PM10 with respect of winter PM10, with IL-6 showing the highest dose-dependent release. The relevance of biogenic components adsorbed onto PM10 in eliciting the proinflammatory mediators release was investigated by inhibition experiments. Polymixin B (Poly) was used to inhibit particle-bind LPS while Toll-like receptor-2 antibody (a-TLR2) to specifically block the activation of this receptor. While cell viability was not modulated in cells coexposed to PM10 and Poly or a-TLR2 or both, inflammatory response did it, with IL-6 release being the most inhibited. In conclusion, Milan PM10-induced seasonal-dependent biological effects, with summer particles showing higher cytotoxic and proinflammatory potential. Cytotoxicity seemed to be unaffected by the PM biogenic components, while inflammation was significantly reduced after the inhibition of some biogenic activated pathways. Besides, the PM-associated biogenic activity does not entirely justify the PM-induced inflammatory effects. © 2010 Wiley Periodicals, Inc. Environ Toxicol 2012.

  17. 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.

  18. 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.

  19. The effect of tyrphostins AG494 and AG1478 on the autocrine growth regulation of A549 and DU145 cells

    Directory of Open Access Journals (Sweden)

    Agnieszka Bojko

    2012-07-01

    Full Text Available We employed two selective EGFR tyrosine kinase inhibitors: AG494 (reversible and AG1478 (irreversible for growth regulation of human lung (A549 and prostate (DU145 cancer cell lines, cultured in chemically defined DMEM/F12 medium. Both tested tyrphostins significantly inhibited autocrine growth of the investigated cell lines. The action of AG494 was dose dependent, and at highest concentrations led to complete inhibition of growth. AG1478 seemed to be more effective at lower concentrations, but was unable to completely inhibit growth of A549 cells. Inhibition of EGFR kinase activity by AG494 in contrast to AG1478 had no effect on the activity of ERK in both cell lines. Both EGFR’s inhibitors induced apoptosis of the investigated lung and prostate cancer cell lines, but the proapoptotic effect of the investigated tyrphostins was greater in A549 than in DU145 cells. The tyrphostins arrested cell growth of DU145 and A549 cells in the G1 phase, similarly to other known inhibitors of EGFR. The influence of AG494 and AG1478 on the activity of two signaling proteins (AKT and ERK was dependent upon the kind of investigated cells. In the case of DU145 cells, there was an evident decline in enzymatic activity of both kinases (stronger for AG1478, while in A549, only AG1478 effectively inhibited the phosphorylation of Akt. Tyrphostins AG494 and AG1478 are ATP-competitors and are supposed to have a similar mechanism of action, but our results suggest that this is not quite true.

  20. DNA double-strand breaks activate ATM independent of mitochondrial dysfunction in A549 cells.

    Science.gov (United States)

    Kalifa, Lidza; Gewandter, Jennifer S; Staversky, Rhonda J; Sia, Elaine A; Brookes, Paul S; O'Reilly, Michael A

    2014-10-01

    Excessive nuclear or mitochondrial DNA damage can lead to mitochondrial dysfunction, decreased energy production, and increased generation of reactive oxygen species (ROS). Although numerous cell signaling pathways are activated when cells are injured, the ataxia telangiectasia mutant (ATM) protein has emerged as a major regulator of the response to both mitochondrial dysfunction and nuclear DNA double-strand breaks (DSBs). Because mitochondrial dysfunction is often a response to excessive DNA damage, it has been difficult to determine whether nuclear and/or mitochondrial DNA DSBs activate ATM independent of mitochondrial dysfunction. In this study, mitochondrial and nuclear DNA DSBs were generated in the A549 human lung adenocarcinoma cell line by infecting with retroviruses expressing the restriction endonuclease PstI fused to a mitochondrial targeting sequence (MTS) or nuclear localization sequence (NLS) and a hemagglutinin antigen epitope tag (HA). Expression of MTS-PstI-HA or NLS-PstI-HA activated the DNA damage response defined by phosphorylation of ATM, the tumor suppressor protein p53 (TP53), KRAB-associated protein (KAP)-1, and structural maintenance of chromosomes (SMC)-1. Phosphorylated ATM and SMC1 were detected in nuclear fractions, whereas phosphorylated TP53 and KAP1 were detected in both mitochondrial and nuclear fractions. PstI also enhanced expression of the cyclin-dependent kinase inhibitor p21 and inhibited cell growth. This response to DNA damage occurred in the absence of detectable mitochondrial dysfunction and excess production of ROS. These findings reveal that DNA DSBs are sufficient to activate ATM independent of mitochondrial dysfunction and suggest that the activated form of ATM and some of its substrates are restricted to the nuclear compartment, regardless of the site of DNA damage.

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

    Science.gov (United States)

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

    2016-12-01

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

  2. Investigation of Gene Expression Profile of A549 Cells after Overexpression of GPC5 
by High Throughput Transcriptome Sequencing

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    Haitian ZHANG

    2016-08-01

    Full Text Available Background and objective Glypican-5 (GPC5 is an important tumor suppressor, while little is known about the impact of GPC5 on proliferation ability and gene expression in lung adenocarcinoma cell lines. Here, we stably overexpressed GPC5 in A549 cells and investigated the impact of cell proliferation ability and gene expression. Methods A549 cells that stably overexpressed GPC5 were constructed by lentivirus. Cell counter kit 8 (CCK8, colony formation, EdU assay were conducted to analyze cell proliferation ability, and transcriptome sequencing was utilized to investigate gene expression profile. Results CCK8 assay showed that compared with empty vector, overexpression of GPC5 significantly inhibited cell proliferation rate in A549 cells and the number of colony was also decreased (181±17 vs 278±23. EdU assay also confirmed the percentage of positive staining cells decreased after GPC5 overexpression. Transcriptome sequencing revealed that 2,108 genes were differentially expressed after GPC5 overexpression. Among these differentially expressed genes, 47 genes of the Gene Ontology item “positive regulation of cell proliferation” were downregulated. Conclusion Overexpression of GPC5 inhibited proliferation ability in lung adenocarcinoma A549 cells and genes with the function of “positive regulation of cell proliferation” were downregulated.

  3. 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

  4. 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.

  5. 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.

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

    Science.gov (United States)

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

    2016-06-01

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

  7. 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.

  8. Activities of Ten Essential Oils towards Propionibacterium acnes and PC-3, A-549 and MCF-7 Cancer Cells

    Directory of Open Access Journals (Sweden)

    Yuangang Zu

    2010-04-01

    Full Text Available Ten essential oils, namely, mint (Mentha spicata L.,Lamiaceae, ginger (Zingiber officinaleRosc.,Zingiberaceae, lemon (Citrus limon Burm.f.,Rutaceae, grapefruit (Citrus paradisi Macf., Rutaceae, jasmine (Jasminum grandiflora L.,Oleaceae, lavender (Mill.,Lamiaceae, chamomile (Matricaria chamomilla L., Compositae, thyme (Thymus vulgaris L., Lamiaceae, rose (Rosa damascena Mill.,Rosaceae and cinnamon (Cinnamomum zeylanicumN. Lauraceae were tested for their antibacterial activities towards Propionibacterium acnes and in vitro toxicology against three human cancer cell lines. Thyme, cinnamon and rose essential oils exhibited the best antibacterial activities towards P. acnes, with inhibition diameters of 40 ± 1.2 mm, 33.5 ± 1.5 mm and 16.5 ± 0.7 mm, and minimal inhibitory concentrations of 0.016% (v/v, 0.016% (v/v and 0.031% (v/v, respectively. Time-kill dynamic procedures showed that thyme, cinnamon, rose, and lavender essential oils exhibited the strongest bactericidal activities at a concentration of 0.25% (v/v, and P. acnes was completely killed after 5 min. The thyme essential oil exhibited the strongest cytotoxicity towards three human cancer cells. Its inhibition concentration 50% (IC50 values on PC-3, A549 and MCF-7 tumor cell lines were 0.010% (v/v, 0.011% (v/v and 0.030% (v/v, respectively. The cytotoxicity of 10 essential oils on human prostate carcinoma cell (PC-3 was significantly stronger than on human lung carcinoma (A549 and human breast cancer (MCF-7 cell lines.

  9. 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.

  10. 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.

  11. 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.

  12. 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.

  13. 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.

  14. 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.

  15. Cytokines from the tumor microenvironment modulate sirtinol cytotoxicity in A549 lung carcinoma cells.

    Science.gov (United States)

    Pal, Shyama; Shankar, Bhavani S; Sainis, Krishna B

    2013-10-01

    Cytokines in tumor microenvironment play an important role in the success or failure of molecular targeted therapies. We have chosen tumor necrosis factor α (TNF-α), TNF related apoptosis inducing ligand (TRAIL), insulin-like growth factor 1 (IGF-1) and transforming growth factor β (TGF-β) as representative pro-inflammatory, pro-apoptotic, anti-apoptotic and anti-inflammatory tumor derived cytokines. Analysis of Oncomine database revealed the differential expression of these cytokines in a subset of cancer patients. The effects of these cytokines on cytotoxicity of FDA approved drugs - cisplatin and taxol and inhibitors of epidermal growth factor receptor - AG658, Janus kinase - AG490 and SIRT1 - sirtinol were assessed in A549 lung cancer cells. TRAIL augmented cytotoxicity of sirtinol and IGF-1 had a sparing effect. Since TRAIL and IGF-1 differentially modulated sirtinol cytotoxicity, further studies were carried out to identify the mechanisms. Sirtinol or knockdown of SIRT1 increased the expression of death receptors DR4 and DR5 and sensitized A549 cells to TRAIL. Increased cell death in presence of TRAIL and sirtinol was caspase independent and demonstrated classical features of necroptosis. Inhibition of iNOS increased caspase activity and switched the mode of cell death to caspase mediated apoptosis. Interestingly, sirtinol or SIRT1 knockdown did not increase IGF-1R expression. Instead, it abrogated ligand induced downregulation of IGF-1R and increased cell survival through PI3K-AKT pathway. In conclusion, these findings reveal that the tumor microenvironment contributes to modulation of cytotoxicity of drugs and that combination therapy, with agents that increase TRAIL signaling and suppress IGF-1 pathway may potentiate anticancer effect.

  16. 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.

  17. 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.

  18. 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.

  19. Synthesis and cytotoxicity evaluation of 4-amino-4-dehydroxylarctigenin derivatives in glucose-starved A549 tumor cells.

    Science.gov (United States)

    Lei, Min; Gan, Xianwen; Zhao, Kun; Yu, Qiang; Hu, Lihong

    2015-02-01

    The natural product arctigenin (ATG) demonstrated preferential cytotoxicity to cancer cells under glucose starvation. A series of 4-amino-4-dehydroxylarctigenin derivatives based on lead compound ATG were designed and synthesized by bioisosteric modifications. Their cytotoxicities were evaluated in glucose-starved A549 tumor cells and the results indicated that the 4-amino-4-dehydroxylarctigenin showed more potent cytotoxicity than arctigenin, and the further substituent group on 4-amino would result in the cytotoxicities decreased significantly. 4-Substituted-arctigenin could selectively target on glucose-starved A549 tumor cells which provide an alternative strategy for anticancer drug development with minimal normal tissue toxicity.

  20. Blocking the NOTCH pathway can inhibit the growth of CD133-positive A549 cells and sensitize to chemotherapy.

    Science.gov (United States)

    Liu, Juntao; Mao, Zhangfan; Huang, Jie; Xie, Songping; Liu, Tianshu; Mao, Zhifu

    2014-02-21

    Cancer stem cells (CSCs) are believed to play an important role in tumor growth and recurrence. These cells exhibit self-renewal and proliferation properties. CSCs also exhibit significant drug resistance compared with normal tumor cells. Finding new treatments that target CSCs could significantly enhance the effect of chemotherapy and improve patient survival. Notch signaling is known to regulate the development of the lungs by controlling the cell-fate determination of normal stem cells. In this study, we isolated CSCs from the human lung adenocarcinoma cell line A549. CD133 was used as a stem cell marker for fluorescence-activated cell sorting (FACS). We compared the expression of Notch signaling in both CD133+ and CD133- cells and blocked Notch signaling using the γ-secretase inhibitor DAPT (GSI-IX). The effect of combining GSI and cisplatin (CDDP) was also examined in these two types of cells. We observed that both CD133+ and CD133- cells proliferated at similar rates, but the cells exhibited distinctive differences in cell cycle progression. Few CD133+ cells were observed in the G2/M phase, and there were half as many cells in S phase compared with the CD133- cells. Furthermore, CD133+ cells exhibited significant resistance to chemotherapy when treated with CDDP. The expression of Notch signaling pathway members, such as Notch1, Notch2 and Hes1, was lower in CD133+ cells. GSI slightly inhibited the proliferation of both cell types and exhibited little effect on the cell cycle. The inhibitory effects of DPP on these two types of cells were enhanced when combined with GSI. Interestingly, this effect was especially significant in CD133+ cells, suggesting that Notch pathway blockade may be a useful CSC-targeted therapy in lung cancer.

  1. 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.

  2. Taxol-induced paraptosis-like A549 cell death is not senescence

    Science.gov (United States)

    Wang, Chao-yang; Chen, Tong-Sheng

    2011-03-01

    Our previous studies have shown that taxol, a potent anticancer agent, induces caspase-independent cell death and cytoplasmic vacuolization in human lung cancer cells. However, the mechanisms of taxol-induced cytoplasmic vacuolization are poorly understood. Cytoplasmic vacuolization have been reported to be a characteristic of cell senescence. Here, we employed confocal fluorescence microscopy imaging to study the reversibility of taxol-induced cytoplasmic vacuolization and whether taxol triggers senescence in A549 cells. We found that taxol-induced cytoplasmic vacuolization at 6 or 9 h after treatment with taxol did not decrease but increase at 24 h or 72 h after refreshing the culture medium without taxol, indicating taxol-induced cytoplasmic vacuolization is irreversible. We used SA-β-Gal (senescence-associated β-galactosidase) to assess whether taxol-induced cell death in cytoplasmic vacuolization fashion is senescence, and found that hydrogen peroxide (H2O2)-treated, but not taxol-treated cells is significantly stained by the SA-β-Gal, a senescence testing kit, indicating that the form of taxol-induced cell death is not senescence.

  3. Effects of green tea extract on lung cancer A549 cells: proteomic identification of proteins associated with cell migration.

    Science.gov (United States)

    Lu, Qing-Yi; Yang, Yanan; Jin, Yu Sheng; Zhang, Zuo-Feng; Heber, David; Li, Frederick P; Dubinett, Steven M; Sondej, Melissa A; Loo, Joseph A; Rao, Jian Yu

    2009-02-01

    Green tea polyphenols exhibit multiple antitumor activities, and the mechanisms of action are not completely understood. Previously, we reported that green tea extract (GTE)-induced actin remolding is associated with increased cell adhesion and decreased motility in A549 lung cancer cells. To identify the cellular targets responsible for green tea-induced actin remodeling, we performed 2-DE LC-MS/MS of A549 cells before and after GTE exposure. We have identified 14 protein spots that changed in expression (> or =2-fold) after GTE treatment. These proteins are involved in calcium-binding, cytoskeleton and motility, metabolism, detoxification, or gene regulation. In particular we found upregulation of several genes that modulate actin remodeling and cell migration, including lamin A/C. Our data indicated that GTE-induced lamin A/C upregulation appears to be at the transcriptional level and the increased expression results in the decrease in cell motility, as confirmed by siRNA. The result of the study demonstrates that GTE alters the levels of many proteins involved in growth, motility and apoptosis of A549 cells and their identification may explain the multiple antitumor activities of GTE.

  4. 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.

  5. The synergistic effect of resveratrol in combination with cisplatin on apoptosis via modulating autophagy in A549 cells.

    Science.gov (United States)

    Hu, Song; Li, Xiaolin; Xu, Rongrong; Ye, Lingyun; Kong, Hui; Zeng, Xiaoning; Wang, Hong; Xie, Weiping

    2016-06-01

    Several studies have shown that combination treatment with natural products and chemotherapy agents can improve the sensitivity and cytotoxicity of chemotherapy agents. Resveratrol, a natural product, has many biological effects including antitumor and antiviral activities, as well as vascular protective effect. The aim of this study is to investigate the synergistic anticancer effect of resveratrol in combination with cisplatin and the potential anticancer mechanisms involved in A549 cells. The results obtained from Cell Counting Kit-8 and isobolographic analysis demonstrated that combination of resveratrol and cisplatin resulted in synergistic cytotoxic effects in A549 cells. Results from Hoechst staining, flow cytometry and western blot analysis suggested that resveratrol enhanced cisplatin-mediated apoptosis. Meanwhile, the changes of LC3-II and P62 levels and formation of autophagosome suggested that resveratrol in combination with cisplatin triggered autophagy. More importantly, inhibiting autophagy by 3-methyladenine markedly attenuated the apoptosis caused by combination of resveratrol and cisplatin in A549 cells. Taken together, our study provides the first evidence that resveratrol combined with cisplatin synergistically induce apoptosis via modulating autophagic cell death in A549 cells. These findings also help us to understand the role of natural products in combination with chemotherapy agents in lung cancer.

  6. Raw and thermally treated cement asbestos exerts different cytotoxicity effects on A549 cells in vitro.

    Science.gov (United States)

    Pugnaloni, Armanda; Lucarini, Guendalina; Rubini, Corrado; Smorlesi, Arianna; Tomasetti, Marco; Strafella, Elisabetta; Armeni, Tatiana; Gualtieri, Alessandro F

    2015-01-01

    Raw cement asbestos (RCA) undergoes a complete solid state transformation when heated at high temperatures. The secondary raw material produced, high temperatures-cement asbestos (HT-CA) is composed of newly-formed crystals in place of the asbestos fibers present in RCA. Our previous study showed that HT-CA exerts lower cytotoxic cell damage compared to RCA. Nevertheless further investigations are needed to deepen our understanding of pathogenic pathways involving oxidative and nitrative damage. Our aim is to deepen the understanding of the biological effects on A549 cells of these materials regarding DNA damage related proteins (p53, its isoform p73 and TRAIL) and nitric oxide (NO) production during inducible nitric oxide synthase (iNOS)-mediated inflammation. Increments of p53/p73 expression, iNOS positive cells and NO concentrations were found with RCA, compared to HT-CA and controls mainly at 48 h. Interestingly, ferrous iron causing reactive oxygen species (ROS)-mediated DNA damage was found in RCA as a contaminant. HT-CA thermal treatment induces a global recrystallization with iron in a crystal form poorly released in media. HT-CA slightly interferes with genome expression and exerts lower inflammatory potential compared to RCA on biological systems. It could represent a safe approach for storing or recycling asbestos and an environmentally friendly alternative to asbestos waste.

  7. Titanium dioxide nanoparticles exhibit genotoxicity and impair DNA repair activity in A549 cells.

    Science.gov (United States)

    Jugan, Mary-Line; Barillet, Sabrina; Simon-Deckers, Angelique; Herlin-Boime, Nathalie; Sauvaigo, Sylvie; Douki, Thierry; Carriere, Marie

    2012-08-01

    Titanium dioxide nanoparticles (TiO(2)-NPs) are produced in large quantities, raising concerns about their impact for human health. The aim of this study was to deeply characterize TiO(2)-NPs genotoxic potential to lung cells, and to link genotoxicity to physicochemical characteristics, e.g., size, specific surface area, crystalline phase. A549 cells were exposed to a panel of TiO(2)-NPs with diameters ranging from 12 to 140 nm, either anatase or rutile. A set of complementary techniques (comet and micronucleus assays, gamma-H2AX immunostaining, 8-oxoGuanine analysis, H2-DCFDA, glutathione content, antioxidant enzymes activities) allowed us to demonstrate that small and spherical TiO(2)-NPs, both anatase and rutile, induce single-strand breaks and oxidative lesions to DNA, together with a general oxidative stress. Additionally we show that these NPs impair cell ability to repair DNA, by inactivation of both NER and BER pathways. This study thus confirms the genotoxic potential of TiO(2)-NPs, which may preclude their mutagenicity and carcinogenicity.

  8. 黄连素在A549细胞中对顺铂抗肿瘤作用的影响及其机制%Inlfuence of Berberine on Cisplatin Antineoplastic Effect in A549 Cells

    Institute of Scientific and Technical Information of China (English)

    蒋国君; 李利; 吴小祥; 董淑英; 童旭辉

    2015-01-01

    Background and objectiveCisplatin is a standard ifrst-line chemotherapeutic agents for treating ad-vanced non-small cell lung cancer. Unfortunately, the clinical application cisplatin is restricted because it induces serious adverse reaction. hTe aim of this study is to investigate the inlfuence and probable mechanism of berberine on cisplatin antineoplastic effect on lung cancer A549 cells.MethodshTe total Cx43 protein amount, localization of Cx43 on cell membrane, and gap junction function were observed atfer the A549 cells were treated with berberine. hTe inlfuence 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. hTe gap junction function between the cells was enhanced through increased Cx43 protein expression and localization of Cx43 on the membrane atfer berberine treatment. hTe 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. hTis effect was associated with the PKC activity. hTe cispla-tin-induced inhibition of colony growth was enhanced when berberine was combined with cisplatin.ConclusionBerberine can obviously increase the antitumor effect of cisplatin by enhancing the function of the gap junction possibly in A549 cells.%背景与目的以顺铂为基础的化疗方案是晚期非小细胞肺癌的一线化疗方案,但是由于顺铂的不良反应严重及耐药性的产生均限制了它的临床应用,本研究采用联合用药的方式观察黄连素对顺铂抗肿瘤作用的影响,并探讨其可能机制。方法分别观察黄连素对肺腺癌细胞A549细胞中总Cx43蛋白、细胞膜Cx43蛋白的表达以及细胞缝隙连接功能的改变,通过标

  9. 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

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

    Directory of Open Access Journals (Sweden)

    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

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

    Directory of Open Access Journals (Sweden)

    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.

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

    Directory of Open Access Journals (Sweden)

    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.

  14. Mechanism of cisplatin combined with zoledronic acid on lung cancer A549 cells%顺铂联合唑来膦酸对肺癌A549细胞增殖的影响及其机制

    Institute of Scientific and Technical Information of China (English)

    茆勇; 马凤锦; 黄朝晖; 许林; 游庆军; 华东

    2011-01-01

    Objective To investigate the function of cisplatin combined with Zoledronic acid on proliferation and mechanisms of A549 cells.Methods ( 1 ) Methyl thiazol tetrazolium (MTT) assay and Annexin V-FITC/PI double-staining flow cytometry were employed to observe the effects of cisplatin combined with Zoledronic acid upon anti-proliferation and apoptosis respectively.Reverse transcription-polymerase chain reaction (RT-PCR) was applied to assay mRNA expression of MDC1 among different groups.Results Inhibition of the cell proliferation was observed under the treatment of combined cisplatin and zoledronic acid (39.16 ±4.94)%,superior to the treatment of zoledronic acid ( 19.66 ±4.57)% or cisplatin ( 16.87 ± 2.50) %.Combined group induced A549 cells apoptosis ( 32.30 ± O.50 ) %,compared with cisplatin (23.90 ± 2.46) %,zoledronic acid ( 18.87 ± 3.04 ) %,the difference was statistically significant; cisplatin after zoledronic acid treatment of A549 cells M DC1 mRNA expression (0.134 ± 0.037 )was significantly decreased compared with single-drug,zoledronic acid ( 0.208 ± 0.040 ) and cisplatin (0.356 ± 0.033) ( P < 0.05 ).Conclusion Zoledronic acid or cisplatin can significantly inhibit A549 cell proliferation and markedly induce the apoptosis.Zoledronic acid and cisplatin in a synergistic way inhibited the cell proliferation and induced apoptosis on A549 cell.The downregulated expression level of MDC1 mRNA may involved in the mechanism of synergistic effect.%目的 观察顺铂联合唑来膦酸对肺癌A-549细胞增殖的影响并探讨其作用机制.方法 以噻唑蓝(MTT)比色法观察顺铂联合唑来膦酸对A549细胞增殖的影响,以Annexin-V/PI双染法检测细胞凋亡,逆转录-聚合酶链反应(RT-PCR)检测DNA损伤检查点蛋白调节因子1(MDC1)mRNA的表达.结果 顺铂联合唑来膦酸对A549细胞增殖的抑制率(39.16±4.94)%高于顺铂(16.87±2.50)%、唑来膦酸(19.66±4.57)%;联合用药诱导A549

  15. 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).

  16. 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.

  17. The effects of dietary phenolic compounds on cytokine and antioxidant production by A549 cells.

    Science.gov (United States)

    Gauliard, Benoit; Grieve, Douglas; Wilson, Rhoda; Crozier, Alan; Jenkins, Carol; Mullen, William D; Lean, Michael

    2008-06-01

    Levels of inflammatory cytokines are raised in chronic obstructive pulmonary disease (COPD). A diet rich in antioxidant vitamins may protect against the development of COPD. This study examined the effects of phenolic compounds and food sources on cytokine and antioxidant production by A549 cells. The effects of the following phenolic compounds on basal and interleukin (IL)-1-stimulated release of IL-8, IL-6, and reduced glutathione (GSH) were examined: resveratrol; Bouvrage, a commercially available raspberry juice (Ella Drinks Ltd., Alloa, Clacksmannanshire, UK); and quercetin 3'-sulfate. Purification of the raspberry juice by high-performance liquid chromatography gave three fractions: Fraction 1 contained phenolic acid and vitamin C, Fraction 2 contained flavonoids and ellagic acid, and Fraction 3 contained anthocyanins and ellagitannins. IL-8 production was increased in the presence of IL-1 (165 vs. 6,011 pg/mL, P or =50 micromol/mL significantly inhibited IL-8 and IL-6 production. Similar findings were made with raspberry juice at concentrations > or =25 microL/mL, and Fractions 1 and 3 were best able to inhibit IL-8 production. Quercetin 3'-sulfate, at 25 micromol/mL, inhibited IL-8 and IL-6 production. The changes observed in IL-8 were paralleled by changes in tumor necrosis factor-alpha. Thus, phenolic compounds can significantly alter cytokine and antioxidant production.

  18. Blocking the NOTCH pathway can inhibit the growth of CD133-positive A549 cells and sensitize to chemotherapy

    Energy Technology Data Exchange (ETDEWEB)

    Liu, Juntao; Mao, Zhangfan; Huang, Jie; Xie, Songping; Liu, Tianshu; Mao, Zhifu, E-mail: 48151660@qq.com

    2014-02-21

    Highlights: • Notch signaling pathway members are expressed lower levels in CD133+ cells. • CD133+ cells are not as sensitive as CD133− cells to chemotherapy. • GSI could inhibit the growth of both CD133+ and CD133− cells. • Blockade of Notch signaling pathway enhanced the effect of chemotherapy with CDDP. • DAPT/CDDP co-therapy caused G2/M arrest and elimination in CD133+ cells. - Abstract: Cancer stem cells (CSCs) are believed to play an important role in tumor growth and recurrence. These cells exhibit self-renewal and proliferation properties. CSCs also exhibit significant drug resistance compared with normal tumor cells. Finding new treatments that target CSCs could significantly enhance the effect of chemotherapy and improve patient survival. Notch signaling is known to regulate the development of the lungs by controlling the cell-fate determination of normal stem cells. In this study, we isolated CSCs from the human lung adenocarcinoma cell line A549. CD133 was used as a stem cell marker for fluorescence-activated cell sorting (FACS). We compared the expression of Notch signaling in both CD133+ and CD133− cells and blocked Notch signaling using the γ-secretase inhibitor DAPT (GSI-IX). The effect of combining GSI and cisplatin (CDDP) was also examined in these two types of cells. We observed that both CD133+ and CD133− cells proliferated at similar rates, but the cells exhibited distinctive differences in cell cycle progression. Few CD133+ cells were observed in the G{sub 2}/M phase, and there were half as many cells in S phase compared with the CD133− cells. Furthermore, CD133+ cells exhibited significant resistance to chemotherapy when treated with CDDP. The expression of Notch signaling pathway members, such as Notch1, Notch2 and Hes1, was lower in CD133+ cells. GSI slightly inhibited the proliferation of both cell types and exhibited little effect on the cell cycle. The inhibitory effects of DPP on these two types of cells were

  19. 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

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

    Science.gov (United States)

    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

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

    Directory of Open Access Journals (Sweden)

    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

  2. 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.

  3. 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.

  4. 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

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

    Science.gov (United States)

    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.

  6. The effects of disodium cromoglycate on enhanced adherence of Haemophilus influenzae to A549 cells infected with respiratory syncytial virus.

    Science.gov (United States)

    Fukasawa, Chie; Ishiwada, Naruhiko; Ogita, Junko; Hishiki, Haruka; Kohno, Yoichi

    2009-08-01

    Nontypeable Haemophilus influenzae (NTHi) secondary infection often complicates respiratory syncytial virus (RSV) infections. Previous studies have revealed that RSV infections enhance NTHi adherence to airway epithelial cells. In this study, we investigated the effects of disodium cromoglycate (DSCG) and corticosteroids, which are frequently used for the treatment of wheezing often related to RSV infections, on the adherence of NTHi to RSV-infected A549 cells. DSCG inhibited enhanced adherence of NTHi to RSV-infected A549 cells, whereas dexamethasone (Dex) and fluticasone propionate (Fp) did not. DSCG suppressed the expression of ICAM-1, which is one of the NTHi receptors. Furthermore, DSCG exhibited an inhibitory effect on RSV infections. It is suggested that DSCG exerts an anti-RSV effect, and consequently attenuates the expression of NTHi receptors.

  7. Role of mechanical stretching and lipopolysaccharide in early apoptosis and IL-8 of alveolar epithelial typeⅡ cells A549

    Institute of Scientific and Technical Information of China (English)

    Qiao-Ming Ning; Xiao-Ning Sun; Xin-Kai Zhao

    2012-01-01

    Objective:To investigate the effects of mechanical stretching and lipopolysaccharide (LPS) on the early apoptosis and IL-8 production of alveolar epithelial typeⅡ cellsA549.Methods:The experimental matrix consisted of three integrated studies.In the first study,A549 cells were subjected to different stretching strain frequency and duration time to see the effects on the early apoptosis.In the second study,A549 cells were subjected to mechanical stretch(15%4 h, 0.5Hz) andLPS(1 or100 ng/mL) to see whether mechanical strain andLPS also have an addictive effect on the early apoptosis.In the third study to investigate whether this addictive effect could be induced byLPS and mechanical stretch onIL-8 production,A549 cells were subjected to LPS(100 ng/mL) and mechanical strain(15%,0.5Hz,4 h).Real timePCR and enzyme linked immunosorbent assay were used to measure mRNA and protein level ofIL-8.The early apoptosis was detected by flow cytometry.Results:Mechanical stretch induced the early apoptosis in a force and frequency and time-dependent manner.In the presence ofLPS, mechanical stretch enhancedLPS-induced early apoptosis, especially in100 ng/mLLPS group compared with1 ng/mLLPS and the control group.Mechanical stretch increasedIL-8 production and enhancedLPS-inducedIL-8 screation both in mRNA and protein levels.Conclusions:Mechanical stretch can induce the early apoptosis andIL-8 secretion.Mechanical stretch andLPS have an addictive effect on the early apoptosis andIL-8 production in alveolar type2 cells, which is one of the mechanisms of ventilator-induced lung injury.

  8. Moringa oleifera Gold Nanoparticles Modulate Oncogenes, Tumor Suppressor Genes, and Caspase-9 Splice Variants in A549 Cells.

    Science.gov (United States)

    Tiloke, Charlette; Phulukdaree, Alisa; Anand, Krishnan; Gengan, Robert M; Chuturgoon, Anil A

    2016-10-01

    Gold nanoparticles (AuNP's) facilitate cancer cell recognition and can be manufactured by green synthesis using nutrient rich medicinal plants such as Moringa oleifera (MO). Targeting dysregulated oncogenes and tumor suppressor genes is crucial for cancer therapeutics. We investigated the antiproliferative effects of AuNP synthesized from MO aqueous leaf extracts (MLAuNP ) in A549 lung and SNO oesophageal cancer cells. A one-pot green synthesis technique was used to synthesise MLAuNP . A549, SNO cancer cells and normal peripheral blood mononuclear cells (PBMCs) were exposed to MLAuNP and CAuNP to evaluate cytotoxicity (MTT assay); apoptosis was measured by phosphatidylserine (PS) externalization, mitochondrial depolarization (ΔΨm) (flow cytometry), caspase-3/7, -9 activity, and ATP levels (luminometry). The mRNA expression of c-myc, p53, Skp2, Fbw7α, and caspase-9 splice variants was determined using qPCR, while relative protein expression of c-myc, p53, SRp30a, Bax, Bcl-2, Smac/DIABLO, Hsp70, and PARP-1 were determined by Western blotting. MLAuNP and CAuNP were not cytotoxic to PBMCs, whilst its pro-apoptotic properties were confirmed in A549 and SNO cells. MLAuNP significantly increased caspase activity in SNO cells while MLAuNP significantly increased PS externalization, ΔΨm, caspase-9, caspase-3/7 activities, and decreased ATP levels in A549 cells. Also, p53 mRNA and protein levels, SRp30a (P = 0.428), Bax, Smac/DIABLO and PARP-1 24 kDa fragment levels were significantly increased. Conversely, MLAuNP significantly decreased Bcl-2, Hsp70, Skp2, Fbw7α, c-myc mRNA, and protein levels and activated alternate splicing with caspase-9a splice variant being significantly increased. MLAuNP possesses antiproliferative properties and induced apoptosis in A549 cells by activating alternate splicing of caspase-9. J. Cell. Biochem. 117: 2302-2314, 2016. © 2016 Wiley Periodicals, Inc.

  9. MiR-200a enhances the migrations of A549 and SK-MES-1 cells by regulating the expression of TSPAN1

    Indian Academy of Sciences (India)

    Yaqing Chen; Wei Peng; Yixiang Lu; Jianxin Chen; York Yuanyuan Zhu; Tao Xi

    2013-09-01

    MicroRNA-200a (miR-200a) has been reported to regulate tumour progression in several tumours; however, little is known about its role in non-small cell lung cancer cells (NSCLCs). Here, we found that miR-200a was up-regulated in A549 and SK-MES-1 cells compared with normal lung cells HELF. By a series of gain-of-function and loss-offunction studies, over-expression of miR-200a was indicated to enhance cells migration, and its knock-down inhibited migration of cells in NSCLC cell lines. Furthermore, miR-200a was identified to induce TSPAN1 expression which was related to migration. TSPAN1 was proved to induce migration, and so up-regulation of TSPAN1 by miR-200a may explain why over-expressing miR-200a promotes NSCLC cells migration.

  10. The effects of perfluorocarbon on ICAM-1 expression in LPS-induced A549 cells and the potential mechanism.

    Science.gov (United States)

    Cao, Lu; Li, Chun-Sun; Chang, Yan; Liang, Zhi-Xin; Chen, Liang-An

    2016-04-01

    Acute lung injury (ALI)/ARDS is a critical clinical syndrome with high mortality, and the effective therapeutic methods for the treatment remain limited. Previous studies have indicated that liquid ventilation with perfluorocarbon (PFC) may be advantageous over conventional mechanical ventilation in the treatment of ALI/ARDS. Additionally, PFC inhibits the inflammatory response caused by ALI/ARDS. However, the anti-inflammatory mechanism remains to be completely elucidated. In the present study, the aim was to determine the anti‑inflammatory mechanism of PFC and the association with microRNA (miR). PFC was used to modulate LPS‑induced A549 cells, with the cells divided into four groups: Untreated control group; LPS group, treated with 10 µg/ml LPS; LPS+PFC group, treated with 10 µg/ml LPS and PFC; and PFC group, treated with PFC alone. The intercellular adhesion molecule‑1 (ICAM‑1) mRNA and protein expression levels of each group were detected by reverse transcription‑quantitative polymerase chain reaction (RT‑qPCR) and western blotting, respectively. A549 cells were transfected with miR‑17‑3p mimics, miR‑17‑3p inhibitors or negative controls to observe the alterations in the anti‑inflammatory effects of PFC. A dual luciferase reporter gene assay was used to determine whether ICAM‑1 is a target gene of miR‑17‑3p. PFC was observed to attenuate the mRNA and protein expression levels of ICAM‑1 in LPS‑induced A549 cells, with no significant effect on the untreated A549 cells. miR‑17‑3p was demonstrated to be regulated by PFC. Transfection with miR‑17‑3p mimics enhanced the anti‑inflammatory effects of PFC, whereas the miR‑17‑3p inhibitor weakened the anti‑inflammatory effects of PFC at early time points. To conclude, the current study indicates that ICAM‑1 was a target gene of miR‑17‑3p, and PFC has anti‑inflammatory effects. Additionally, the present study is the first report, to the best of our knowledge, that

  11. Apoptotic induction of lung adenocarcinoma A549 cells infected by recombinant RVG Newcastle disease virus (rL-RVG) in vitro.

    Science.gov (United States)

    Yan, Yulan; Liang, Bing; Zhang, Jin; Liu, Yang; Bu, Xuefeng

    2015-01-01

    Newcastle disease virus (NDV) is a member of the genus Avulavirus in the Paramyxoviridae family and its antitumor properties depend on its ability to kill malignant cells while not affecting normal cells. The present study investigated a recombinant avirulent NDV LaSota strain (wild-type NDV strain) expressing the rabies virus glycoprotein (rL-RVG), examined its oncolytic effect on the lung adenocarcinoma A549 cell line and evaluated its potential to serve as a vaccine against lung cancer. A549 cells were infected with the rL-RVG virus and analyzed by MTT, western blot, polymerase chain reaction (PCR), immunofluorescence, terminal deoxynucleotidyl transferase dUTP nick end labeling and flow-cytometric analyses. PCR, western blot and immunofluorescence showed that the RVG gene and protein were stably expressed in A549 cells following infection with rL-RVG. The growth of A549 cells in the rL-RVG group was inhibited more effectively compared to those infected with the wild-type NDV strain. MTT results showed that cell growth inhibition rates in the rL-RVG group were significantly higher than those in the NDV group (PrL-RVG group was also more evident, with the apoptotic index being increased in rL-RVG group. The expression of the pro-apoptotic proteins caspase-3, -8 and -9 increased. The expression of caspase-3 decreased following application of the broad-specificity caspase inhibitor Z-VAD-FMK. However, the expression of the inhibitory apoptosis protein B-cell lymphoma 2 (bcl-2) did not change, but bcl-2-associated X/bcl-2 ratio was higher in the rL-RVG group than that in the NDV group. The rL-RVG strain was able to suppress lung cancer cell growth and promote lung cancer cell apoptosis to a greater extent than the wild-type NDV strain. Therefore, the rL-RVG strain is a potent antitumor agent.

  12. 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.

  13. 黄芩苷对人肺泡上皮细胞A549氧化损伤的保护作用研究%Study on the protective effect of baicalin on oxidation damage in A549 cell

    Institute of Scientific and Technical Information of China (English)

    隋妍; 李威; 杨云芬; 许秀东; 李文芳

    2012-01-01

    目的 研究黄芩苷对人肺泡上皮细胞的抗氧化作用.方法 用过氧化氢制作肺泡Ⅱ型上皮细胞A549氧化损伤模型,再加入不同浓度的黄芩苷进行处理,MTT法测定细胞活力、测定细胞培养上清液中谷胱甘肽过氧化物酶(GSH-PX)、丙二醛(MDA)、超氧化物歧化酶(SOD)、一氧化氮合酶(NOS)的活性.结果 氧化损伤组的细胞活力显著降低、MDA水平显著上升、SOD、GSH-PX、NOS活性下降(P<0.05).黄芩苷处理各组与损伤组比较,细胞活性显著升高、MDA水平显著下降、SOD、GSH-PX、NOS活性和蛋白含量显著上升(P<0.05).结论 黄芩苷对过氧化氢引起的A549细胞氧化损伤具有保护作用.%Objective To investigate the baicalin's protective effect on oxidation damage in A549. Methods A549 cells were treated with H2O2 as oxidative stress damage cell model and different concentrations of baicalin were added. Cell viability was evaluated by MIT assay and the level of MDA and SOD?NOS?CSH-PX in cell culture supernatant was measured with antioxidation testing cassete. Results The MTT assay results showed that H2 O2 decreased the viability of A549 cells (P <0. 05) ,the protein level decreased and oxidative damage index rose on the group of H2O2;. Baicalin inhibited the decrease of A549 cell viability and protein induced by H2O2(P <0.05). The decrease of SOD?NOS?GSH-PX was improved (P<0. 05). The viability of SOD,NOS?CSH-PX and protein content increased significantly. Conclusions Baca-lin could inhibit the injury caused by H2O2 and reduce the oxidative damage effect induced by H2O2 (P<0. 05 ).

  14. Platinum(II) phenanthroimidazole G-quadruplex ligand induces selective telomere shortening in A549 cancer cells.

    Science.gov (United States)

    Mancini, Johanna; Rousseau, Philippe; Castor, Katherine J; Sleiman, Hanadi F; Autexier, Chantal

    2016-02-01

    Telomere maintenance, achieved by the binding of protective shelterin capping proteins to telomeres and by either telomerase or a recombination-based alternative lengthening of telomere (ALT) mechanism, is critical for cell proliferation and survival. Extensive telomere shortening or loss of telomere integrity activates DNA damage checkpoints, leading to cell senescence or death. Although telomerase upregulation is an attractive target for anti-cancer therapy, the lag associated with telomere shortening and the potential activation of ALT pose a challenge. An alternative approach is to modify telomere interactions with binding proteins (telomere uncapping). G-quadruplex ligands stabilize structures generated from single-stranded G-rich 3'-telomere end (G-quadruplex) folding, which in principle, cannot be elongated by telomerase, thus leading to telomere shortening. Ligands can also mediate rapid anti-proliferative effects by telomere uncapping. We previously reported that the G-quadruplex ligand, phenylphenanthroimidazole ethylenediamine platinum(II) (PIP), inhibits telomerase activity in vitro[47]. In the current study, a long-term seeding assay showed that PIP significantly inhibited the seeding capacity of A549 lung cancer cells and to a lesser extent primary MRC5 fibroblast cells. Importantly, treatment with PIP caused a significant dose- and time-dependent decrease in average telomere length of A549 but not MRC5 cells. Moreover, cell cycle analysis revealed a significant increase in G1 arrest upon treatment of A549 cells, but not MRC5 cells. Both apoptosis and cellular senescence may contribute to the anti-proliferative effects of PIP. Our studies validate the development of novel and specific therapeutic ligands targeting telomeric G-quadruplex structures in cancer cells.

  15. Intracellular dynamics and fate of polystyrene nanoparticles in A549 Lung epithelial cells monitored by image (cross-) correlation spectroscopy and single particle tracking.

    Science.gov (United States)

    Deville, Sarah; Penjweini, Rozhin; Smisdom, Nick; Notelaers, Kristof; Nelissen, Inge; Hooyberghs, Jef; Ameloot, Marcel

    2015-10-01

    Novel insights in nanoparticle (NP) uptake routes of cells, their intracellular trafficking and subcellular targeting can be obtained through the investigation of their temporal and spatial behavior. In this work, we present the application of image (cross-) correlation spectroscopy (IC(C)S) and single particle tracking (SPT) to monitor the intracellular dynamics of polystyrene (PS) NPs in the human lung carcinoma A549 cell line. The ensemble kinetic behavior of NPs inside the cell was characterized by temporal and spatiotemporal image correlation spectroscopy (TICS and STICS). Moreover, a more direct interpretation of the diffusion and flow detected in the NP motion was obtained by SPT by monitoring individual NPs. Both techniques demonstrate that the PS NP transport in A549 cells is mainly dependent on microtubule-assisted transport. By applying spatiotemporal image cross-correlation spectroscopy (STICCS), the correlated motions of NPs with the early endosomes, late endosomes and lysosomes are identified. PS NPs were equally distributed among the endolysosomal compartment during the time interval of the experiments. The cotransport of the NPs with the lysosomes is significantly larger compared to the other cell organelles. In the present study we show that the complementarity of ICS-based techniques and SPT enables a consistent elaborate model of the complex behavior of NPs inside biological systems.

  16. 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.

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

    Directory of Open Access Journals (Sweden)

    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.

  18. AS1411对紫杉醇耐药肺腺癌A549细胞凋亡的影响%Effects of AS1411 on the apoptosis of taxol-resistant lung adenocarcinoma A549 cell

    Institute of Scientific and Technical Information of China (English)

    周卫; 赵梓彤; 刘玲燕; 詹启敏; 宋咏梅

    2014-01-01

    目的 探讨核酸适配子AS1411对紫杉醇耐药肺腺癌A549细胞(A549/T细胞)凋亡的影响.方法 采用0 ~ 20.0 μmol/L浓度的AS1411处理A549/T细胞,甲基甲苯基硫细胞检测(MTS)实验、平板克隆形成实验检测细胞活性[吸光度值(A490nm)]及增殖能力变化,流式细胞仪检测对细胞凋亡的影响,同时应用蛋白免疫印迹法检测凋亡信号通路相关蛋白表达的变化.结果 A549/T细胞呈现部分上皮细胞间质化(EMT)、表皮生长因子受体(EGFR)表达缺失等特征.经5.0 μmol/L的AS1411处理后,与对照序列相比,A549/T细胞的细胞活性显著降低(A490nm:0.185±0.009比0.272±0.006,P<0.001)、克隆形成数显著减少(74±13比120±12,P=0.010);随AS1411浓度的增加,细胞活性明显降低,呈剂量依赖性.用20.0 μmol/L的AS1411处理48 h后,A549/T细胞凋亡率显著高于对照序列[(19.9±2.6)%比(8.8±1.3)%,P=0.002],同时蛋白激酶B(AKT)、细胞外信号调节激酶1/2(ERK1/2)和B细胞淋巴瘤因子2(Bcl-2)表达均显著低于对照序列处理组(0.353±0.003、0.432±0.015、0.294 ±0.015比0.688±0.003、0.911±0.019、0.422±0.018,均P<0.001).结论 AS1411可通过抑制AKT-ERK通路而促进A549/T细胞凋亡.%Objective To explore the effects of AS1411 on the apoptosis of taxol-resistant lung adenocarinoma A549 cell (A549/T cell).Methods A549/T cells were treated with AS1411 at a concentration gradient of 0-20.0 μmol/L.The assays of methyl tolyl sulfide (MTS) and colony formation were used to detect the cellular vitality (absorbance value (A490,nm)) and proliferation.The apoptotic effects were detected by flow cytometer and the relevant apoptotic signaling proteins detected by Western blot.Results A549/T cells exhibited some characteristics of epithelial mesenchymal transition (EMT) and a negative expression of epidermal growth factor receptor (EGFR).After a treatment of 5.0 μmol/L AS1411,compared to the control sequence,cell vitality was inhibited

  19. 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.

  20. 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

    the preparation process of DC , should be selected as optimum extraction time DC ; while DC and human lung cancer cell line A -549 with a ratio of 1∶1 shows the highest percentage combined vaccine .

  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. Radiosensitization of TPGS-emulsified docetaxel-loaded poly(lactic-co-glycolic acid) nanoparticles in CNE-1 and A549 cells.

    Science.gov (United States)

    Shi, Wei; Yuan, Yin; Chu, Min; Zhao, Shuang; Song, Qingle; Mu, Xiaoqian; Xu, Shuangbing; Zhang, Zhiping; Yang, Kunyu

    2016-03-01

    Docetaxel is among the most effective radiosensitizers. It is widely used as radiosensitizer in many tumors, including head and neck carcinoma. Nevertheless, poor solubility and severe hypersensitivity limit its clinical use and its therapeutic effect remains to be improved. In this study, docetaxel-loaded polymeric nanoparticles were prepared by nanoprecipitation method to be new radiosensitizer with lower side effects and higher efficacy. The physiochemical characteristics of the nanoparticles were studied. Two human tumor cell lines which are resistant to radiotherapy were used in this research. We have compared the radioenhancement efficacy of docetaxel-loaded nanoparticles with docetaxel in A549 and CNE-1 cells. Compared with docetaxel, radiosensitization of docetaxel-loaded nanoparticles was improved significantly (sensitization enhancement ratio in A549 increased 1.24-fold to 1.68-fold when the radiation was applied 2 h after the drug, p nanoparticles. The improved radiosensitization efficacy was associated with enhanced G2/M arrest, promoted apoptosis and the role of D-alpha-tocopheryl polyethylene glycol 1000 succinate which will enhance the cell uptake and inhibit the multiple drug resistance. Moreover, the radiosensitization efficacy of docetaxel-loaded nanoparticles was more prominent than docetaxel. In conclusion, tocopheryl polyethylene glycol 1000 succinate-emulsified docetaxel-loaded PLGA nanoparticles were more efficacious and fewer adverse effects were observed than with the commercial docetaxel formulation. Thus, PLGA nanoparticles hold promise as a radiosensitizing agent.

  3. The surface charge of liposomal adjuvants is decisive for their interactions with the Calu-3 and A549 airway epithelial cell culture models

    DEFF Research Database (Denmark)

    Ingvarsson, Pall Thor; Rasmussen, Ida Svahn; Viaene, Michelle

    2014-01-01

    potential for mucosal vaccination via the airways. The purpose of this study was to investigate the importance of the liposomal surface charge on the interaction with lung epithelial cells. Thus, the cationic DDA in the liposomes was subjected to a step-wise replacement with the zwitterionic...... distearoylphosphatidylcholine (DSPC). The liposomes were tested with the model protein antigen ovalbumin for the mucosal deposition, the effect on cellular viability and the epithelial integrity by using the two cell lines A549 and Calu-3, representing cells from the alveolar and the bronchiolar epithelium, respectively...... and viability of the mucus-covered Calu-3 cells. Our in vitro results thus indicate that DDA/TDB liposomes might be efficiently and safely used as an adjuvant system for vaccines targeting the mucus-covered epithelium of the upper respiratory tract and the conducting airways....

  4. [Influence of Berberine on Cisplatin Antineoplastic Effect in A549 Cells].

    Science.gov (United States)

    Jiang, Guojun; Li, Li; Wu, Xiaoxiang; Dong, Shuying; Tong, Xuhui

    2015-08-01

    背景与目的 以顺铂为基础的化疗方案是晚期非小细胞肺癌的一线化疗方案,但是由于顺铂的不良反应严重及耐药性的产生均限制了它的临床应用,本研究采用联合用药的方式观察黄连素对顺铂抗肿瘤作用的影响,并探讨其可能机制。方法 分别观察黄连素对肺腺癌细胞A549细胞中总Cx43蛋白、细胞膜Cx43蛋白的表达以及细胞缝隙连接功能的改变,通过标准细胞集落克隆实验观察黄连素对顺铂细胞毒性的影响;并观察PKC激酶的表达。结果 黄连素在0 μM-10 μM浓度范围内对细胞无毒性,通过增加细胞内总Cx43蛋白和胞膜Cx43蛋白的表达而增强细胞缝隙连接功能,0.1 μM、1 μM、10 μM黄连素可以显著增强细胞间的荧光传递,与空白对照组相比,黄连素预处理后的细胞间荧光传递功能分别增加了33.3% (P=0.002,3)、67.0% (P<0.001)、160.0% (P<0.001),这种作用与PKC的活性被抑制相关,抑制PKC活性可以进一步增加顺铂对A549细胞的毒性作用。结论 黄连素可通过增加A549细胞的缝隙连接功能而明显增强顺铂的细胞毒性。.

  5. Proteomic analysis of cellular response induced by multi-walled carbon nanotubes exposure in A549 cells.

    Directory of Open Access Journals (Sweden)

    Li Ju

    Full Text Available The wide application of multi-walled carbon nanotubes (MWCNT has raised serious concerns about their safety on human health and the environment. However, the potential harmful effects of MWCNT remain unclear and contradictory. To clarify the potentially toxic effects of MWCNT and to elucidate the associated underlying mechanisms, the effects of MWCNT on human lung adenocarcinoma A549 cells were examined at both the cellular and the protein level. Cytotoxicity and genotoxicity were examined, followed by a proteomic analysis (2-DE coupled with LC-MS/MS of the cellular response to MWCNT. Our results demonstrate that MWCNT induces cytotoxicity in A549 cells only at relatively high concentrations and longer exposure time. Within a relatively low dosage range (30 µg/ml and short time period (24 h, MWCNT treatment does not induce significant cytotoxicity, cell cycle changes, apoptosis, or DNA damage. However, at these low doses and times, MWCNT treatment causes significant changes in protein expression. A total of 106 proteins show altered expression at various time points and dosages, and of these, 52 proteins were further identified by MS. Identified proteins are involved in several cellular processes including proliferation, stress, and cellular skeleton organization. In particular, MWCNT treatment causes increases in actin expression. This increase has the potential to contribute to increased migration capacity and may be mediated by reactive oxygen species (ROS.

  6. 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.

  7. Acrolein activates cell survival and apoptotic death responses involving the endoplasmic reticulum in A549 lung cells.

    Science.gov (United States)

    Tanel, André; Pallepati, Pragathi; Bettaieb, Ahmed; Morin, Patrick; Averill-Bates, Diana A

    2014-05-01

    Acrolein, a highly reactive α,β-unsaturated aldehyde, is a product of endogenous lipid peroxidation. It is a ubiquitous environmental pollutant that is generated mainly by smoke, overheated cooking oil and vehicle exhaust. Acrolein damages cellular proteins, which could lead to accumulation of aberrantly-folded proteins in the endoplasmic reticulum (ER). This study determines the mechanisms involved in acrolein-induced apoptosis mediated by the ER and possible links with the ER stress response in human A549 lung cells. The exposure of cells to acrolein (15-50μM) for shorter times of 15 to 30min activated several ER stress markers. These included the ER chaperone protein BiP and the three ER sensors: (i) the survival/rescue molecules protein kinase RNA (PKR)-like ER kinase (PERK) and eukaryotic initiation factor 2 alpha (eIF2α) were phosphorylated; (ii) cleavage of activating transcription factor 6 (ATF6) occurred, and (iii) inositol-requiring protein-1 alpha (IRE1α) was phosphorylated. Acrolein (25-50μM) caused apoptotic cell death mediated by the ER after 2h, which was characterised by the induction of CHOP and activation of ER proteases calpain and caspase-4. Calpain and caspase-7 were the initiating factors for caspase-4 activation in acrolein-induced apoptosis. These results increase our knowledge about cellular responses to acrolein in lung cells, which have implications for human health.

  8. 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.

    Science.gov (United States)

    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.

  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.

    Directory of Open Access Journals (Sweden)

    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.

  10. Cellular and spectroscopic characterization of cancer stem cell-like cells derived from A549 lung carcinoma

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    Murali M. S. Balla

    2016-01-01

    Conclusions and General Significance: Overall, these observations provide novel FT-IR and CD spectroscopy signatures in A549 clones enriched with CSCs, which may have implications in the quantifying magnitude of CSCs as prognostic markers in cancer therapy.

  11. The Effect of Vitamin A on Secretion of IFN-γ and IL-4 in A549 Cells Induced by Mycoplasma Pneumoniae

    Institute of Scientific and Technical Information of China (English)

    Xiaolan WU; Xianzhou LIU; Jilu TANG

    2008-01-01

    In order to investigate the effect of vitamin A (VA) on the secretion of IFN-γ and IL-4 in Mycoplasma Pneumoniae (MP)-induced A549 cells, A549 cells were co-cultured with MP for different time lengths and then the levels of IFN-γ and IL-4 in the cell culture supematants were detected before and after treatment with different concentrations of VA by using the enzyme-linked immunosorbent assay (ELISA). The results showed that the level of IFN-γ and IL-4 in the supernatants of MP-induced A549 cells was much higher than that in non-induced cells (P<0.01). After application of VA, IL-4 level was not increased until the concentration of VA was up to 0.5 × 10-5 mol/L (P<0.01).However, with concentration of VA increased up to 1 × 10-4 mol/L, IL-4 was significantly suppressed (P<0.01). It was concluded that MP could induce the secretion of IFN-y and IL-4 in A549 cells. VA could inhibit the secretion of IFN-γ, and increase the IL-4 level in MP-induced A549 cells. However,high concentration of VA had an inhibitory effect on the secretion of IL-4 as well as on the IFN-γ.These data provided a theoretical basis for the application of VA in MP pneumonia in the clinical practice.

  12. 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.

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

    Energy Technology Data Exchange (ETDEWEB)

    Yun, Hong Shik; Hong, Eun-Hee [Division of Radiation Cancer Biology, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Department of Chemistry, College of Natural Sciences, Hanyang University, Seoul 133-791 (Korea, Republic of); Lee, Su-Jae [Department of Chemistry, College of Natural Sciences, Hanyang University, Seoul 133-791 (Korea, Republic of); Baek, Jeong-Hwa [Division of Radiation Cancer Biology, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon 440-746 (Korea, Republic of); Lee, Chang-Woo [Department of Molecular Cell Biology, Samsung Biomedical Research Institute, Sungkyunkwan University School of Medicine, Suwon 440-746 (Korea, Republic of); Yim, Ji-Hye; Um, Hong-Duck [Division of Radiation Cancer Biology, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of); Hwang, Sang-Gu, E-mail: sgh63@kcch.re.kr [Division of Radiation Cancer Biology, Korea Institute of Radiological and Medical Sciences, Seoul 139-706 (Korea, Republic of)

    2013-09-27

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

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

    Directory of Open Access Journals (Sweden)

    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

  15. IFN-gamma Impairs Release of IL-8 by IL-1beta-stimulated A549 Lung Carcinoma Cells

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    Pfeilschifter Josef

    2008-09-01

    Full Text Available Abstract Background Production of interferon (IFN-γ is key to efficient anti-tumor immunity. The present study was set out to investigate effects of IFNγ on the release of the potent pro-angiogenic mediator IL-8 by human A549 lung carcinoma cells. Methods A549 cells were cultured and stimulated with interleukin (IL-1β alone or in combination with IFNγ. IL-8 production by these cells was analyzed with enzyme linked immuno sorbent assay (ELISA. mRNA-expression was analyzed by real-time PCR and RNase protection assay (RPA, respectively. Expression of inhibitor-κ Bα, cellular IL-8, and cyclooxygenase-2 was analyzed by Western blot analysis. Results Here we demonstrate that IFNγ efficiently reduced IL-8 secretion under the influence of IL-1β. Surprisingly, real-time PCR analysis and RPA revealed that the inhibitory effect of IFNγ on IL-8 was not associated with significant changes in mRNA levels. These observations concurred with lack of a modulatory activity of IFNγ on IL-1β-induced NF-κB activation as assessed by cellular IκB levels. Moreover, analysis of intracellular IL-8 suggests that IFNγ modulated IL-8 secretion by action on the posttranslational level. In contrast to IL-8, IL-1β-induced cyclooxygenase-2 expression and release of IL-6 were not affected by IFNγ indicating that modulation of IL-1β action by this cytokine displays specificity. Conclusion Data presented herein agree with an angiostatic role of IFNγ as seen in rodent models of solid tumors and suggest that increasing T helper type 1 (Th1-like functions in lung cancer patients e.g. by local delivery of IFNγ may mediate therapeutic benefit via mechanisms that potentially include modulation of pro-angiogenic IL-8.

  16. Heat-modified citrus pectin induces apoptosis-like cell death and autophagy in HepG2 and A549 cancer cells.

    Directory of Open Access Journals (Sweden)

    Lionel Leclere

    Full Text Available Cancer is still one of the leading causes of death worldwide, and finding new treatments remains a major challenge. Previous studies showed that modified forms of pectin, a complex polysaccharide present in the primary plant cell wall, possess anticancer properties. Nevertheless, the mechanism of action of modified pectin and the pathways involved are unclear. Here, we show that citrus pectin modified by heat treatment induced cell death in HepG2 and A549 cells. The induced cell death differs from classical apoptosis because no DNA cleavage was observed. In addition, Z-VAD-fmk, a pan-caspase inhibitor, did not influence the observed cell death in HepG2 cells but appeared to be partly protective in A549 cells, indicating that heat-modified citrus pectin might induce caspase-independent cell death. An increase in the abundance of the phosphatidylethanolamine-conjugated Light Chain 3 (LC3 protein and a decrease in p62 protein abundance were observed in both cell types when incubated in the presence of heat-modified citrus pectin. These results indicate the activation of autophagy. To our knowledge, this is the first time that autophagy has been revealed in cells incubated in the presence of a modified form of pectin. This autophagy activation appears to be protective, at least for A549 cells, because its inhibition with 3-methyladenine increased the observed modified pectin-induced cytotoxicity. This study confirms the potential of modified pectin to improve chemotherapeutic cancer treatments.

  17. Glucosamine, a naturally occurring amino monosaccharide, inhibits A549 and H446 cell proliferation by blocking G1/S transition.

    Science.gov (United States)

    Ju, Yinghua; Yu, Aiming; Sun, Xiuhua; Wu, Didi; Zhang, Hongkai

    2013-09-01

    Uncontrolled proliferation is important in tumorigenesis. In the present study, the effects of glucosamine on lung cancer cell proliferation were investigated. The expression of cyclin E, one of the key cyclins in the G1/S transition, and Skp2, the ubiquitin ligase subunit that targets the negative cell cycle regulator, p27Kip1, were also assessed. Moreover, the underlying mechanisms of action of glucosamine were investigated in lung cancer cells. A549 and H446 cells were synchronized using thymidine in the presence or absence of glucosamine. The effect of glucosamine on lung cancer cell proliferation was determined by MTT assay. Cyclin E and p27Kip1 proteins and their phosphorylation levels were detected by western blot analysis. Furthermore, the effect of glucosamine on the cell cycle was evaluated by flow cytometry. Glucosamine was found to inhibit lung cancer cell proliferation and to suppress Skp2 and cyclin E expression. Notably, the phosphorylation levels of cyclin E (Thr62) and p27Kip1 (Thr187) were downregulated by glucosamine, and negatively correlated with degradation. Glucosamine was also found to arrest lung cancer cells in the G1/S phase. Thus, glucosamine may inhibit lung cancer cell proliferation by blocking G1/S transition through the inhibition of cyclin E and Skp2 protein expression.

  18. Telomerase Cajal body protein 1 depletion inhibits telomerase trafficking to telomeres and induces G1 cell cycle arrest in A549 cells.

    Science.gov (United States)

    Yuan, Ping; Wang, Zhitian; Lv, Wang; Pan, Hui; Yang, Yunhai; Yuan, Xiaoshuai; Hu, Jian

    2014-09-01

    Telomerase Cajal body protein 1 (TCAB1) is a telomerase holoenzyme, which is markedly enriched in Cajal bodies (CBs) and facilitates the recruitment of telomerase to CBs in the S phase of the cell cycle. This recruitment is dependent on TCAB1 binding to a telomerase RNA component. The majority of cancer cells are able to grow indefinitely due to telomerase and its mechanism of trafficking to telomeres. In the present study, a certain level of TCAB1 expression in A549 human lung cells was identified and TCAB1 knockdown exhibited a potent antiproliferative effect on these cells, which was coupled with a decrease in the cell density and activity of the cellular enzymes. In addition, TCAB1-depletion was demonstrated to inhibit telomerase trafficking to telomeres in the A549 cells, leading to subsequent G1 cell cycle arrest without inducing apoptotic cell death. Overall, these observations indicated that TCAB1 may be essential for A549 cell proliferation and cell cycle regulation, and may be a potential candidate for the development of a therapeutic target for lung adenocarcinomas.

  19. 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.

  20. Apoptosis-Inducing Activity of Marine Sponge Haliclona sp. Extracts Collected from Kosrae in Nonsmall Cell Lung Cancer A549 Cells

    Directory of Open Access Journals (Sweden)

    Woori Bae

    2015-01-01

    Full Text Available Although various anticancer drugs have been developed for the treatment of nonsmall cell lung cancer, chemotherapeutic efficacy is still limited. Natural products such as phytochemicals have been screened as novel alternative materials, but alternative funds such as marine bioresources remain largely untapped. Of these resources, marine sponges have undergone the most scrutiny for their biological activities, including antiinflammatory, antiviral, and anticancer properties. However, the biological mechanisms of the activities of these marine sponges are still unclear. We investigated the anticancer activity of marine sponges collected from Kosrae in Micronesia and examined their mechanisms of action using nonsmall cell lung cancer A549 cells as a model system. Of 20 specimens, the Haliclona sp. (KO1304-328 showed both dose- and time-dependent cytotoxicity. Further, methanol extracts of Haliclona sp. significantly inhibited cell proliferation and cell viability. A549 cells treated with Haliclona sp. demonstrated induced expression of c-Jun N-terminal kinase (JNK, p53, p21, caspase-8, and caspase-3. The percentage of apoptotic cells significantly increased in A549 cultures treated with Haliclona sp. These results indicate that Haliclona sp. induces apoptosis via the JNK-p53 pathway and caspase-8, suggesting that this marine sponge is a good resource for the development of drugs for treatment of nonsmall cell lung cancer.

  1. 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.

  2. Impact of CHK2-small interfering RNA on CpG ODN7909-enhanced radiosensitivity in lung cancer A549 cells

    OpenAIRE

    Chen W; Liu XQ; Qiao TK; Yuan SJ

    2012-01-01

    Wei Chen,* Xiaoqun Liu,* Tiankui Qiao, Sujuan Yuan Department of Oncology, Jinshan Hospital, Fudan University, Shanghai, People's Republic of China*These authors contributed equally to this workObjective: To investigate the impact of checkpoint kinase 2 (CHK2)-small interfering RNA (CHK2-siRNA) on the enhancement of radiosensitivity by CpG oligodeoxynucleotide (ODN) 7909 in lung cancer A549 cells.Methods: The A549 cells were randomly divided into five groups: control, CpG, X-ray, CpG ...

  3. 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.

  4. 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

  5. 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.

  6. Study of simulated microgravity affecting antitumor response of mes-enchymal stem cells against A549 transplantation tumor%模拟微重力对骨髓间充质干细胞全细胞瘤苗干预A549移植瘤增殖的研究

    Institute of Scientific and Technical Information of China (English)

    李静; 陈军; 李秀玉; 牛潞芳

    2016-01-01

    目的:比较骨髓间充质干细胞(MSCs)于不同重力环境下获得全细胞抗原(WCAs)对于人肺腺癌细胞株A549荷瘤的影响。方法全骨髓贴壁法原代培养小鼠MSCs,采用2D回转模式,以30 r/min水平回转模拟微重力(MMG)培养条件,并以正常重力(NG)为对照培养 MSCs;随后以15 Gy的 X线灭活 MSCs获取 WCAs,将BALB/c小鼠随机分成4组,实验组(MMG组)皮下接种WCAs为(1次/3 d,共2周);对照组分别为PBS组,NG组,A549组;各组均采用A549细胞系皮下移植荷瘤,观察4组小鼠肿瘤生长情况;测量肿瘤直径、计算肿瘤体积,并采用免疫组化法检测增殖细胞核抗原(PCNA)表达。结果肺腺癌A549细胞皮下移植使小鼠成功荷瘤, MMG可促进WCAs抑制移植瘤的生长。MMG组肿瘤体积显著小于PBS组与NG组(P<0.05);NG组及MMG组的肿瘤至第6天方见生长,MMG组移植瘤体积显著小于NG组(P<0.05)。同时,A459组的抑制作用最强;PBS组肿瘤最大;12 d对各组移植瘤称重,其中NG组为(458.7±21.6)g,MMG组为(315.6±18.5)g,MMG组的抑制作用显著高于NG组(P<0.05)。HRP标记染色提示MMG可增强WCAs的免疫刺激,使得PCNA表达下调(P<0.05)。结论采用MSCs获得WCAs进行免疫应激可产生抑制肿瘤生长作用,MMG可加强其抑瘤作用,能显著观察到PCNA表达下调,其内在的免疫分子机制有待深层次的探索。%Objective To estimate the whole cell antigens (WCAs) of mesenchymal stem cells (MSCs) cultured in dif-ferent gravity can generate different immune response against cancer in mice which induced by lung cancer cell line A549. Methods The MSCs was isolated and cultured adherent cells from marrow and a 2-dimensional clinostat (2D clinostat) was used to keep cells in continuous 2D rotation at a speed of 30 r/min around the horizontal axis, to mimc the microgravity (MMG), and the normal gravity (NG) was used as control. And then MSCs was to irradiate to X-ray (15 Gy) to get the WCAs. The

  7. 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

  8. Wheatgrass extract inhibits hypoxia-inducible factor-1-mediated epithelial-mesenchymal transition in A549 cells

    Science.gov (United States)

    Do, Nam Yong; Shin, Hyun-Jae

    2017-01-01

    BACKGROUND/OBJECTIVES Epithelial-mesenchymal transition (EMT) is involved in not only cancer development and metastasis but also non-cancerous conditions. Hypoxia is one of the proposed critical factors contributing to formation of chronic rhinosinusitis or nasal polyposis. Wheatgrass (Triticum aestivum) has antioxidant, anti-aging, and anti-inflammatory effects. In this study, we analyzed whether wheatgrass has an inhibitory effect on the EMT process in airway epithelial cells. MATERIALS/METHODS A549 human lung adenocarcinoma cells were incubated in hypoxic conditions (CO2 5%/O2 1%) for 24 h in the presence of different concentrations of wheatgrass extract (50, 75, 100, and 150 µg/mL) and changes in expression of epithelial or mesenchymal markers were evaluated by immunoblotting and immunofluorescence. Accordingly, associated EMT-related transcriptional factors, Snail and Smad, were also evaluated. RESULTS Hypoxia increased expression of N-cadherin and reduced expression of E-cadherin. Mechanistically, E-cadherin levels were recovered during hypoxia by silencing hypoxia inducible factor (HIF)-1α or administering wheatgrass extract. Wheatgrass inhibited the hypoxia-mediated EMT by reducing the expression of phosphorylated Smad3 (pSmad3) and Snail. It suppressed the hypoxia-mediated EMT processes of airway epithelial cells via HIF-1α and the pSmad3 signaling pathway. CONCLUSION These results suggest that wheatgrass has potential as a therapeutic or supplementary agent for HIF-1-related diseases.

  9. Cyto- and genotoxicity assessment of Gold nanoparticles obtained by laser ablation in A549 lung adenocarcinoma cells

    Energy Technology Data Exchange (ETDEWEB)

    Bucchianico, Sebastiano Di [Karolinska Institutet, Institute of Environmental Medicine (Sweden); Migliore, Lucia [University of Pisa, Department of Translational Research and New Technologies in Medicine and Surgery, Division of Medical Genetics (Italy); Marsili, Paolo [Institute of Complex Systems (ISC-CNR) (Italy); Vergari, Chiara [Plasma Diagnostics and Technologies s.r.l. (Italy); Giammanco, Francesco [University of Pisa, Department of Physics “E. Fermi” (Italy); Giorgetti, Emilia, E-mail: emilia.giorgetti@fi.isc.cnr.it [Institute of Complex Systems (ISC-CNR) (Italy)

    2015-05-15

    Gold nanoparticles have attracted enormous interest in biomedical applications, based on their unique optical properties. However, their toxicity on human tissues is still an open issue. Beyond the potential intrinsic toxicity of nanostructured gold, a non-negligible contribution of stabilizers or reaction by-products related to current wet chemical synthesis procedures can be expected. Aimed at isolating gold contribution from that of any other contaminant, we produced colloidal suspensions of Gold nanoparticles having average size <10 nm in deionized water or acetone by pulsed laser ablation, that permits preparation of uncoated and highly stable Gold nanoparticles in pure solvents. Subsequently, we investigated the role of surface chemistry, size, and dispersivity of synthesized Gold nanoparticles in exerting toxicity in a cell model system of deep respiratory tract, representing the main route of exposure to NPs, namely adenocarcinoma epithelial A549 cells. Gold nanoparticles prepared in water showed no particular signs of cytotoxicity, cytostasis, and/or genotoxicity as assessed by MTT colorimetric viability test and Cytokinesis-block micronucleus cytome assay up to concentrations of the order of 5 μg/mL. In contrast, Gold nanoparticles produced in pure acetone and then transferred into deionized water showed impaired cell viability, apoptosis responses, micronuclei, and dicentric chromosomes induction as well as nuclear budding, as a function of the amount of surface contaminants like amorphous carbon and enolate ions.

  10. 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.

  11. Tamarind seed coat ameliorates fluoride induced cytotoxicity, oxidative stress, mitochondrial dysfunction and apoptosis in A549 cells.

    Science.gov (United States)

    Ameeramja, Jaishabanu; Panneerselvam, Lakshmikanthan; Govindarajan, Vimal; Jeyachandran, Sivakamavalli; Baskaralingam, Vaseeharan; Perumal, Ekambaram

    2016-01-15

    Fluoride (F) is an environmental contaminant and industrial pollutant. Molecular mechanisms remain unclear in F induced pulmonary toxicity even after numerous studies. Tamarind fruits act as defluoridating agents, but no study was conducted in in vitro systems. Hence, we aimed to assess the ameliorative impact of the tamarind seed coat extract (TSCE) against F toxicity utilizing lung epithelial cells, A549. Cells were exposed to sodium fluoride (NaF-5 mM) alone and in combination with TSCE (750 ng/ml) or Vitamin C (positive control) for 24 h and analyzed for F content, intracellular calcium ([Ca(2+)]i) level, oxidative stress, mitochondrial integrity and apoptotic markers. TSCE treatment prevented the F induced alterations in [Ca(2+)]i overload, F content, oxidant (reactive oxygen species generation, lipid peroxidation, protein carbonyl content and nitric oxide) and antioxidant (superoxide dismutase, catalase, glutathione peroxidase and glutathione) parameters. Further, TSCE modulates F activated changes in mitochondrial membrane potential, permeability transition pore opening, cytochrome-C release, Bax/Bcl-2 ratio, caspase-3 and PARP-1 expressions. In conclusion, our study demonstrated that TSCE as a potential protective agent against F toxicity, which can be utilized as a neutraceutical.

  12. 肺腺癌A549细胞葡萄糖代谢与紫杉醇耐药的关系%Glucose metabolism of lung adenocarcinoma A549 cells and its correlation with taxol-resistance

    Institute of Scientific and Technical Information of China (English)

    赵士艳; 周翔; 李佳津; 黄钢

    2013-01-01

    目的:探讨人肺腺癌细胞株A549及其紫杉醇耐药细胞株A549/taxol之间葡萄糖代谢的差异,以及二氯乙酸盐(dichloroacetate,DCA)对这2种细胞葡萄糖代谢的影响.方法:首先采用CCK-8法检测A549A549/taxol细胞对紫杉醇的耐药性.再用液体闪烁仪检测A549A549/taxol细胞摄取14C-葡萄糖后CO2的产生和脂质的生成情况.另外,分别用γ计数器和乳酸测定试剂盒检测18氟-2-脱氧-β-D-葡萄糖(18F-2-deoxy-β-D-glucose,18F-FDG)摄取和乳酸产生情况.结果:A549/taxol细胞摄取6-14C-葡萄糖后CO2释放量、18F-FDG摄取率和乳酸生成量均低于A549细胞.A549细胞经DCA处理后6-14C-葡萄糖释放的CO2水平升高,而A549/taxol细胞经DCA处理后6-14C-葡萄糖释放的CO2量无变化.结论:A549/taxol细胞有一定的线粒体氧化呼吸抑制作用.DCA能促进A549细胞线粒体的氧化呼吸作用,而对其耐药株A549/taxol细胞的氧化呼吸作用不大.

  13. Resveratrol reduces IL-6 and VEGF secretion from co-cultured A549 lung cancer cells and adipose-derived mesenchymal stem cells.

    Science.gov (United States)

    Sahin, Erhan; Baycu, Cengiz; Koparal, Ayse Tansu; Burukoglu Donmez, Dilek; Bektur, Ezgi

    2016-06-01

    Stem cell therapies are important treatment methodologies used in many areas of experimental or clinical medicine. In recent studies of cancer models, Mesenchymal stem cells (MSCs) suppressed the growth of cancer cells. However, also in some studies, stem cell treatments have been shown to induce cancer formation, increase tumor volume, induce the formation of new vessels, and lead to cancer invasion. The presence of MSC-secreted cytokines and their effects on cancer cells limits the reliability of MSC-based treatments. Resveratrol (trans-3,5,4'-trihydroxystilbene), an antioxidant found in red wine, has been shown to have therapeutic effects against several cancers. The aim of this study was to co-culture MSCs with A549 cancer cells to suppress the release of cancer-promoting cytokines from MSCs and to increase the applicability and reliability of stem cell therapies with resveratrol. MTT (3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) and neutral red cell viability assays were used to find safety dose of resveratrol. The MSCs secreted the cytokines IL-6 and VEGF, and the effect of resveratrol on these cytokines was analyzed by ELISA and western blot analysis of conditioned medium. One μM of resveratrol was found to be the safety dose for the A549 cancer cells and MSCs. We observed the highest release of IL-6 and VEGF from the co-cultured A549 cells and MSCs, and resveratrol was found to significantly decrease the release of these cytokines. Our study suggests that resveratrol exerts a positive effect on the release of cytokines. The safety dose of resveratrol can be administered together with stem cells during stem cell treatment.

  14. Effects of Alpha Particle and Proton Beam Irradiation as Putative Cross-Talk between A549 Cancer Cells and the Endothelial Cells in a Co-Culture System

    Directory of Open Access Journals (Sweden)

    Hélène Riquier

    2015-03-01

    Full Text Available Background: High-LET ion irradiation is being more and more often used to control tumors in patients. Given that tumors are now considered as complex organs composed of multiple cell types that can influence radiosensitivity, we investigated the effects of proton and alpha particle irradiation on the possible radioprotective cross-talk between cancer and endothelial cells. Materials and Methods: We designed new irradiation chambers that allow co-culture study of cells irradiated with a particle beam. A549 lung carcinoma cells and endothelial cells (EC were exposed to 1.5 Gy of proton beam or 1 and 2 Gy of alpha particles. Cell responses were studied by clonogenic assays and cell cycle was analyzed by flow cytometry. Gene expression studies were performed using Taqman low density array and by RT-qPCR. Results: A549 cells and EC displayed similar survival fraction and they had similar cell cycle distribution when irradiated alone or in co-culture. Both types of irradiation induced the overexpression of genes involved in cell growth, inflammation and angiogenesis. Conclusions: We set up new irradiation chamber in which two cell types were irradiated together with a particle beam. We could not show that tumor cells and endothelial cells were able to protect each other from particle irradiation. Gene expression changes were observed after particle irradiation that could suggest a possible radioprotective inter-cellular communication between the two cell types but further investigations are needed to confirm these results.

  15. Effects of Alpha Particle and Proton Beam Irradiation as Putative Cross-Talk between A549 Cancer Cells and the Endothelial Cells in a Co-Culture System

    Science.gov (United States)

    Riquier, Hélène; Abel, Denis; Wera, Anne-Catherine; Heuskin, Anne-Catherine; Genard, Géraldine; Lucas, Stéphane; Michiels, Carine

    2015-01-01

    Background: High-LET ion irradiation is being more and more often used to control tumors in patients. Given that tumors are now considered as complex organs composed of multiple cell types that can influence radiosensitivity, we investigated the effects of proton and alpha particle irradiation on the possible radioprotective cross-talk between cancer and endothelial cells. Materials and Methods: We designed new irradiation chambers that allow co-culture study of cells irradiated with a particle beam. A549 lung carcinoma cells and endothelial cells (EC) were exposed to 1.5 Gy of proton beam or 1 and 2 Gy of alpha particles. Cell responses were studied by clonogenic assays and cell cycle was analyzed by flow cytometry. Gene expression studies were performed using Taqman low density array and by RT-qPCR. Results: A549 cells and EC displayed similar survival fraction and they had similar cell cycle distribution when irradiated alone or in co-culture. Both types of irradiation induced the overexpression of genes involved in cell growth, inflammation and angiogenesis. Conclusions: We set up new irradiation chamber in which two cell types were irradiated together with a particle beam. We could not show that tumor cells and endothelial cells were able to protect each other from particle irradiation. Gene expression changes were observed after particle irradiation that could suggest a possible radioprotective inter-cellular communication between the two cell types but further investigations are needed to confirm these results. PMID:25794049

  16. Effects of Alpha Particle and Proton Beam Irradiation as Putative Cross-Talk between A549 Cancer Cells and the Endothelial Cells in a Co-Culture System

    Energy Technology Data Exchange (ETDEWEB)

    Riquier, Hélène; Abel, Denis [URBC-NARILIS, University of Namur, 61 rue de Bruxelles, Namur 5000 (Belgium); Wera, Anne-Catherine; Heuskin, Anne-Catherine [LARN-PMR, NARILIS, University of Namur, Namur 5000 (Belgium); Genard, Géraldine [URBC-NARILIS, University of Namur, 61 rue de Bruxelles, Namur 5000 (Belgium); Lucas, Stéphane [LARN-PMR, NARILIS, University of Namur, Namur 5000 (Belgium); Michiels, Carine, E-mail: carine.michiels@unamur.be [URBC-NARILIS, University of Namur, 61 rue de Bruxelles, Namur 5000 (Belgium)

    2015-03-18

    Background: High-LET ion irradiation is being more and more often used to control tumors in patients. Given that tumors are now considered as complex organs composed of multiple cell types that can influence radiosensitivity, we investigated the effects of proton and alpha particle irradiation on the possible radioprotective cross-talk between cancer and endothelial cells. Materials and Methods: We designed new irradiation chambers that allow co-culture study of cells irradiated with a particle beam. A549 lung carcinoma cells and endothelial cells (EC) were exposed to 1.5 Gy of proton beam or 1 and 2 Gy of alpha particles. Cell responses were studied by clonogenic assays and cell cycle was analyzed by flow cytometry. Gene expression studies were performed using Taqman low density array and by RT-qPCR. Results: A549 cells and EC displayed similar survival fraction and they had similar cell cycle distribution when irradiated alone or in co-culture. Both types of irradiation induced the overexpression of genes involved in cell growth, inflammation and angiogenesis. Conclusions: We set up new irradiation chamber in which two cell types were irradiated together with a particle beam. We could not show that tumor cells and endothelial cells were able to protect each other from particle irradiation. Gene expression changes were observed after particle irradiation that could suggest a possible radioprotective inter-cellular communication between the two cell types but further investigations are needed to confirm these results.

  17. Effect of Cone Beam CT on the Cell-cycle and Adaptive Response of A-549 Cells%CBCT对A-549肿瘤细胞周期变化及适应性反应的影响

    Institute of Scientific and Technical Information of China (English)

    李文艺; 金雏凤; 曹瑞芬; 裴曦; 吴宜灿; FDS团队

    2013-01-01

    目的:研究临床图像引导放射治疗模式下锥形束CT(Cone Beam CT,CBCT)对肿瘤细胞A-549的细胞周期影响,并观察CBCT辐照后采用大剂量照射是否引起肿瘤细胞适应性反应.方法:将体外培养的A-549细胞分为对照组(不照射)、CBCT组(行CBCT照射)、2 Gy组(单纯行2 Gy照射)和CBCT+2 Gy组(CBCT后30min行2 Gy照射),各组照射后经过6h、12h、24h孵育,收集细胞,流式细胞仪检测细胞周期.结果:与对照组相比,CBCT组未发现A-549细胞周期的统计学变化(P>0.05);CBCT+2 Gy组与对照组比较出现了G1期阻滞;CBCT组与2 Gy组比较,未出现A-549肿瘤细胞适应性反应(P>0.05).结论:CBCT剂量不会引起A-549细胞的周期改变,并且临床治疗模式下CBCT剂量不会引起A-549肿瘤细胞适应性反应,从而不会降低对该肿瘤细胞的杀灭作用.

  18. Molecular analysis of hepatitis E virus from farm rabbits in Inner Mongolia, China and its successful propagation in A549 and PLC/PRF/5 cells.

    Science.gov (United States)

    Jirintai, Suljid; Jinshan; Tanggis; Manglai, Dugarjavin; Mulyanto; Takahashi, Masaharu; Nagashima, Shigeo; Kobayashi, Tominari; Nishizawa, Tsutomu; Okamoto, Hiroaki

    2012-12-01

    Rabbit hepatitis E virus (HEV) strains have recently been isolated in several areas of China and in the US and France. However, the host range, distribution and zoonotic potential of these HEV strains remain unknown and their propagation in cultured cells has not yet been reported. A total of 211 4-month-old rabbits raised on a farm in Inner Mongolia were tested for the presence of anti-HEV antibodies and HEV RNA. Overall, 121 rabbits (57.3%) tested positive for anti-HEV antibodies, and 151 (71.6%) had detectable HEV RNA. The 174 HEV strains recovered from these viremic rabbits, including two distinct strains each from 23 rabbits, differed from each other by up to 13.6% in a 412-nucleotide (nt) sequence within ORF2, and were 89.3-95.9% identical to the reported rabbit HEV strains in other provinces of China. Three representative Inner Mongolian strains, one each from three phylogenetic clusters, whose entire genomic sequences were determined, shared 79.6-96.7% identities with reported rabbit HEV strains within the entire or 242- to 1349-nt partial genomic sequence. Rabbit HEV strains recovered from liver tissues of rabbits with a high HEV load propagated efficiently in human cell lines (A549 and PLC/PRF/5 cells), suggesting the potential zoonotic risk of rabbit HEV.

  19. 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

  20. Inactivation of A549 cancer cells by a helium-oxygen vlasma needle%氦-氧等离子体针灭活肺癌A549细胞

    Institute of Scientific and Technical Information of China (English)

    陈维; 黄骏; 李辉; 吕国华; 王兴权; 张国平; 王鹏业; 杨思泽

    2012-01-01

    研究了介质阻挡放电等离子体针对肺癌A549细胞的灭活机制,探讨从不锈钢管注入到氦等离子体尾流区域的氧气含量对杀灭肺癌细胞A549的影响.利用中性红吸收测试法定性观察了等离子体处理后死亡的细胞和活着的细胞的形态区别,并且定量测定了不同条件下的细胞存活率.在固定功率24w的处理过程中,氦一氧等离子体的灭活效率主要取决于等离子体曝光时间以及氦气中添加氧气的百分含量.实验结果显示最好的处理参数为:处理时间150S,800mL/min的氦气添加3%氧气,保持针距样品的距离为3nlnl.根据氦.氧等离子体发射光谱,可以推断在细胞灭活过程中,氦.氧等离子体中的活性粒子(如羟基和氧自由基)起主要作用.%An inactivation mechanism of A549 cancer cells is studied by using a dielectric barrier discharge (DBD) plasma needle. The influence of oxygen concentration, which is injected into helium plasma afterglow region through a stainless steel tube, is investigated. The neutral red uptake assay provides a qualitative observation of morphological differences between the dead cells and the viable ceils after plasma treatment and a quantitative estimation of cell viability under different conditions. In the treatment process at a fixed power of 24 W, the inactivation efficiency of helium-oxygen plasma depends mainly on the exposure time and percentage of added oxygen in helium plasma. Experimental results show that the best parameters of the process are 150 s treatment time, 800 mL/min He with 3% Oz addition and separation of needle-to-sample 3 mm. According to the helium-oxygen emission spectra of the plasma jet, it is concluded that the reactive species (for example, OH and O) in the helium-oxygen plasma play a major role in the cell deactivation.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  2. 大麻受体激动剂对肺癌A549细胞凋亡和增殖的影响%Effect of Cannabinoid Receptor Activation by THC on Proliferation and Apoptosis of Lung Cancer A549 Cells

    Institute of Scientific and Technical Information of China (English)

    朱晓琴; 胡景鑫; 周于婷; 白红波; 赵青

    2011-01-01

    目的 研究大麻受体激动剂(delta9-tetrahydrocannabinol,THC)对肺癌A549细胞凋亡和增殖的影响.方法 MTT法测定THC对A549细胞增殖的影响;苏木精-伊红染色、扫描电镜观察细胞的形态学变化;Western blot 法分析大麻受体CB1、CB2的蛋白表达;DNA梯度电泳检测A549细胞凋亡;流式细胞仪分析细胞凋亡率变化.结果 THC预处理后,MTT检测表明THC对A549细胞增殖有明显抑制作用,随着药物浓度增大,抑制作用更加明显;苏木精-伊红染色、扫描电镜观察显示:肺癌A549细胞有典型的细胞凋亡形态;Western blot检测显示:A549细胞大麻受体CB1、CB2水平较正常气道上皮细胞株16HBE升高;DNA梯度电泳法及流式细胞仪检测显示:THC能抑制A549细胞生长,诱导其凋亡,并具有剂量依赖性.结论 大麻受体激动剂THC能抑制肺癌细胞的增殖,并诱导肺癌细胞凋亡,此效应可能与大麻受体CB1、CB2作用有关.%Objective To study the effect of the cannabinoid receptor activation by THC on the proliferation and apoptosis of lung cancer A549 cells- Methods The effects of THC on proliferation of A549 cells were measured by using MTT assay,and morphological changes of A549 cells after HE staining were observed under an electron microscopy. Protein expression of can nabinoid receptors CB1 and CB2 was detected by using Western blot. Apoptosis of A549 cells was examined by using DNA gra dient gel electrophoresis,and the change of apoptosis rate was analyzed by using flow cytometry. Results After pretreatment with THC,MTT assay revealed that THC could significantly suppress proliferation of A549 cells in a dose dependent man ner. HE staining and electron microscopy displayed that A549 cells had the typical apoptotic morphology. Western blot showed that cannabinoid receptors CB1 and CB2 were increased A549 cells as compared with those in normal airway epithelial cells 16HBE. DNA gradient electrophoresis and flow cytometry demonstrated

  3. Iron release and ROS generation from mineral particles are not related to cytokine release or apoptosis in exposed A549 cells.

    Science.gov (United States)

    Ovrevik, J; Hetland, R B; Schins, R P; Myran, T; Schwarze, P E

    2006-08-01

    The generation of reactive oxygen species (ROS) by mineral particles is believed to be central to their toxicity and their ability to induce inflammation. Surface bound or soluble iron may contribute to the particle-effects by enhancing the ROS generation through the Fenton reaction. Nevertheless, the importance of ROS and transition metals to mineral particle-induced effects is still unclear and further investigations are needed. In the present study we have investigated different mineral particles for their total iron content, amount of soluble iron at pH 7.0 and 4.0, their ability to generate ROS in a cell-free environment, and their ability to induce cytokine release and apoptosis in a human alveolar epithelial cell line (A549). All the investigated parameters varied considerably between the different particles, with the exception of ability to induce apoptosis. Total iron content did not reflect the amount of soluble iron, and neither total nor soluble iron was correlated with ROS generation. Moreover, iron content and ROS was not correlated with the ability of particles to induce cytokine release or apoptosis. The present results suggest that there is no clear relationship between the particles iron content and ability to generate ROS. Moreover, neither iron content nor the ability to induce ROS generation appears to be a prerequisite for the inflammatory potential or cytotoxicity of mineral particles.

  4. 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.

  5. 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.

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

    Energy Technology Data Exchange (ETDEWEB)

    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.

  7. 卷烟烟气暴露下A549和BEAS-2B细胞促炎性因子的释放变化%Release of proinflammatory cytokines from A549 cells and BEAS-2B cells exposed to mainstream cigarette smoke

    Institute of Scientific and Technical Information of China (English)

    李翔; 张世敏; 赵俊伟; 尚平平; 彭斌; 谢复炜; 刘惠民; 谢剑平

    2016-01-01

    为比较不同类型细胞对卷烟烟气诱导的促炎症反应的应答差异,采用中性红细胞毒性试验测试了卷烟烟气总粒相物(TPM)和全烟气(WS)对人肺腺癌细胞A549和人支气管上皮细胞BEAS-2B的细胞毒性效应;采用酶联免疫法(ELISA)分析了TPM和WS暴露后,A549和BEAS-2B促炎性细胞因子白细胞介素-6(IL-6)和白细胞介素-8(IL-8)的释放水平。结果表明:BEAS-2B细胞较A549细胞对卷烟烟气的细胞毒性效应敏感。卷烟烟气暴露下A549细胞没有增加促炎性细胞因子IL-6和IL-8的释放;而BEAS-2B细胞在TPM和WS暴露下其IL-6和IL-8的释放显著增加。卷烟烟气诱导的促炎症反应存在细胞类型的差异。%In order to compare the differences of response to proinflammatory reaction induced by mainstream cigarette smoke among cells of different types, neutral red uptake (NRU) assay was employed to assess the cytotoxicity of cigarette smoke total particulate matter (TPM) or whole smoke (WS) to A549 cells (human adenocarcinoma cells) and BEAS-2B cells (human bronchial epithelial cells). The levels of proinflammatory cytokines interleukin-6 and interleukin-8 (IL-6 and IL-8) released from A549 and BEAS-2B cells exposed to TPM and WS were analyzed by ELISA. The results showed that BEAS-2B cells were more sensitive to smoke-induced cytotoxicity than A549 cells. The induction of IL-6 and IL-8 released from A549 cells was not impacted by cigarette smoke exposure, while TPM and WS all caused dose-dependently increase of IL-6 and IL-8 released from BEAS-2B cells. The proinflammatory response to cigarette smoke thus might be different depending on cell types.

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

    Directory of Open Access Journals (Sweden)

    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.

  9. Atrial natriuretic peptide: A novel mediator for TGF-β1-induced epithelial-mesenchymal transition in 16HBE-14o and A549 cells.

    Science.gov (United States)

    Chu, Shuyuan; Zhang, Xiufeng; Sun, Yabing; Yu, Yuanyuan; Liang, Yaxi; Jiang, Ming; Huang, Jianwei; Ma, Libing

    2017-02-13

    Atrial natriuretic peptide (ANP) is increasingly expressed on airway and inhibits pulmonary arterial remodeling. However, the role of ANP in remodeling of respiratory system is still unclear. The role of ANP on airway remodeling and the possible mechanism was explored in this study. Both human bronchial epithelial 16HBE-14o cells and alveolar epithelial A549 cells were stimulated by TGF-β1, ANP, cGMP inhibitor, PKG inhibitor, and cGMP analogue. The expressions of epithelial markers, mesenchymal markers, and Smad3 were assessed by quantitative real-time PCR and western blotting. Immunohistochemical staining was employed to assess Smad3 expression once it was silenced by siRNA in 16HBE-14o or A549 cells. Our results showed that the mRNA and protein expressions of E-Cadherin were decreased, whereas α-SMA expressions were increased after induction by TGF-β1 in 16HBE-14o and A549 cells. The E-Cadherin expressions were increased and α-SMA expressions were decreased after ANP stimulation. Inhibition of cGMP or PKG decreased E-Cadherin expression but increased α-SMA expression, which could be reversed by cGMP analogue. Moreover, the phosphorylated Smad3 expression was consistent with α-SMA expression. After smad3 was silenced, Smad3 was mostly expressed in cytoplasm instead of nucleus as non-silenced cells during epithelial-mesenchymal transition (EMT). In conclusion, ANP inhibits TGF-β1-induced EMT in 16HBE-14o and A549 cells through cGMP/PKG signaling, by which it targets TGF-β1/Smad3 via attenuating phosphorylation of Smad3. These findings suggest the potential of ANP in the treatment on pulmonary diseases with airway remodeling.

  10. 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.

  11. Study of the Effects of Betaine and/or C-Phycocyanin on the Growth of Lung Cancer A549 Cells In Vitro and In Vivo

    Directory of Open Access Journals (Sweden)

    Rea Bingula

    2016-01-01

    Full Text Available We investigated the effects of betaine, C-phycocyanin (C-PC, and their combined use on the growth of A549 lung cancer both in vitro and in vivo. When cells were coincubated with betaine and C-PC, an up to 60% decrease in viability was observed which is significant compared to betaine (50% or C-PC treatment alone (no decrease. Combined treatment reduced the stimulation of NF-κB expression by TNF-α and increased the amount of the proapoptotic p38 MAPK. Interestingly, combined treatment induced a cell cycle arrest in G2/M phase for ~60% of cells. In vivo studies were performed in pathogen-free male nude rats injected with A549 cells in their right flank. Their daily food was supplemented with either betaine, C-PC, both, or neither. Compared to the control group, tumour weights and volumes were significantly reduced in either betaine- or C-PC-treated groups and no additional decrease was obtained with the combined treatment. This data indicates that C-PC and betaine alone may efficiently inhibit tumour growth in rats. The synergistic activity of betaine and C-PC on A549 cells growth observed in vitro remains to be further confirmed in vivo. The reason behind the nature of their interaction is yet to be sought.

  12. 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.

  13. 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.

  14. miRNA-126靶向抑制A549细胞迁移和侵袭能力的研究%miR-126 may inhibit A549 cell migration and invasion by targeting EGFL7

    Institute of Scientific and Technical Information of China (English)

    孙艳芹; 刘建强

    2013-01-01

    目的 研究microRNA-126(miR-126)对非小细胞肺癌(NSCLC)细胞迁移和侵袭能力的影响. 方法 利用脂质体介导法将构建的miR 126过表达质粒转染至NSCLC细胞株A549细胞(A549/ miR-126组),并设空质粒转染组(A549/MOCK组)和空白对照组(A549组).利用RT-PCR技术检测3组细胞中EGFL7(miR-126的靶标)的表达水平,采用细胞划痕实验观察细胞迁移能力差异,采用Transwell小室法分析3组细胞侵袭能力的差异. 结果 RT-PCR检测A549/miR-126组、A549/MOCK组和A549组细胞中EGFL7 mRNA分别为2.32±0.088、1.43±0.026和1.00±0.000,差异有统计学意义(P<0.01);细胞划痕实验显示A549/ miR-126组、A549/MOCK组和A549组细胞平均迁移距离分别为3.0 μm、2.65 μm和0.5 μm,平均抑制率分别为0、11.25%和83.75%,差异有统计学意义(P<0.05);Transwell小室实验显示,A549/miR-126组24 hr侵袭细胞数为(28.6±2.322)个,36 h侵袭细胞数为(29.2±3.7683)个,A549/MOCK组24 h为(49.8±3.7014)个,差异有统计学意义(P<0.05). 结论 miR-126可上调EGFL7 mRNA的表达,并可能通过表达产物EGFL7蛋白抑制A549细胞的迁移和侵袭能力.

  15. 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.

  16. Marsdenia tenacissima extract suppresses A549 cell migration through regulation of CCR5-CCL5 axis, Rho C, and phosphorylated FAK.

    Science.gov (United States)

    Lin, Sen-Sen; Li, Fang-Fang; Sun, Li; Fan, Wei; Gu, Ming; Zhang, Lu-Yong; Qin, Song; Yuan, Sheng-Tao

    2016-03-01

    Marsdenia tenacissima, a traditional Chinese medicine, is long been used to treat various diseases including asthma, cancer, trachitis, tonsillitis, pharyngitis, cystitis, and pneumonia. Although Marsdenia tenacissima has been demonstrated to have strong anti-tumor effects against primary tumors, its effect on cancer metastasis remains to be defined, and the molecular mechanism underlying the anti-metastatic effect is unknown. In the present study, we investigated the effects of XAP (an extract of Marsdenia tenacissima) on A549 lung cancer cell migration and explored the role of CCR5-CCL5 axis in the anti-metastatic effects of XAP. Our resutls showed that XAP inhibited A549 lung cancer cell migration and invasion in a dose-dependent manner. The protein levels of CCR5, but not CCR9 and CXCR4, were decreased by XAP. The secretion of CCL5, the ligand of CCR5, was reduced by XAP. XAP down-regulated Rho C expression and FAK phosphorylation. In conclusion, XAP inhibited A549 cell migration and invasion through down-regulation of CCR5-CCL5 axis, Rho C, and FAK.

  17. 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.

  18. Genistein enhances the effect of trichostatin A on inhibition of A549 cell growth by increasing expression of TNF receptor-1

    Energy Technology Data Exchange (ETDEWEB)

    Wu, Tzu-Chin [Chest Clinic, Chung Shan Medical University Hospital, Taichung, Taiwan, ROC (China); Yang, Ying-Chihi; Huang, Pei-Ru [Department of Nutritional Science, Chung Shan Medical University, Taichung, Taiwan, ROC (China); Wen, Yu-Der [Department of Biology, National Changhua University of Education, Changhua, Taiwan, ROC (China); Yeh, Shu-Lan, E-mail: suzyyeh@csmu.edu.tw [Department of Nutritional Science, Chung Shan Medical University, Taichung, Taiwan, ROC (China); Department of Nutrition, Chung Shan Medical University Hospital, Taichung, Taiwan, ROC (China)

    2012-08-01

    Our previous study has shown that genistein enhances apoptosis in A549 lung cancer cells induced by trichostatin A (TSA). The precise molecular mechanism underlying the effect of genistein, however, remains unclear. In the present study, we investigated whether genistein enhances the anti-cancer effect of TSA through up-regulation of TNF receptor-1 (TNFR-1) death receptor signaling. We incubated A549 cells with TSA (50 ng/mL) alone or in combination with genistein and then determined the mRNA and protein expression of TNFR-1 as well as the activation of downstream caspases. Genistein at 5 and 10 μM significantly enhanced the TSA-induced decrease in cell number and apoptosis in a dose-dependent manner. The combined treatment significantly increased mRNA and protein expression of TNFR-1 at 6 and 12 h, respectively, compared with that of the control group; while TSA alone had no effect. TSA in combination with 10 μM of genistein increased TNFR-1 mRNA and protein expression by about 70% and 40%, respectively. The underlying mechanism for this effect of genistein may be partly associated with the estrogen receptor pathway. The combined treatment also increased the activation of caspase-3 and ‐10 as well as p53 protein expression in A549 cells. The enhancing effects of genistein on the TSA-induced decrease in cell number and on the expression of caspase-3 in A549 cells were suppressed by silencing TNFR-1 expression. These data demonstrated that the upregulation of TNFR-1 death receptor signaling plays an important role, at least in part, in the enhancing effect of genistein on TSA-induced apoptosis in A549 cells. -- Highlights: ► TSA combined with genistein rather than TSA alone increases the expression of TNFR-1. ► Genistein may exert such an effect partly through estrogen receptor pathway. ► The combined treatment increases the activation of caspase-10 and caspase-3. ► The combined treatment also increases the expression of p53 protein. ► TNFR-1 si

  19. 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.

    Science.gov (United States)

    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.

  20. Meloxicam increases intracellular accumulation of doxorubicin via downregulation of multidrug resistance-associated protein 1 (MRP1) in A549 cells.

    Science.gov (United States)

    Chen, S F; Zhang, Z Y; Zhang, J L

    2015-11-19

    It has been suggested that selected COX inhibitors can overcome multidrug resistance through the inhibition of ATP‑binding cassette-transporter proteins thereby enhancing the inhibitory effect of doxorubicin on human tumor growth and promoting the actions of cytostatics. However, their effect on lung cancer and the molecular mechanisms involved in the overcoming of multidrug resistance are unclear. In the present study, the ability of meloxicam, a COX-2-specific inhibitor to enhance doxorubicin‑mediated inhibition was investigated in human A549 lung cancer in vivo and in vitro. In order to unravel the molecular mechanisms involved in doxorubicin accumulation, we measured the levels of multidrug resistance-associated protein (MRP)-transporter protein activity and expression by western blotting, since this has been implicated in meloxicam action as well as in chemoresistance. We found that, in A549 cells, meloxicam could increase intracellular accumulation of doxorubicin, a substrate for MRP, through inhibition of cellular export. Western blot analysis indicated that meloxicam reduced the expression of MRP1 and MRP4. The results reported in the present study demonstrate for the first time that the specific COX-2 inhibitor meloxicam can increase the intracellular accumulation of doxorubicin and enhance doxorubicin-induced cytotoxicity in A549 cancer cells by reducing the expression of MRP1 and MRP4.

  1. Genome-wide transcriptional analysis of apoptosis-related genes and pathways regulated by H2AX in lung cancer A549 cells.

    Science.gov (United States)

    Lu, Chengrong; Xiong, Min; Luo, Yuan; Li, Jing; Zhang, Yanjun; Dong, Yaqiong; Zhu, Yanjun; Niu, Tianhui; Wang, Zhe; Duan, Lianning

    2013-09-01

    Histone H2AX is a novel tumor suppressor protein and plays an important role in apoptosis of cancer cells. However, the role of H2AX in lung cancer cells is unclear. The detailed mechanism and epigenetic regulation by H2AX remain elusive in cancer cells. We showed that H2AX was involved in apoptosis of lung cancer A549 cells as in other tumor cells. Knockdown of H2AX strongly suppressed apoptosis of A549 cells. We clarified the molecular mechanisms of apoptosis regulated by H2AX based on genome-wide transcriptional analysis. Microarray data analysis demonstrated that H2AX knockdown in A549 cells affected expression of 3,461 genes, including upregulation of 1,435 and downregulation of 2,026. These differentially expressed genes were subjected to bioinformatic analysis for exploring biological processes regulated by H2AX in lung cancer cells. Gene ontology analysis showed that H2AX affected expression of many genes, through which, many important functions including response to stimuli, gene expression, and apoptosis were involved in apoptotic regulation of lung cancer cells. Pathway analysis identified the mitogen-activated protein kinase signaling pathway and apoptosis as the most important pathways targeted by H2AX. Signal transduction pathway networks analysis and chromatin immunoprecipitation assay showed that two core genes, NFKB1 and JUN, were involved in apoptosis regulated by H2AX in lung cancer cells. Taken together, these data provide compelling clues for further exploration of H2AX function in cancer cells.

  2. New geranylated flavanones from the fruits of Paulownia catalpifolia Gong Tong with their anti-proliferative activity on lung cancer cells A549.

    Science.gov (United States)

    Gao, Tian-yang; Jin, Xing; Tang, Wen-zhao; Wang, Xiao-jing; Zhao, Yun-xue

    2015-09-01

    Three new geranylated flavanones, named as paucatalinone A (1), B (2), and isopaucatalinone B (3), were isolated from the fruits of Paulownia catalpifolia Gong Tong (Scrophulariaceae). Their structures were well determined by means of IR, MS, 1D and 2D NMR, and CD techniques. Paucatalinone A (1) is the first sample as a dimeric geranylated flavanone derivative isolated from natural products. Paucatalinone A (1) displayed good antiproliferative effects on human lung cancer cells A549 and resulted in a clear increase of the percentage of cells in G1 phase and a decrease in the percentage of cells in S and G2/M phases in comparison with control cells.

  3. Inhibitory effect of new copper (Ⅱ) complex with coumarin derivatives on lung cancer cells A549 in vivo and vitro%新型香豆素类酰腙-铜配合物对肺腺癌A549细胞的体内外抑制作用

    Institute of Scientific and Technical Information of China (English)

    陆勤; 欧秋霞; 朱文娇; 朱涛峰

    2015-01-01

    目的:观察新型香豆素类酰腙—铜配合物(以下缩写为CCCD)在体内外对肺癌A549细胞的抑制作用,并探讨其机制。方法培养肺癌A549细胞,分别加入5、10、20、30、50、80、120、160μmol/L的CCCD,干预72 h后,采用MTT法,计算细胞生长抑制率( IR )。将A549细胞分为干预组、对照组,干预组分别加入10、20、40μmol/L CCCD,对照组加入PBS,采用流式细胞术检测各组细胞凋亡情况并计算细胞凋亡率,采用Western blot法检测各组细胞Caspase-3蛋白表达。取18只裸鼠建立肺癌荷瘤鼠模型,分为观察1组、观察2组、对照组,每组各6只,分别予尾静脉注射4、8 mg/kg CCCD及PBS,1次/周,共干预3周,干预结束后测算各组肿瘤体积并计算抑瘤率。随后处死各组裸鼠,取瘤体组织,应用TUNEL法检测各组肿瘤细胞凋亡情况并计算凋亡指数( AD)。结果加入5、10、20、30、50、80、120、160μmol/L CCCD 后, A549细胞 IR 分别为8.80%、16.52%、37.24%、55.75%、77.22%、87.16%、95.25%、98.70%,随着药物浓度增高,IR呈增高趋势。干预组加入10、20、40μmol/L CCCD后,细胞凋亡率均高于对照组(P均<0.05)。干预组Caspase-3蛋白表达高于对照组(P<0.05)。观察1组、观察2组、对照组AD分别为16.83%±8.44%、24.65%±11.24%、3.30%±2.12%,各组间比较P均<0.05。观察1组、观察2组抑瘤率分别为51.08%、56.78%。结论 CCCD在体内外均可抑制肺癌A549细胞的生长,促进细胞凋亡,其作用机制可能与经Caspase-3途径诱导细胞凋亡有关。%Objective To observe the inhibitory effect of a new copper (Ⅱ) complex with coumarin derivatives ( CCCD) on lung cancer cell line A549 in vivo and in vitro and to investigate the mechanism .Methods The lung cancer A549 cells were cultured and were treated with 5, 10

  4. 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.

  5. 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.

  6. Effect of 3-bromopyruvate combined with cisplatin on inhibiting growth of lung cancer A549 cells%3-溴丙酮酸联合顺铂抑制肺癌细胞株A549的生长

    Institute of Scientific and Technical Information of China (English)

    张梦娇; 张明; 胡义德

    2016-01-01

    目的 探讨3-溴丙酮酸(3-bromopyruvate,3-BrPA)联合顺铂体外抗肺癌A549细胞的作用及其可能机制.方法 采用CCK-8检测不同浓度的3-BrPA、顺铂单用及联用对A549细胞的增殖抑制;选择低于半数抑制浓度(IC50)的40 μmol/L 3-BrPA与1 mg/L顺铂单独和联合作用A549细胞48 h后,用倒置相差显微镜观察细胞形态变化,流式细胞术检测细胞凋亡;不同浓度的3-BrPA作用A549细胞48 h后,流式细胞术检测细胞周期变化,己糖激酶(hexokinase,HK)活性检测试剂盒检测3-BrPA对细胞内HK活性的影响.结果 CCK-8结果显示,与阴性对照组比较,3-BrPA浓度大于20 μmol/L时对A549细胞有明显抑制作用(P<0.01),与单用顺铂组比较,3-BrPA联合顺铂组可显著增强顺铂对A549细胞的毒性作用(P<0.01);3-BrPA作用A549细胞48 h后,倒置显微镜观察到3-BrPA组大多数细胞出现凋亡,而联合用药组细胞凋亡更加明显,部分区域出现细胞碎片等坏死征象;流式细胞术结果显示,3-BrPA和顺铂单用组及联合用药组细胞凋亡率均明显高于阴性对照组(P<0.01),且联合用药组细胞凋亡率明显高于单独用药组(P<0.01);细胞周期检测结果显示3-BrPA可将A549细胞阻滞于G1期;HK活性结果显示:与阴性对照组比较,3-BrPA组HK活性明显降低(P<0.01).结论 3-BrPA有抑制肺癌A549细胞增殖、诱导其凋亡的作用,且与顺铂具有协同抗肺癌A549细胞增殖作用,其机制可能为抑制肺癌细胞糖酵解,进而影响肺癌细胞能量代谢.

  7. Antisense Oligonucleotides targeting protein kinase Cα inhibits the proliferation of A549 cells%PKCα反义核酸对肺癌A549细胞增殖的影响

    Institute of Scientific and Technical Information of China (English)

    严玉霞; 蒋建伟; 黄志宏; 吴志慧; 林春兰; 吴风云

    2009-01-01

    目的 探讨聚乙烯亚胺(PEI)介导的人蛋白激酶Cα(PKCα)基因反义寡核苷酸(ASODN)对人肺癌A549细胞生长增殖及PKCα基因表达的影响.方法 以PEI介导PKCα ASODN,随机寡核苷酸(RODN)分别转染A549细胞,采用WST法和克隆形成抑制实验分析细胞生长增殖抑制效果;RT-PCR检测PKcd mRNA的表达水平;Western blotting检测PKCα蛋白的表达水平.结果 PEI介导的PKCaASODN转染A549细胞后,细胞牛长增殖和克隆形成均受到抑制(P<0.05),并呈浓度依赖关系,其抑制率明显高于PEI或PEI介导的RODN组;RT-PCR和Westernblotting结果均表明:PEI 介导的PKCαASODN转染A549细胞能明显抑制PKCα的表达,与空白对照组、PEI或PEI介导的RODN转染组相比,有统计学差异(P<0.05).结论 PEI介导的PKCα反义核酸能下调PKCα基因的表达,显著抑制A549细胞生长和增殖.

  8. Noxa基因的克隆及其诱导A549细胞凋亡的研究%Apoptosis of A549 Cells Induced by Cloned Noxa Gene

    Institute of Scientific and Technical Information of China (English)

    陈登榜; 曹康; 杨靖; 潘渠; 陈恬; 李晋川; 王昕; 殷建华; 邹毅

    2013-01-01

    目的 克隆人Noxa基因,构建真核表达质粒pcDN A-Noxa,将重组质粒pcDN A-Noxa转染人肺癌A549细胞株,并观察重组质粒pcDN A-Noxa诱导A549细胞凋亡的作用.方法 使用RT-PCR的方法扩增出Noxa基因,克隆至真核表达质粒pcDNA(一),构建出重组质粒pcDNA-Noxa.将重组质粒pcDN A-Noxa转染人肺癌A549细胞株,使用Western blot的方法检测Noxa基因的过表达;使用Hoechst 33258染色,观察A549细胞的凋亡;使用MTT法检测细胞活力.结果 经酶切和测序鉴定,重组质粒pcDN A-Noxa构建成功.Western blot检测出Noxa基因的过表达;Hoechst 33258染色和MTT实验发现:重组质粒pcDNA-Noxa能明显诱导A549细胞出现凋亡现象.结论 本实验初步探索了Noxa基因的促凋亡作用,为以后进一步深入研究Noxa基因的促凋亡功能基础.

  9. Inhibition of mitogen activated protein kinases increases the sensitivity of A549 lung cancer cells to the cytotoxicity induced by a kava chalcone analog.

    Science.gov (United States)

    Warmka, Janel K; Solberg, Eric L; Zeliadt, Nicholette A; Srinivasan, Balasubramanian; Charlson, Aaron T; Xing, Chengguo; Wattenberg, Elizabeth V

    2012-08-03

    We are interested in investigating the biological activity of chalcones, a major class of compounds found in the beverage kava, in order to develop potent and selective chemopreventive candidates. Consumption of kava in the South Pacific Islands is inversely correlated with cancer incidence, even among smokers. Accordingly, chalcones have anti-cancer activities in animal and cell culture models. To investigate signaling pathways that affect chalcone action we studied a potent analog, (E)-3-(3-hydroxy-4-methoxyphenyl)-1-(3,4,5-trimethoxyphenyl)prop-2-en-1-one (chalcone-24). Chalcone-24 was selected from a series of chalcone analogs that were synthesized based on the structures derived from flavokawain compounds found in kava, and screened in A549 lung cancer cells for induction of cytotoxicity and inhibition of NF-κB, a transcription factor associated with cell survival. Incubation of A549 cells with chalcone-24 resulted in a dose-dependent inhibition of cell viability, inhibition of NF-κB, activation of caspases, and activation of extracellular signal regulated kinase 1/2 (ERK1/2) and c-Jun N-terminal kinase (JNK); ERK1/2 and JNK are mitogen activated protein kinases that play central roles in regulating cell fate. Pharmacological inhibitors of ERK1/2 or JNK increased the sensitivity of A549 cells to chalcone-24-induced cytotoxicity, without affecting NF-κB or caspase activity. These results will help refine the synthesis of chalcone analogs to maximize the combination of actions required to prevent and treat cancer.

  10. Evaluation of 5-HT7 Receptor Trafficking on In Vivo and In Vitro Model of Lipopolysaccharide (LPS)-Induced Inflammatory Cell Injury in Rats and LPS-Treated A549 Cells.

    Science.gov (United States)

    Ayaz, Gulsen; Halici, Zekai; Albayrak, Abdulmecit; Karakus, Emre; Cadirci, Elif

    2017-02-01

    This study aimed to investigate the effects of the 5-HT7 receptor agonist (LP44) and antagonist (SB269970) on LPS-induced in vivo tissue damage and cell culture by molecular methods. This study was conducted in two steps. For in vivo studies, 24 female rats were divided into four groups. Group I: healthy; II (2nd h): LPS 5 mg/kg administered intraperitoneally (i.p.); III (4th h): LPS 5 mg/kg administered i.p.; IV (8th h): LPS 5 mg/kg administered i.p. For in vitro studies, we used the A549 cell line. Groups: I control (healthy) (2-4 h); II LPS: 1 µg/ml E. Coli O55:B5 strain (2-4 h); III agonist (LP44) 10(-9) M (2-4 h); IV antagonist (SB269970) 10(-9) M (2-4 h); V LPS+agonist 10(-9) M (LP44 1 µg/ml) (2-4 h); VI LPS+antagonist 10(-9) M (2-4 h). In molecular analyses, we determined increased TNF-α, IL-1β, NF-κB, and 5-HT7 mRNA expressions in rat lung tissues and increased TNF-α, iNOS, and 5-HT7 mRNA expressions in the A549 cell line. In in vitro parameters, LP44 agonist administration-related decrease was observed. Our study showed that lung 5-HT7 receptor expression is increased in LPS-induced endotoxemia. All this data suggest that 5-HT7 receptor overexpression is an important protective mechanism during LPS-induced sepsis-related cell damage.

  11. TGF-β and Hypoxia/Reoxygenation Promote Radioresistance of A549 Lung Cancer Cells through Activation of Nrf2 and EGFR

    Directory of Open Access Journals (Sweden)

    Sae-lo-oom Lee

    2016-01-01

    Full Text Available Although many studies have examined the roles of hypoxia and transforming growth factor- (TGF- β separately in the tumor microenvironment, the effects of simultaneous treatment with hypoxia/reoxygenation and TGF-β on tumor malignancy are unclear. Here, we investigated the effects of redox signaling and oncogenes on cell proliferation and radioresistance in A549 human lung cancer cells in the presence of TGF-β under hypoxia/reoxygenation conditions. Combined treatment with TGF-β and hypoxia activated epidermal growth factor receptor (EGFR and nuclear factor (erythroid-derived 2-like 2 (Nrf2, a redox-sensitive transcription factor. Interestingly, Nrf2 knockdown suppressed the effects of combined treatment on EGFR phosphorylation. In addition, blockade of EGFR signaling also suppressed induction of Nrf2 following combined treatment with hypoxia and TGF-β, indicating that the combined treatment induced positive crosstalk between Nrf2 and EGFR. TGF-β and hypoxia/reoxygenation increased the accumulation of reactive oxygen species (ROS, while treatment with N-acetyl-L-cysteine abolished the activation of Nrf2 and EGFR. Treatment with TGF-β under hypoxic conditions increased the proliferation of A549 cells compared with that after vehicle treatment. Moreover, cells treated with the combined treatment exhibited resistance to ionizing radiation (IR, and knockdown of Nrf2 increased IR-induced cell death under these conditions. Thus, taken together, our findings suggested that TGF-β and hypoxia/reoxygenation promoted tumor progression and radioresistance of A549 cells through ROS-mediated activation of Nrf2 and EGFR.

  12. Expressions and Significances of PRL-3 and RhoC in A549 Cell%PRL-3和RhoC在A549细胞中的表达及意义

    Institute of Scientific and Technical Information of China (English)

    张平; 张志培; 李香敏; 雷杰; 苏凯; 李小飞; 王小平

    2010-01-01

    背景与目的 在前期研究中,我们发现在非小细胞肺癌中PRL-3和RhoC的表达具有相关性,提示两者可能存在相互作用.本研究通过检测两者在A549细胞迁移中的作用以及是否存在相互作用,为进一步研究它们在肿瘤发生发展中的作用机理提供实验依据.方法 应用PRL-3抗体、RhoC抗体分别阻断A549细胞中PRL-3和RhoC的功能,采用细胞划痕实验检测两者对其迁移能力的影响;应用RT-PCR检测PRL-3和RhoC表达的变化.结果 PRL-3和RhoC功能被阻断后,A549细胞迁移能力明显降低;阻断PRL-3的A549细胞中 RhoC表达降低,然而阻断RhoC的A549细胞中 PRL-3表达无明显改变.结论 PRL-3和RhoC 可能具有促进A549细胞远处转移的功能;PRL-3可能具有上调RhoC表达的功能;PRL-3促进癌细胞远处转移的功能可能是通过RhoC及其下游因子实现的,而RhoC对PRL-3的表达可能无反馈性调节作用.

  13. 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

  14. Impact of CHK2-small interfering RNA on CpG ODN7909-enhanced radiosensitivity in lung cancer A549 cells

    Directory of Open Access Journals (Sweden)

    Chen W

    2012-12-01

    Full Text Available Wei Chen,* Xiaoqun Liu,* Tiankui Qiao, Sujuan Yuan Department of Oncology, Jinshan Hospital, Fudan University, Shanghai, People's Republic of China*These authors contributed equally to this workObjective: To investigate the impact of checkpoint kinase 2 (CHK2-small interfering RNA (CHK2-siRNA on the enhancement of radiosensitivity by CpG oligodeoxynucleotide (ODN 7909 in lung cancer A549 cells.Methods: The A549 cells were randomly divided into five groups: control, CpG, X-ray, CpG + X-ray, and CHK2-siRNA + CpG + X-ray. Cell colonization was observed using inverted microscopy. Cell cycle and apoptosis were analyzed by flow cytometry. CHK2 expression was detected by Western blot. CHK2-siRNA was adopted to silence the expression of CHK2.Results: The level of CHK2 phosphorylation was higher in the CpG + X-ray group than in the X-ray group. Increases in G2/mitotic (M phase arrest and apoptosis and a decrease of cell survival rate in the CpG + X-ray group were statistically significant (P < 0.05 when compared with the CHK2-siRNA + CpG + X-ray group in which the expression of CHK2 was obviously inhibited. The combination of CpG ODN7909 and X-ray irradiation was found to enhance the mitotic death of A549 cells. The sensitization enhancement ratio of mean death dose (D0 was 1.42 in the CpG + X-ray group, which was higher than that of the CHK2-siRNA + CpG + X-ray group, in which D0 was 1.05.Conclusion: To a certain extent, the impact of a combination of CpG ODN7909 and X-ray on G2/M phase arrest, apoptosis, and rate of cell survival was attenuated by CHK2-siRNA in human lung adenocarcinoma A549 cells, indicating that increased phosphorylation of CHK2 might be a radiosensitive pathway.Keywords: oligodeoxynucleotide, checkpoint kinase 2, mitotic death, apoptosis, X-ray

  15. Rho GTPases RhoA and Rac1 mediate effects of dietary folate on metastatic potential of A549 cancer cells through the control of cofilin phosphorylation.

    Science.gov (United States)

    Oleinik, Natalia V; Helke, Kristi L; Kistner-Griffin, Emily; Krupenko, Natalia I; Krupenko, Sergey A

    2014-09-19

    Folate, an important nutrient in the human diet, has been implicated in cancer, but its role in metastasis is not established. We have shown previously that the withdrawal of medium folate leads to the inhibition of migration and invasion of A549 lung carcinoma cells. Here we have demonstrated that medium folate regulates the function of Rho GTPases by enabling their carboxyl methylation and translocation to plasma membrane. Conversely, the lack of folate leads to the retention of these proteins in endoplasmic reticulum. Folate also promoted the switch from inactive (GDP-bound) to active (GTP-bound) GTPases, resulting in the activation of downstream kinases p21-activated kinase and LIM kinase and phosphorylation of the actin-depolymerizing factor cofilin. We have further demonstrated that in A549 cells two GTPases, RhoA and Rac1, but not Cdc42, are immediate sensors of folate status: the siRNA silencing of RhoA or Rac1 blocked effects of folate on cofilin phosphorylation and cellular migration and invasion. The finding that folate modulates metastatic potential of cancer cells was confirmed in an animal model of lung cancer using tail vein injection of A549 cells in SCID mice. A folate-rich diet enhanced lung colonization and distant metastasis to lymph nodes and decreased overall survival (35 versus 63 days for mice on a folate-restricted diet). High folate also promoted epithelial-mesenchymal transition in cancer cells and experimental mouse tumors. Our study provides experimental evidence for a mechanism of metastasis promotion by dietary folate and highlights the interaction between nutrients and metastasis-related signaling.

  16. Effect of Quercetin on Caspase-3 of A549 Cell Induced by Influenza Virus H1N1%槲皮素对甲型H1N1流感病毒诱导的A549细胞凋亡效应酶Caspase-3的影响

    Institute of Scientific and Technical Information of China (English)

    万巧凤; 吴莉; 杨美玲; 马锐; 梁军; 顾立刚

    2011-01-01

    Objective To investigate the influence of quercetin on Caspase-3 of lung epithelial tumour A549 infected with H1N1. Methods MTT method was adopted to determine H1N1 virulence, quercetin cytotoxicity, inhibitory effect of quercetin on A549 cytopathic with H1N1 cause. H1N1 of 100 TCID50 was used to infect A549 for 2 h and then change quercetin fluid containing 10 mg/L to cultivate. After 4, 12, 24, 48 h, cells were collected to extract total protein. By adopting the Western-blot method and applying Image-Pro Plus, the grayscale value of Caspase-3 and fl-actin were determined. The gray ratio was relative expression quantity of Caspase-3. Results The TCIDso of H1N1 to A549 was IO-4175, the highest non-toxic concentration of quercetin to A549 (Tco) was 40 mg/L, the least effective concentration (MEC) of quercetin inhibition to A549 cytopathic with H1N1 cause was 10 mg/L. After the infection of H1N1, quercetin significantly inhibited Caspase-3 expression within 4-48 h, showing that quercetin plays an obvious antiapoptotic role. Conclusion Quercetin can play antiviral infectious functions by inhibiting the content or activity of Caspase-3.%目的 观察银杏叶主要活性成分槲皮素对甲型H1N1流感病毒感染的人肺上皮瘤细胞A549凋亡效应酶Caspase-3的影响.方法 采用MTT法测定H1N1毒力、槲皮素细胞毒性作用、槲皮素对H1N1致A549细胞病变的抑制作用.然后用100 TCID50甲型H1N1感染A549 2 h后换用含10 mg/L槲皮素维持液继续培养,于感染后4、12、24、48 h收集细胞,提取细胞总蛋白,采用Western blot方法,应用Image-Pro Plus测定Caspase-3与β-actin灰度值,两者灰度比值为Caspase-3的相对表达量.结果 甲型H1N1 A549的TCID50为10-4.75,槲皮素A549的最大无毒浓度为40 mg/L,槲皮素抑制甲型H1N1致A549细胞病变的最小有效浓度为10 mg/L.在甲型H1N1感染A549细胞后4~ 48 h内,槲皮素可显著抑制Caspase-3蛋白表达,具有明显的抗凋亡作用.结论

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

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    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.

  18. DAPI染色对A549肺癌细胞弹性模量的影响%Effect of DAPI Stain on Elastic Properties of A549 Cells

    Institute of Scientific and Technical Information of China (English)

    叶志义; 张丽; 范霞

    2010-01-01

    原子力显微镜(AFM)能测量活细胞的力学性质.DAPI染色A549肺癌细胞核后用AFM测量细胞的弹性模量,分析DAPI染色对细胞力学性质的影响.通过Hertz模型拟合AFM测量压痕细胞的力-压痕曲线,得到细胞的弹性模量.实验结果:用DAPI染色细胞后其弹性模量为3.66±2.08kPa,而对照组未染色的细胞弹性模量为3.21±1.73kPa,数据比较显示,DAPI染色对A549肺癌细胞弹性模量没有显著性影响.

  19. Reversal of multidrug resistance by cisplatin-loaded magnetic Fe3O4 nanoparticles in A549/DDP lung cancer cells in vitro and in vivo.

    Science.gov (United States)

    Li, Ke; Chen, Baoan; Xu, Lin; Feng, Jifeng; Xia, Guohua; Cheng, Jian; Wang, Jun; Gao, Feng; Wang, Xuemei

    2013-01-01

    The purpose of this study was to explore whether magnetic Fe(3)O(4) nanoparticles (Fe(3)O(4)-MNP) loaded with cisplatin (Fe(3)O(4)-MNP-DDP) can reverse DDP resistance in lung cancer cells and to investigate mechanisms of multidrug resistance in vitro and in vivo. MTT assay showed that DDP inhibited both A549 cells and DDP-resistant A549 cells in a time-dependent and dose-dependent manner, and that this inhibition was enhanced by Fe(3)O(4)-MNP. An increased rate of apoptosis was detected in the Fe(3)O(4)-MNP-DDP group compared with a control group and the Fe(3)O(4)-MNP group by flow cytometry, and typical morphologic features of apoptosis were confirmed by confocal microscopy. Accumulation of intracellular DDP in the Fe(3)O(4)-MNP-DDP group was greater than that in the DDP group by inductively coupled plasma mass spectrometry. Further, lower levels of multidrug resistance-associated protein-1, lung resistance-related protein, Akt, and Bad, and higher levels of caspase-3 genes and proteins, were demonstrated by reverse transcriptase polymerase chain reaction and Western blotting in the presence of Fe(3)O(4)-MNP-DDP. We also demonstrated that Fe(3)O(4)-MNP enhanced the effect of DDP on tumor growth in BALB/c nude mice bearing DDP-resistant human A549 xenografts by decreasing localization of lung resistance-related protein and Ki-67 immunoreactivity in cells. There were no apparent signs of toxicity in the animals. Overall, these findings suggest potential clinical application of Fe(3)O(4)-MNP-DDP to increase cytotoxicity in lung tumor xenografts.

  20. 全反式维甲酸联合顺铂对A549细胞增殖及PDCD5和TP53 mRNA表达的影响%Effects of ATRA combined with cisplatin on A549 cell proliferation and expression of PDCD5 and TP53 mRNA

    Institute of Scientific and Technical Information of China (English)

    刘红; 李海燕; 李伟国; 康燕; 陈红杰; 黄永杰; 王静

    2013-01-01

    目的 观察全反式维甲酸(ATRA)联合顺铂对肺癌A549细胞增殖的影响,探索ATRA应用的最适浓度,研究各组肺癌A549细胞PDCD5、TP53基因表达情况.方法 以不同浓度的ATRA(10-5 mol/L、10-6 mol/L、10-7 mol/L)与顺铂(10-2 μmol/L)联合作用于肺癌A549细胞,采用MTT法检测细胞增殖情况.利用RT-PCR检测ATRA、顺铂作用于肺癌细胞A549细胞后,凋亡相关基因PDCD5、TP53 mRNA的表达.结果 ①MTT研究可观察到,在24 h、48 h、72 h各时间点,ATRA组、顺铂组及两药联合组A549细胞增殖率与阴性对照组比较明显降低(P<0.05);在同一时间点,ATRA不同的浓度间,抑制率随浓度的增加而增加(P<0.05).②与阴性对照组相比,药物干预组PDCD5、TP53mRNA含量明显增加(P<0.05).结论 ATRA对肺癌A549的生长具有抑制作用,有明显的浓度和时间依赖性;ATRA及其与顺铂联用,可以诱导PDCD5及TP53的mRNA的表达.PDCD5与TP53的表达呈正相关.%Objective To observe the functions of all-trans retinoic acid (ATRA) combined with cisplatin on proliferation of A549 cells and to explore the optimal concentration of ATRA and the expressions of PDCD5,TP53 genes of A549 cells in different groups.Methods A549 cells were treated by different concentrations of ATRA (10-5 mol/L,10-6 mol/L,10-7 mol/L) and cisplatin (10-2 μmol/L).MTT assay was used to detect cell proliferation in different groups.The expressions of PDCD5 and TP53 mRNA were detected by RT-PCR.Results ①MTT assay showed the cell proliferation rate in ATRA group,cisplatin group,and two-drug group was lower than that in control group (P <0.05) in 24 h,48 h,and 72 h after treatment.At the same time-point,cell inhibition rate increased with the increase of the concentration of ATRA (P <0.05).②Compared with control group,mRNA of PDCD5 and TP53 in groups treated with drugs increased (P <0.05).Conclusions The growth of lung cancer A549 cells can be inhibited by ATRA in a concentration

  1. 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.

  2. 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.

  3. Reversal of multidrug resistance by cisplatin-loaded magnetic Fe3O4 nanoparticles in A549/DDP lung cancer cells in vitro and in vivo

    Directory of Open Access Journals (Sweden)

    Li K

    2013-05-01

    Full Text Available Ke Li,1 Baoan Chen,1,2 Lin Xu,3 Jifeng Feng,3 Guohua Xia,1,2 Jian Cheng,1,2 Jun Wang,1,2 Feng Gao,1,2 Xuemei Wang,41Department of Hematology, Key Medical Disciplines of Jiangsu Province, Zhongda Hospital, Medical School, Southeast University, Nanjing, 2Faculty of Oncology, Medical School, Southeast University, Nanjing, 3Department of Thoracic Surgery, Jiangsu Province Cancer Hospital, Jiangsu Province, 4State Key Laboratory of Bioelectronics, Southeast University, Nanjing, People’s Republic of ChinaAbstract: The purpose of this study was to explore whether magnetic Fe3O4 nanoparticles (Fe3O4-MNP loaded with cisplatin (Fe3O4-MNP-DDP can reverse DDP resistance in lung cancer cells and to investigate mechanisms of multidrug resistance in vitro and in vivo. MTT assay showed that DDP inhibited both A549 cells and DDP-resistant A549 cells in a time-dependent and dose-dependent manner, and that this inhibition was enhanced by Fe3O4-MNP. An increased rate of apoptosis was detected in the Fe3O4-MNP-DDP group compared with a control group and the Fe3O4-MNP group by flow cytometry, and typical morphologic features of apoptosis were confirmed by confocal microscopy. Accumulation of intracellular DDP in the Fe3O4-MNP-DDP group was greater than that in the DDP group by inductively coupled plasma mass spectrometry. Further, lower levels of multidrug resistance-associated protein-1, lung resistance-related protein, Akt, and Bad, and higher levels of caspase-3 genes and proteins, were demonstrated by reverse transcriptase polymerase chain reaction and Western blotting in the presence of Fe3O4-MNP-DDP. We also demonstrated that Fe3O4-MNP enhanced the effect of DDP on tumor growth in BALB/c nude mice bearing DDP-resistant human A549 xenografts by decreasing localization of lung resistance-related protein and Ki-67 immunoreactivity in cells. There were no apparent signs of toxicity in the animals. Overall, these findings suggest potential clinical application of

  4. Enhanced sensitivity of A549 cells to the cytotoxic action of anticancer drugs via suppression of Nrf2 by procyanidins from Cinnamomi Cortex extract

    Energy Technology Data Exchange (ETDEWEB)

    Ohnuma, Tomokazu; Matsumoto, Takashi; Itoi, Ayano; Kawana, Ayako; Nishiyama, Takahito; Ogura, Kenichiro [Department of Drug Metabolism and Molecular Toxicology, School of Pharmacy, Tokyo University of Pharmacy and Life Sciences, 1432-1 Horinouchi, Hachioji-shi, Tokyo 192-0392 (Japan); Hiratsuka, Akira, E-mail: hiratuka@toyaku.ac.jp [Department of Drug Metabolism and Molecular Toxicology, School of Pharmacy, Tokyo University of Pharmacy and Life Sciences, 1432-1 Horinouchi, Hachioji-shi, Tokyo 192-0392 (Japan)

    2011-10-07

    Highlights: {yields} We found a novel inhibitor of Nrf2 known as a chemoresistance factor. {yields} Overexpressed Nrf2 in lung cancer cells was suppressed by Cinnamomi Cortex extract. {yields} Cytotoxic action of anticancer drugs in cells treated with the extract was enhanced. {yields} Procyanidin tetramers and pentamers were active components in suppressing Nrf2. -- Abstract: Nuclear factor-E2-related factor 2 (Nrf2) is an important cytoprotective transcription factor because Nrf2-regulated enzymes play a key role in antioxidant and detoxification processes. Recent studies have reported that lung cancer cells overexpressing Nrf2 exhibit increased resistance to chemotherapy. Suppression of overexpressed Nrf2 is needed for a new therapeutic approach against lung cancers. In the present study, we found that Cinnamomi Cortex extract (CCE) has an ability to suppress Nrf2-regulated enzyme activity and Nrf2 expression in human lung cancer A549 cells with high Nrf2 activity. Moreover, we demonstrated that CCE significantly enhances sensitivity of A549 cells to the cytotoxic action of doxorubicin and etoposide as well as increasing the intracellular accumulation of both drugs. These results suggest that CCE might be an effective concomitant agent to reduce anticancer drug resistance derived from Nrf2 overexpression. Bioactivity-guided fractionation revealed that procyanidin tetramers and pentamers contained in CCE were active components in suppressing Nrf2.

  5. Legionella pneumophila infection induces programmed cell death, caspase activation, and release of high-mobility group box 1 protein in A549 alveolar epithelial cells: inhibition by methyl prednisolone

    Directory of Open Access Journals (Sweden)

    Koide Michio

    2008-05-01

    Full Text Available Abstract Background Legionella pneumophila pneumonia often exacerbates acute lung injury (ALI and acute respiratory distress syndrome (ARDS. Apoptosis of alveolar epithelial cells is considered to play an important role in the pathogenesis of ALI and ARDS. In this study, we investigated the precise mechanism by which A549 alveolar epithelial cells induced by L. pneumophila undergo apoptosis. We also studied the effect of methyl prednisolone on apoptosis in these cells. Methods Nuclear deoxyribonucleic acid (DNA fragmentation and caspase activation in L. pneumophila-infected A549 alveolar epithelial cells were assessed using the terminal deoxyribonucleotidyl transferase-mediated triphosphate (dUTP-biotin nick end labeling method (TUNEL method and colorimetric caspase activity assays. The virulent L. pneumophila strain AA100jm and the avirulent dotO mutant were used and compared in this study. In addition, we investigated whether methyl prednisolone has any influence on nuclear DNA fragmentation and caspase activation in A549 alveolar epithelial cells infected with L. pneumophila. Results The virulent strain of L. pneumophila grew within A549 alveolar epithelial cells and induced subsequent cell death in a dose-dependent manner. The avirulent strain dotO mutant showed no such effect. The virulent strains of L. pneumophila induced DNA fragmentation (shown by TUNEL staining and activation of caspases 3, 8, 9, and 1 in A549 cells, while the avirulent strain did not. High-mobility group box 1 (HMGB1 protein was released from A549 cells infected with virulent Legionella. Methyl prednisolone (53.4 μM did not influence the intracellular growth of L. pneumophila within alveolar epithelial cells, but affected DNA fragmentation and caspase activation of infected A549 cells. Conclusion Infection of A549 alveolar epithelial cells with L. pneumophila caused programmed cell death, activation of various caspases, and release of HMGB1. The dot/icm system, a

  6. 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。

  7. Artemisinin induces A549 cell apoptosis dominantly via a reactive oxygen species-mediated amplification activation loop among caspase-9, -8 and -3.

    Science.gov (United States)

    Gao, Weijie; Xiao, Fenglian; Wang, Xiaoping; Chen, Tongsheng

    2013-10-01

    This report is designed to explore the roles of caspase-8, -9 and -3 in artemisinin (ARTE)-induced apoptosis in non-small cell lung cancer cells (A549 cells). ARTE induced reactive oxygen species (ROS)-mediated apoptosis in dose- and time-dependent fashion. Although ARTE treatment did not induce Bid cleavage and significant loss of mitochondrial membrane potential, it induced release of Smac and AIF but not cytochrome c from mitochondria, and silencing of Bak but not Bax significantly prevented ARTE-induced cytotoxicity. Moreover, ARTE treatment induced ROS-dependent activation of caspase-9, -8 and -3. Of the utmost importance, silencing or inhibiting any one of caspase-8, -9 and -3 almost completely prevented ARTE-induced activation of all the three caspases and remarkably abrogated the cytotoxicity of ARTE, suggesting that ARTE triggered an amplification activation loop among caspase-9, -8 and -3. Collectively, our data demonstrate that ARTE induces a ROS-mediated amplification activation loop among caspase-9, -8 and -3 to dominantly mediate the apoptosis of A549 cells.

  8. Growth arrest of lung carcinoma cells (A549) by polyacrylate-anchored peroxovanadate by activating Rac1-NADPH oxidase signalling axis.

    Science.gov (United States)

    Chatterjee, Nirupama; Anwar, Tarique; Islam, Nashreen S; Ramasarma, T; Ramakrishna, Gayatri

    2016-09-01

    Hydrogen peroxide is often required in sublethal, millimolar concentrations to show its oxidant effects on cells in culture as it is easily destroyed by cellular catalase. Previously, we had shown that diperoxovanadate, a physiologically stable peroxovanadium compound, can substitute H2O2 effectively in peroxidation reactions. We report here that peroxovanadate when anchored to polyacrylic acid (PAPV) becomes a highly potent inhibitor of growth of lung carcinoma cells (A549). The early events associated with PAPV treatment included cytoskeletal modifications, increase in GTPase activity of Rac1, accumulation of the reactive oxygen species, and also increase in phosphorylation of H2AX (γH2AX), a marker of DNA damage. These effects persisted even at 24 h after removal of the compound and culminated in increased levels of p53 and p21 together with growth arrest. The PAPV-mediated growth arrest was significantly abrogated in cells pre-treated with the N-acetylcysteine, Rac1 knocked down by siRNA and DPI an inhibitor of NADPH oxidase. In conclusion, our results show that polyacrylate derivative of peroxovanadate efficiently arrests growth of A549 cancerous cells by activating the axis of Rac1-NADPH oxidase leading to oxidative stress and DNA damage.

  9. PI3K Mediates the Effect of Resolvin D1 on the Protein Expression of Epithelial Sodium Channel in A549 Cells Treated with Lipopolysaccharide%PI3K介导消退素D1上调脂多糖刺激的A549细胞钠离子通道

    Institute of Scientific and Technical Information of China (English)

    杨艺; 程杨; 金胜威; 高防

    2013-01-01

    目的 研究促炎症消退介质消退素D1(resolvin D1,RvD1)对脂多糖(lipopolysaccharide,LPS)刺激的肺泡上皮A549细胞钠离子通道α亚基(epithelial sodium channel α-subunit,α-ENaC)和γ亚基(epithelial sodium channel-γ-subunit,γ-ENaC)蛋白表达的影响并探讨其机制.方法 不同时间点和不同剂量的LPS刺激A549细胞建立LPS刺激A549细胞模型.将A549细胞分为:空白对照组;LPS(1μg/ml)组;LPS+RvD1(100nmol/L)组;LPS+LY294002(10μmol/L)组.用Western blot检测A549细胞中α-ENaC和γ-ENaC蛋白表达及磷酸化的磷脂酰肌醇-3-激酶(PI3K)水平.结果 LPS下调A549细胞α-ENaC和γ-ENaC蛋白表达,消退素D1抑制LPS对ENaC的下调作用,抑制LPS刺激的磷酸化PI3K.结论 消退素D1通过下调磷酸化PI3K,抑制LPS对A549细胞α-ENaC和γ-ENaC蛋白表达的下调作用.%Objective To study the effect of resolvin D1 (RvD1) on the protein expression of epithelial sodium channel αt-subunit (αt-ENaC) and epithelial sodium channel γ-subunit (γ-ENaC) in A549 cells treated with lipopolysaccharide (LPS),and to explore the molecular mechanisms of signal pathway in RvD1 actions.Methods To establish the model of LPS-induced injury,A549 cells were treated with different concentrations of LPS and simulated by LPS at different time points.A549 cells were divided into four groups:control group; LPS (LPS,1μg/ml) group; LPS +RvD1 (100nmol/L) group and LPS + LY294002 (10μmoL/L) group.Protein expression of ENaC and phosphorylation of phosphoinositide 3-kinase (PI3K) were detected by western blot.Results Protein expression of α-ENaC and γ-ENaC was found to be markedly decreased in the LPS group as compared with control group.This decrease was significantly reduced by RvD1 and LY294002.RvD1 inhibited phosphorylation of PI3K induced by LPS.Conclusion RvD1 increases the protein expression of α-ENaC and γ-ENaC stimulated by LPS via PI3 K pathway in A549 cells.

  10. 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.

  11. 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.

  12. Role of miR-155 in invasion and metastasis of lung adenocarcinoma A549 cells%miR-155在肺腺癌 A549细胞侵袭和转移中的作用

    Institute of Scientific and Technical Information of China (English)

    程田力; 胡成平; 李敏; 顾其华; 安健

    2016-01-01

    拟物对照组、miR-155抑制剂组和 miR-155抑制剂对照组的 PTEN 蛋白相对表达水平分别为0.4±0.1、1.0±0.3、2.8±0.2和1.4±0.1。 miR-155模拟物组与miR-155模拟物对照组、miR-155抑制剂组与 miR-155抑制剂对照组的 PTEN mRNA 和蛋白表达差异均有统计学意义(均 P<0.05)。结论miR-155可能是通过下调靶基因 PTEN 的表达而促进肺腺癌的侵袭和转移。%Objective To investigate the role and mechanism of miR-155 in invasion and metastasis of lung adenocarcinoma A549 cells.Methods Real-time PCR and fluorescence in situ hybridization were used to detect the miR-155 expression in patients′lung adenocarcinoma and adjacent tissue and lymph nodes.Scratch test and Transwell migration assay were used to assess the effect of miR-155 on the A549 cell migration and invasion capability.Bioinformatics software was used to predict the target genes of miR-155, and using luciferase to assay the target gene.Western blot and real-time PCR were performed to confirm the role of miR-155 expression in the regulation of target gene PTEN.Results The real-time quantitative PCR showed that the miR-155 expression levels in adjacent normal tissue, lung adenocarcinoma and metastatic lymph nodes were 4.1±0.5, 9.6±3.1 and 7.8±2.2, respectively.The in situ hybridization showed that the expression rates of miR-155 in the adjacent normal tissue, lung adenocarcinoma and metastatic lymph nodes were (23.2±15.3)%, (75.4±20.2 )% and (60.4±25.1)%,respectively.The Scratch assay showed that the wound healing rates in the miR-155 mimics group, miR-155 mimics NC group, miR-155 inhibitor group and miR-155 inhibitor NC group at 24 h were (43.2±2.2)%, (21.3±4.2)%, (24.3±5.3)%, and (35.2± 5.1)%, and that at 48 h were (75.2±4.5)%, (52.6±5.2)%, (39.4±4.2)%, and( 51.5±4.3)%, respectively.Dual luciferase reporter gene assay showed that the value of the luciferase in the miR -155 mimics group co-transfected with PTEN 3

  13. 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.

  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. 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.

    Science.gov (United States)

    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

  16. 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.

  17. 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.

  18. 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

  19. 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.

  20. 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

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

    Directory of Open Access Journals (Sweden)

    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.

  2. 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

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

    Directory of Open Access Journals (Sweden)

    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.

  4. Cell cycle synchronization and BrdU incorporation as a tool to study the possible selective elimination of ErbB1 gene in the micronuclei in A549 cells

    Directory of Open Access Journals (Sweden)

    C. Lauand

    2015-05-01

    Full Text Available Lung cancer often exhibits molecular changes, such as the overexpression of the ErbB1 gene that encodes epidermal growth factor receptor (EGFR. ErbB1 amplification and mutation are associated with tumor aggressiveness and low response to therapy. The aim of the present study was to design a schedule to synchronize the cell cycle of A549 cell line (a non-small cell lung cancer and to analyze the possible association between the micronuclei (MNs and the extrusion of ErbB1 gene extra-copies. After double blocking, by the process of fetal bovine serum deprivation and vincristine treatment, MNs formation was monitored with 5-bromo-2-deoxyuridine (BrdU incorporation, which is an S-phase marker. Statistical analyses allowed us to infer that MNs may arise both in mitosis as well as in interphase. The MNs were able to replicate their DNA and this process seemed to be non-synchronous with the main cell nuclei. The presence of ErbB1 gene in the MNs was evaluated by fluorescent in situ hybridization (FISH. ErbB1 sequences were detected in the MNs, but a relation between the MNs formation and extrusion of amplified ErbB1could not be established. The present study sought to elucidate the meaning of MNs formation and its association with the elimination of oncogenes or other amplified sequences from the tumor cells.

  5. Cell cycle synchronization and BrdU incorporation as a tool to study the possible selective elimination of ErbB1 gene in the micronuclei in A549 cells

    Energy Technology Data Exchange (ETDEWEB)

    Lauand, C.; Niero, E.L.; Dias, V.M.; Machado-Santelli, G.M. [Departamento de Biologia Celular e do Desenvolvimento, Instituto de Ciências Biomédicas, Universidade de São Paulo, São Paulo, SP (Brazil)

    2015-03-06

    Lung cancer often exhibits molecular changes, such as the overexpression of the ErbB1 gene that encodes epidermal growth factor receptor (EGFR). ErbB1 amplification and mutation are associated with tumor aggressiveness and low response to therapy. The aim of the present study was to design a schedule to synchronize the cell cycle of A549 cell line (a non-small cell lung cancer) and to analyze the possible association between the micronuclei (MNs) and the extrusion of ErbB1 gene extra-copies. After double blocking, by the process of fetal bovine serum deprivation and vincristine treatment, MNs formation was monitored with 5-bromo-2-deoxyuridine (BrdU) incorporation, which is an S-phase marker. Statistical analyses allowed us to infer that MNs may arise both in mitosis as well as in interphase. The MNs were able to replicate their DNA and this process seemed to be non-synchronous with the main cell nuclei. The presence of ErbB1 gene in the MNs was evaluated by fluorescent in situ hybridization (FISH). ErbB1 sequences were detected in the MNs, but a relation between the MNs formation and extrusion of amplified ErbB1could not be established. The present study sought to elucidate the meaning of MNs formation and its association with the elimination of oncogenes or other amplified sequences from the tumor cells.

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

    Science.gov (United States)

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

    2015-01-01

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

  7. 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.

  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)

    LIANG; Xingjie; (

    2001-01-01

    , 1146(1): 136.[12] Howlett, N. G., Avery, S. V., Relationship between cadmium sensitivity and degree of plasma membrane fatty acid unsatu-ration in Saccharomyces cerevisiae, Appl. Microbiol. Biotechnol., 1997, 48(4): 539.[13] Petriz, J., Oconnor, J. E., Carmona, M. et al., Is Rhodamine-123 an appropriate fluorescent probe to assess P-glycoprotein mediated multidrug resistance in vinblastine-resistant CHO cells? Analytical Cellular Pathology, 1997, 14(3): 129.[14] Leonce, S., Burbridge, M., Flow cytometry: a useful technique in the study of multidrug resistance, J. Bio. Cell, 1993, 78(1-2): 63.[15] Le Moyec, L., Tatoud, R., Degorges, A. et al., Proton nuclear magnetic resonance spectroscopy reveals cellular lipids in-volved in resistance to Adriamycin and Taxol by the K562 Leukemia cell line, Cancer Res., 1996, 56: 3461.[16] Callaghan, R., Stafford, A., Epand, R. M., Increased accumulation of drugs in a multidrug resistant cell line by alteration of membrane biophysical properties, Biochim. Biophys. Acta, 1993, 1175(3): 277.[17] Sinicrope, F. A., Dudeia, P. K., Bissommette, B. M. et al., Modulation of P-glycoprotein-mediated drug transport by al-terations in lipid fluidity of rat liver canlicular membrane vesicles, J. Biol. Chem., 1992, 267(35): 24995.[18] Romsicki, Y., Sharom, F. J., The membrane lipid environment modulates drug interactions with the P-glycoprotein multi-drug transporter, Biochemistry, 1999, 38(21): 6887.[19] Garel, O., Lecureur, V., Guillouzo, A., The P-glycoprotein multidrug transporter, Gen. Pharmacol., 1996, 27(8): 1283.[20] Aran, J. M., Pastan, I., Gottesman, M. M., Therapeutic strategies involving the multidrug resistance phenotype: the MDR1 gene as target, chemoprotectant, and selectable marker in gene therapy, Adv. Pharmacol., 1999, 46: 1.[21] Zaman, G. J., Flens, M. J., Vanleusden, M. R. et al., The human multidrug resistance-associated protein (MRP) is a plasma membrane drug efflux pump, Proc. Natl. Acad

  9. Antitumor activity of paclitaxel or/and cisplatin drug delivery system against lung cancer cells A549 in vitro%紫杉醇-顺铂联合药物控释系统对肺腺癌细胞系 A549细胞生长的抑制作用

    Institute of Scientific and Technical Information of China (English)

    崔永; 柳明亮; 吴炳群; 段新春; 龚民

    2014-01-01

    Objective To observe paclitaxel and/ or cisplatin loaded microfiber by electrospinning technique, deliver this system to lung cancer cell A549 in vitro and observe the inhibition of cancer cell and to research effectiveness of controlled drugs delivered by electrospinning technique in antitumor field. Methods Lung cancer cell A549 was cultivated in vitro and incubated on 96-well plates with density of 1×104 per well. The plates were incubated at 37 ℃ and saturated humidity for 24 hours. The plates were taken out and drugs were delivered at different concentrations in each group. There were controlled groups. Plates were incubated for 48 hours. Add in MTT(20 μL/ well) and incubated for 4 hours. The medium containing MTT was discarded thoroughly and 150 μL DMSO was added, gently shaken to get a clear solution 10 ~ 15 min later. OD 490 was determined. Inhibition rate of drugs was calculated. Results Poly propylene carbonate loading paclitaxel and cisplatin controlled delivery system by electrospinning technique could inhibit cancer cell in vitro, stronger than naked paclitaxel and cisplatin and their single drug-loaded microfiber. Poly propylene carbonate loading paclitaxel or cisplatin has stronger inhibition to A549 lung cancer cells than naked paclitaxel or cisplatin. Blank poly propylene carbonate showed no inhibitory effect on the cancer cells. Conclusion Poly propylene carbonate loading paclitaxel and/ or cisplatin by electrospinning technique could inhibit lung cancer cells in vitro significantly. Controlled drug-delivery system by electrospinning technique could implant antitumor drugs locally, reduce toxicity and side effect of chemotherapeutics and have a great application potential.%目的:观察以聚碳酸亚丙酯乳液作为纺丝液,采用静电纺丝技术,负载紫杉醇和顺铂制备的载药纤维控释系统对体外培养的肺腺癌细胞系 A549的抑制率,为进一步的动物实验奠定基础,并探讨用于肺癌治疗的

  10. Long-term exposure of A549 cells to titanium dioxide nanoparticles induces DNA damage and sensitizes cells towards genotoxic agents.

    Science.gov (United States)

    Armand, Lucie; Tarantini, Adeline; Beal, David; Biola-Clier, Mathilde; Bobyk, Laure; Sorieul, Sephanie; Pernet-Gallay, Karin; Marie-Desvergne, Caroline; Lynch, Iseult; Herlin-Boime, Nathalie; Carriere, Marie

    2016-09-01

    Titanium dioxide nanoparticles (TiO2-NPs) are one of the most produced NPs in the world. Their toxicity has been studied for a decade using acute exposure scenarios, i.e. high exposure concentrations and short exposure times. In the present study, we evaluated their genotoxic impact using long-term and low concentration exposure conditions. A549 alveolar epithelial cells were continuously exposed to 1-50 μg/mL TiO2-NPs, 86% anatase/14% rutile, 24 ± 6 nm average primary diameter, for up to two months. Their cytotoxicity, oxidative potential and intracellular accumulation were evaluated using MTT assay and reactive oxygen species measurement, transmission electron microscopy observation, micro-particle-induced X-ray emission and inductively-coupled plasma mass spectroscopy. Genotoxic impact was assessed using alkaline and Fpg-modified comet assay, immunostaining of 53BP1 foci and the cytokinesis-blocked micronucleus assay. Finally, we evaluated the impact of a subsequent exposure of these cells to the alkylating agent methyl methanesulfonate. We demonstrate that long-term exposure to TiO2-NPs does not affect cell viability but causes DNA damage, particularly oxidative damage to DNA and increased 53BP1 foci counts, correlated with increased intracellular accumulation of NPs. In addition, exposure over 2 months causes cellular responses suggestive of adaptation, characterized by decreased proliferation rate and stabilization of TiO2-NP intracellular accumulation, as well as sensitization to MMS. Taken together, these data underline the genotoxic impact and sensitization effect of long-term exposure of lung alveolar epithelial cells to low levels of TiO2-NPs.

  11. 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.

  12. 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.

  13. 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.

  14. 缺氧诱导因子-1α对A549细胞株中生存素表达及其生物学特性的影响%Effect of hypoxia inducible factor-1αon the biological and the transcription of survivin in A549 cells

    Institute of Scientific and Technical Information of China (English)

    舒红梅; 赵成岭; 孙艳; 李伟; 陈余清

    2014-01-01

    Objective To evaluate the effect of hypoxia inducible factor-1α(HIF-1α) on the transcriptional regulation of survivin and the proliferation, apoptosis and migration of A549 cells. Methods The binding of nuclear protein to the survivin promoter sequences was detected using electrophoretic mobility shift assay (EMSA). It was divided into 5 groups in EMSA: negative control reaction, binding reaction, probe cold competing reaction, mutant probe cold competing reaction Super-shift reaction. The plasmid pcDNATM6.2-GW/EmGFP miRNA HIF-1α was stably transfected into adenocarcinoma of lung A549 cell by LipofectamineTM 2000. Cells were divided into 4 groups in trsnsfection:untreated group, HIF-1αmiRNA group, negative control group,β-actin group .The expression of HIF-1α and survivin in A549 cells were detected by RT-PCR and Western blot, to screen the best silencing vector A549/HIF-1α-miRNA. The transfected cells were exposed to normoxia and hypoxia, cell proliferation, apotosis and migration were measured by CCK8, flow cytometry (FCM) and transwell chambers methods. Results DNA-neucleoprotein bands were observed when A549 nuclear extracts incubating with theγ-32P labeled 18-bp probe (nucleotides-26to-9) of survivin promoter in EMSA assay. The specific bands were competed away by the cold 18-bp probe, but not the mutated cold probe in competition assay. The bands could bind the antibody of HIF-1α, exhibited in super-shift reaction. Under the normal condition, in the HIF-1αmiRNA group, the mRNA of HIF-1αand survivin were 0.313±0.051 and 0.060±0.008, respectively, which were significantly decreased compared with those of the control group (1.042±0.036 and 1.071±0.016, respectively)(F=436.433, 0.002, all P<0.0.5). HIF-1α and survivin protin in the HIF-1α miRNA group(0.559±0.051 and 0.051±0.003) were significantly lower than those in control group(F=53.525 and 331.646, all P<0.05).Under hypoxia, HIF-1α, survivin mRNA and protein in the transfection of HIF-1

  15. Pneumocystis carinii major surface glycoprotein induces interleukin-8 and monocyte chemoattractant protein-1 release from a human alveolar epithelial cell line

    DEFF Research Database (Denmark)

    Benfield, T L; Lundgren, Bettina; Shelhamer, J H

    1999-01-01

    (IL-8) and monocyte chemoattractant protein-1 (MCP-1) from an alveolar epithelial cell line (A549). RESULTS: Incubation of A549 cells with MSG in concentrations from 0.4 to 10 microg mL-1 for 24 h caused dose-dependent increases in IL-8 release (3.4-fold above control, P

  16. 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.

  17. 转化生长因子β1对A549细胞晚期糖基化终产物受体与细胞外基质表达的影响%Expression of RAGE and extracellular matrix induced by transforming growth factor beta 1 in A549 cells

    Institute of Scientific and Technical Information of China (English)

    丁辉; 冯艳; 陈如华

    2013-01-01

    目的 研究转化生长因子β1 (transforming growth factor beta 1,TGF-β1)对人A549细胞的晚期糖基化终产物受体(receptor for advanced glycation end-products,RAGE)和胶原-Ⅰ、α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)表达的作用.方法 体外培养人A549细胞,以不加TGF-β1刺激为对照组,不同终浓度TGF-β1(2 μg/L、5 μg/L、10 μg/L)处理A549细胞为实验组,RT-PCR和Western blotting法检测TGF-β1刺激12 h、24 h、36 h后对照组和实验组RAGE mRNA和RAGE、胶原-Ⅰ及α-SMA的蛋白表达.结果 ①随着TGF-β1浓度的增加,刺激时间的延长,A549细胞的RAGE mRNA和蛋白的表达逐渐降低,呈时间-浓度依赖关系.②与对照组相比,A549在TGF-β1(5 μg/L)刺激24 h后,RAGE mRNA的表达明显降低(0.387±0.088 vs 1.345±0.132,P<0.01).RAGE蛋白表达较对照组明显降低(0.174±0.061 vs 1.229±0.112,P<0.01).③TGF-β1(5μg/L)刺激24 h后,A549的胶原-Ⅰ及α-SMA蛋白表达(0.916±0.071和0.899±0.022)分别较对照组(0.295±0.041和0.134±0.028)升高3.1和6.7倍,差异有统计学意义(P<0.01).④相关分析显示,A549的RAGE蛋白表达与胶原-Ⅰ(r=-0.843,P<0.05)、α-SMA蛋白的表达(r=-0.897,P<0.05)呈负相关.结论 TGF-β1下调A549细胞RAGE的mRNA和蛋白表达,与胶原-Ⅰ及α-SMA蛋白表达上调呈负相关,提示RAGE的表达下调可能与肺纤维化的发生和发展有关.%Objective To investigate the expressions of receptor for advanced glycation endproducts (RAGE),collagen-Ⅰ,and α-smooth muscle actin (α-SMA) in human A549 cells induced by transforming growth factor beta 1 (TGF-β1).Methods With no TGF-β1-stimulated cells for the control group,human A549 cells were cultured in vitro.Cultured cells were exposed to TGF-β1 with different final concentration (2 μg/L,5 μg/L,10 μg/L) for different time (12 h,24 h,36 h).RAGE mRNA and protein,collagen-Ⅰ and α-SMA in cells were detected at different points after the treatment of TGF

  18. Airborne particulate matter in vitro exposure induces cytoskeleton remodeling through activation of the ROCK-MYPT1-MLC pathway in A549 epithelial lung cells.

    Science.gov (United States)

    Chirino, Yolanda I; García-Cuellar, Claudia María; García-García, Carlos; Soto-Reyes, Ernesto; Osornio-Vargas, Álvaro Román; Herrera, Luis A; López-Saavedra, Alejandro; Miranda, Javier; Quintana-Belmares, Raúl; Pérez, Irma Rosas; Sánchez-Pérez, Yesennia

    2017-03-06

    Airborne particulate matter with an aerodynamic diameter ≤10μm (PM10) is considered a risk factor for the development of lung cancer. Little is known about the cellular mechanisms by which PM10 is associated with cancer, but there is evidence that its exposure can lead to an acquired invasive phenotype, apoptosis evasion, inflammasome activation, and cytoskeleton remodeling in lung epithelial cells. Cytoskeleton remodeling occurs through actin stress fiber formation, which is partially regulated through ROCK kinase activation, we aimed to investigate if this protein was activated in response to PM10 exposure in A549 lung epithelial cells. Results showed that 10μg/cm(2) of PM10 had no influence on cell viability but increased actin stress fibers, cytoplasmic ROCK expression, and phosphorylation of myosin phosphatase-targeting 1 (MYPT1) and myosin light chain (MLC) proteins, which are targeted by ROCK. The inhibition of ROCK prevented actin stress fiber formation and the phosphorylation of MYPT1 and MLC, suggesting that PM10 activated the ROCK-MYPT1-MLC pathway in lung epithelial cells. The activation of ROCK1 has been involved in the acquisition of malignant phenotypes, and its induction by PM10 exposure could contribute to the understanding of PM10 as a risk factor for cancer development through the mechanisms associated with invasive phenotype.

  19. 艾烟冷凝物对肺泡Ⅱ型上皮细胞A549活性及凋亡的影响%Influences of condensate of moxa smoke on viability and apoptosis of type Ⅱ alveolar epithelial cells (A549)

    Institute of Scientific and Technical Information of China (English)

    胡海; 赵百孝; 邬继红; 杨陟华; 韩丽; 蔡虹; 朱茂祥

    2012-01-01

    目的 观察艾烟冷凝物(PM10采取人体可吸入的艾烟部分)对肺泡Ⅱ型上皮细胞(A549)活性及凋亡的影响.方法 体外培养A549细胞,加入不同浓度的艾烟冷凝物,应用四甲基偶氮唑蓝(MTT)法、荧光显微镜来观察其对A549的细胞活性以及细胞凋亡的影响.结果 MTT法结果显示:与对照组相比,A549的细胞存活率随浓度和时间发生改变,具有明显的时间浓度依赖性;0.12g/L浓度的艾烟冷凝物作用于细胞12h后可显著提高细胞存活率(P=0.005<0.01);荧光显微镜下观察发现艾烟冷凝物可以引起细胞发生凋亡,且具有浓度依赖性.结论 A549细胞随着艾烟冷凝物浓度的增加、刺激时间的增加而引起细胞活力逐渐下降,表明一定浓度的艾烟冷凝物和一定的刺激时间对细胞具有毒性作用;一定浓度的艾烟冷凝物在较短刺激时间内具有细胞增殖作用,故认为艾烟对细胞的增殖作用可能是艾烟发挥有效作用的重要因素之一;能够引起细胞的凋亡可能是毒性作用的重要因素之一.%Objective To observe the influences of condensate of moxa smoke (PM10, taken inhalable portion of moxa smoke) on viability and apoptosis of type Ⅱ alveolar epithelial cells ( A549 ). Methods A546 cells were cultured in vitro and then condensate of moxa smoke was added in different concentration. The influences of condensate of moxa smoke on the viability and apoptosis of A546 cells were observed by applying MTT assay and fluorescence microscope. Results The results of MTT assay showed that compared with control group, the survival rate of A546 cells changed with the changes of concentration and time showing an obvious time-dependence and a concentration-dependence. The condensate in the dose of 0. 12 mg/mL significantly improved the survival rate of A546 cells after acting on the cells for 12 hours (P =0. 005 <0. 01). The observation of fluorescence microscope showed that the apoptosis of A

  20. 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

    Directory of Open Access Journals (Sweden)

    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

  1. 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

    Energy Technology Data Exchange (ETDEWEB)

    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

  2. 四嗪二甲酰胺抑制肺癌细胞株A549、EBC-1、MA增殖并诱导细胞凋亡的研究%Effects or N, N'-di-(m-methylphenyi) -3,6-dimethyl-l, 4-dlhydro-l ,2,4,5-tetrazine-l, 4-dicarboam-ide on proliferation and apoptosis of A549, EBC-1, MA cells in vitro

    Institute of Scientific and Technical Information of China (English)

    许武林; 周永列; 王珍妮; 吕亚萍; 胡惟孝

    2010-01-01

    目的 观察四嗪二甲酰胺(ZGDHu-1)体外抑制肺癌细胞株A549、EBC-1、MA增殖并诱导细胞凋亡作用.方法 将不同浓度的ZGDHu-1与A549、EBC-1、MA细胞在体外培养,用锥虫蓝染色、噻唑蓝(MTT)比色法、5'-溴-2脱氧尿苷(HrdU)-酶联免疫吸附试验(ELISA)法观察ZGDHu-1对A549、EBC-1、MA细胞增殖的抑制作用;用细胞形态学、DNA含量及细胞周期分析、Annexin-V/PI双标记、Hoechst33258荧光染色观察细胞凋亡.结果 ZGDHu-1能抑制A549、EBC-1、MA细胞增殖和活力,呈现作用时间和剂量的量效关系.A549、EBC-1、MA细胞经ZGDHu-1作用48 h后,其IC50分别为165、106、210μg/L;72h后的IC50分别为42、18、90μg/L.大部分细胞阻滞于G2~M期;出现典型的细胞形态改变,亚G1峰检出并显著增加,Annexin-V+/PI-表达升高,细胞内DNA片段含量增加,Hoechst33258荧光染色后出现凋亡细胞的特征性改变,证实ZGDHu-1能诱导A549、EBC-1、MA细胞凋亡.结论 ZGDHu-1能抑制A549、EBC-1、MA细胞增殖,并可诱导其细胞凋亡.%Objective To study the effect of N, N-di-( m-methylphenyi) -3,6-dimethyl-l ,4-di-hydro-1,2,4,5-tetrazine-1,4-dicarboamide (ZGDHu-1) on proliferation and apoptosis of A549, EBC-1 and MA cells in vitro. Methods Different concentrations of ZGDHu-1 were incubated with A549, EBC-1, or MA cells for different durations. The proliferation inhibition was analyzed by alive cell count, Methyl thi-azolyl tetrazolium (MTT) assay and BrdU-ELISA. Cell apoptosis was measured by cell morphology, DNA content, Annexin-V/PI and Hoechst33258 labeling method. Results ZGDHu-1 could inhibit proliferation of A549, EBC-1 and MA cells within a certain range of treating duration and does, with a 48 h IC50 of 165, 106, 210μLg/L, 72 h of 42, 18, 90μg/L, and a majority of A549, EBC-1 and MA cells were arrested in G2/M phase. The apoptosis of A549, EBC-1 and MA cells was confirmed by type cell morphology, DNA fragment, sub-G1 phase and Hoechst33258

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

    Science.gov (United States)

    Xiong, Ying

    2017-01-01

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

  4. Natural iron chelators: Protective role in A549 cells of flavonoids-rich extracts of Citrus juices in Fe(3+)-induced oxidative stress.

    Science.gov (United States)

    Ferlazzo, Nadia; Visalli, Giuseppa; Cirmi, Santa; Lombardo, Giovanni Enrico; Laganà, Pasqualina; Di Pietro, Angela; Navarra, Michele

    2016-04-01

    Exogenous iron in particulate matter and imbalanced iron homeostasis cause deleterious effects on health. Natural and synthetic iron chelators may be of therapeutic benefit, therefore we evaluated the protective effect of Citrus flavonoids-rich extracts from bergamot and orange juices in iron overloaded human lung epithelial cells. Cytofluorimetric, biochemical and genotoxic analyses were performed in Fe2(SO4)3 exposed A549, pretreated with each extract whose chemical composition was previously detected. Chelating activity was assessed in cells by a calcein ester. Both extracts reduced the generation of reactive oxygen species and membrane lipid peroxidation, improved mitochondrial functionality, and prevented DNA-oxidative damage in iron-exposed cells. Antioxidant effects were attributed to the chelating property, blocking upstream the redox activity of iron. Flavonoid-rich extracts also induced antioxidant catalase. The bergamot and orange juice extracts had a broad-spectrum protective effect. Their use prevents iron oxidative injury and these natural iron chelators could be used as therapeutic agents.

  5. Nanoparticle abraxane possesses impaired proliferation in A549 cells due to the underexpression of glucosamine 6-phosphate N-acetyltransferase 1 (GNPNAT1/GNA1)

    Science.gov (United States)

    Zhao, Minzhi; Li, Haiyun; Ma, Yan; Gong, He; Yang, Shu; Fang, Qiaojun; Hu, Zhiyuan

    2017-01-01

    Abraxane (Abr), a US Food and Drug Administration-approved albumin-bound nanoparticle applied for the treatment of non-small-cell lung cancer, has been reported to be more effective than paclitaxel (PTX). To further understand the molecular mechanisms that produce this superior drug efficacy of Abr, a quantitative proteomic approach has been applied to investigate the global protein expression profiles of lung cancer cell A549 treated with Abr and PTX. Only one protein, namely, glucosamine 6-phosphate N-acetyltransferase 1 (GNA1), showed significant differential expression (P<0.05) in the cutoff of 2.0 fold, suggesting that Abr can be used safely as a substitute for PTX. GNA1 is a key enzyme in the biosynthesis of uridine diphosphate-N-acetylglucosamine, which is an important donor substrate for N-linked glycosylation and has several important functions such as embryonic development and growth. Albumin plays a major role in the regulation of this protein. In summary, this study first shows that the superior drug effect of Abr is mainly due to the downregulation of GNA1, which causes proliferative delay and cell adhesion defect. It is also noteworthy that the deficiency of GNA1 might reduce insulin secretion which correlates with type 2 diabetes.

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

    Science.gov (United States)

    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.

  7. Perfluorocarbon inhibits lipopolysaccharide-induced macrophage inflammatory protein-2 expression and activation of ATF-2 and c-Jun in A549 pulmonary epithelial cells.

    Science.gov (United States)

    Hu, Y; Li, C S; Li, Y Q; Liang, Y; Cao, L; Chen, L A

    2016-04-30

    The signaling pathway that mediates the anti-inflammatory effects of perfluorocarbon (PFC) in alveolar epithelial cells treated with lipopolysaccharide (LPS) remains unclear. To evaluate the role of macrophage-inflammatory protein-2 (MIP-2), four A549 treatment groups were utilized: (1) untreated control, (2) 10 μg/mL of LPS, (3) 10 μg/mL of LPS+30% PFC and (4) 30% PFC. MIP-2 mRNA expression was determined by qPCR and ELISA. Mitogen-activated protein kinase (MAPK) activation was determined by Western blot analysis, and MIP-2 expression was determined by qPCR following treatment with MAPK inhibitors. PFC suppressed LPS-induced MIP-2 mRNA levels (P≤0.035) and MIP-2 secretion (P≤0.046). LPS induced ATF-2 and c-Jun phosphorylation, which was suppressed by PFC. Finally, inhibitors of ERK, JNK, and p38 suppressed LPS-induced MIP-2 mRNA expression. Thus, PFC inhibits LPS-induced MIP-2 expression and ATF-2 and c-Jun phosphorylation. To fully explore the therapeutic potential of PFC for acute lung injury (ALI), in vivo analyses are required to confirm these effects.

  8. Phenazine-1-carboxamide (PCN) from Pseudomonas sp. strain PUP6 selectively induced apoptosis in lung (A549) and breast (MDA MB-231) cancer cells by inhibition of antiapoptotic Bcl-2 family proteins.

    Science.gov (United States)

    Kennedy, R Kamaraj; Veena, V; Naik, P Ravindra; Lakshmi, Pragna; Krishna, R; Sudharani, S; Sakthivel, N

    2015-06-01

    Phenazine-1-carboxamide (PCN), a naturally occurring simple phenazine derivative isolated from Pseudomonas sp. strain PUP6, exhibited selective cytotoxic activity against lung (A549) and breast (MDA-MB-231) cancer cell lines in differential and dose-dependent manner compared to normal peripheral blood mononuclear cells. PCN-treated cancer cells showed the induction of apoptosis as evidenced by the release of low level of LDH, morphological characteristics, production of reactive oxygen species, loss of mitochondrial membrane potential (ΔΨm) and induction of caspase-3. At molecular level, PCN instigates apoptosis by mitochondrial intrinsic apoptotic pathway via the overexpression of p53, Bax, cytochrome C release and activation of caspase-3 with the inhibition of oncogenic anti-apoptotic proteins such as PARP and Bcl-2 family proteins (Bcl-2, Bcl-w and Bcl-xL). The in silico docking studies of PCN targeted against the anti-apoptotic members of Bcl-2 family proteins revealed the interaction of PCN with the BH3 domain, which might lead to the induction of apoptosis due to the inhibition of antiapoptotic proteins. Due to its innate inhibition potential of antiapoptotic Bcl-2 family proteins, PCN may be used as potent anticancer agent against both lung and breast cancer.

  9. 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

  10. Nanoparticle abraxane possesses impaired proliferation in A549 cells due to the underexpression of glucosamine 6-phosphate N-acetyltransferase 1 (GNPNAT1/GNA1

    Directory of Open Access Journals (Sweden)

    Zhao MZ

    2017-03-01

    Full Text Available Minzhi Zhao,* Haiyun Li,* Yan Ma, He Gong, Shu Yang, Qiaojun Fang, Zhiyuan Hu Chinese Academy of Sciences Key Laboratory for Biomedical Effects of Nanomaterials and Nanosafety, CAS Center for Excellence in Nanoscience, National Center for Nanoscience and Technology, Beijing, People’s Republic of China *These authors contributed equally to this work Abstract: Abraxane (Abr, a US Food and Drug Administration-approved albumin-bound nanoparticle applied for the treatment of non-small-cell lung cancer, has been reported to be more effective than paclitaxel (PTX. To further understand the molecular mechanisms that produce this superior drug efficacy of Abr, a quantitative proteomic approach has been applied to investigate the global protein expression profiles of lung cancer cell A549 treated with Abr and PTX. Only one protein, namely, glucosamine 6-phosphate N-acetyltransferase 1 (GNA1, showed significant differential expression (P<0.05 in the cutoff of 2.0 fold, suggesting that Abr can be used safely as a substitute for PTX. GNA1 is a key enzyme in the biosynthesis of uridine diphosphate-N-acetylglucosamine, which is an important donor substrate for N-linked glycosylation and has several important functions such as embryonic development and growth. Albumin plays a major role in the regulation of this protein. In summary, this study first shows that the superior drug effect of Abr is mainly due to the downregulation of GNA1, which causes proliferative delay and cell adhesion defect. It is also noteworthy that the deficiency of GNA1 might reduce insulin secretion which correlates with type 2 diabetes. Keywords: quantitative proteomics, nano-drug, drug efficacy, lung cancer, molecular mechanisms, abraxane

  11. DNA damage and cytotoxicity in type II lung epithelial (A549 cell cultures after exposure to diesel exhaust and urban street particles

    Directory of Open Access Journals (Sweden)

    Møller Peter

    2008-04-01

    Full Text Available Abstract Background Exposure to air pollution particles has been acknowledged to be associated with excess generation of oxidative damage to DNA in experimental model systems and humans. The use of standard reference material (SRM, such as SRM1650 and SRM2975, is advantageous because experiments can be reproduced independently, but exposure to such samples may not mimic the effects observed after exposure to authentic air pollution particles. This study was designed to compare the DNA oxidizing effects of authentic street particles with SRM1650 and SRM2975. The authentic street particles were collected at a traffic intensive road in Copenhagen, Denmark. Results All of the particles generated strand breaks and oxidized purines in A549 lung epithelial cells in a dose-dependent manner and there were no overt differences in their potency. The exposures also yielded dose-dependent increase of cytotoxicity (as lactate dehydrogenase release and reduced colony forming ability with slightly stronger cytotoxicity of SRM1650 than of the other particles. In contrast, only the authentic street particles were able to generate 8-oxo-7,8-dihydro-2'-deoxyguanosine (8-oxodG in calf thymus DNA, which might be due to the much higher level of transition metals. Conclusion Authentic street particles and SRMs differ in their ability to oxidize DNA in a cell-free environment, whereas cell culture experiments indicate that the particle preparations elicit a similar alteration of the level of DNA damage and small differences in cytotoxicity. Although it cannot be ruled out that SRMs and authentic street particles might elicit different effects in animal experimental models, this study indicates that on the cellular level, SRM1650 and SRM2975 are suitable surrogate samples for the study of authentic street particles.

  12. 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

  13. 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.

  14. Sulforaphane derived from broccoli inhibit proliferation and invasion of lung cancer A549 cells in vitro%西兰花提取物萝卜硫素抑制肺癌细胞的生长和侵袭

    Institute of Scientific and Technical Information of China (English)

    贾侃; 贺云冲; 洪姣; 黄春琦; 任军; 许健

    2014-01-01

    Sulforaphane was a multifunction compound derived from brassicaceous vegetable such as broccoli, reports showed that Sulforaphane provided with effection of antitumor and antioxidant. Lung cancer is an aggressive malignancy with a tendency of early distant metastases, the antitumor function of sulforaphane was corroborated by numerous lines of evidence, but the anticancer mechanism of this compound has not been wel obsvered. In this work, we analyzed vitality and invasion of A549 cels treated with sulforaphane by cellcounting kit (CCK8) and transwel, then measure the half maximal (50%) inhibitory concentration (IC50) of sulforaphane for A549 cels. The cels cycle, apoptosis and DNA fragment were analyzed using Flow Cytometry Analysis and agarose electrophoresis, TGF-βand NF-κB were analyzed by western blot after treatment with 3μg/mL sulforaphane. Results showed that A549 cels proliferate and invade were inhibited by sulforaphane with a dose-dependent manner, IC50 of sulforaphane was 3μg/mL, and the cellcycle were arrested at G2/M phase. 3μg/mL sulforaphane induced apoptosis , DNA fragment, decreased the expression of TGF-βand NF-κB in A549 cels. Our results pointed out that sulforaphane inhibited proliferation and invasion of lung cancer A549 cels in vitro, decreased the expression of inflammation proteins, maybe a novel chemotherapy for lung cancer.%萝卜硫素是从十字花科蔬菜中提取的多功能物质,研究已证实其具有抗癌、抗氧化等功效。肺癌是恶性程度高、具有转移倾向的恶性肿瘤,萝卜硫素抗肺癌的机制尚不是十分清楚。本研究通过CCK-8和transwel侵袭实验分析初步判断萝卜硫素对A549肺癌细胞活性和转移侵袭的影响,计算体外干预A549的IC50,流式细胞学分析IC50浓度萝卜硫素对细胞周期和凋亡的影响,电泳分析DNA片段化改变。结果显示A549细胞活性对萝卜硫素剂量依赖性下降,萝卜硫素作用于A549细胞的IC50为3μg

  15. 肺癌细胞A549抗原相关旋毛虫Tsp06172基因的克隆及原核表达%Cloning and prokaryotic expression of the Tsp06172 gene of Trichinella spiralis in A549 lung cancer cells

    Institute of Scientific and Technical Information of China (English)

    高江明; 徐晓芳; 吕萌; 左绍志; 宫鹏涛; 杨举; 李赫; 李建华; 张国才

    2013-01-01

    目的 克隆旋毛虫(Trichinella spiralis)与肺癌细胞A549相关抗原Tsp06172基因,并进行原核表达. 方法 采用RT-PCR方法扩增Tsp06172基因,连接原核表达载体pET-28a,转化入感受态细胞BL21,IPTG诱导表达,经SDS-PAGE和Western blot鉴定表达产物. 结果 重组表达质粒经双酶切及测序鉴定正确.表达分子质量单位约为16 ku的融合蛋白.Western blot检测融合蛋白能被抗A549细胞的多克隆抗体识别. 结论 构建的原核表达载体pET-28a Tsp06172表达具有A549细胞反应原性的蛋白,为旋毛虫Tsp06172重组蛋白功能的研究了奠定基础.%Objective To clone and express the Trichinella spiralis Tsp06172 gene in BL21. Methods The Tsp06172 gene was amplified with RT-PCR and then subcloned into the prokaryotic expression vector pET-28a. BL21 containing the recombinant plasmid pET-28a-Tsp06172 was induced with IPTG. The fusion protein was detected and i-dentified with SDS-PAGE and Western blotting. Results The recombinant expression plasmid was successfully constructed. After induction in an E. coli system. SDS-PAGE results showed that a fusion protein of about 16 ku was successfully expressed. Western blotting indicated that the fusion protein was readily recognized by polyclonal antibodies from A549 cells. Conclusion The recombinant expression plasmid pET-28a-Tsp06172 expressed the corresponding protein in BL21. This finding lays the foundation for research into the function of the Tsp06172 protein.

  16. 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.

  17. The telomeric protein TRF2 is critical for the protection of A549 cells from both telomere erosion and DNA double-strand breaks driven by salvicine.

    Science.gov (United States)

    Zhang, Yong-Wei; Zhang, Zhi-Xiang; Miao, Ze-Hong; Ding, Jian

    2008-03-01

    Telomere repeat binding factor 2 (TRF2) has been increasingly recognized to be involved in DNA damage response and telomere maintenance. Our previous report found that salvicine (SAL), a novel topoisomerase II poison, elicited DNA double-strand breaks and telomere erosion in separate experimental systems. However, it remains to be clarified whether they share a common response to these two events and in particular whether TRF2 is involved in this process. In this study, we found that SAL concurrently induced DNA double-strand breaks, telomeric DNA damage, and telomere erosion in lung carcinoma A549 cells. It was unexpected to find that SAL led to disruption of TRF2, independently of either its transcription or proteasome-mediated degradation. By overexpressing the full-length trf2 gene and transfecting TRF2 small interfering RNAs, we showed that TRF2 protein protected both telomeric and genomic DNA from the SAL-elicited events. It is noteworthy that although both the Ataxia-telangiectasia-mutated (ATM) and the ATM- and Rad3-related (ATR) kinases responded to the SAL-induced DNA damages, only ATR was essential for the telomere erosion. The study also showed that the activated ATR augmented the SAL-triggered TRF2 disruption, whereas TRF2 reduction in turn enhanced ATR function. All of these findings suggest the emerging significance of TRF2 protecting both telomeric DNA and genomic DNA on the one hand and reveal the mutual modulation between ATR and TRF2 in sensing DNA damage signaling during cancer development on the other hand.

  18. Curcumin extract influence on tumor angiogenesis of A549 cell subsets SP and NON-SP cells%姜黄素对A549细胞亚群SP和NON-SP的NF-kB、VEGF及Notch通路的影响

    Institute of Scientific and Technical Information of China (English)

    李小江; 贾英杰; 张文治; 张莹; 许文婷

    2016-01-01

    [Objective] Through the animal experiment further understand the main ingredient in turmeric extract curcumin-of tumor angiogenesis of the specific mechanism. [Methods] The 40 BALB/C mice were divided into four groups of male nude, one group is only 10. group A (SP and group of cells curcumin group), group B (SP and group of cells tumor-burdened control group), group C (as SP and group of cells curcumin group), group D (as SP and group of cells tumor-burdened control group). A and B two groups in lung adenocarcinoma before the establishment A549SP cells subsets tumor-burdened model, C, D two groups establish lung adenocarcinoma A549 cells subsets as SP tumor-burdened model, and after 16 days the model are observated, in group A, C intraperitoneal injection of mice curcumin, the next day 1, eight times, B and D two groups of injection saline. 16 days later executed mice, will tumor piece of tissue dissection, comparison of the heavy; immunohistochemical method to detect tumor tissue VEGF, CD34, NF-kB expression; PT-PCR detected Notch1 mRNA content. [Results] The group of cells SP curcumin group of suppression effect was better than the group of cells as SP curcumin group that cancer stem cells and tumor angiogenesis closely related. [Conclusion] Curcumin can inhibit tumor growth, and may inhibit the expression of NF-kB, down regulate the content of mRNA Notch, block the Notch1 signaling pathway and inhibit the expression of VEGF in tumor tissues.%[目的]通过动物实验了解姜黄提取物-姜黄素抗肿瘤血管生成的具体作用机制.[方法]将40只BALB/C雄性裸小鼠分为4组,每组10只.分别为A组(SP亚群细胞姜黄素组)、B组(SP亚群细胞荷瘤对照组)、C组(NON-SP亚群细胞姜黄素组)、D组(NON-SP亚群细胞荷瘤对照组).A、B两组于实验前建立肺腺癌A549 SP细胞亚群荷瘤模型,C、D两组建立肺腺癌A549 NON-SP细胞亚群荷瘤模型,建立模型后观察16 d,于A组、C组小鼠腹腔注射姜黄素,隔天1

  19. Influence of thermalization on A549 cells growth, c-Jun N-terminal kinase phosphorylation and expression of heat shock protein 70 in patients with lung cancer%热化联合对肺癌患者A549细胞生长、c-Jun N-末端激酶磷酸化及热休克蛋白70表达的影响

    Institute of Scientific and Technical Information of China (English)

    吴海乔; 田甜; 胡君程; 林蓁

    2015-01-01

    目的:观察热化联合对肺癌患者A549细胞生长的影响及机制探讨。方法对A549细胞分别进行单独热疗、单独化疗,热化联合干预及热化联合并SP600125干预,同时选取未做任何处理的A549细胞作为对照组。观察各组细胞增殖率、细胞侵袭力的变化。同时采用蛋白免疫印记法(Western Bolt)检测JNK磷酸化以及热休克蛋白70(HSP70)的表达。结果热化联合组的A549细胞增值率明显低于单独热疗、单独化疗和热化联合并SP600125组(P<0.05)。热化联合组JNK磷酸化表达明显高于对照组及单独化疗组(P<0.05),热化联合组HSP70表达明显低于单独热疗组(P<0.05)。热化联合干预下,p-JNK表达水平出现上升,与对照组、单独热疗组和单独化疗组相比,差异均具有统计学意义(P<0.05);热化联合并SP600125组的p-JNK的表达水平较热化联合组显著下降(P<0.05)。结论热化联合抑制A549细胞增殖的效果优于单独热疗或单独化疗,作用机制可能与激活JNK信号通路或抑制HSP70表达有关。%Objective To investigate the effect of thermalization on A549 cells growth in patients with lung cancer and its mechanism. Methods A549 cells were given thermotherapy alone (group A), chemotherapy alone (group B), and thermotherapy combined with chemotherapy (group C), thermotherapy combined with chemotherapy and SP600125 intervention (group D). Untreated A549 cells were selected as the control group. The changes of cell in-vasion, proliferation rate of the cells in each group were observed. Phosphorylated JNK and expression of heat shock protein 70 (HSP70) were detected by Western blot. Results A549 cell proliferation rate of group C was significantly lower than that of group A, group B and group D (P<0.05). The expression of group C was significantly higher than that of control group and group B (P<0.05), and the expression of HSP70 in group C was significantly lower than that in group A

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

    Directory of Open Access Journals (Sweden)

    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. 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

  2. 红霉素抑制A549细胞中鼻病毒14介导的白介素8和MUC5AC的分泌%Inhibition of erythromycin on rhinovirus-14-induced interleukin-8 and MUC5AC secretion in A549 cells

    Institute of Scientific and Technical Information of China (English)

    吴超民; 罗志兵; 沈策

    2009-01-01

    Objective To examine the effect of erythromycin on rhinovirus-14 (RV14)-induced cytokines and airway mucin hypersecretion. Methods A549 cells were pre-incubated with medium containing 10 μmol/L erythromycin for three hours before RV14 infection. Immunoreactive interleukin-8 (IL-8) and MUC5AC were quantitated using dual antibody ELISA kits according to the manufacturer's protocol. And activated p44/42MAPK was tested by Western blot. Results Erythromycin blocked the hypersecretion of IL-8 and MUC5AC protein, and the activation of p44/42MAPK induced by RV14 in the A549 cells.Conclusions Erythromyein may inhibit RV14-indueed IL-8 and MUC5AC hypersecretion by blocking the p44/42MAPK pathway or its upstream regulators.%目的 探讨红霉素是否对鼻病毒14(RV14)介导的细胞因子产生和气道黏液高分泌有抑制作用.方法 EM预处理肺泡Ⅱ型上皮细胞A549 3h后,用RV14刺激细胞,然后采用酶联免疫吸附试验法检测细胞培养上清中细胞因子白介素8和细胞裂解液中黏液蛋白MUC5AC的浓度,并用Western blot检测磷酸化p44/42MAPK信号分子变化情况.结果 红霉索明显抑制RV14介导的白介素8和MUC5AC的产生和分泌,并且对RV14介导的p44/42MAPK的激活也有抑制作用.结论 红霉素可有效抑制RV14介导的细胞因子和黏液蛋白的产生和分泌,并且这种抑制作用可能是通过阻断p44/42MAPK信号分子的激活来实现的.

  3. 香烟对大鼠肺及A549细胞IL-1,6,8表达影响的实验研究%Influence of cigarette smoke on the expression of IL-1, 6, 8 in rats' lungs and A549 cell

    Institute of Scientific and Technical Information of China (English)

    李文芳; 万丹; 刘福荣; 石梦蝶

    2012-01-01

    目的 研究香烟烟气对肺部炎性介质IL-1,IL-6,IL-8表达的影响.方法 (1)建立动物长期吸烟模型,用放射免疫法检测肺灌洗液(BALF)中IL-1,IL-6,IL-8的表达水平.(2)制备香烟烟气提取物(CSE)并用它染毒A549细胞,检测细胞上清液中IL-1,IL-6,Ⅱ-8的含量.结果 动物吸烟各剂量组IL-1表达水平与对照组相比差异无统计学意义(P>0.05),低、中、高剂量组IL-6,IL-8水平显著升高,差异有统计学意义(P<0.05).CSE染毒的A549细胞培养上清液中IL-1表达水平与对照组相比差异无统计学意义(P>0.05),20%、50%、100%CSE组IL-6的表达水平明显升高,IL-8表达水平下降,差异有统计学意义(P<0.05).结论 香烟对肺部炎性介质IL-6,IL-8的表达有影响.%OBJECTIVE To investigate the influence of cigarette smoke on the expression of inflammatory mediators in lungs. METHODS The animal model of smoking was established. Each group of rats was given smoking by the respective doses for 12 weeks. IL-1, IL-6 and IL-8 in bronchoalveolar lavage fluid (BALF) were detected by radiation immune method. Human lung adenocarcinoma A549 cells were cultured. Mainstream smoke was collected by using dimethyl sulfoxide (DMSO) as absorbent AS49 cells were incubated with different concentrations of CSE (cigarette smoke extract) . After 4 hours the level of 1L-1, IL-6 and IL-8 was measured with radioimmunoassay. RESULTS The expression of IL-1 in A549 cells with different concentrations of CSE had no difference with the control cells (P > 0.05). Compared with the control group, the IL-6 expressions of 20%, 50%, 100% concentrations increased (P 0.05). The levels of IL-6 and IL-8 in BALF increased significantly in high-dose group, middle-dose group and low-dose group respectively compared to that of Control group (P < 0.05). CONCLUSION Cigarette smoke could influence the expression of IL-6 and IL-8 in lungs.

  4. CpG-ODN 7909 increases radiation sensitivity of radiation-resistant human lung adenocarcinoma cell line by overexpression of Toll-like receptor 9.

    Science.gov (United States)

    Yan, Li; Xu, Guoxiong; Qiao, Tiankui; Chen, Wei; Yuan, Sujuan; Li, Xuan

    2013-09-01

    Radioresistance is one of the main reasons for the failure of radiotherapy in lung cancer. The aim of this study was to establish a radiation-resistant lung cancer cell line, to evaluate whether CpG oligodeoxyribonucleotide (CpG-ODN) 7909 could increase its radiosensitivity and to explore the relevant mechanisms. The radioresistant cell line, referred to as R-A549, was generated by reduplicative fractionated irradiation from the human lung adenocarcinoma cell line A549. The radioresistance of R-A549 cells were confirmed by the Cell Counting Kit-8 (CCK-8), cell viability assay, and clonogenic assay. Cell growth kinetics, morphological feature, and radiosensitivity were compared between the original A549 cells and R-A549 cells treated with or without CpG-ODN 7909 or radiation. To further explore the potential mechanisms of radiosensitivity, the cell cycle distributions and the expression of Toll-like receptor 9 (TLR-9) were examined by Western blot and flow cytometry. The R-A549 cell line was generated and its radioresistance was further confirmed. CpG-ODN 7909 was found to increase much more radiosensitivity of R-A549 cells under combined treatments with CpG-ODN 7909 and radiation compared with its control group without any treatments. They presented their respective D0 1.33 ± 0.20 Gy versus 1.76 ± 0.25 Gy with N 3.44 ± 1.01 versus 4.96 ± 0.32. Further, there was a larger cell population of R-A549 cells under combined treatment in the G2/M phase compared with the control group after treatment with CpG-ODN7909 or radiation alone at 24 and 48 hour. The expression level of TLR-9 in R-A549 cells was found higher than in A549 cells. These results suggested that CpG-ODN 7909 increased the radiosensitivity of R-A549 cells, which might be mediated via the upregulated TLR-9 and prolonged cell cycle arrest in the G2/M phase compared with A549 cells.

  5. EXPRESSION AND REVERSION OF DRUG RESISTANCE-AND APOPTOSIS-RELATED GENES OF A DDP-RESISTANT LUNG ADENOCARCINOMA CELL LINE

    Institute of Scientific and Technical Information of China (English)

    2000-01-01

    Objective: To investigate the co-expression of drug resistance- and apoptosis-related genes of cisplatin (CDDP)-selected lung adenocarcinoma cell line A549DDP for compared to the parental cell line A549, and reverse of drug resistance by antisense s-oligodeoxynucleotides (S-ODNs) of differentially expressed genes. Methods: Sense and antisense S-ODN were transferred into A549DDP cells by lipofectin. The expression of drug resistance and apoptosis related genes was examined by RT-PCR, immunocytochemistry and flow cytometry, respectively. Apoptostic cells were identified by DNA electrophoresis and terminal deoxynucleotidyl transferase (TdT)-mediated biotin dUTP nick end-labeling(TUNEL). Drug resistance of tumor cells was detected by a cell viability (MTT) assay. Results: The expression of bc1-2 was positive and that of multidrug resistance-associated protein (MRP) at mRNA and protein level was increased in A549DDP compared to A549 cells. MDR1, c-myc and topoisomeras II (TOPO II) were similarly co-expressed in two cell lines. Both cell lines were negative for c-erbB-2 expression. In A549DDP cells, the expression of bc1-2 and MRP was significantly inhibited by their respective antisense S-ODNs. Antisense S-ODNs could also decrease significantly drug resistance of A549DDP cells to CDDP by promoting cell apoptosis. Conclusion: Both intrinsic and acquired drug resistance were involved in co-expression of multiple MDR-related genes in lung adenocarcinoma. Co- operation of bc1-2 and MRP genes appeared to play an important action to confer the resistance of A549DDP cells to CDDP. Their antisense S-ODNs are responsible for the decrease of drug resistance of this cell line by promoting apoptosis.

  6. Synergistic Effect of Subtoxic-dose Cisplatin and TRAIL to Mediate Apoptosis by Down-regulating Decoy Receptor 2 and Up-regulating Caspase-8, Caspase-9 and Bax Expression on NCI-H460 and A549 Cells

    Directory of Open Access Journals (Sweden)

    Xiaoyan Zhang

    2013-05-01

    Full Text Available Objective(s: Although tumor necrosis factor-related apoptosis-inducing ligand (TRAIL can selectively induce apoptosis in tumor cells, more than half of tumors including non-small cell lung cancer (NSCLC exhibit TRAIL-resistance. The purpose of this study was to determine whether subtoxic-dose cisplatin and TRAIL could synergistically enhance apoptosis on NSCLC cells and investigate its underlying mechanisms. Materials and Methods:NCI-H460 and A549 cells were treated with TRAIL alone, cisplatin alone or combination treatment in this study. The cytotoxicity was evaluated according to Sulforhodamine B assay, and apoptosis was examined using Hoechst 33342 staining and flow cytometry. The mRNA and protein levels of TRAIL receptors and apoptotic proteins including caspase-8, caspase-9, Bcl-2 and Bax were determined by RT-PCR and Western blotting, respectively. Results:Our results showed that NCI-H460 cells were sensitive to TRAIL, whereas A549 cells were resistant. However, subtoxic-dose cisplatin could enhance the both cells to TRAIL-mediated cell proliferation inhibition and apoptosis. The underlying mechanisms might be associated with the down-regulation of DcR2 and up-regulation of Caspase-8, Caspase-9 and Bax. Conclusion:Subtoxic-dose cisplatin could enhance both TRAIL- sensitive and TRAIL- resistant NSCLC cells to TRAIL-mediated apoptosis. These findings motivated further studies to evaluate such a combinatory therapeutic strategy against NSCLC in the animal models.

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

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    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

  8. Pneumocystis carinii major surface glycoprotein induces interleukin-8 and monocyte chemoattractant protein-1 release from a human alveolar epithelial cell line

    DEFF Research Database (Denmark)

    Benfield, T L; Lundgren, Bettina; Shelhamer, J H;

    1999-01-01

    (IL-8) and monocyte chemoattractant protein-1 (MCP-1) from an alveolar epithelial cell line (A549). RESULTS: Incubation of A549 cells with MSG in concentrations from 0.4 to 10 microg mL-1 for 24 h caused dose-dependent increases in IL-8 release (3.4-fold above control, P ..., suggesting that MSG stimulates A549 cells in part through carbohydrate moieties. Dexamethasone significantly inhibited MSG-induced IL-8 release in concentrations of 10-6-10-8 mol L-1 compared with control experiments (P

  9. 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.

  10. 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.

  11. 下调miR-18a和miR-328抑制肺腺癌A549细胞的侵袭和迁移%Downregulation of miR-18a or miR-328 inhibits the invasion and migration of lung adenocarcinoma A549 cells

    Institute of Scientific and Technical Information of China (English)

    杜传冲; 郑金旭; 陆小威; 汪毅

    2016-01-01

    目的 分析miR-18a和miR-328在肺腺癌A549细胞中的表达,并探讨下调miR-18a或miR-328的表达对其侵袭和迁移能力的影响.方法 用荧光定量PCR检测A549细胞中miR-18a和miR-328的表达;通过转染miR-18a抑制物(miR-18ainhibitor)或miR-328 inhibitor,下调miR-18a或miR-328的表达,采用TranswellTM侵袭、迁移实验分析A549细胞侵袭和迁移能力的变化.结果 A549细胞中miR-18a和miR-328均呈高表达;而转染相应抑制物后miR-18a、miR-328表达下降,A549细胞的侵袭、迁移能力均明显降低.结论 下调A549细胞miR-18a或miR-328水平可有效抑制肿瘤细胞的侵袭、迁移.